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Self-assembly of alpha-repeat brick and staple proteins into macroscopic superhelices	A versatile strategy to create an inducible protein assembly with predefined geometry is demonstrated. The assembly is triggered by a binding protein that staples two identical protein bricks together in a predictable spatial conformation. The brick and staple proteins are designed for mutual directional affinity and engineered by directed evolution from a synthetic modular repeat protein library. As a proof of concept, this article reports on the spontaneous, extremely fast and quantitative self-assembly of two designed alpha-repeat (αRep) brick and staple proteins into macroscopic tubular superhelices at room temperature. Small-angle X-ray scattering (SAXS) and transmission electron microscopy (TEM with staining agent and cryoTEM) elucidate the resulting superhelical arrangement that precisely matches the a priori intended 3D assembly. The highly ordered, macroscopic biomolecular construction sustains temperatures as high as 75 °C thanks to the robust αRep building blocks. Since the α-helices of the brick and staple proteins are highly programmable, their design allows encoding the geometry and chemical surfaces of the final supramolecular protein architecture. This work opens routes toward the design and fabrication of multiscale protein origami with arbitrarily programmed shapes and chemical functions.	[ "In nature, assembled protein structures offer the most complex functional structures. The understanding of the mechanisms ruling protein-protein interactions opens the door to manipulate protein assemblies in a rational way. Proteins are versatile scaffolds with great potential as tools in nanotechnology and biomedicine because of their chemical, structural, and functional versatility. Currently, bottom-up self-assembly based on biomolecular interactions of small and well-defined components, is an attractive approach to biomolecular engineering and biomaterial design. Specifically, repeat proteins are simplified systems for this purpose. In this work, we provide an overview of fundamental concepts of the design of new protein interfaces. We describe an experimental approach to form higher order architectures by a bottom-up assembly of repeated building blocks. For this purpose, we use designed consensus tetratricopeptide repeat proteins (CTPRs). CTPR arrays contain multiple identical repeats that interact through a single inter-repeat interface to form elongated superhelices. Introducing a novel interface along the CTPR superhelix allows two CTPR molecules to assemble into protein nanotubes. We apply three approaches to form protein nanotubes: electrostatic interactions, hydrophobic interactions, and π-π interactions. We isolate and characterize the stability and shape of the formed dimers and analyze the nanotube formation considering the energy of the interaction and the structure in the three different models. These studies provide insights into the design of novel protein interfaces for the control of the assembly into more complex structures, which will open the door to the rational design of nanostructures and ordered materials for many potential applications in nanotechnology.", "The association of amphipathic α helices in water leads to α-helical-bundle protein structures. However, the driving force for this-the hydrophobic effect-is not specific and does not define the number or the orientation of helices in the associated state. Rather, this is achieved through deeper sequence-to-structure relationships, which are increasingly being discerned. For example, for one structurally extreme but nevertheless ubiquitous class of bundle-the α-helical coiled coils-relationships have been established that discriminate between all-parallel dimers, trimers, and tetramers. Association states above this are known, as are antiparallel and mixed arrangements of the helices. However, these alternative states are less well understood. Here, we describe a synthetic-peptide system that switches between parallel hexamers and various up-down-up-down tetramers in response to single-amino-acid changes and solution conditions. The main accessible states of each peptide variant are characterized fully in solution and, in most cases, to high resolution with X-ray crystal structures. Analysis and inspection of these structures helps rationalize the different states formed. This navigation of the structural landscape of α-helical coiled coils above the dimers and trimers that dominate in nature has allowed us to design rationally a well-defined and hyperstable antiparallel coiled-coil tetramer (apCC-Tet). This robust de novo protein provides another scaffold for further structural and functional designs in protein engineering and synthetic biology.", "Protein engineering has enabled the design of molecular scaffolds that display a wide variety of sizes, shapes, symmetries and subunit compositions. Symmetric protein-based nanoparticles that display multiple protein domains can exhibit enhanced functional properties due to increased avidity and improved solution behavior and stability. Here we describe the creation and characterization of a computationally designed circular tandem repeat protein (cTRP) composed of 24 identical repeated motifs, which can display a variety of functional protein domains (cargo) at defined positions around its periphery. We demonstrate that cTRP nanoparticles can self-assemble from smaller individual subunits, can be produced from prokaryotic and human expression platforms, can employ a variety of cargo attachment strategies and can be used for applications (such as T-cell culture and expansion) requiring high-avidity molecular interactions on the cell surface.", "Not all adaptive immune systems use the immunoglobulin fold as the basis for specific recognition molecules: sea lampreys, for example, have evolved an adaptive immune system that is based on leucine-rich repeat proteins. Additionally, many other proteins, not necessarily involved in adaptive immunity, mediate specific high-affinity interactions. Such alternatives to immunoglobulins represent attractive starting points for the design of novel binding molecules for research and clinical applications. Indeed, through progress and increased experience in library design and selection technologies, gained not least from working with synthetic antibody libraries, researchers have now exploited many of these novel scaffolds as tailor-made affinity reagents. Significant progress has been made not only in the basic science of generating specific binding molecules, but also in applications of the selected binders in laboratory procedures, proteomics, diagnostics and therapy. Challenges ahead include identifying applications where these novel proteins can not only be an alternative, but can enable approaches so far deemed technically impossible, and delineate those therapeutic applications commensurate with the molecular properties of the respective proteins.", "Ordered two-dimensional arrays such as S-layers1,2 and designed analogues3-5 have intrigued bioengineers6,7, but with the exception of a single lattice formed with flexible linkers8, they are constituted from just one protein component. Materials composed of two components have considerable potential advantages for modulating assembly dynamics and incorporating more complex functionality9-12. Here we describe a computational method to generate co-assembling binary layers by designing rigid interfaces between pairs of dihedral protein building blocks, and use it to design a p6m lattice. The designed array components are soluble at millimolar concentrations, but when combined at nanomolar concentrations, they rapidly assemble into nearly crystalline micrometre-scale arrays nearly identical to the computational design model in vitro and in cells without the need for a two-dimensional support. Because the material is designed from the ground up, the components can be readily functionalized and their symmetry reconfigured, enabling formation of ligand arrays with distinguishable surfaces, which we demonstrate can drive extensive receptor clustering, downstream protein recruitment and signalling. Using atomic force microscopy on supported bilayers and quantitative microscopy on living cells, we show that arrays assembled on membranes have component stoichiometry and structure similar to arrays formed in vitro, and that our material can therefore impose order onto fundamentally disordered substrates such as cell membranes. In contrast to previously characterized cell surface receptor binding assemblies such as antibodies and nanocages, which are rapidly endocytosed, we find that large arrays assembled at the cell surface suppress endocytosis in a tunable manner, with potential therapeutic relevance for extending receptor engagement and immune evasion. Our work provides a foundation for a synthetic cell biology in which multi-protein macroscale materials are designed to modulate cell responses and reshape synthetic and living systems.", "A major challenge of protein design is to create useful new proteins that interact specifically with biological targets in living cells. Such binding modules have many potential applications, including the targeted perturbation of protein networks. As a general approach to create such modules, we designed a library with approximately 10(9) different binding specificities based on a small 3-tetratricopeptide repeat (TPR) motif framework. We employed a novel strategy, based on split GFP reassembly, to screen the library for modules with the desired binding specificity. Using this approach, we identified modules that bind tightly and specifically to Dss1, a small human protein that interacts with the tumor suppressor protein BRCA2. We showed that these modules also bind the yeast homologue of Dss1, Sem1. Furthermore, we demonstrated that these modules inhibit Sem1 activity in yeast. This strategy will be generally applicable to make novel genetically encoded tools for systems/synthetic biology applications.", "Microtubules are cytoskeletal filaments of eukaryotic cells made of αβ-tubulin heterodimers. Structural studies of non-microtubular tubulin rely mainly on molecules that prevent its self-assembly and are used as crystallization chaperones. Here we identified artificial proteins from an αRep library that are specific to α-tubulin. Turbidity experiments indicate that these αReps impede microtubule assembly in a dose-dependent manner and total internal reflection fluorescence microscopy further shows that they specifically block growth at the microtubule (-) end. Structural data indicate that they do so by targeting the α-tubulin longitudinal surface. Interestingly, in one of the complexes studied, the α subunit is in a conformation that is intermediate between the ones most commonly observed in X-ray structures of tubulin and those seen in the microtubule, emphasizing the plasticity of tubulin. These α-tubulin-specific αReps broaden the range of tools available for the mechanistic study of microtubule dynamics and its regulation." ]
The Vestibulo-Ocular Reflex in Healthy Children	The video Head Impulse Test is routinely used to assess semicircular canal function in adults, but to date, pediatric reference values are scarce. This study aimed to explore the vestibulo-ocular reflex (VOR) in healthy children at different development stages and to compare the obtained gain values with reference to those in an adult population.	[ "Dysfunction of the vestibular organs has a great deal of influence on children's balance. Children with sensorineural hearing loss (SNHL) may often have accompanying abnormal vestibular responses. Video head impulse test (vHIT) combined with a test battery of cervical and ocular vestibular evoked myogenic potentials (cVEMPs, oVEMPs) make it possible to determine the prevalence of abnormal vestibular response in any of the five paired vestibular organs amongst children with SNHL. Prospective cross-sectional study including children aged 3-17 years with uni- or bilateral SNHL (bone-conduction (BC) pure tone average (PTA4) above 20 dB). Assessments included vHIT, cVEMP, and oVEMP. Descriptive data were analyzed. T-tests were performed to detect any correlation between the degree of SNHL and abnormal vestibular responses. 42 children (27 males), a total of 63 ears with SNHL, mean BC PTA4 SNHL of 42.3 dB (32.0; 58.4 IR) were included. 28.6% had at least one ear with abnormal vestibular response. 97.6% (41/42) completed one and 75.0% (27/36) completed all vestibular examinations. Ears with two pathological examinations had significantly more severe SNHL than ears with one pathological or normal vestibular examinations (p = 0.008, p = 0.005). All test methods used with this study were easy, fast, and comfortable for children (as young as 3 years of age) to undergo. Even a moderate SNHL might be associated with abnormal vestibular response. Structural damage to the organs is the most likely cause of vestibular deficit. Consensus on criteria defining pathological examinations with both vHIT and VEMPs are required for more accurate comparison with previous studies.", "In 1988, we introduced impulsive testing of semicircular canal (SCC) function measured with scleral search coils and showed that it could accurately and reliably detect impaired function even of a single lateral canal. Later we showed that it was also possible to test individual vertical canal function in peripheral and also in central vestibular disorders and proposed a physiological mechanism for why this might be so. For the next 20 years, between 1988 and 2008, impulsive testing of individual SCC function could only be accurately done by a few aficionados with the time and money to support scleral search-coil systems-an expensive, complicated and cumbersome, semi-invasive technique that never made the transition from the research lab to the dizzy clinic. Then, in 2009 and 2013, we introduced a video method of testing function of each of the six canals individually. Since 2009, the method has been taken up by most dizzy clinics around the world, with now close to 100 refereed articles in PubMed. In many dizzy clinics around the world, video Head Impulse Testing has supplanted caloric testing as the initial and in some cases the final test of choice in patients with suspected vestibular disorders. Here, we consider seven current, interesting, and controversial aspects of video Head Impulse Testing: (1) introduction to the test; (2) the progress from the head impulse protocol (HIMPs) to the new variant-suppression head impulse protocol (SHIMPs); (3) the physiological basis for head impulse testing; (4) practical aspects and potential pitfalls of video head impulse testing; (5) problems of vestibulo-ocular reflex gain calculations; (6) head impulse testing in central vestibular disorders; and (7) to stay right up-to-date-new clinical disease patterns emerging from video head impulse testing. With thanks and appreciation we dedicate this article to our friend, colleague, and mentor, Dr Bernard Cohen of Mount Sinai Medical School, New York, who since his first article 55 years ago on compensatory eye movements induced by vertical SCC stimulation has become one of the giants of the vestibular world." ]
Place of death among individuals with chronic respiratory disease in China	Chronic respiratory disease (CRD) is a common cause of mortality in China, but little is known about the place of death (POD) among individuals with CRD.	[ "This study examined the features of informal end-of-life care of older people living in the community and the association between informal care characteristics and dying at home. Retrospective data were obtained from interviews and self-administered questionnaires of 56 persons who had been primary caregivers of older relatives in the last three months of their lives. Results showed that informal caregivers of terminally ill older people living in the community provided a considerable amount of personal, household, and management care. Secondary informal caregivers and formal caregivers assisted resident primary caregivers less often than nonresident primary caregivers. Primary caregivers who felt less burdened, who gave personal care more intensively, and/or who were assisted by secondary caregivers, were more likely to provide informal end-of-life care at home until the time of death. Our study showed that informal care at the end of life of older people living in the community is complex, since the care required is considerable and highly varied, and involves assistance from secondary informal caregivers, formal home caregivers as well as institutional care. Burden of informal care is one of the most important factors associated with home death. More attention is needed to help ease the burden on informal caregivers, specifically with regard to resident caregivers and spouses. Since these resident caregivers were disadvantaged in several respects (i.e., health, income, assistance from other carers) compared to nonresident caregivers, interventions by formal caregivers should also be directed towards these persons, enabling them to bear the burden of end-of-life care.", "Several surveys in Japan have indicated that most terminally ill Japanese patients would prefer to die at home or in a homelike setting. However, there is a great disparity between this stated preference and the reality, since most Japanese die in hospital. We report here national changes in home deaths in Japan over the last 5 decades. Using prefecture data, we also examined the factors in the medical service associated with home death in Japan. Published data on place of death was obtained from the vital statistics compiled by the Ministry of Health, Labor and Welfare of Japan. We analyzed trends of home deaths from 1951 to 2002, and describe the changes in the proportion of home deaths by region, sex, age, and cause of death. Joinpoint regression analysis was used for trend analysis. Logistic regression analysis was performed to identify secular trends in home deaths, and the impact of age, sex, year of deaths and cause of deaths on home death. We also examined the association between home death and medical service factors by multiple regression analysis, using home death rate by prefectures in 2002 as a dependent variable. A significant decrease in the percentage of patients dying at home was observed in the results of joinpoint regression analysis. Older patients and males were more likely to die at home. Patients who died from cancer were less likely to die at home. The results of multiple regression analysis indicated that home death was related to the number of beds in hospital, ratio of daily occupied beds in general hospital, the number of families in which the elderly were living alone, and dwelling rooms. The pattern of the place of death has not only been determined by social and demographic characteristics of the decedent, but also associated with the medical service in the community." ]
Herb Diversity and Multifunctionality in Dryland Ecosystems	Considerable attention has been given to how different aspects of biodiversity sustain ecosystem functions. Herbs are a critical component of the plant community of dryland ecosystems, but the importance of different life form groups of herbs is often overlooked in experiments on biodiversity-ecosystem multifunctionality. Hence, little is known about how the multiple attributes of diversity of different life form groups of herbs affect changes to the multifunctionality of ecosystems.	[ "The biodiversity-ecosystem functioning concept asserts that processes in ecosystems are markedly influenced by species richness and other facets of biodiversity. However, biodiversity-ecosystem functioning studies have been largely restricted to single ecosystems, ignoring the importance of functional links - such as the exchange of matter, energy, and organisms - between coupled ecosystems. Here we present a basic concept and outline three pathways of cross-boundary biodiversity effects on ecosystem processes and propose an agenda to assess such effects, focusing on terrestrial-aquatic linkages to illustrate the case. This cross-boundary perspective of biodiversity-ecosystem functioning relationships presents a promising frontier for biodiversity and ecosystem science with repercussions for the conservation, restoration, and management of biodiversity and ecosystems from local to landscape scales.", "Accelerating rates of species extinction have prompted a growing number of researchers to manipulate the richness of various groups of organisms and examine how this aspect of diversity impacts ecological processes that control the functioning of ecosystems. We summarize the results of 44 experiments that have manipulated the richness of plants to examine how plant diversity affects the production of biomass. We show that mixtures of species produce an average of 1.7 times more biomass than species monocultures and are more productive than the average monoculture in 79% of all experiments. However, in only 12% of all experiments do diverse polycultures achieve greater biomass than their single most productive species. Previously, a positive net effect of diversity that is no greater than the most productive species has been interpreted as evidence for selection effects, which occur when diversity maximizes the chance that highly productive species will be included in and ultimately dominate the biomass of polycultures. Contrary to this, we show that although productive species do indeed contribute to diversity effects, these contributions are equaled or exceeded by species complementarity, where biomass is augmented by biological processes that involve multiple species. Importantly, both the net effect of diversity and the probability of polycultures being more productive than their most productive species increases through time, because the magnitude of complementarity increases as experiments are run longer. Our results suggest that experiments to date have, if anything, underestimated the impacts of species extinction on the productivity of ecosystems.", "Relationships between biodiversity and multiple ecosystem functions (that is, ecosystem multifunctionality) are context-dependent. Both plant and soil microbial diversity have been reported to regulate ecosystem multifunctionality, but how their relative importance varies along environmental gradients remains poorly understood. Here, we relate plant and microbial diversity to soil multifunctionality across 130 dryland sites along a 4,000 km aridity gradient in northern China. Our results show a strong positive association between plant species richness and soil multifunctionality in less arid regions, whereas microbial diversity, in particular of fungi, is positively associated with multifunctionality in more arid regions. This shift in the relationships between plant or microbial diversity and soil multifunctionality occur at an aridity level of ∼0.8, the boundary between semiarid and arid climates, which is predicted to advance geographically ∼28% by the end of the current century. Our study highlights that biodiversity loss of plants and soil microorganisms may have especially strong consequences under low and high aridity conditions, respectively, which calls for climate-specific biodiversity conservation strategies to mitigate the effects of aridification.", "Plants can respond to environmental impacts by variation in functional traits, thereby increasing their performance relative to neighbors. We hypothesized that trait adjustment should also occur in response to influences of the biotic environment, in particular different plant diversity of the community. We used 12 legume species as a model and assessed their variation in morphological, physiological, life-history and performance traits in experimental grasslands of different plant species (1, 2, 4, 8, 16 and 60) and functional group (1-4) numbers. Mean trait values and their variation in response to plant diversity varied among legume species and from trait to trait. The tall-growing Onobrychis viciifolia showed little trait variation in response to increasing plant diversity, whereas the species with shorter statures responded in apparently adaptive ways. The formation of longer shoots with elongated internodes, increased biomass allocation to supporting tissue at the cost of leaf mass, reduced branching, higher specific leaf areas and lower foliar δ(13)C values indicated increasing efforts for light acquisition in more diverse communities. Although leaf nitrogen concentrations and shoot biomass:nitrogen ratios were not affected by increasing plant diversity, foliar δ(15)N values of most legumes decreased and the application of the (15)N natural abundance method suggested that they became more reliant on symbiotic N(2) fixation. Some species formed fewer inflorescences and delayed flowering with increasing community diversity. The observed variation in functional traits generally indicated strategies of legumes to optimize light and nutrient capturing, but they were largely species-dependent and only partly attributable to increasing canopy height and community biomass with increasing plant diversity. Thus, the analysis of individual plant species and their adjustment to growth conditions in communities of increasing plant diversity is essential to get a deeper insight into the mechanisms behind biodiversity-ecosystem functioning relationships.", "Biodiversity loss decreases ecosystem functioning at the local scales at which species interact, but it remains unclear how biodiversity loss affects ecosystem functioning at the larger scales of space and time that are most relevant to biodiversity conservation and policy. Theory predicts that additional insurance effects of biodiversity on ecosystem functioning could emerge across time and space if species respond asynchronously to environmental variation and if species become increasingly dominant when and where they are most productive. Even if only a few dominant species maintain ecosystem functioning within a particular time and place, ecosystem functioning may be enhanced by many different species across many times and places (β-diversity). Here, we develop and apply a new approach to estimate these previously unquantified insurance effects of biodiversity on ecosystem functioning that arise due to species turnover across times and places. In a long-term (18-year) grassland plant diversity experiment, we find that total insurance effects are positive in sign and substantial in magnitude, amounting to 19% of the net biodiversity effect, mostly due to temporal insurance effects. Species loss can therefore reduce ecosystem functioning both locally and by eliminating species that would otherwise enhance ecosystem functioning across temporally fluctuating and spatially heterogeneous environments.", "Bringing together leaf trait data spanning 2,548 species and 175 sites we describe, for the first time at global scale, a universal spectrum of leaf economics consisting of key chemical, structural and physiological properties. The spectrum runs from quick to slow return on investments of nutrients and dry mass in leaves, and operates largely independently of growth form, plant functional type or biome. Categories along the spectrum would, in general, describe leaf economic variation at the global scale better than plant functional types, because functional types overlap substantially in their leaf traits. Overall, modulation of leaf traits and trait relationships by climate is surprisingly modest, although some striking and significant patterns can be seen. Reliable quantification of the leaf economics spectrum and its interaction with climate will prove valuable for modelling nutrient fluxes and vegetation boundaries under changing land-use and climate.", "Crop rotation schemes are believed to work by preventing specialist soil-borne pests from depressing the future yields of similar crops. In ecology, such negative plant-soil feedbacks may be viewed as a type of Janzen-Connell effect, which promotes species coexistence and diversity by preventing the same species from repeatedly occupying a particular site. In a controlled greenhouse experiment with 24 plant species and using soils from established field monocultures, we reveal community-wide soil-based Janzen-Connell effects between the three major functional groups of plants in temperate European grasslands. The effects are much stronger and more prevalent if plants are grown in interspecific competition. Using several soil treatments (gamma irradiation, activated carbon, fungicide, fertilizer) we show that the mechanism of the negative feedback is the buildup of soil pathogens which reduce the competitive ability of nearly all species when grown on soils they have formerly occupied. We further show that the magnitude of the change in competitive outcome is sufficient to stabilize observed fitness differences between functional groups in reasonably large communities. The generality and strength of this negative feedback suggests that Janzen-Connell effects have been underestimated as drivers of plant diversity in temperate ecosystems." ]
Fibrin gel as a cell carrier enhances cellular proliferation and production of extracellular matrix within trilayer polycaprolactone substrates for heart valve tissue engineering	Efficient cell seeding and subsequent support from a substrate ensure optimal cell growth and neotissue development during tissue engineering, including heart valve tissue engineering. Fibrin gel as a cell carrier may provide high cell seeding efficiency and adhesion property, improved cellular interaction, and structural support to enhance cellular growth in trilayer polycaprolactone (PCL) substrates that mimic the structure of native heart valve leaflets. This cell carrier gel coupled with a trilayer PCL substrate may enable the production of native-like cell-cultured leaflet constructs suitable for heart valve tissue engineering. In this study, we seeded valvular interstitial cells onto trilayer PCL substrates with fibrin gel as a cell carrier and cultured them for 1 month in vitro to determine if this gel can improve cell proliferation and production of extracellular matrix within the trilayer cell-cultured constructs. We observed that the fibrin gel enhanced cellular proliferation, their vimentin expression, and collagen and glycosaminoglycan production, leading to improved structure and mechanical properties of the developing PCL cell-cultured constructs. Fibrin gel as a cell carrier significantly improved the orientations of the cells and their produced tissue materials within trilayer PCL substrates that mimic the structure of native heart valve leaflets and, thus, may be highly beneficial for developing functional tissue-engineered leaflet constructs.	[ "It is estimated that the number of patients requiring heart valve replacement will triple over the next five decades. None of the current replacement valves can fully restore native valve function because they lack growth and remodeling capabilities. Heart valve tissue engineering is a promising technology to overcome these limitations. Various approaches are being employed, either aimed at development of the valve substitute in vitro or at the use of the regenerative potential of the body (in situ) for the tissue culture phase. This review provides an overview of the progress within both the in vitro and in situ tissue engineering approaches for trileaflet heart valve tissue engineering. Current challenges with these approaches are discussed, focusing in particular on the use of synthetic scaffold materials.", "Tissue-engineered heart valves are a promising alternative solution to prosthetic valves. However, long-term functionalities of tissue-engineered heart valves depend on the ability to mimic the trilayered, oriented structure of native heart valve leaflets. In this study, using electrospinning, we developed trilayered microfibrous leaflet substrates with morphological characteristics similar to native leaflets. The substrates were implanted subcutaneously in rats to study the effect of their trilayered oriented structure on in vivo tissue engineering. The tissue constructs showed a well-defined structure, with a circumferentially oriented layer, a randomly oriented layer and a radially oriented layer. The extracellular matrix, produced during in vivo tissue engineering, consisted of collagen, glycosaminoglycans, and elastin, all major components of native leaflets. Moreover, the anisotropic tensile properties of the constructs were sufficient to bear the valvular physiological load. Finally, the expression of vimentin and α-smooth muscle actin, at the gene and protein level, was detected in the residing cells, revealing their growing state and their transdifferentiation to myofibroblasts. Our data support a critical role for the trilayered structure and anisotropic properties in functional leaflet tissue constructs, and indicate that the leaflet substrates have the potential for the development of valve scaffolds for heart valve replacements.", "Heart valves are elaborate and highly heterogeneous structures of the circulatory system. Despite the well accepted relationship between the structural and mechanical anisotropy and the optimal function of the valves, most approaches to create tissue-engineered heart valves (TEHVs) do not try to mimic this complexity and rely on one homogenous combination of cells and materials for the whole construct. The aim of this study was to establish an easy and versatile method to introduce spatial diversity into a heart valve fibrin scaffold. We developed a multiple-step injection molding process that enables the fabrication of TEHVs with heterogeneous composition (cell/scaffold material) of wall and leaflets without the need of gluing or suturing components together, with the leaflets firmly connected to the wall. The integrity of the valves and their functionality was proved by either opening/closing cycles in a bioreactor (proof of principle without cells) or with continuous stimulation over 2 weeks. We demonstrated the potential of the method by the two-step molding of the wall and the leaflets containing different cell lines. Immunohistology after stimulation confirmed tissue formation and demonstrated the localization of the different cell types. Furthermore, we showed the proof of principle fabrication of valves using different materials for wall (fibrin) and leaflets (hybrid gel of fibrin/elastin-like recombinamer) and with layered leaflets. The method is easy to implement, does not require special facilities, and can be reproduced in any tissue-engineering lab. While it has been demonstrated here with fibrin, it can easily be extended to other hydrogels.", "Optimal cell migration rate in a given direction (velocity) is a function of speed and directional persistence. Migration speed has been reported to be a function of adhesion strength such that optimal cell migration occurs where the cell is able to form enough stable attachments for good traction while allowing attachments at the trailing end to be broken during locomotion. This is particularly important in peripheral nerve regeneration where rapid Schwann cell recruitment across the injury site will lead to better functional recovery and reduced end organ atrophy. The aim here was to investigate the effects of changing adhesion properties of Fn materials by adding fibrinogen in order to design an optimal material for repair processes. Cell migration on Fn/Fg-cables increased with increasing content of %Fg to a peak cell migration velocity (Schwann cells) of 49 microm/h, at 50% Fg. Further increases in Fg content hindered cell migration. Vinculin-rich attachment plaques were reduced in a dose-dependent manner as the content of %Fg was increased whilst cells at the optimum Fg proportion for cell migration were moderately well spread. These results support the idea that optimum cell migration rates occur at intermediate attachment conditions, in this case at 50% Fg. These results show that incorporation of Fg into Fn-based materials will enhance the speed of Schwann cell migration and this would be likely to improve peripheral nerve regeneration. Indeed, directionally aligned Fn-based materials can now be engineered to give optimal cell velocity during repair cell recruitment in a range of tissue repair or tissue engineering applications.", "A tissue-engineered heart valve can be an alternative to current mechanical or bioprosthetic valves that face limitations, especially in pediatric patients. However, it remains challenging to produce a functional tissue-engineered heart valve with three leaflets mimicking the trilayered, oriented structure of a native valve leaflet. In our previous study, a flat, trilayered nanofibrous substrate mimicking the orientations of three layers in a native leaflet-circumferential, random and radial orientations in fibrosa, spongiosa and ventricularis layers, respectively, was developed through electrospinning. In this study, we sought to develop a trilayered tissue structure mimicking the orientations of a native valve leaflet through in vivo tissue engineering, a practical regenerative medicine technology that can be used to develop an autologous heart valve. Thus, the nanofibrous substrate was placed inside the closed trileaflet-shaped cavity of a mold and implanted subcutaneously in a rat model for in vivo tissue engineering. After two months, the explanted tissue construct had a trilayered structure mimicking the orientations of a native valve leaflet. The infiltrated cells and their deposited collagen fibrils were oriented along the nanofibers in each layer of the substrate. Besides collagen, presence of glycosaminoglycans and elastin in the construct was observed.", "Heart valve leaflet substrates with native trilayer and anisotropic structures are crucial for successful heart valve tissue engineering. In this study, we used the electrospinning technique to produce trilayer microfibrous leaflet substrates using two biocompatible and biodegradable polymers-poly (L-lactic acid) (PLLA) and polycaprolactone (PCL), separately. Different polymer concentrations for each layer were applied to bring a high degree of mechanical and structural anisotropy to the substrates. PCL leaflet substrates exhibited lower unidirectional tensile properties than PLLA leaflet substrates. However, the PLLA substrates exhibited a lower flexural modulus than the PCL substrates. These substrates were seeded with porcine valvular interstitial cells (PVICs) and cultured for one month in static conditions. Both substrates exhibited cellular adhesion and proliferation, resulting in the production of tissue-engineered constructs. The PLLA tissue-engineered constructs had more cellular growth than the PCL tissue-engineered constructs. The PLLA substrates showed higher hydrophilicity, lower crystallinity, and more significant anisotropy than PCL substrates, which may have enhanced their interactions with PVICs. Analysis of gene expression showed higherα-smooth muscle actin and collagen type 1 expression in PLLA tissue-engineered constructs than in PCL tissue-engineered constructs. The differences in anisotropic and flexural properties may have accounted for the different cellular behaviors in these two individual polymer substrates.", "The fibrosa layer of a cardiac aortic valve is composed mostly of a dense network of type I collagen fibers oriented in circumferential direction. This main layer bears the tensile load and responds to the high stress on a leaflet. The inner fibrosa layer is also the site of pathophysiologic changes that result in valvular dysfunction, including stenosis and regurgitation. In vitro studies of these changes are limited by the absence of a substrate that mimics the circumferentially oriented structure of the fibrosa layer. In heart valve tissue engineering, generation of this layer is challenging. This study aimed to develop an artificial fibrosa layer of a native aortic leaflet. A unique morphologically biomimicked, pliable, but standalone substrate with circumferentially oriented nanofibers was fabricated by electrospinning on a novel collector designed for this study. The substrate had low-bulk tensile stiffness and ultimate strength; thus, cultured valvular interstitial cells (VICs) showed a fibroblast phenotype that is generally observed in a healthy aortic leaflet. Furthermore, gene and protein expression and morphology of VICs in substrates were close to those in the fibrosa layer of a native aortic leaflet. This artificial fibrosa layer can be useful for in vitro studies of valvular dysfunctions." ]
Health visiting services in the UK during the COVID-19 pandemic	Health visiting services, providing support to under 5s and their families, are organised and delivered in very different ways in different parts of the UK. While there has been attention to the key components of health visiting practice and what works well and how, there is little research on how health visiting services are organised and delivered and how that affects their ability to meet their objectives. The COVID-19 pandemic rapidly disrupted service delivery from March 2020. This realist review aims to synthesise the evidence on changes during the pandemic to identify the potential for improving health visiting services and their delivery.	[ "Clinicians often have to make treatment decisions based on the likelihood that an individual patient will benefit. In this article we consider the relevance of relative and absolute risk reductions, and draw attention to the importance of expressing the results of trials and subgroup analyses in terms of absolute risk. We describe the limitations of univariate subgroup analysis in situations in which there are several determinants of treatment effect, and review the potential for targeting treatments with risk models, especially when benefit is probably going to be dependent on the absolute risk of adverse outcomes with or without treatment. The ability to systematically take into account the characteristics of an individual patient and their interactions, to consider the risks and benefits of interventions separately if needed, and to provide patients with personalised estimates of their likelihood of benefit is shown using the example of endarterectomy for symptomatic carotid stenosis.", "Physicians today are confronted with increasing demand to ensure and improve care of their patients. A variety of approaches claim to provide solutions to the problems of health care delivery. These approaches represent different perspectives on optimal care and the best method for improving care. By summarizing recent reviews and debates in this field, this article critically reflects on the value of some of the approaches that have gained popularity during the last decades: evidence-based medicine and clinical practice guidelines, professional development, assessment and accountability, patient empowerment, and total quality management. Evidence regarding the impact and feasibility of the various approaches is mixed or simply lacking. In particular, the health care community lacks an understanding of which approaches are most appropriate for what types of improvement in what settings and of the determinants of successful performance change. Given the complexity of improvement and change in patient care, it is not realistic to expect that one approach can solve all the problems in health care delivery. None of the popular models for improving clinical performance appear to be superior. Therefore, bridges must be built and models must be integrated to be truly effective.", "Despite widespread agreement that stakeholder engagement is needed in patient-centered outcomes research (PCOR), no taxonomy exists to guide researchers and policy makers on how to address this need. We followed an iterative process, including several stages of stakeholder review, to address three questions: (1) Who are the stakeholders in PCOR? (2) What roles and responsibilities can stakeholders have in PCOR? (3) How can researchers start engaging stakeholders? We introduce a flexible taxonomy called the 7Ps of Stakeholder Engagement and Six Stages of Research for identifying stakeholders and developing engagement strategies across the full spectrum of research activities. The path toward engagement will not be uniform across every research program, but this taxonomy offers a common starting point and a flexible approach.", "Large pragmatic trials provide the most reliable data about the effects of treatments, but should be designed, analysed, and reported to enable the most effective use of treatments in routine practice. Subgroup analyses are important if there are potentially large differences between groups in the risk of a poor outcome with or without treatment, if there is potential heterogeneity of treatment effect in relation to pathophysiology, if there are practical questions about when to treat, or if there are doubts about benefit in specific groups, such as elderly people, which are leading to potentially inappropriate undertreatment. Analyses must be predefined, carefully justified, and limited to a few clinically important questions, and post-hoc observations should be treated with scepticism irrespective of their statistical significance. If important subgroup effects are anticipated, trials should either be powered to detect them reliably or pooled analyses of several trials should be undertaken. Formal rules for the planning, analysis, and reporting of subgroup analyses are proposed." ]
is peripheral ischemia a comorbidity	Peripheral ischemia caused by peripheral artery disease is associated with systemic inflammation, which may aggravate underlying comorbidities such as atherosclerosis and heart failure. However, the mechanisms of increased inflammation and inflammatory cell production in patients with peripheral artery disease remain poorly understood.	[ "Reconstruction of the molecular pathways controlling organ development has been hampered by a lack of methods to resolve embryonic progenitor cells. Here we describe a strategy to address this problem that combines gene expression profiling of large numbers of single cells with data analysis based on diffusion maps for dimensionality reduction and network synthesis from state transition graphs. Applying the approach to hematopoietic development in the mouse embryo, we map the progression of mesoderm toward blood using single-cell gene expression analysis of 3,934 cells with blood-forming potential captured at four time points between E7.0 and E8.5. Transitions between individual cellular states are then used as input to develop a single-cell network synthesis toolkit to generate a computationally executable transcriptional regulatory network model of blood development. Several model predictions concerning the roles of Sox and Hox factors are validated experimentally. Our results demonstrate that single-cell analysis of a developing organ coupled with computational approaches can reveal the transcriptional programs that underpin organogenesis.", "To determine whether the G(-174)C interleukin 6 (IL-6) polymorphism influences the development of peripheral arterial disease (PAD) in individuals with type 2 diabetes. This was investigated by comparing the distribution of G(-174)C genotypes between patients with type 2 diabetes and PAD (PAD+) and those with type 2 diabetes but without PAD (PAD-). Plasma concentrations of IL-6, fibrinogen, C reactive protein (CRP), and vascular endothelial growth factor (VEGF) were also compared in PAD+ and PAD- patients. Blood samples were collected from 146 PAD+ and 144 PAD- patients. SfaNI was used to determine the G(-174)C genotype. Plasma concentrations of IL-6, fibrinogen, CRP, and VEGF were measured by an enzyme linked immunosorbent assay. The GG genotype was more common in PAD+ patients than in PAD- patients. PAD+ patients also had increased mean plasma concentrations of IL-6, fibrinogen, CRP, and VEGF compared with PAD- patients. Mean plasma concentrations of IL-6, fibrinogen, and CRP in both PAD+ and PAD- patients were higher in those with the GG genotype than in those with the GC or CC genotypes. In contrast, mean plasma concentrations of VEGF in PAD+ and PAD- patients were not significantly different between those with different G(-174)C genotypes. These results support a model in which the GG genotype promotes PAD development among individuals with type 2 diabetes by inducing increased release of IL-6. Higher concentrations of IL-6 among those with the GG genotype is associated with increased plasma concentrations of fibrinogen and CRP.", "Maintenance of the blood system requires balanced cell fate decisions by hematopoietic stem and progenitor cells (HSPCs). Because cell fate choices are executed at the individual cell level, new single-cell profiling technologies offer exciting possibilities for mapping the dynamic molecular changes underlying HSPC differentiation. Here, we have used single-cell RNA sequencing to profile more than 1600 single HSPCs, and deep sequencing has enabled detection of an average of 6558 protein-coding genes per cell. Index sorting, in combination with broad sorting gates, allowed us to retrospectively assign cells to 12 commonly sorted HSPC phenotypes while also capturing intermediate cells typically excluded by conventional gating. We further show that independently generated single-cell data sets can be projected onto the single-cell resolution expression map to directly compare data from multiple groups and to build and refine new hypotheses. Reconstruction of differentiation trajectories reveals dynamic expression changes associated with early lymphoid, erythroid, and granulocyte-macrophage differentiation. The latter two trajectories were characterized by common upregulation of cell cycle and oxidative phosphorylation transcriptional programs. By using external spike-in controls, we estimate absolute messenger RNA (mRNA) levels per cell, showing for the first time that despite a general reduction in total mRNA, a subset of genes shows higher expression levels in immature stem cells consistent with active maintenance of the stem-cell state. Finally, we report the development of an intuitive Web interface as a new community resource to permit visualization of gene expression in HSPCs at single-cell resolution for any gene of choice.", "Experimental and clinical data suggest that reducing inflammation without affecting lipid levels may reduce the risk of cardiovascular disease. Yet, the inflammatory hypothesis of atherothrombosis has remained unproved. We conducted a randomized, double-blind trial of canakinumab, a therapeutic monoclonal antibody targeting interleukin-1β, involving 10,061 patients with previous myocardial infarction and a high-sensitivity C-reactive protein level of 2 mg or more per liter. The trial compared three doses of canakinumab (50 mg, 150 mg, and 300 mg, administered subcutaneously every 3 months) with placebo. The primary efficacy end point was nonfatal myocardial infarction, nonfatal stroke, or cardiovascular death. At 48 months, the median reduction from baseline in the high-sensitivity C-reactive protein level was 26 percentage points greater in the group that received the 50-mg dose of canakinumab, 37 percentage points greater in the 150-mg group, and 41 percentage points greater in the 300-mg group than in the placebo group. Canakinumab did not reduce lipid levels from baseline. At a median follow-up of 3.7 years, the incidence rate for the primary end point was 4.50 events per 100 person-years in the placebo group, 4.11 events per 100 person-years in the 50-mg group, 3.86 events per 100 person-years in the 150-mg group, and 3.90 events per 100 person-years in the 300-mg group. The hazard ratios as compared with placebo were as follows: in the 50-mg group, 0.93 (95% confidence interval [CI], 0.80 to 1.07; P=0.30); in the 150-mg group, 0.85 (95% CI, 0.74 to 0.98; P=0.021); and in the 300-mg group, 0.86 (95% CI, 0.75 to 0.99; P=0.031). The 150-mg dose, but not the other doses, met the prespecified multiplicity-adjusted threshold for statistical significance for the primary end point and the secondary end point that additionally included hospitalization for unstable angina that led to urgent revascularization (hazard ratio vs. placebo, 0.83; 95% CI, 0.73 to 0.95; P=0.005). Canakinumab was associated with a higher incidence of fatal infection than was placebo. There was no significant difference in all-cause mortality (hazard ratio for all canakinumab doses vs. placebo, 0.94; 95% CI, 0.83 to 1.06; P=0.31). Antiinflammatory therapy targeting the interleukin-1β innate immunity pathway with canakinumab at a dose of 150 mg every 3 months led to a significantly lower rate of recurrent cardiovascular events than placebo, independent of lipid-level lowering. (Funded by Novartis; CANTOS ClinicalTrials.gov number, NCT01327846 .).", "Splenic hematopoiesis is crucial to the pathogenesis of diseases including myocardial infarction and atherosclerosis. The spleen acts as a reservoir of myeloid cells, which are quickly expelled out in response to acute inflammation. In contrast to the well-defined bone marrow hematopoiesis, the cellular and molecular components sustaining splenic hematopoiesis are poorly understood. Surprisingly, we found that, unlike quiescent bone marrow hematopoietic stem cells (HSC), most of splenic HSC are in the G1 phase in C57BL/6 mice. Moreover, splenic HSC were enriched for genes involved in G0-G1 transition and expressed lower levels of genes responsible for G1-S transition. These data indicate that, at steady state, splenic HSC are pre-activated, which may expedite their cell cycle entry in emergency conditions. Consistently, in the acute phase of septic shock induced by LPS injection, splenic HSC entered the S-G2-M phase, whereas bone marrow HSC did not. Mobilization and transplantation experiments displayed that bone marrow HSC, once in the spleen, acquired cell cycle status similar to splenic HSC, strongly suggesting that the splenic microenvironment plays an important role in HSC pre-activation. In addition, we found that myeloid translocation gene 16 (Mtg16) deficiency in C57BL/6 mice resulted in significantly increased S-G2-M entry of splenic but not bone marrow HSC, suggesting that Mtg16 is an intrinsic negative regulator of G1-S transition in splenic HSC. Altogether, this study demonstrates that compared to bone marrow, splenic HSC are in a pre-activated state, which is driven by extracellular signals provided by splenic microenvironment and HSC intrinsic factor Mtg16.", "It has long been the dream of biologists to map gene expression at the single-cell level. With such data one might track heterogeneous cell sub-populations, and infer regulatory relationships between genes and pathways. Recently, RNA sequencing has achieved single-cell resolution. What is limiting is an effective way to routinely isolate and process large numbers of individual cells for quantitative in-depth sequencing. We have developed a high-throughput droplet-microfluidic approach for barcoding the RNA from thousands of individual cells for subsequent analysis by next-generation sequencing. The method shows a surprisingly low noise profile and is readily adaptable to other sequencing-based assays. We analyzed mouse embryonic stem cells, revealing in detail the population structure and the heterogeneous onset of differentiation after leukemia inhibitory factor (LIF) withdrawal. The reproducibility of these high-throughput single-cell data allowed us to deconstruct cell populations and infer gene expression relationships. VIDEO ABSTRACT.", "Stem cell biology is scientifically, clinically, and politically a current topic. The hematopoietic stem cell, the common ancestor of all types of blood cells, is one of the best-characterized stem cells in the body and the only stem cell that is clinically applied in the treatment of diseases such as breast cancer, leukemias, and congenital immunodeficiencies. Multicolor cell sorting enables the purification not only of hematopoietic stem cells, but also of their downstream progenitors such as common lymphoid progenitors and common myeloid progenitors. Recent genetic approaches including gene chip technology have been used to elucidate the gene expression profile of hematopoietic stem cells and other progenitors. Although the mechanisms that control self-renewal and lineage commitment of hematopoietic stem cells are still ambiguous, recent rapid advances in understanding the biological nature of hematopoietic stem and progenitor cells have broadened the potential application of these cells in the treatment of diseases." ]
Genetic architecture of cortical global expansion and regionalization	Human cortical expansion has occurred non-uniformly across the brain. We assessed the genetic architecture of cortical global expansion and regionalization by comparing two sets of genome-wide association studies of 24 cortical regions with and without adjustment for global measures (i.e., total surface area, mean cortical thickness) using a genetically informed parcellation in 32,488 adults. We found 393 and 756 significant loci with and without adjusting for globals, respectively, where 8% and 45% loci were associated with more than one region. Results from analyses without adjustment for globals recovered loci associated with global measures. Genetic factors that contribute to total surface area of the cortex particularly expand anterior/frontal regions, whereas those contributing to thicker cortex predominantly increase dorsal/frontal-parietal thickness. Interactome-based analyses revealed significant genetic overlap of global and dorsolateral prefrontal modules, enriched for neurodevelopmental and immune system pathways. Consideration of global measures is important in understanding the genetic variants underlying cortical morphology.	[ "Quantitative measurement of brain size, shape, and temporal change (for example, in order to estimate atrophy) is increasingly important in biomedical image analysis applications. New methods of structural analysis attempt to improve robustness, accuracy, and extent of automation. A fully automated method of longitudinal (temporal change) analysis, SIENA, was presented previously. In this paper, improvements to this method are described, and also an extension of SIENA to a new method for cross-sectional (single time point) analysis. The methods are fully automated, robust, and accurate: 0.15% brain volume change error (longitudinal): 0.5-1% brain volume accuracy for single-time point (cross-sectional). A particular advantage is the relative insensitivity to differences in scanning parameters. The methods provide easy manual review of their output by the automatic production of summary images which show the results of the brain extraction, registration, tissue segmentation, and final atrophy estimation.", "We report the dynamic anatomical sequence of human cortical gray matter development between the age of 4-21 years using quantitative four-dimensional maps and time-lapse sequences. Thirteen healthy children for whom anatomic brain MRI scans were obtained every 2 years, for 8-10 years, were studied. By using models of the cortical surface and sulcal landmarks and a statistical model for gray matter density, human cortical development could be visualized across the age range in a spatiotemporally detailed time-lapse sequence. The resulting time-lapse \"movies\" reveal that (i) higher-order association cortices mature only after lower-order somatosensory and visual cortices, the functions of which they integrate, are developed, and (ii) phylogenetically older brain areas mature earlier than newer ones. Direct comparison with normal cortical development may help understanding of some neurodevelopmental disorders such as childhood-onset schizophrenia or autism.", "Transcription factors provide the link between early membrane-proximal signalling events and changes in gene expression. NF-kappa B is one of the best-characterized transcription factors. It is expressed ubiquitously and regulates the expression of many genes, most of which encode proteins that play an important and often determining role in the processes of immunity and inflammation. Apart from its role in these events, evidence has begun to accumulate that NF-kappa B is involved in brain function, particularly following injury and in neurodegenerative conditions such as Alzheimer's disease. NF-kappa B might also be important for viral replication in the CNS. An involvement of NF-kappa B in neuronal development is suggested from studies that demonstrate its activation in neurones in certain regions of the brain during neurogenesis. Brain-specific activators of NF-kappa B include glutamate (via both AMPA/KA and NMDA receptors) and neurotrophins, pointing to an involvement in synaptic plasticity. NF-kappa B can therefore be considered as one of the most important transcription factors characterized in brain to date and it might be as crucial for neuronal and glial cell function as it is for immune cells.", "Regional brain morphology has a complex genetic architecture, consisting of many common polymorphisms with small individual effects. This has proven challenging for genome-wide association studies (GWAS). Due to the distributed nature of genetic signal across brain regions, multivariate analysis of regional measures may enhance discovery of genetic variants. Current multivariate approaches to GWAS are ill-suited for complex, large-scale data of this kind. Here, we introduce the Multivariate Omnibus Statistical Test (MOSTest), with an efficient computational design enabling rapid and reliable inference, and apply it to 171 regional brain morphology measures from 26,502 UK Biobank participants. At the conventional genome-wide significance threshold of α = 5 × 10-8, MOSTest identifies 347 genomic loci associated with regional brain morphology, more than any previous study, improving upon the discovery of established GWAS approaches more than threefold. Our findings implicate more than 5% of all protein-coding genes and provide evidence for gene sets involved in neuron development and differentiation.", "Data quality is increasingly recognized as one of the most important confounding factors in brain imaging research. It is particularly important for studies of brain development, where age is systematically related to in-scanner motion and data quality. Prior work has demonstrated that in-scanner head motion biases estimates of structural neuroimaging measures. However, objective measures of data quality are not available for most structural brain images. Here we sought to identify quantitative measures of data quality for T1-weighted volumes, describe how these measures relate to cortical thickness, and delineate how this in turn may bias inference regarding associations with age in youth. Three highly-trained raters provided manual ratings of 1840 raw T1-weighted volumes. These images included a training set of 1065 images from Philadelphia Neurodevelopmental Cohort (PNC), a test set of 533 images from the PNC, as well as an external test set of 242 adults acquired on a different scanner. Manual ratings were compared to automated quality measures provided by the Preprocessed Connectomes Project's Quality Assurance Protocol (QAP), as well as FreeSurfer's Euler number, which summarizes the topological complexity of the reconstructed cortical surface. Results revealed that the Euler number was consistently correlated with manual ratings across samples. Furthermore, the Euler number could be used to identify images scored \"unusable\" by human raters with a high degree of accuracy (AUC: 0.98-0.99), and out-performed proxy measures from functional timeseries acquired in the same scanning session. The Euler number also was significantly related to cortical thickness in a regionally heterogeneous pattern that was consistent across datasets and replicated prior results. Finally, data quality both inflated and obscured associations with age during adolescence. Taken together, these results indicate that reliable measures of data quality can be automatically derived from T1-weighted volumes, and that failing to control for data quality can systematically bias the results of studies of brain maturation.", "The folding of the human cerebral cortex is a highly genetically regulated process that allows for a much larger surface area to fit into the cranial vault and optimizes functional organization. Sulcal depth is a robust yet understudied measure of localized folding, previously associated with multiple neurodevelopmental disorders. Here, we report the first genome-wide association study of sulcal depth. Through the multivariate omnibus statistical test (MOSTest) applied to vertex-wise measures from 33,748 U.K. Biobank participants (mean age, 64.3 years; 52.0% female), we identified 856 genome-wide significant loci (P < 5 × 10−8). Comparisons with cortical thickness and surface area indicated that sulcal depth has higher locus yield, heritability, and effective sample size. There was a large amount of genetic overlap between these traits, with gene-based analyses indicating strong associations with neurodevelopmental processes. Our findings demonstrate sulcal depth is a promising neuroimaging phenotype that may enhance our understanding of cortical morphology.", "Functional magnetic resonance imaging (fMRI) is a noninvasive method for measuring brain function by correlating temporal changes in local cerebral blood oxygenation with behavioral measures. fMRI is used to study individuals at single time points, across multiple time points (with or without intervention), as well as to examine the variation of brain function across normal and ill populations. fMRI may be collected at multiple sites and then pooled into a single analysis. This paper describes how fMRI data is analyzed at each of these levels and describes the noise sources introduced at each level." ]
Generalization of Fear Memory in Posttraumatic Stress Disorder	Fear memory generalization is regarded as the core characteristic of posttraumatic stress disorder (PTSD) development. However, the mechanism that contributes to the generalization of conditioned fear memory is still unclear. The generalization is generally considered to be a mismatch that occurs during memory consolidation.	[ "Learning is accompanied by modulation of postsynaptic signal transduction pathways in neurons. Although the neuronal protein kinase cyclin-dependent kinase 5 (Cdk5) has been implicated in cognitive disorders, its role in learning has been obscured by the perinatal lethality of constitutive knockout mice. Here we report that conditional knockout of Cdk5 in the adult mouse brain improved performance in spatial learning tasks and enhanced hippocampal long-term potentiation and NMDA receptor (NMDAR)-mediated excitatory postsynaptic currents. Enhanced synaptic plasticity in Cdk5 knockout mice was attributed to reduced NR2B degradation, which caused elevations in total, surface and synaptic NR2B subunit levels and current through NR2B-containing NMDARs. Cdk5 facilitated the degradation of NR2B by directly interacting with both it and its protease, calpain. These findings reveal a previously unknown mechanism by which Cdk5 facilitates calpain-mediated proteolysis of NR2B and may control synaptic plasticity and learning.", "A hippocampal pyramidal neuron receives more than 10(4) excitatory glutamatergic synapses. Many of these synapses contain the molecular machinery for messenger RNA translation, suggesting that the protein complement (and thus function) of each synapse can be regulated on the basis of activity. Here, local postsynaptic protein synthesis, triggered by synaptic activation of metabotropic glutamate receptors, was found to modify synaptic transmission within minutes.", "Activation of the N-methyl-D-aspartate receptor (NMDAR) glycine site has been shown to accelerate adaptive forms of learning that may benefit psychopathologies involving cognitive and perseverative disturbances. In this study, the effects of increasing the brain levels of the endogenous NMDAR glycine site agonist D-serine, through the genetic inactivation of its catabolic enzyme D-amino acid oxidase (DAO), were examined in behavioral tests of learning and memory. In the Morris water maze task (MWM), mice carrying the hypofunctional Dao1(G181R) mutation demonstrated normal acquisition of a single platform location but had substantially improved memory for a new target location in the subsequent reversal phase. Furthermore, Dao1(G181R) mutant animals exhibited an increased rate of extinction in the MWM that was similarly observed following pharmacological administration of D-serine (600 mg/kg) in wild-type C57BL/6J mice. In contextual and cued fear conditioning, no alterations were found in initial associative memory recall; however, extinction of the contextual fear memory was facilitated in mutant animals. Thus, an augmented level of D-serine resulting from reduced DAO activity promotes adaptive learning in response to changing conditions. The NMDAR glycine site and DAO may be promising therapeutic targets to improve cognitive flexibility and inhibitory learning in psychiatric disorders such as schizophrenia and anxiety syndromes.", "Previous results indicate that intra-amygdala infusions of NMDA receptor antagonists block the extinction of conditioned fear. Mitogen-activated protein kinase (MAPK) can be activated by NMDA receptor stimulation and is involved in excitatory fear conditioning. Here, we evaluate the role of MAPK within the basolateral amygdala in the extinction of conditioned fear. Rats received 10 light-shock pairings. After 24 hr, fear was assessed by eliciting the acoustic startle reflex in the presence of the conditioned stimulus (CS) (CS-noise trials) and also in its absence (noise-alone trials). Rats subsequently received an intra-amygdala or intrahippocampal infusion of either 20% DMSO or the MAPK inhibitor PD98059 (500 ng/side) followed 10 min later by 30 presentations of the light CS without shock (extinction training). After 24 hr, they were again tested for fear-potentiated startle. PD98059 infusions into the basolateral amygdala but not the hippocampus significantly reduced extinction, which was otherwise evident in DMSO-infused rats. Control experiments indicated that the effect of intra-amygdala PD98059 could not be attributed to lasting damage to the amygdala or to state dependency. These results suggest that a MAPK-dependent signaling cascade within or very near the basolateral amygdala plays an important role in the extinction of conditioned fear.", "The mechanisms that contribute to the extinction of previously acquired memories are not well understood. These processes, often referred to as inhibitory learning, are thought to be parallel learning mechanisms that require a reacquisition of new information and suppression of previously acquired experiences in order to adapt to novel situations. Using newly generated metabotropic glutamate receptor 5 (mGluR5) knock-out mice, we investigated the role of mGluR5 in the acquisition and reversal of an associative conditioned task and a spatial reference task. We found that acquisition of fear conditioning is partially impaired in mice lacking mGluR5. More markedly, we found that extinction of both contextual and auditory fear was completely abolished in mGluR5 knock-out mice. In the Morris Water Maze test (MWM), mGluR5 knock-out mice exhibited mild deficits in the rate of acquisition of the regular water maze task, but again had significant deficits in the reversal task, despite overall spatial memory being intact. Together, these results demonstrate that mGluR5 is critical to the function of neural circuits that are required for inhibitory learning mechanisms, and suggest that targeting metabotropic receptors may be useful in treating psychiatric disorders in which aversive memories are inappropriately retained.", "A substantial and growing body of evidence from cognitive neuroscience supports the concept of multiple memory systems (MMS). However, the existence of multiple systems has been questioned by theorists who instead propose that dissociations can be accounted for within a single memory system. We present convergent evidence from neuroimaging and neuropsychological studies of category learning in favor of the existence of MMS for category learning and declarative knowledge. Whereas single-system theorists have argued that their approach is more parsimonious because it only postulates a single form of memory representation, we show that the MMS approach is superior in its ability to account for a broad range of data from psychology and neuroscience." ]
Predictive Model of Necrotizing Apoptosis-Related Genes in Lower Grade Gliomas	The purpose of this research is to develop a predictive model based on necroptosis-related genes to predict the prognosis and survival of lower grade gliomas (LGGs) efficiently. To achieve this goal, we searched for differentially expressed necrotizing apoptosis-related genes using the TCGA and CGGA databases. To construct a prognostic model, LASSO Cox and COX regression analyses were conducted on the differentially expressed genes. In this study, three genes were used to develop a prognostic model of necrotizing apoptosis, and all samples were split into high- and low-risk groups. We observed that patients with a high-risk score had a worse overall survival rate (OS) than those with a low-risk score. In the TCGA and CGGA cohorts, the nomogram plot showed a high capacity to predict overall survival of LGG patients. GSEA analysis revealed that the high-risk group was enriched for inflammatory responses, tumor-related pathways, and pathological processes. Additionally, the high-risk score was associated with invading immune cell expression. In conclusion, our predictive model based on necroptosis-related genes in LGG was shown to be effective in the diagnosis and could predict the prognosis of LGG. In addition, we identified possible targets related to necroptosis-related genes for glioma therapy in this study.	[ "Over the last decade, our picture of cell death signals involved in experimental disease models totally shifted. Indeed, in addition to apoptosis, multiple forms of regulated necrosis have been associated with an increasing number of pathologies such as ischemia-reperfusion injury in brain, heart and kidney, inflammatory diseases, sepsis, retinal disorders, neurodegenerative diseases and infectious disorders. Especially necroptosis is currently attracting the attention of the scientific community. However, the in vivo identification of ongoing necroptosis in experimental disease conditions remains troublesome, mainly due to the lack of specific biomarkers. Initially, Receptor-Interacting Protein Kinase 1 (RIPK1) and RIPK3 kinase activity were uniquely associated with induction of necroptosis, however recent evidence suggests pleiotropic functions in cell death, inflammation and survival, obscuring a clear picture. In this review, we will present the last methodological advances for in vivo necroptosis identification and discuss past and recent data to provide an update of the so-called \"necroptosis-associated pathologies\".", "Expression of the DNA repair protein O6-alkylguanine-DNA-alkyltransferase (AGT), encoded by the O6-methylguanine (O6-mG) -DNA-methyltransferase (MGMT) DNA repair gene, results in resistance to alkylating agents, and hypermethylation of the MGMT promoter is associated with chemosensitivity as it prevents AGT expression. As the interpretation of the results of immunohistochemistry to evaluate AGT expression proved to be difficult, the aim of our present study is to establish a feasible, reliable, and robust method for MGMT promoter hypermethylation testing that can be easily implemented in a diagnostic setting and is applicable to routinely processed tissue. MGMT hypermethylation analysis using methylation-specific (MS-) multiplex ligation-dependent probe amplification (MLPA) was performed on 62 glioma samples of 55 individual tumors (including 12 cell lines) and compared to the more conventionally used, but improved, MS-polymerase chain reaction (PCR). In contrast to MS-PCR, MS-MLPA (i) is not based on bisulfite conversion of unmethylated cytosines (a somewhat troublesome step in MS-PCR), (ii) provided methylation status of all samples, (iii) proved to be semiquantitative, (iv) can be used to evaluate methylation status of multiple sequences (CpG dinucleotides) simultaneously, and (v) allows for a combined copy number detection and methylation specific analysis. The potential therapeutic value of MGMT hypermethylation evaluation using MS-MLPA was shown in a group of 20 glioblastoma patients receiving temozolomide chemotherapy. We conclude that MS-MLPA is a robust and reliable method that can be easily applied to differently processed tissues, including those fixed in formalin and embedded in paraffin. The semiquantitative aspect of MS-MLPA may prove to be of great value, especially in predicting response to alkylating agents, not only for gliomas as evaluated in this study but also for tumors in general.", "IAPs were named as inhibitors of apoptosis, programmed cell death, but it has become apparent that they are regulators of other types of cell death too. Because they inhibit cell death in cancer cells there has been an intense interest in developing inhibitors of these proteins to induce or sensitise cancer cells to death. In this article, we will discuss the involvement of IAPs in the apoptosis, necroptosis and pyroptosis programmed cell death paradigms. All these types of cell death are intimately involved with causing or repressing inflammation and it should perhaps therefore come as no surprise that IAPs are also involved in regulating inflammation directly. To come full circle, the IAP antagonist drugs that were developed to sensitise cancer cells to apoptosis have led to some of these insights.", "Cell death research was revitalized by the understanding that necrosis can occur in a highly regulated and genetically controlled manner. Although RIPK1 (receptor-interacting protein kinase 1)- and RIPK3-MLKL (mixed lineage kinase domain-like)-mediated necroptosis is the most understood form of regulated necrosis, other examples of this process are emerging, including cell death mechanisms known as parthanatos, oxytosis, ferroptosis, NETosis, pyronecrosis and pyroptosis. Elucidating how these pathways of regulated necrosis are interconnected at the molecular level should enable this process to be therapeutically targeted.", "Necroptosis is a tightly regulated form of necrosis that requires the activation of receptor-interacting protein (RIP) kinases RIPK1 and RIPK3, as well as the RIPK3 substrate mixed lineage kinase domain-like protein (MLKL). Because of membrane rupture, necroptotic cells release damage-associated molecular patterns (DAMPs) that evoke immune responses. Necroptosis is emerging as an important cellular response in the modulation of cancer initiation, progression, and metastasis. Necroptosis of cancer cells is considered to be an immunogenic cell death capable of activating anti-tumor immunity. Necroptosis also participates in the promotion of myeloid cell-induced adaptive immune suppression and thus contributes to oncogenesis. In addition, necroptosis of endothelial cells and tumor cells is conducive to tumor metastasis. In this review, we summarize the current knowledge of the complex role of necroptosis in cancer and discuss the potential of targeting necroptosis components for cancer therapies.", "O(6)-methylguanine-DNA methyltransferase (MGMT) is a repair enzyme that removes promutagenic O(6)-methylguanine adducts in DNA, to protect cells from acquisition of G:C--> A:T mutations. MGMT promoter methylation and polymorphisms may affect MGMT expression and activity. In the present study, we assessed MGMT promoter methylation and polymorphisms (Leu84Phe, Ile143Val, c.-56C>T) in 371 glioblastomas diagnosed at the population level. MGMT methylation was observed in 165 (44%) glioblastomas, with a higher frequency in females than males (53 vs. 39%; p = 0.0106) and in secondary than primary glioblastomas (73 vs. 43%; p = 0.0074). The frequency of TP53 G:C-->A:T mutations in glioblastomas with MGMT methylation was 25%, which was significantly higher than that in glioblastomas with MGMT methylation (16%; Fisher exact test; p = 0.0385). MGMT 143 Val allele in glioblastomas was significantly less frequent than in a healthy European Caucasian population, and was associated with longer survival than those with the MGMT 143 Ile allele (hazard ratio 0.70; 95% CI 0.48-1.01). These results suggest that MGMT methylation may be associated with susceptibility to acquire TP53 G:C-->A:T mutations, and that MGMT polymorphisms may affect the risk and prognosis of glioblastomas.", "O6-methylguanine-methyltransferase (MGMT) promoter methylation has been shown to predict survival of patients with glioblastomas if temozolomide is added to radiotherapy (RT). It is unknown if MGMT promoter methylation is also predictive to outcome to RT followed by adjuvant procarbazine, lomustine, and vincristine (PCV) chemotherapy in patients with anaplastic oligodendroglial tumors (AOT). In the European Organisation for the Research and Treatment of Cancer study 26951, 368 patients with AOT were randomly assigned to either RT alone or to RT followed by adjuvant PCV. From 165 patients of this study, formalin-fixed, paraffin-embedded tumor tissue was available for MGMT promoter methylation analysis. This was investigated with methylation specific multiplex ligation-dependent probe amplification. In 152 cases, an MGMT result was obtained, in 121 (80%) cases MGMT promoter methylation was observed. Methylation strongly correlated with combined loss of chromosome 1p and 19q loss (P = .00043). In multivariate analysis, MGMT promoter methylation, 1p/19q codeletion, tumor necrosis, and extent of resection were independent prognostic factors. The prognostic significance of MGMT promoter methylation was equally strong in the RT arm and the RT/PCV arm for both progression-free survival and overall survival. In tumors diagnosed at central pathology review as glioblastoma, no prognostic effect of MGMT promoter methylation was observed. In this study, on patients with AOT MGMT promoter methylation was of prognostic significance and did not have predictive significance for outcome to adjuvant PCV chemotherapy. The biologic effect of MGMT promoter methylation or pathogenetic features associated with MGMT promoter methylation may be different for AOT compared with glioblastoma.", "Necroptosis is a form of regulated cell death, which is induced by ligand binding to TNF family death domain receptors, pattern recognizing receptors and virus sensors. The common feature of these receptor systems is the implication of proteins, which contain a receptor interaction protein kinase (RIPK) homology interaction motif (RHIM) mediating recruitment and activation of receptor-interacting protein kinase 3 (RIPK3), which ultimately activates the necroptosis executioner mixed lineage kinase domain-like (MLKL). In case of the TNF family members, the initiator is the survival- and cell death-regulating RIPK1 kinase, in the case of Toll-like receptor 3/4 (TLR3/4), a RHIM-containing adaptor, called TRIF, while in the case of Z-DNA-binding protein ZBP1/DAI, the cytosolic viral sensor itself contains a RHIM domain. In this review, we discuss the different protein complexes that serve as nucleation platforms for necroptosis and the mechanism of execution of necroptosis. Transgenic models (knockout, kinase-dead knock-in) and pharmacologic inhibition indicate that RIPK1, RIPK3 or MLKL are implicated in many inflammatory, degenerative and infectious diseases. However, the conclusion of necroptosis being solely involved in the etiology of diseases is blurred by the pleiotropic roles of RIPK1 and RIPK3 in other cellular processes such as apoptosis and inflammasome activation.", "Programmed necrosis or necroptosis is controlled by the action of two serine/threonine kinases, RIP1 (receptor-interacting serine/threonine protein kinase 1; also known as RIPK1) and RIP3. The phosphorylation of RIP1 and RIP3 is critical for assembly of the necrosome, an amyloid-like complex that initiates transmission of the pro-necrotic signal. In the present study, we used site-directed mutagenesis to systematically examine the effects of putative phosphoacceptor sites on RIP1 and RIP3 on TNF (tumour necrosis factor)-induced programmed necrosis. We found that mutation of individual serine residues in the kinase domain of RIP1 had little effect on RIP1 kinase activity and TNF-induced programmed necrosis. Surprisingly, an alanine residue substitution for Ser(89) enhanced RIP1 kinase activity and TNF-induced programmed necrosis without affecting RIP1-RIP3 necrosome formation. This indicates that Ser(89) is an inhibitory phosphoacceptor site that can dampen the pro-necrotic function of RIP1. In addition, we show that a phosphomimetic mutant of RIP3, S204D, led to programmed necrosis that was refractory to RIP1 siRNA and insensitive to necrostatin-1 inhibition. Our results show that programmed necrosis is regulated by positive and inhibitory phosphorylation events.", "Engagement of tumor necrosis factor receptor 1 signals two diametrically opposed pathways: survival-inflammation and cell death. An additional switch decides, depending on the cellular context, between caspase-dependent apoptosis and RIP kinase (RIPK)-mediated necrosis, also termed necroptosis. We explored the contribution of both cell death pathways in TNF-induced systemic inflammatory response syndrome (SIRS). Deletion of apoptotic executioner caspases (caspase-3 or -7) or inflammatory caspase-1 had no impact on lethal SIRS. However, deletion of RIPK3 conferred complete protection against lethal SIRS and reduced the amounts of circulating damage-associated molecular patterns. Pretreatment with the RIPK1 kinase inhibitor, necrostatin-1, provided a similar effect. These results suggest that RIPK1-RIPK3-mediated cellular damage by necrosis drives mortality during TNF-induced SIRS. RIPK3 deficiency also protected against cecal ligation and puncture, underscoring the clinical relevance of RIPK kinase inhibition in sepsis and identifying components of the necroptotic pathway that are potential therapeutic targets for treatment of SIRS and sepsis." ]
Eating disorder care clinicians' perceptions of facilitators of and barriers to effective eating disorder treatment for transgender and gender diverse patients	The prevalence of eating disorders is higher in transgender and non-binary compared to cisgender people. Gender diverse people who seek eating disorder treatment often report struggling to find affirming and inclusive treatment from healthcare clinicians. We sought to understand eating disorder care clinicians' perceptions of facilitators of and barriers to effective eating disorder treatment for transgender and gender diverse patients.	[ "We estimated the prevalence of diagnosed eating disorders, overall and by select demographics, among commercially insured individuals identified as transgender in a national claims database. From the 2018 IBM® MarketScan® Commercial Database, there were 10,415 people identifiable as transgender based on International Classification of Disease (ICD-10) codes and procedure codes, specific to gender-affirming care, from inpatient and outpatient claims. Eating disorders were identified from ICD-10 codes and included anorexia nervosa, bulimia nervosa, binge eating disorder, eating disorder not otherwise specified, avoidant restrictive feeding and intake disorder, and other specified feeding and eating disorders. We estimated the prevalence of specific eating disorders diagnoses by selecting patient characteristics. Of individuals receiving some form of gender-affirming care, 2.43% (95% confidence interval: 2.14%-2.74%) were diagnosed with an eating disorder: 0.84% anorexia nervosa, 0.36% bulimia nervosa, 0.36% binge eating disorder, 0.15% avoidant restrictive feeding and intake disorder, 0.41% other specified feeding and eating disorders, and 1.37% with an unspecified eating disorder. Among transgender-identifiable patients aged 12-15 years, 5.60% had an eating disorder diagnosis, whereas 0.52% had an eating disorder diagnosis in patients aged 45-64 years. In patients identifiable as transgender, with receipt of gender-affirming care, the prevalence of diagnosed eating disorders was low compared to extant self-reported data for eating disorder diagnosis in transgender individuals. Among this population, eating disorders were highest in adolescents and young adults. Clinically verified prevalence estimates for eating disorder diagnosis in transgender people with a history of gender-affirming care warrant further investigation. The present study aims to provide clinically validated, contemporary prevalence estimates for diagnosed eating disorders among a medically affirmed population of transgender adults and children in the United States. We report low prevalence of having any eating disorder relative to prevalence estimates reported in prior literature without clinical validation. These findings may be explained by access to affirming care and medical care generally.", "Purpose: To investigate whether the prevalence of eating disorders (EDs) differs across diverse gender identity groups in a transgender sample. Methods: Secondary analysis of data from Project VOICE, a cross-sectional study of stress and health among 452 transgender adults (ages 18-75 years) residing in Massachusetts. Age-adjusted logistic regression models were fit to compare the prevalence of self-reported lifetime EDs in female-to-male (FTM), male-to-female (MTF), and gender-nonconforming participants assigned male at birth (MBGNC) to gender-nonconforming participants assigned female at birth (FBGNC; referent). Results: The age-adjusted odds of self-reported ED in MTF participants were 0.14 times the odds of self-reported ED in FBGNC participants (p=0.022). In FTM participants, the age-adjusted odds of self-reported ED were 0.46 times the odds of self-reported ED in FBGNC participants, a marginally significant finding (p=0.068). No statistically significant differences in ED prevalence were found for MBGNC individuals. Conclusions: Gender nonconforming individuals assigned a female sex at birth appear to have heightened lifetime risk of EDs relative to MTF participants. Further research into specific biologic and psychosocial ED risk factors and gender-responsive intervention strategies are urgently needed. Training clinical providers and ensuring competency of treatment services beyond the gender binary will be vital to addressing this disparity.", "The present review addresses the outcome of anorexia nervosa and whether it changed over the second half of the 20th century. A total of 119 study series covering 5,590 patients suffering from anorexia nervosa that were published in the English and German literature were analyzed with regard to mortality, global outcome, and other psychiatric disorders at follow-up. There were large variations in the outcome parameters across studies. Mortality estimated on the basis of both crude and standardized rates was significantly high. Among the surviving patients, less than one-half recovered on average, whereas one-third improved, and 20% remained chronically ill. The normalization of the core symptoms, involving weight, menstruation, and eating behaviors, was slightly better when each symptom was analyzed in isolation. The presence of other psychiatric disorders at follow-up was very common. Longer duration of follow-up and, less strongly, younger age at onset of illness were associated with better outcome. There was no convincing evidence that the outcome of anorexia nervosa improved over the second half of the last century. Several prognostic features were isolated, but there is conflicting evidence. Most clearly, vomiting, bulimia, and purgative abuse, chronicity of illness, and obsessive-compulsive personality symptoms are unfavorable prognostic features. Anorexia nervosa did not lose its relatively poor prognosis in the 20th century. Advances in etiology and treatment may improve the course of patients with anorexia nervosa in the future.", "Transgender men and women may be at risk for eating disorders, but prior community norms of the Eating Disorders Examination Questionnaire (EDE-Q) are based on presumed cisgender men and woman and have not intentionally included transgender people. The objective of this study was to develop community norms for eating disorder attitudes and disordered eating behaviors in transgender men and women using the EDE-Q. Participants were 312 transgender men and 172 transgender women participants in The PRIDE Study, an existing cohort study of sexual and gender minority people. We present mean scores, standard deviations, and percentile ranks for the Global score and four subscale scores of the EDE-Q in transgender men and women. Transgender men and women reported any occurrence (≥1/week) of dietary restraint (25.0% and 27.9%), objective binge episodes (11.2% and 12.8%), excessive exercise (8.0% and 8.1%), self-induced vomiting (1.6% and 1.7%), and laxative misuse (.3% and .6%), respectively. Compared to a prior study of presumed cisgender men 18-26 years (Lavender, De Young, & Anderson, 2010), our age-matched subsample of transgender men reported lower rates of objective binge episodes and excessive exercise. Compared to a prior study of presumed cisgender women 18-42 years (Mond, Hay, Rodgers, & Owen, 2006), we found that an age-matched sample of transgender women reported higher rates of dietary restraint but lower rates of excessive exercise. These norms should aid clinicians in applying and researchers in investigating and interpreting the EDE-Q scores of transgender men and women.", "Minority stress theory posits that transgender and gender diverse (TGD) individuals exhibit greater rates of depression and suicidality due to internal (proximal) and external (distal) stressors related to their TGD identity. Yet, the magnitude of the relationship between minority stress processes and mental health outcomes has not been quantitatively summarized. The current research meta-analyzed the relationship between minority stress constructs and depression, suicidal ideation, and suicide attempt. Results from 85 cross-sectional quantitative studies indicate that distal stress, expectations of rejection, internalized transphobia, and concealment are significantly associated with increased depression, suicidal ideation, and suicide attempt. Greater effect sizes were observed for expectations of rejection and internalized transphobia when compared to distal stress and concealment. Future research on the relationship between minority stress, depression, and suicide would benefit from longitudinal designs and more diverse and representative samples of TGD individuals.", "This paper discusses the findings of qualitative research that examined the accounts of five \"mostly recovered\" ex-patients who had experienced transition between two or more eating disorder diagnoses. This study found that, in the minds of participants, the different diagnostic labels were associated with various good or bad character traits. This contributed to the belief in a diagnostic hierarchy, whereby individuals diagnosed with anorexia nervosa were viewed as morally better than those diagnosed with bulimia nervosa or binge eating disorder. Consequently, diagnostic crossover from a \"better\" to a \"worse\" eating disorder was often experienced as shameful moral failing, and a new diagnosis impacted the individual's sense of self-identity. These findings are of significance for both ethicists and clinicians; the paper concludes by outlining the relevance and possible clinical implications of shame in diagnostic crossover and suggesting avenues for future research.", "Synthesizing both objectification theory (Fredrickson & Roberts, 1997) and minority stress theory (Meyer, 2003), the present study used a pantheoretical model of dehumanization (Moradi, 2013) to examine body image concerns and disordered eating symptomatology with 205 transgender women from the United States. Objectification theory constructs (i.e., sexual objectification, internalization of sociocultural standards of attractiveness, body surveillance, body dissatisfaction) and minority stress-related variables (i.e., antitransgender discrimination) were examined as direct and indirect predictors of disordered eating. Results of a latent variable SEM (with a higher-order dehumanization factor comprised of sexual objectification and discrimination) generally provided support for our hypothesized direct and indirect relations. As expected, dehumanization was related directly to internalization and disordered eating and had significant indirect links to body surveillance, body dissatisfaction, and disordered eating via internalization. Potential implications of a pantheoretical model for future research with transgender women are discussed. (PsycINFO Database Record (c) 2019 APA, all rights reserved)." ]
Epidemiology of hereditary von Willebrand disease	Although hereditary von Willebrand disease (VWD) is the most common bleeding disorder, its epidemiology is not well understood. A systematic review (PROSPERO CRD42020197674/CRD42021244374) on the epidemiology/burden of illness of VWD was conducted to better understand patients' unmet needs.	[ "The aim of this study was to elucidate the fall in von Willebrand factor (VWF) and factor VIII activity (FVIII) after childbirth in women with and without von Willebrand disease (VWD). VWF:RCo, VWF:Ag, and FVIII were obtained in the third trimester of pregnancy, on admission for childbirth, and 10 times postpartum. Specimens were processed within 4 h and analysed centrally. Means were calculated at each time point. Forty women (40 pregnancies) without VWD and 32 women (35 pregnancies) with VWD were enrolled. 15/32 with VWD were treated (30% of those with type 1 and all of those with type 2) in 17 pregnancies. Treatments prior to delivery consisted of desmopressin (2/17), VWF concentrate (15/17) and after delivery VWF concentrate (16/17). Duration of treatment was 0-21 days (median 6). VWF levels peaked at 250% of baseline--4 h postpartum in women with VWD and 12 h postpartum in women without VWD. Thereafter, VWF levels fell rapidly, approached baseline at 1 week and reached baseline at 3 weeks. Except immediately postpartum, when the levels among treated cases were higher, levels among women with VWD appeared to parallel, but were lower than those among women without VWD. Levels were lowest among those who received treatment. VWF levels fall rapidly after childbirth. Except immediately postpartum, current treatment strategies do not raise VWF levels to the levels of women without VWD or even to the levels of women with milder, untreated VWD. Consequently, women with VWD may be at risk of postpartum haemorrhage despite treatment.", "Joint bleeds (JB) are reported in a minority of patients with von Willebrand disease (VWD) but may lead to structural joint damage. Prevalence, severity and impact of JB in VWD are largely unknown. The aim of this study was to assess JB prevalence, onset, treatment and impact on health-related quality of life (HR-QoL) and joint integrity in moderate and severe VWD. In the Willebrand in the Netherlands study 804 moderate and severe VWD patients [von Willebrand factor (VWF) activity ≤30U dL(-1)] completed a questionnaire on occurrence, sites and consequences of JB. To analyse JB number, onset, treatment and impact on joint integrity we additionally performed a patient-control study on medical file data comparing patients with JB to age, gender, factor VIII (FVIII)- and VWF activity matched VWD patients without JB. Of all VWD patients 23% (184/804) self-reported JB. These 184 patients reported joint damage more often (54% vs. 18%, P < 0.001) and had lower HR-QoL (SF36, P < 0.05) compared to VWD patients not reporting JB. Of 55 patients with available JB data, 65% had the first JB before age 16. These 55 patients used more clotting factor concentrate (CFC; median dose 43 vs. 0 IE FVIII kg(-1) year(-1) , P < 0.001), more often had X-ray joint damage (44% vs. 11%, P = 0.001] and chronic joint pain (44% vs. 18%, P = 0.008) compared to 55 control VWD patients without JB. In conclusion, joint bleeds are reported by 23% of moderate and severe VWD patients, mostly start in childhood, are associated with more CFC use, joint pain, lower HR-QoL and significantly more radiological and self-reported joint damage.", "The prevalence of von Willebrand's disease (VWD) in Western European countries and Israel was assessed. Possible patients were identified initially by questionnaire and those with plasma levels of von Willebrand factor antigen ( WFAg ; factor VIII related antigen) less than 1 u/dl (less than 1%) were confirmed by immunoradiometric assay performed at four reference centres. The prevalences of severe VWD in Scandinavian countries were similar to each other and about 10 times greater than those in other Western European countries. The prevalence in Israel was intermediate. The Registry will form the basis for future non- invasive epidemiological studies of atherosclerosis in these individuals.", "von Willebrand disease (VWD) is a bleeding disorder caused by inherited defects in the concentration, structure, or function of von Willebrand factor (VWF). VWD is classified into three primary categories. Type 1 includes partial quantitative deficiency, type 2 includes qualitative defects, and type 3 includes virtually complete deficiency of VWF. VWD type 2 is divided into four secondary categories. Type 2A includes variants with decreased platelet adhesion caused by selective deficiency of high-molecular-weight VWF multimers. Type 2B includes variants with increased affinity for platelet glycoprotein Ib. Type 2M includes variants with markedly defective platelet adhesion despite a relatively normal size distribution of VWF multimers. Type 2N includes variants with markedly decreased affinity for factor VIII. These six categories of VWD correlate with important clinical features and therapeutic requirements. Some VWF gene mutations, alone or in combination, have complex effects and give rise to mixed VWD phenotypes. Certain VWD types, especially type 1 and type 2A, encompass several pathophysiologic mechanisms that sometimes can be distinguished by appropriate laboratory studies. The clinical significance of this heterogeneity is under investigation, which may support further subdivision of VWD type 1 or type 2A in the future.", "von Willebrand disease (VWD) is a hereditary bleeding disorder, caused by a deficiency in the levels and/or function of von Willebrand factor (VWF). Women with VWD appear to be at increased risk of experiencing postpartum hemorrhage (PPH), though the levels of VWF increase during pregnancy. There is limited knowledge of how PPH is associated with the subtype of VWD, plasma levels of other coagulations factors than VWF and given hemostatic treatment. The aims were to investigate the incidence of PPH in women with VWD and to analyse the correlation between PPH and: (1) type of VWD, (2) laboratory monitoring of VWF and FVIII and (3) hemostatic drug treatment. This was a retrospective observational study. The study participants (n = 34) were recruited from the Coagulation Unit, Karolinska University hospital. Fifty-nine deliveries, which occurred in 14 different obstetrics units (years 1995-2012) were included in the study. The incidence of primary PPH was 44%, severe primary PPH 20% and secondary PPH 12%. VWD type 3 was associated with a higher risk of experiencing severe primary PPH compared to other subtypes. FVIII:C in pregnancy was inversely correlated to blood loss during delivery. There was a significantly higher incidence of secondary PPH when the VWD diagnosis was unknown at time of delivery. The women with VWD are at higher risk of PPH, especially those with type 3 VWD or when diagnosis is unknown prior to delivery. Identification of pregnant women with undiagnosed VWD may be of importance in order to prevent PPH.", "von Willebrand factor (VWF) plays a critical role in platelet adhesion and aggregation after vascular injury and at sites of high shear rate. Elevated VWF levels are associated with an increased risk of ischemic cardiovascular events; however, it is unclear whether VWF deficiency is protective against atherosclerosis. We aimed to compare the prevalence of cardiovascular disease (CVD) among patients with and without von Willebrand disease (VWD). A cross-sectional analysis was performed on discharge data for adults from the Nationwide Inpatient Sample (NIS) between the years 2009 and 2011. CVD was defined as ischemic heart disease, myocardial infarction, ischemic cerebrovascular disease, or peripheral vascular disease. For prevalence calculations and statistical analyses, we used discharge-level weights provided by the NIS to reflect national estimates. CVD was compared across groups by use of the Rao-Scott chi-square test. Multivariable logistic regression was used to estimate the likelihood of CVD in VWD patients after adjustment for age, gender, and CVD-related risk factors. The prevalence of CVD in VWD patients was less than the prevalence of CVD in non-VWD patients (15.0% versus 26.0%). VWD was associated with a decreased likelihood of CVD after adjustment for age, gender, and CVD-related risk factors (odds ratio 0.85; 95% confidence interval 0.79-0.92). These findings indicate that the risk of CVD is decreased among VWD patients, and that VWF deficiency may be protective against CVD.", "von Willebrand disease (VWD) is the commonest inherited bleeding disorder. Management of major surgery or bleeding often requires treatment with a plasma-derived (pd) VWF/FVIII containing concentrate. Wilate® is a dual-virally inactivated pd-concentrate, produced specifically for the treatment of VWD, containing physiological (1:1) ratios of VWF: FVIII. We reviewed efficacy and safety of Wilate® usage (2007-2012) at our centre including 2 years following product switching the majority of patients. Clinical and laboratory data of all adult patients treated with Wilate® during the study period were evaluated. Fifty four patients used 3 972 150 IU of Wilate® (1378 infusions) between 1/3/07 and 1/5/12. Efficacy was rated as being excellent or good in 94% of surgical episodes (n = 70; 34 patients) and 98% of bleeding/traumatic episodes (n = 46; 25 patients). Eight patients (2 636 100 IU) were managed on home treatment regimens. Two patients switched to Wilate® prophylaxis in the evaluation period, demonstrating similar efficacy to a previous product. Incremental recoveries (n = 37) were 2.24 IU dL(-1) per IU kg(-1) for FVIII:C, 2.39 IU dL(-1) per IU kg(-1) for VWF:Ag and 1.88 IU dL(-1) per IU kg(-1) for VWF:RCo. Six adverse events occurred in six patients (11.1% patients) over 1378 infusions (0.44%). Half of these were retrospectively felt to be infusion speed related. No notable accumulation of FVIII was seen in patients treated for ≥3 days. There was no treatment failure, thrombosis, transfusion transmitted infection or inhibitory VWF antibodies seen. Our findings confirm safety and efficacy of Wilate® in an adult VWD population with lack of notable FVIII accumulation." ]
Trends in Pregnancy Outcomes in Indian Women, 2015-2021	Pregnancy outcome is an important health indicator of the quality of maternal health. Adverse pregnancy outcomes is a major public health problem, which can lead to poor maternal and neonatal outcomes. This study investigates the trends in pregnancy outcomes prevalent during 2015-2021 in Indian women.	[ "Overweight, obesity, hypertension, and diabetes increase the risk of non-communicable diseases and all-cause mortality worldwide. Previous studies have not determined the prevalence of these conditions/diseases throughout India. Therefore, this study was aimed to address this limitation. Data on these conditions/diseases among men and women aged ≥ 18 years were obtained from the fourth National Family Health Survey conducted throughout India between January 2015 and December 2016. The prevalence and prevalence rate per 100,000 population were calculated at the national level and by age group, sex, and type of residence for each state and union territory. The national prevalence of overweight, obesity, hypertension, and diabetes were 14.6%, 3.4%, 5.2%, and 7.1%, respectively. The highest prevalence of these conditions/diseases at the national level was seen among those aged 35-49 years (54 years for men), especially women living in urban areas. In India, 1 out of every 7, 29, 19, and 14 individuals at the national level had overweight, obesity, hypertension, and diabetes, respectively-between 2015 and 2016. These results are important for the healthcare system and government policies in the future. Moreover, targeted efforts are required to establish public health strategies for the prevention, management, and treatment of these conditions/diseases throughout India.", "The metabolic syndrome is a common metabolic disorder that results from the increasing prevalence of obesity. The disorder is defined in various ways, but in the near future a new definition(s) will be applicable worldwide. The pathophysiology seems to be largely attributable to insulin resistance with excessive flux of fatty acids implicated. A proinflammatory state probably contributes to the syndrome. The increased risk for type 2 diabetes and cardiovascular disease demands therapeutic attention for those at high risk. The fundamental approach is weight reduction and increased physical activity; however, drug treatment could be appropriate for diabetes and cardiovascular disease risk reduction.", "Despite the rising number of patients with diabetes and hypertension in India, there is a dearth of nationwide, comprehensive prevalence data on these diseases. Our study aimed at collecting data on the prevalence of diabetes and hypertension and the underlying risk factors in various outpatient facilities throughout India. This cross-sectional study was planned to be conducted in 10 Indian states, one state at a time. It was targeted to enroll about 2,000 patients from 100 centers in each state. Each center enrolled the first 10 patients (≥18 years of age, not pregnant, signed consent) per day on two consecutive days. \"Diabetes\" and \"hypertension\" were defined by the 2008 American Diabetes Association and the Joint National Committee's 7(th) Report guidelines, respectively. Patient data (demographics, lifestyle factors, medical history, and laboratory diagnostic results) were collected and analyzed. During 2009-2010, in total, 15,662 eligible patients (54.8% males; mean age, 48.9±13.9 years) from eight states were enrolled. Diabetes was prevalent in 5,427 (34.7%) patients, and 7,212 (46.0%) patients had hypertension. Diabetes and hypertension were coexistent in 3,227 (20.6%) patients. Among those whose disease status was not known at enrollment, 7.2% (793 of 11,028) and 22.2% (2,408 of 10,858) patients were newly diagnosed with diabetes and hypertension, respectively; additionally, 18.4% (2,031 of 11,028) were classified as having prediabetes and 60.1% (6,521 of 10,858) as having prehypertension. A positive association (P<0.05) was observed between diabetes/hypertension and age, familial history of either, a medical history of cardiovascular disorders, alcohol consumption, and diet. Our study demonstrates that the substantial burden of diabetes and hypertension is on the rise in India. Patient awareness and timely diagnosis and intervention hold the key to limiting this twin epidemic.", "To evaluate the prevalence, awareness, treatment and control of hypertension among elderly individuals in Bangladesh and India. A community-based sample of 1203 elderly individuals (670 women; mean age, 70 years) was selected using a multistage cluster sampling technique from two sites in Bangladesh and three sites in India. The overall prevalence of hypertension (WHO-International Society for Hypertension criteria) was 65% (95% confidence interval = 62-67%). The prevalence was higher in urban than rural areas, but did not differ significantly between the sexes. Multiple logistic regression analyses identified a higher body mass index, higher education status and prevalent diabetes mellitus as important correlates of the prevalence of hypertension. Physical activity, rural residence, and current smoking were inversely related to the prevalence of hypertension. Among study subjects who had hypertension, 45% were aware of their condition, 40% were taking anti-hypertensive medications, but only 10% achieved the level established by the US Sixth Joint National Committee on Detection, Evaluation and Treatment of Hypertension (JNC VI)/WHO criteria. A visit to a physician in the previous year, higher educational attainment and being female emerged as important correlates of hypertension awareness. Our findings emphasize the need to implement effective and low cost management regimens based on absolute levels of cardiovascular risk appropriate for the economic context. From a public health perspective, the only sustainable approach to the high prevalence of hypertension in the Indian subcontinent is through a strategy to reduce the average blood pressure in the population." ]
Transversely-isotropic with anisotropic damping MR elastography of the brain	Measuring tissue parameters from increasingly sophisticated mechanical property models may uncover new contrast mechanisms with clinical utility. Building on previous work on in vivo brain MR elastography (MRE) with a transversely-isotropic with isotropic damping (TI-ID) model, we explore a new transversely-isotropic with anisotropic damping (TI-AD) model that involves six independent parameters describing direction-dependent behavior for both stiffness and damping. The direction of mechanical anisotropy is determined by diffusion tensor imaging and we fit three complex-valued moduli distributions across the full brain volume to minimize differences between measured and modeled displacements. We demonstrate spatially accurate property reconstruction in an idealized shell phantom simulation, as well as an ensemble of 20 realistic, randomly-generated simulated brains. We characterize the simulated precisions of all six parameters across major white matter tracts to be high, suggesting that they can be measured independently with acceptable accuracy from MRE data. Finally, we present in vivo anisotropic damping MRE reconstruction data. We perform t-tests on eight repeated MRE brain exams on a single-subject, and find that the three damping parameters are statistically distinct for most tracts, lobes and the whole brain. We also show that population variations in a 17-subject cohort exceed single-subject measurement repeatability for most tracts, lobes and whole brain, for all six parameters. These results suggest that the TI-AD model offers new information that may support differential diagnosis of brain diseases.	[ "The mechanical properties of brain tissue, particularly those of white matter (WM), need to be characterized accurately for use in finite element (FE) models of brain biomechanics and traumatic brain injury (TBI). Magnetic resonance elastography (MRE) is a powerful tool for non-invasive estimation of the mechanical properties of soft tissues. While several studies involving direct mechanical tests of brain tissue have shown mechanical anisotropy, most MRE studies of brain tissue assume an isotropic model. In this study, an incompressible transversely isotropic (TI) material model parameterized by minimum shear modulus (μ2), shear anisotropy parameter (ϕ), and tensile anisotropy parameter (ζ) is applied to analyze MRE measurements of ex vivo porcine white matter (WM) brain tissue. To characterize shear anisotropy, \"slow\" (pure transverse) shear waves were propagated at 100, 200 and 300Hz through sections of ex vivo brain tissue including both WM and gray matter (GM). Shear waves were found to propagate with elliptical fronts, consistent with TI material behavior. Shear wave fields were also analyzed within regions of interest (ROI) to find local shear wavelengths parallel and perpendicular to fiber orientation. FE simulations of a TI material with a range of plausible shear modulus (μ2) and shear anisotropy parameters (ϕ) were run and the results were analyzed in the same fashion as the experimental case. Parameters of the FE simulations which most closely matched each experiment were taken to represent the mechanical properties of that particular sample. Using this approach, WM in the ex vivo porcine brain was found to be mildly anisotropic in shear with estimates of minimum shear modulus (actuation frequencies listed in parenthesis): μ2= 1.04 ± 0.12 kPa (at 100Hz), μ2= 1.94 ± 0.29 kPa (at 200Hz), and μ2= 2.88 ± 0.34 kPa (at 300Hz) and corresponding shear anisotropy factors of ϕ= 0.27 ± 0.09 (at 100Hz), ϕ= 0.29 ± 0.14 (at 200Hz) and ϕ= 0.34 ± 0.13 (at 300Hz). Future MRE studies will focus on tensile anisotropy, which will require both slow and fast shear waves for accurate estimation.", "Magnetic resonance elastography (MRE) is a phase-contrast-based MRI imaging technique that can directly visualize and quantitatively measure propagating acoustic strain waves in tissue-like materials subjected to harmonic mechanical excitation. The data acquired allows the calculation of local quantitative values of shear modulus and the generation of images that depict tissue elasticity or stiffness. This is significant because palpation, a physical examination that assesses the stiffness of tissue, can be an effective method of detecting tumors, but is restricted to parts of the body that are accessible to the physician's hand. MRE shows promise as a potential technique for 'palpation by imaging', with possible applications in tumor detection (particularly in breast, liver, kidney and prostate), characterization of disease, and assessment of rehabilitation (particularly in muscle). We describe MRE in the context of other recent techniques for imaging elasticity, discuss the processing algorithms for elasticity reconstruction and the issues and assumptions they involve, and present recent ex vivo and in vivo results.", "A nuclear magnetic resonance imaging (MRI) method is presented for quantitatively mapping the physical response of a material to harmonic mechanical excitation. The resulting images allow calculation of regional mechanical properties. Measurements of shear modulus obtained with the MRI technique in gel materials correlate with independent measurements of static shear modulus. The results indicate that displacement patterns corresponding to cyclic displacements smaller than 200 nanometers can be measured. The findings suggest the feasibility of a medical imaging technique for delineating elasticity and other mechanical properties of tissue.", "Magnetic resonance elastography (MRE) has emerged as a sensitive imaging technique capable of providing a quantitative understanding of neural microstructural integrity. However, a reliable method for the quantification of the anisotropic mechanical properties of human white matter is currently lacking, despite the potential to illuminate the pathophysiology behind neurological disorders and traumatic brain injury. In this study, we examine the use of multiple excitations in MRE to generate wave displacement data sufficient for anisotropic inversion in white matter. We show the presence of multiple unique waves from each excitation which we combine to solve for parameters of an incompressible, transversely isotropic (ITI) material: shear modulus, μ, shear anisotropy, ϕ, and tensile anisotropy, ζ. We calculate these anisotropic parameters in the corpus callosum body and find the mean values as μ = 3.78 kPa, ϕ = 0.151, and ζ = 0.099 (at 50 Hz vibration frequency). This study demonstrates that multi-excitation MRE provides displacement data sufficient for the evaluation of the anisotropic properties of white matter.", "Alzheimer's disease is a personally devastating neurodegenerative disorder and a major public health concern. There is an urgent need for medical imaging techniques that better characterize the early stages and monitor the progression of the disease. Magnetic resonance elastography (MRE) is a relatively new and highly sensitive MRI technique that can non-invasively assess tissue microstructural integrity via measurement of brain viscoelastic mechanical properties. For the first time, we use high-resolution MRE methods to conduct a voxel-wise MRE investigation and state-of-the-art post hoc region of interest analysis of the viscoelastic properties of the cerebral cortex in patients with Alzheimer's disease (N = 11) compared with cognitively healthy older adults (N = 12). We replicated previous findings that have reported significant volume and stiffness reductions at the whole-brain level. Significant reductions in volume were also observed in Alzheimer's disease when white matter, cortical grey matter and subcortical grey matter compartments were considered separately; lower stiffness was also observed in white matter and cortical grey matter, but not in subcortical grey matter. Voxel-based morphometry of both cortical and subcortical grey matter revealed localized reductions in volume due to Alzheimer's disease in the hippocampus, fusiform, middle, superior temporal gyri and precuneus. Similarly, voxel-based MRE identified lower stiffness in the middle and superior temporal gyri and precuneus, although the spatial distribution of these effects was not identical to the pattern of volume reduction. Notably, MRE additionally identified stiffness deficits in the operculum and precentral gyrus located within the frontal lobe; regions that did not undergo volume loss identified through voxel-based morphometry. Voxel-based-morphometry and voxel-based MRE results were confirmed by a complementary post hoc region-of-interest approach in native space where the viscoelastic changes remained significant even after statistically controlling for regional volumes. The pattern of reduction in cortical stiffness observed in Alzheimer's disease patients raises the possibility that MRE may provide unique insights regarding the neural mechanisms which underlie the development and progression of the disease. The measured mechanical property changes that we have observed warrant further exploration to investigate the diagnostic usefulness of MRE in cases of Alzheimer's disease and other dementias.", "Neurological disorders are one of the most important public health concerns in developed countries. Established brain imaging techniques such as magnetic resonance imaging (MRI) and x-ray computerised tomography (CT) have been essential in the identification and diagnosis of a wide range of disorders, although usually are insufficient in sensitivity for detecting subtle pathological alterations to the brain prior to the onset of clinical symptoms-at a time when prognosis for treatment is more favourable. The mechanical properties of biological tissue provide information related to the strength and integrity of the cellular microstructure. In recent years, mechanical properties of the brain have been visualised and measured non-invasively with magnetic resonance elastography (MRE), a particularly sensitive medical imaging technique that may increase the potential for early diagnosis. This review begins with an introduction to the various methods used for the acquisition and analysis of MRE data. A systematic literature search is then conducted to identify studies that have specifically utilised MRE to investigate the human brain. Through the conversion of MRE-derived measurements to shear stiffness (kPa) and, where possible, the loss tangent (rad), a summary of results for global brain tissue and grey and white matter across studies is provided for healthy participants, as potential baseline values to be used in future clinical investigations. In addition, the extent to which MRE has revealed significant alterations to the brain in patients with neurological disorders is assessed and discussed in terms of known pathophysiology. The review concludes by predicting the trends for future MRE research and applications in neuroscience.", "To define thresholds for detecting significant change in liver viscoelasticity with magnetic resonance (MR) elastography, both for whole-liver measurements and for voxel-wise measurements in relation to spatial resolution. This prospective study was approved by the institutional review board, and all participants provided written informed consent. Thirty participants (16 volunteers and 14 patients with hepatitis B or C; 18 men; median age, 30.4 years; age range, 18.9-58.6 years) underwent imaging twice while in the same position (intraimage reproducibility), after repositioning (within-day reproducibility), and 1-4 weeks later (between-weeks reproducibility). MR elastography parameters comprised elasticity, viscosity, attenuation parameter α, and propagation parameter β. Bland-Altman analysis was used to calculate repeatability indexes for each parameter. Analyses were performed in a region-of-interest and a voxel-by-voxel level. Voxel-wise results were calculated in relation to spatial resolution by applying Gaussian filtering to establish the optimal trade-off point between resolution and reproducibility. For elasticity, α, and β, within-day and between-weeks results were significantly lower than intraimage results (P ≤ .018 for all). Within-day and between-weeks results did not differ significantly. Over-time changes of more than 22.2% for elasticity, 26.3% for viscosity, 26.8% for α, and 10.1% for β represented thresholds for significant change. The optimal trade-off between spatial resolution and reproducibility was found at a filter size of 8-mm full width at half maximum (FWHM) for elasticity and propagation parameter β and at 16-mm FWHM for viscosity and attenuation parameter α. Repositioning causes a significant decrease in the reproducibility of MR elastography. The propagation parameter β is the most reliable parameter, with an over-time threshold for significant change of 10.1% and the ability to reproduce viscoelasticity up to a resolution of 8-mm FWHM. Online supplemental material is available for this article." ]
Mediterranean-DASH Intervention for Neurodegenerative Delay (MIND) and Mediterranean dietary patterns with -amyloid load, phosphorylated tau tangles, and global Alzheimer disease pathology in postmortem brain tissue of older adults	Diet may reduce Alzheimer dementia risk and slow cognitive decline, but the understanding of the relevant neuropathologic mechanisms remains limited. The association of dietary patterns with Alzheimer disease (AD) pathology has been suggested using neuroimaging biomarkers. This study examined the association of Mediterranean-DASH Intervention for Neurodegenerative Delay (MIND) and Mediterranean dietary patterns with β-amyloid load, phosphorylated tau tangles, and global AD pathology in postmortem brain tissue of older adults.	[ "To systematically review all the prospective cohort studies that have analysed the relation between adherence to a Mediterranean diet, mortality, and incidence of chronic diseases in a primary prevention setting. Meta-analysis of prospective cohort studies. English and non-English publications in PubMed, Embase, Web of Science, and the Cochrane Central Register of Controlled Trials from 1966 to 30 June 2008. Studies reviewed Studies that analysed prospectively the association between adherence to a Mediterranean diet, mortality, and incidence of diseases; 12 studies, with a total of 1 574,299 subjects followed for a time ranging from three to 18 years were included. The cumulative analysis among eight cohorts (514,816 subjects and 33,576 deaths) evaluating overall mortality in relation to adherence to a Mediterranean diet showed that a two point increase in the adherence score was significantly associated with a reduced risk of mortality (pooled relative risk 0.91, 95% confidence interval 0.89 to 0.94). Likewise, the analyses showed a beneficial role for greater adherence to a Mediterranean diet on cardiovascular mortality (pooled relative risk 0.91, 0.87 to 0.95), incidence of or mortality from cancer (0.94, 0.92 to 0.96), and incidence of Parkinson's disease and Alzheimer's disease (0.87, 0.80 to 0.96). Greater adherence to a Mediterranean diet is associated with a significant improvement in health status, as seen by a significant reduction in overall mortality (9%), mortality from cardiovascular diseases (9%), incidence of or mortality from cancer (6%), and incidence of Parkinson's disease and Alzheimer's disease (13%). These results seem to be clinically relevant for public health, in particular for encouraging a Mediterranean-like dietary pattern for primary prevention of major chronic diseases.", "The Mediterranean diet has been shown to have beneficial effects on cardiovascular risk factors. To compare the short-term effects of 2 Mediterranean diets versus those of a low-fat diet on intermediate markers of cardiovascular risk. Substudy of a multicenter, randomized, primary prevention trial of cardiovascular disease (Prevención con Dieta Mediterránea [PREDIMED] Study). Primary care centers affiliated with 10 teaching hospitals. 772 asymptomatic persons 55 to 80 years of age at high cardiovascular risk who were recruited from October 2003 to March 2004. Participants were assigned to a low-fat diet (n = 257) or to 1 of 2 Mediterranean diets. Those allocated to Mediterranean diets received nutritional education and either free virgin olive oil, 1 liter per week (n = 257), or free nuts, 30 g/d (n = 258). The authors evaluated outcome changes at 3 months. Body weight, blood pressure, lipid profile, glucose levels, and inflammatory molecules. The completion rate was 99.6%. Compared with the low-fat diet, the 2 Mediterranean diets produced beneficial changes in most outcomes. Compared with the low-fat diet, the mean changes in the Mediterranean diet with olive oil group and the Mediterranean diet with nuts group were -0.39 mmol/L (95% CI, -0.70 to -0.07 mmol/L) and -0.30 mmol/L (CI, -0.58 to -0.01 mmol/L), respectively, for plasma glucose levels; -5.9 mm Hg (CI, -8.7 to -3.1 mm Hg) and -7.1 mm Hg (CI, -10.0 to -4.1 mm Hg), respectively, for systolic blood pressure; and -0.38 (CI, -0.55 to -0.22) and - 0.26 (CI, -0.42 to -0.10), respectively, for the cholesterol-high-density lipoprotein cholesterol ratio. The Mediterranean diet with olive oil reduced C-reactive protein levels by 0.54 mg/L (CI, 1.04 to 0.03 mg/L) compared with the low-fat diet. This short-term study did not focus on clinical outcomes. Nutritional education about low-fat diet was less intense than education about Mediterranean diets. Compared with a low-fat diet, Mediterranean diets supplemented with olive oil or nuts have beneficial effects on cardiovascular risk factors.", "To test the hypothesis that the association of amyloid load with clinical Alzheimer disease (AD) and cognitive impairment is mediated through neurofibrillary tangles. Longitudinal clinicopathologic cohort study. Forty-four individuals with clinically diagnosed AD and 53 without dementia who participated in the Religious Orders Study underwent a uniform structured clinical evaluation for AD and cognitive testing about 8 months prior to death, and brain autopsy at death. The percent area occupied by amyloid-beta and the density of neurofibrillary tangles were quantified from 6 brain regions and averaged to yield summary measures of amyloid load and neurofibrillary tangles. Multivariate regression analyses were used to simultaneously examine the effects of amyloid load and neurofibrillary tangles on clinically diagnosed AD and level of cognition. Clinically diagnosed AD and level of global cognitive function proximate to death. In separate logistic regression analyses, each 1% increase in amyloid load was associated with about a 50% increase in the odds of clinical AD (P =.002), and each neurofibrillary tangle was associated with a greater than 20% increase in the odds of clinical AD (P<.001). When a term for tangles was added to the regression model with amyloid, the association of amyloid load with clinical disease was reduced by more than 60% and was no longer significant, whereas the association of tangles with clinical disease was essentially unchanged. Similar results were found in analyses of global cognitive function. These findings are consistent with a sequence of pathologic events whereby the effect of amyloid deposition on clinical disease is mediated by neurofibrillary tangles.", "Higher adherence to the Mediterranean diet (MeDi) may protect from Alzheimer disease (AD), but its association with mild cognitive impairment (MCI) has not been explored. To investigate the association between the MeDi and MCI. In a multiethnic community study in New York, we used Cox proportional hazards to investigate the association between adherence to the MeDi (0-9 scale; higher scores indicate higher adherence) and (1) the incidence of MCI and (2) the progression from MCI to AD. All of the models were adjusted for cohort, age, sex, ethnicity, education, APOE genotype, caloric intake, body mass index, and duration between baseline dietary assessment and baseline diagnosis. Incidence of MCI and progression from MCI to AD. There were 1393 cognitively normal participants, 275 of whom developed MCI during a mean (SD) follow-up of 4.5 (2.7) years (range, 0.9-16.4 years). Compared with subjects in the lowest MeDi adherence tertile, subjects in the middle tertile had 17% less risk (hazard ratio [HR] = 0.83; 95% confidence interval [CI], 0.62-1.12; P = .24) of developing MCI and those in the highest tertile had 28% less risk (HR = 0.72; 95% CI, 0.52-1.00; P = .05) of developing MCI (trend HR = 0.85; 95% CI, 0.72-1.00; P for trend = .05). There were 482 subjects with MCI, 106 of whom developed AD during a mean (SD) follow-up of 4.3 (2.7) years (range, 1.0-13.8 years). Compared with subjects in the lowest MeDi adherence tertile, subjects in the middle tertile had 45% less risk (HR = 0.55; 95% CI, 0.34-0.90; P = .01) of developing AD and those in the highest tertile had 48% less risk (HR = 0.52; 95% CI, 0.30-0.91; P = .02) of developing AD (trend HR = 0.71; 95% CI, 0.53-0.95; P for trend = .02). Higher adherence to the MeDi is associated with a trend for reduced risk of developing MCI and with reduced risk of MCI conversion to AD.", "To determine whether following a Mediterranean-like diet (MeDi) relates to cognitive functions and in vivo biomarkers for Alzheimer disease (AD), we analyzed cross-sectional data from the German DZNE-Longitudinal Cognitive Impairment and Dementia Study. METHOD: The sample (n=512, mean age: 69.5±5.9 years) included 169 cognitively normal participants and subjects at higher AD risk (53 with relatives with AD, 209 with subjective cognitive decline, and 81 with mild cognitive impairment). We defined MeDi adherence based on the Food Frequency Questionnaire. Brain volume outcomes were generated via voxel-based morphometry on T1-MRI and cognitive performance with an extensive neuropsychological battery. AD-related biomarkers (Aβ42/40 ratio, pTau181) in cerebrospinal fluid were assessed in n=226 individuals. We analyzed the associations between MeDi and the outcomes with linear regression models controlling for several covariates. Additionally, we applied hypothesis-driven mediation and moderation analysis. Higher MeDi adherence related to larger mediotemporal gray matter volume (p<0.05 FWE corrected), better memory (β±SE = 0.03 ± 0.02; p=0.038), and less amyloid (Aβ42/40 ratio, β±SE = 0.003 ± 0.001; p=0.008) and pTau181 pathology (β±SE = -1.96±0.68; p=0.004). Mediotemporal volume mediated the association between MeDi and memory (40% indirect mediation). Finally, MeDi favorably moderated the associations between Aβ42/40 ratio, pTau181 and mediotemporal atrophy. Results were consistent correcting for ApoE-ε4 status. Our findings corroborate the view of MeDi as a protective factor against memory decline and mediotemporal atrophy. Importantly, they suggest that these associations might be explained by a decrease of amyloidosis and tau-pathology. Longitudinal and dietary intervention studies should further examine this conjecture and its treatment implications.", "A Physical Activity Scale for the Elderly (PASE) was evaluated in a sample of community-dwelling, older adults. Respondents were randomly assigned to complete the PASE by mail or telephone before or after a home visit assessment. Item weights for the PASE were derived by regressing a physical activity principal component score on responses to the PASE. The component score was based on 3-day motion sensor counts, a 3-day physical activity dairy and a global activity self-assessment. Test-retest reliability, assessed over a 3-7 week interval, was 0.75 (95% CI = 0.69-0.80). Reliability for mail administration (r = 0.84) was higher than for telephone administration (r = 0.68). Construct validity was established by correlating PASE scores with health status and physiologic measures. As hypothesized, PASE scores were positively associated with grip strength (r = 0.37), static balance (r = +0.33), leg strength (r = 0.25) and negatively correlated with resting heart rate (r = -0.13), age (r = -0.34) and perceived health status (r = -0.34); and overall Sickness Impact Profile score (r = -0.42). The PASE is a brief, easily scored, reliable and valid instrument for the assessment of physical activity in epidemiologic studies of older people.", "Specific immunocytochemical methods (AT8) permit evaluation of neuronal changes well before the actual formation of neurofibrillary tangles and neuropil threads. Initial changes are found in the transentorhinal region (temporal lobe). From here the destructive process encroaches upon the entorhinal region, Ammon's horn, and neocortex. Initial changes occur in comparatively young individuals and can also be observed at the same predilection sites in a few species of old aged domestic animals. In a later state of destruction, AT8 immunoreactive neurons develop typical argyrophilic neurofibrillary tangles and neuropil threads. Six stages of disease propagation can be distinguished with respect to the location of the tangle-bearing neurons and the severity of changes (transentorhinal stages I-II: clinically silent cases; limbic stages III-IV: incipient Alzheimer's disease; neocortical stages V-VI: fully developed Alzheimer's disease). Whole mount techniques reveal the lesional pattern of the particularly severely involved superficial entorhinal layer as seen from the free surface of the parahippocampal gyrus. This approach facilitates recognition of even subtle pathologic changes throughout the entire extent of cortical territories such as the transentorhinal and entorhinal regions." ]
Target protein degradation: a novel therapeutic modality	Target protein degradation (TPD) provides a novel therapeutic modality, other than inhibition, through the direct depletion of target proteins. Two primary human protein homeostasis mechanisms are exploited: the ubiquitin-proteasome system (UPS) and the lysosomal system. TPD technologies based on these two systems are progressing at an impressive pace.	[ "The current inhibitor-based approach to therapeutics has inherent limitations owing to its occupancy-based model: 1) there is a need to maintain high systemic exposure to ensure sufficient in vivo inhibition, 2) high in vivo concentrations bring potential for off-target side effects, and 3) there is a need to bind to an active site, thus limiting the drug target space. As an alternative, induced protein degradation lacks these limitations. Based on an event-driven model, this approach offers a novel catalytic mechanism to irreversibly inhibit protein function by targeting protein destruction through recruitment to the cellular quality control machinery. Prior protein degrading strategies have lacked therapeutic potential. However, recent reports of small-molecule-based proteolysis-targeting chimeras (PROTACs) have demonstrated that this technology can effectively decrease the cellular levels of several protein classes.", "The catalytic properties of proteolysis targeting chimeras (PROTACs) may lead to uncontrolled off-tissue target degradation that causes potential toxicity, limiting their clinical applications. The precise control of this technology in a tissue-selective manner can minimize the potential toxicity. Hypoxia is a hallmark of most solid tumors, accompanied by elevated levels of nitroreductase (NTR). Based on this character, we presented a type of NTR-responsive PROTACs to selectively degrade proteins of interest (POI) in tumor tissues. Compound 17-1 was the first NTR-responsive PROTAC synthesized by incorporating the caging group on the Von Hippel-Lindau (VHL) E3 ubiquitin ligase ligand. It could be activated by NTR to release the active PROTAC 17 to efficiently degrade the EGFR protein and subsequently exert antitumor efficacy. Thus, a general strategy for the precise control of PROTAC to induce POI degradation in tumor tissues by NTR was established, which provided a generalizable platform for the development of NTR-controlled PROTACs to achieve selective degradation.", "Target-protein degradation is an emerging field in drug discovery and development. In particular, the substrate-receptor proteins of the cullin-ubiquitin ligase system play a key role in selective protein degradation, which is an essential component of the anti-myeloma activity of immunomodulatory drugs (IMiDs), such as lenalidomide. Here, we demonstrate that a series of anticancer sulfonamides NSC 719239 (E7820), indisulam, and NSC 339004 (chloroquinoxaline sulfonamide, CQS) induce proteasomal degradation of the U2AF-related splicing factor coactivator of activating protein-1 and estrogen receptors (CAPERα) via CRL4DCAF15 mediated ubiquitination in human cancer cell lines. Both CRISPR-Cas9-based knockout of DCAF15 and a single amino acid substitution of CAPERα conferred resistance against sulfonamide-induced CAPERα degradation and cell-growth inhibition. Thus, these sulfonamides represent selective chemical probes for disrupting CAPERα function and designate DCAFs as promising drug targets for promoting selective protein degradation in cancer therapy.", "Hijacking the cellular protein degradation system offers unique opportunities for drug discovery, as exemplified by proteolysis-targeting chimeras. Despite their great promise for medical chemistry, so far, it has not been possible to reprogram the bacterial degradation machinery to interfere with microbial infections. Here, we develop small-molecule degraders, so-called BacPROTACs, that bind to the substrate receptor of the ClpC:ClpP protease, priming neo-substrates for degradation. In addition to their targeting function, BacPROTACs activate ClpC, transforming the resting unfoldase into its functional state. The induced higher-order oligomer was visualized by cryo-EM analysis, providing a structural snapshot of activated ClpC unfolding a protein substrate. Finally, drug susceptibility and degradation assays performed in mycobacteria demonstrate in vivo activity of BacPROTACs, allowing selective targeting of endogenous proteins via fusion to an established degron. In addition to guiding antibiotic discovery, the BacPROTAC technology presents a versatile research tool enabling the inducible degradation of bacterial proteins.", "Immunomodulatory drugs bind to cereblon (CRBN) to confer differentiated substrate specificity on the CRL4(CRBN) E3 ubiquitin ligase. Here we report the identification of a new cereblon modulator, CC-885, with potent anti-tumour activity. The anti-tumour activity of CC-885 is mediated through the cereblon-dependent ubiquitination and degradation of the translation termination factor GSPT1. Patient-derived acute myeloid leukaemia tumour cells exhibit high sensitivity to CC-885, indicating the clinical potential of this mechanism. Crystallographic studies of the CRBN-DDB1-CC-885-GSPT1 complex reveal that GSPT1 binds to cereblon through a surface turn containing a glycine residue at a key position, interacting with both CC-885 and a 'hotspot' on the cereblon surface. Although GSPT1 possesses no obvious structural, sequence or functional homology to previously known cereblon substrates, mutational analysis and modelling indicate that the cereblon substrate Ikaros uses a similar structural feature to bind cereblon, suggesting a common motif for substrate recruitment. These findings define a structural degron underlying cereblon 'neosubstrate' selectivity, and identify an anti-tumour target rendered druggable by cereblon modulation.", "Thalidomide and its derivatives, lenalidomide and pomalidomide, are immune modulatory drugs (IMiDs) used in the treatment of haematologic malignancies. IMiDs bind CRBN, the substrate receptor of the CUL4-RBX1-DDB1-CRBN (also known as CRL4(CRBN)) E3 ubiquitin ligase, and inhibit ubiquitination of endogenous CRL4(CRBN) substrates. Unexpectedly, IMiDs also repurpose the ligase to target new proteins for degradation. Lenalidomide induces degradation of the lymphoid transcription factors Ikaros and Aiolos (also known as IKZF1 and IKZF3), and casein kinase 1α (CK1α), which contributes to its clinical efficacy in the treatment of multiple myeloma and 5q-deletion associated myelodysplastic syndrome (del(5q) MDS), respectively. How lenalidomide alters the specificity of the ligase to degrade these proteins remains elusive. Here we present the 2.45 Å crystal structure of DDB1-CRBN bound to lenalidomide and CK1α. CRBN and lenalidomide jointly provide the binding interface for a CK1α β-hairpin-loop located in the kinase N-lobe. We show that CK1α binding to CRL4(CRBN) is strictly dependent on the presence of an IMiD. Binding of IKZF1 to CRBN similarly requires the compound and both, IKZF1 and CK1α, use a related binding mode. Our study provides a mechanistic explanation for the selective efficacy of lenalidomide in del(5q) MDS therapy. We anticipate that high-affinity protein-protein interactions induced by small molecules will provide opportunities for drug development, particularly for targeted protein degradation.", "Reversibly altering endogenous protein levels are persistent issues. Herein, we designed photoswitchable azobenzene-proteolysis targeting chimeras (Azo-PROTACs) by including azobenzene moieties between ligands for the E3 ligase and the protein of interest. Azo-PROTACs are light-controlled small-molecule tools for protein knockdown in cells. The light-induced configuration change can switch the active state to induce protein degradation activity, which can be reversely controlled by light exposure in intact cells. We compared the protein degradation abilities of Azo-PROTACs with different configurations and linker lengths. Using the stable form with the best degradation ability against the BCR-ABL fusion and ABL proteins in myelogenous leukemia K562 cells, we showed that Azo-PROTAC combines the potent protein knockdown and facile cell uptake properties of the small-molecule PROTAC with a reversible photoswitchability, offering a promising chemical knockdown strategy based on the light-induced reversible on/off properties." ]
Cellular kinetics and signaling pathway understanding of episodic permanent tumor regression	Based on the well-documented studies, numerous tumors episodically regress permanently without treatment. Knowing the host tissue-initiated causative factors would offer considerable translational applicability, as a permanent regression process may be therapeutically replicated on patients. For this, we developed a systems biological formulation of the regression process with experimental verification and identified the relevant candidate biomolecules for therapeutic utility. We devised a cellular kinetics-based quantitative model of tumor extinction in terms of the temporal behavior of three main tumor-lysis entities: DNA blockade factor, cytotoxic T-lymphocyte and interleukin-2. As a case study, we analyzed the time-wise biopsy and microarrays of spontaneously regressing melanoma and fibrosarcoma tumors in mammalian/human hosts. We analyzed the differentially expressed genes (DEGs), signaling pathways, and bioinformatics framework of regression. Additionally, prospective biomolecules that could cause complete tumor regression were investigated. The tumor regression process follows a first-order cellular dynamics with a small negative bias, as verified by experimental fibrosarcoma regression; the bias is necessary to eliminate the residual tumor. We identified 176 upregulated and 116 downregulated DEGs, and enrichment analysis showed that the most significant were downregulated cell-division genes: TOP2A-KIF20A-KIF23-CDK1-CCNB1. Moreover, Topoisomerase-IIA inhibition might actuate spontaneous regression, with collateral confirmation provided from survival and genomic analysis of melanoma patients. Candidate molecules such as Dexrazoxane/Mitoxantrone, with interleukin-2 and antitumor lymphocytes, may potentially replicate permanent tumor regression process of melanoma. To conclude, episodic permanent tumor regression is a unique biological reversal process of malignant progression, and signaling pathway understanding, with candidate biomolecules, may plausibly therapeutically replicate the regression process on tumors clinically.	[ "Cyclin-dependent kinase 1 (CDK1) plays an important role in cancer development, progression, and the overall process of tumorigenesis. However, no pan-cancer analysis has been reported for CDK1, and the predictive role of CDK1 in immune checkpoint inhibitors (ICIs) therapy response remains unexplored. Thus, in this study, we first investigated the potential oncogenic role of CDK1 in 33 tumors by multidimensional bioinformatics analysis based on The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) datasets. Bioinformatic analysis and immunohistochemical experiments confirmed that CDK1 is significantly upregulated in most common cancers and is strongly associated with prognosis. Further analysis indicated that CDK1 may influence tumor immunity mainly by mediating the degree of tumor infiltration of immune-associated cells, and the effect of CDK1 on immunity is diverse across tumor types in tumor microenvironment. CDK1 was also positively correlated with tumor mutational burden (TMB) and microsatellite instability (MSI) in certain cancer types, linking its expression to the assessment of possible treatment response. The results of the pan-cancer analysis study showed that the CDK1 gene was positively associated with the expression of three classes of RNA methylation regulatory proteins, and affects RNA function through multiple mechanisms of action and plays an important role in the posttranscriptional regulation of the tumor microenvironment. These findings shed light on the role of the CDK1 gene in cancer progression and provide information to further study the CDK1 gene as a potential target for pan-cancer.", "Background: Tumor mutation burden (TMB) is considered as a novel biomarker of response to immunotherapy and correlated with survival outcomes in various malignancies. Here, TMB-related genes (TRGs) expression signatures were constructed to investigate the association between TMB and prognosis in epithelial ovarian cancer (EOC), and the potential mechanism in immunoregulation was also explored. Methods: Based on somatic mutation data of 436 EOC samples from The Cancer Genome Atlas database, we examined the relationship between TMB level and overall survival (OS), as well as disease-free survival (DFS). Next, the TRGs signatures were constructed and validated. Differential abundance of immune cell infiltration, expression levels of immunomodulators and functional enrichment in high- and low-risk groups were also analyzed. Results: Higher TMB level revealed better OS and DFS, and correlated with earlier clinical stages in EOCs (P = 2.796e-04). The OS-related prognostic model constructed based on seven TRGs (B3GALT1, LIN7B, ANGPT2, D2HGDH, TAF13, PFDN4 and DNAJC19) significantly stratified EOC patients into high- and low-risk groups (P < 0.001). The AUC values of the seven-gene prognostic signature at 1 year, 3 years, and 5 years were 0.703, 0.758 and 0.777. While the DFS-related prognostic model was constructed based on the 4 TRGs (LPIN3, PXYLP1, IGSF23 and B3GALT1), with AUCs of 0.617, 0.756, and 0.731, respectively. Functional analysis indicated that immune-related pathways were enriched in low-risk groups. When considering the infiltration patterns of immune cells, we found higher proportions of follicular helper T (Tfh) cell and M1 macrophage, while lower infiltration of M0 macrophage in low-risk groups (P < 0.05). Accordingly, TMB levels of low-risk patients were significantly higher both in OS and DFS model (P < 0.01). Conclusions: Our TRGs-based models are reliable predictive tools for OS and DFS. High TMB may confer with an immunogenic microenvironment and predict favorable outcomes in EOCs.", "Although remission rates for metastatic melanoma are generally very poor, some patients can survive for prolonged periods following metastasis. We used gene expression profiling, mitotic index (MI), and quantification of tumor infiltrating leukocytes (TILs) and CD3+ cells in metastatic lesions to search for a molecular basis for this observation and to develop improved methods for predicting patient survival. We identified a group of 266 genes associated with postrecurrence survival. Genes positively associated with survival were predominantly immune response related (e.g., ICOS, CD3d, ZAP70, TRAT1, TARP, GZMK, LCK, CD2, CXCL13, CCL19, CCR7, VCAM1) while genes negatively associated with survival were cell proliferation related (e.g., PDE4D, CDK2, GREF1, NUSAP1, SPC24). Furthermore, any of the 4 parameters (prevalidated gene expression signature, TILs, CD3, and in particular MI) improved the ability of Tumor, Node, Metastasis (TNM) staging to predict postrecurrence survival; MI was the most significant contributor (HR = 2.13, P = 0.0008). An immune response gene expression signature and presence of TILs and CD3+ cells signify immune surveillance as a mechanism for prolonged survival in these patients and indicate improved patient subcategorization beyond current TNM staging.", "Cancer-associated fibroblasts (CAF) are a poorly characterized cell population in the context of liver cancer. Our study investigates CAF functions in intrahepatic cholangiocarcinoma (ICC), a highly desmoplastic liver tumor. Genetic tracing, single-cell RNA sequencing, and ligand-receptor analyses uncovered hepatic stellate cells (HSC) as the main source of CAF and HSC-derived CAF as the dominant population interacting with tumor cells. In mice, CAF promotes ICC progression, as revealed by HSC-selective CAF depletion. In patients, a high panCAF signature is associated with decreased survival and increased recurrence. Single-cell RNA sequencing segregates CAF into inflammatory and growth factor-enriched (iCAF) and myofibroblastic (myCAF) subpopulations, displaying distinct ligand-receptor interactions. myCAF-expressed hyaluronan synthase 2, but not type I collagen, promotes ICC. iCAF-expressed hepatocyte growth factor enhances ICC growth via tumor-expressed MET, thus directly linking CAF to tumor cells. In summary, our data demonstrate promotion of desmoplastic ICC growth by therapeutically targetable CAF subtype-specific mediators, but not by type I collagen." ]
Chronological Changes in Physical and Cognitive Functions in Middle-Aged and Older Adults with and Without Rheumatoid Arthritis	This study evaluated the chronological changes in physical and cognitive functions in middle-aged and older adults with and without rheumatoid arthritis (RA).	[ "Sarcopenia is defined as the loss of skeletal muscle mass and quality, which accelerates with aging and is associated with functional decline. Rising obesity prevalence has led to a high-risk group with both disorders. We assessed mortality risk associated with sarcopenia and sarcopenic obesity in elders. A subsample of 4652 subjects ≥60 years of age was identified from the National Health and Nutrition Examination Survey III (1988-1994), a cross-sectional survey of non-institutionalized adults. National Death Index data were linked to this data set. Sarcopenia was defined using a bioelectrical impedance formula validated using magnetic resonance imaging-measured skeletal mass by Janssen et al. Cutoffs for total skeletal muscle mass adjusted for height(2) were sex-specific (men: ≤5.75 kg/m(2); females ≤10.75 kg/m(2)). Obesity was based on % body fat (males: ≥27%, females: ≥38%). Modeling assessed mortality adjusting for age, sex, ethnicity (model 1), comorbidities (hypertension, diabetes, congestive heart failure, osteoporosis, cancer, coronary artery disease and arthritis), smoking, physical activity, self-reported health (model 2) and mobility limitations (model 3). Mean age was 70.6±0.2 years and 57.2% were female. Median follow-up was 14.3 years (interquartile range: 12.5-16.1). Overall prevalence of sarcopenia was 35.4% in women and 75.5% in men, which increased with age. Prevalence of obesity was 60.8% in women and 54.4% in men. Sarcopenic obesity prevalence was 18.1% in women and 42.9% in men. There were 2782 (61.7%) deaths, of which 39.0% were cardiovascular. Women with sarcopenia and sarcopenic obesity had a higher mortality risk than those without sarcopenia or obesity after adjustment (model 2, hazard ratio (HR): 1.35 (1.05-1.74) and 1.29 (1.03-1.60)). After adjusting for mobility limitations (model 3), sarcopenia alone (HR: 1.32 ((1.04-1.69) but not sarcopenia with obesity (HR: 1.25 (0.99-1.58)) was associated with mortality. For men, the risk of death with sarcopenia and sarcopenic obesity was nonsignificant in both model-2 (HR: 0.98 (0.77-1.25), and HR: 0.99 (0.79-1.23)) and model 3 (HR: 0.98 (0.77-1.24) and HR: 0.98 (0.79-1.22)). Older women with sarcopenia have an increased all-cause mortality risk independent of obesity.", "Slow gait speed may be associated with premature mortality, cardiovascular disease (CVD), and cancer, although a comprehensive meta-analysis is lacking. In this systematic review and meta-analysis, we explored potential associations between gait speed and mortality, incident CVD, and cancer. A systematic search in major databases was undertaken from inception until March 15, 2018 for prospective cohort studies reporting data on gait speed and mortality, incident CVD, and cancer. All available. The adjusted hazard ratios (HRs) and 95% confidence intervals (CIs), based on the model with the maximum number of covariates for each study between gait speed (categorized as decrease in 0.1 m/s) and mortality, incident CVD, and cancer, were meta-analyzed with a random effects model. Among 7026 articles, 44 articles corresponding to 48 independent cohorts were eligible. The studies followed up on a total of 101,945 participants (mean age 72.2 years; 55% women) for a median of 5.4 years. After adjusting for a median of 9 potential confounders and the presence of publication bias, each reduction of 0.1 m/s in gait speed was associated with a 12% increased risk of earlier mortality (45 studies; HR = 1.12, 95% CI: 1.09-1.14; I2 = 90%) and 8% increased risk of CVD (13 studies; HR = 1.08, 95% CI: 1.03-1.13; I2 = 81%), but no relationship with cancer was observed (HR = 1.00, 95% CI: 0.97-1.04; I2 = 15%). Slow gait speed may be a predictor of mortality and CVD in older adults. Because gait speed is a quick and inexpensive measure to obtain, our study suggests that it should be routinely used and may help identify people at risk of premature mortality and CVD.", "Sarcopenia is a risk-factor for all-cause mortality among older adults, but it is unknown if sarcopenia predisposes older adults to specific causes of death. Further, it is unknown if the prognostic role of sarcopenia differs between males and females, and obese and non-obese individuals. A population-based cohort study among 4425 older adults from the Third National Health and Nutrition Survey (1988-1994). Muscle mass was quantified using bioimpedance analysis, and muscle function was quantified using gait speed. Multivariable-adjusted Cox regression analysis examined the relationship between sarcopenia and mortality outcomes. The mean age of study participants was 70.1 years. The prevalence of sarcopenia was 36.5%. Sarcopenia associated with an increased risk of all-cause mortality [hazard ratio (HR): 1.29 (95% confidence interval (95% CI): 1.13-1.47); P < 0.001] among males and females. Sarcopenia associated with an increased risk of cardiovascular-specific mortality among females [HR: 1.61 (95% CI: 1.22-2.12); P = 0.001], but not among males [HR: 1.07 (95% CI: 0.81-1.40; P = .643); P interaction = 0.079]. Sarcopenia was not associated with cancer-specific mortality among males and females [HR: 1.07 (95% CI: 0.78-1.89); P = 0.672]. Sarcopenia associated with an increased risk of mortality from other causes (i.e. non-cardiovascular and non-cancer) among males and females [HR: 1.32 (95% CI: 1.07-1.62); P = 0.008]. Obesity, defined using body mass index (P interaction = 0.817) or waist circumference (P interaction = 0.219) did not modify the relationship between sarcopenia and all-cause mortality. Sarcopenia is a prevalent syndrome that is associated with premature mortality among community-dwelling older adults. The prognostic value of sarcopenia may vary by cause-specific mortality and differ between males and females.", "In older adults reduced mobility is common and is an independent risk factor for morbidity, hospitalization, disability, and mortality. Limited evidence suggests that physical activity may help prevent mobility disability; however, there are no definitive clinical trials examining whether physical activity prevents or delays mobility disability. To test the hypothesis that a long-term structured physical activity program is more effective than a health education program (also referred to as a successful aging program) in reducing the risk of major mobility disability. The Lifestyle Interventions and Independence for Elders (LIFE) study was a multicenter, randomized trial that enrolled participants between February 2010 and December 2011, who participated for an average of 2.6 years. Follow-up ended in December 2013. Outcome assessors were blinded to the intervention assignment. Participants were recruited from urban, suburban, and rural communities at 8 centers throughout the United States. We randomized a volunteer sample of 1635 sedentary men and women aged 70 to 89 years who had physical limitations, defined as a score on the Short Physical Performance Battery of 9 or below, but were able to walk 400 m. Participants were randomized to a structured, moderate-intensity physical activity program (n = 818) conducted in a center (twice/wk) and at home (3-4 times/wk) that included aerobic, resistance, and flexibility training activities or to a health education program (n = 817) consisting of workshops on topics relevant to older adults and upper extremity stretching exercises. The primary outcome was major mobility disability objectively defined by loss of ability to walk 400 m. Incident major mobility disability occurred in 30.1% (246 participants) of the physical activity group and 35.5% (290 participants) of the health education group (hazard ratio [HR], 0.82 [95% CI, 0.69-0.98], P = .03).Persistent mobility disability was experienced by 120 participants (14.7%) in the physical activity group and 162 participants (19.8%) in the health education group (HR, 0.72 [95% CI, 0.57-0.91]; P = .006). Serious adverse events were reported by 404 participants (49.4%) in the physical activity group and 373 participants (45.7%) in the health education group (risk ratio, 1.08 [95% CI, 0.98-1.20]). A structured, moderate-intensity physical activity program compared with a health education program reduced major mobility disability over 2.6 years among older adults at risk for disability. These findings suggest mobility benefit from such a program in vulnerable older adults. clinicaltrials.gov Identifier: NCT01072500.", "Sarcopenia is associated with increased risk of adverse outcomes in older people. Aim of the study was to explore the predictive value of the European Working Group on Sarcopenia in Older People (EWGSOP) diagnostic algorithm in terms of disability, hospitalization, and mortality and analyze the specific role of grip strength and walking speed as diagnostic criteria for sarcopenia. Longitudinal analysis of 538 participants enrolled in the InCHIANTI study. Sarcopenia was defined as having low muscle mass plus low grip strength or low gait speed (EWGSOP criteria). Muscle mass was assessed using bioimpedance analysis. Cox proportional and logistic regression models were used to assess risk of death, hospitalization, and disability for sarcopenic people and to investigate the individual contributions of grip strength and walking speed to the predictive value of the EWGSOP's algorithm. Prevalence of EWGSOP-defined sarcopenia at baseline was 10.2%. After adjusting for potential confounders, sarcopenia was associated with disability (odds ratio 3.15; 95% confidence interval [CI] 1.41-7.05), hospitalization (hazard ratio [HR] 1.57; 95% CI 1.03-2.41), and mortality (HR 1.88; 95% CI 0.91-3.91). The association between an alternative sarcopenic phenotype, defined only by the presence of low muscle mass and low grip strength, and both disability and mortality were similar to the association with the phenotypes defined by low muscle mass and low walking speed or by the EWGSOP algorithm. The EWGSOP's phenotype is a good predictor of incident disability, hospitalization and death. Assessment of only muscle weakness, in addition to low muscle mass, provided similar predictive value as compared to the original algorithm." ]
Microbiology and immunomics in male infertility	Up to 50% of infertility is caused by the male side. Varicocele, orchitis, prostatitis, oligospermia, asthenospermia, and azoospermia are common causes of impaired male reproductive function and male infertility. In recent years, more and more studies have shown that microorganisms play an increasingly important role in the occurrence of these diseases. This review will discuss the microbiological changes associated with male infertility from the perspective of etiology, and how microorganisms affect the normal function of the male reproductive system through immune mechanisms. Linking male infertility with microbiome and immunomics can help us recognize the immune response under different disease states, providing more targeted immune target therapy for these diseases, and even the possibility of combined immunotherapy and microbial therapy for male infertility.	[ "A total of 93 infertile and 70 fertile men attending various urology and gynecology clinics in Jordan were investigated in this prospective study. First void urine and the corresponding semen specimens were collected from 96% of the patients. Presence of Neisseria gonorrhoeae (NG), Chlamydia trachomatis (CT), Ureaplasma urealyticum (UU), and Mycoplasma genitalium (MG) DNA in specimen was detected using polymerase chain reaction. The distribution of NG, CT, UU, and MG in semen and FVU specimens among infertile versus fertile men was 6.5% versus 0%, 4.3% versus 1.4%, 10.8% versus 5.7%, and 3.2% versus 1.4%, respectively. Two of infertile and 1 of fertile men harbored mixed pathogens. The highest number of positive potential pathogens was found among young men aged 20-29 years old. The present study found a very high concordance between the detection of CT, UU, and MG DNA in semen and the corresponding FVU specimens, while NG DNA found only in semen and not in the corresponding FVU specimens. This study also revealed that Ureaplasma parvum species is more prevalent than Ureaplasma urealyticum in specimens of infertile men (90%). The study demonstrates that infertile men have higher prevalence of NG, CT, UU, and MG compared with fertile men and NG as significantly associated with infertile men.", "Testicular germ cell tumours of young adults, seminoma or non-seminomas, are preceded by a pre-invasive precursor, carcinoma in situ (CIS), understood to arise through differentiation arrest of embryonic germ cells. Knowledge about the malignant transformation of germ cells is currently limited by the lack of experimental models. The aim of this study was to establish an experimental tissue culture model to maintain normal and malignant germ cells within their niche and allow investigation of treatment effects. Human testis and testis cancer specimens from orchidectomies were cultured in 'hanging drops' and effects of activin A and follistatin treatment were investigated in seminoma cultures. Testis fragments with normal spermatogenesis or CIS cells were cultured for 14 days with sustained proliferation of germ cells and CIS cells and without increased apoptosis. Seminoma cultures survived 7 days, with proliferating cells detectable during the first 5 days. Activin A treatment significantly reduced KIT transcript and protein levels in seminoma cultures, thereby demonstrating a specific treatment response. Hanging drop cultures of human testis and testis cancer samples can be employed to delineate mechanisms governing growth of normal, CIS and tumorigenic germ cells retained within their niche.", "Acute and chronic infections of the seminal tract are among the most common causes of male infertility. As at least half of male infertility cases are classified as idiopathic, some of these cases might be attributed to asymptomatic infection. The detection and quantification of Epstein-Barr virus (EBV), cytomegalovirus (CMV) and human herpes virus type 6 (HHV-6) DNA in semen samples were performed. A total of 232 patients were divided into five groups: (i) infertile men with varicocoele; (ii) men with idiopathic infertility; (iii) infertile men with chronic inflammatory urogenital tract diseases (IUTD); (iv) fertile men with IUTD and (v) men whose partners had a history of pregnancy loss. In the study population, the prevalence of viral DNA was 17.7, 3.4% for EBV, 5.2% for CMV, 6.5% for HHV-6, 0.43% for EBV + CMV, 0.87% for EBV + HHV-6 and 1.3% for CMV + HHV-6. The median viral loads for EBV, CMV and HHV-6 were 500, 2250 and 250 copies/mL respectively. Of the sperm cell fractions, derived from infected samples 87.5% contained viral DNA. No association between EBV and fertility disorders or IUTD was found. CMV detection was much higher in the group of patients with infertility and concomitant IUTD compared with the other groups combined (18.5% vs. 5.4%, p = 0.03) and associated with reduced sperm cell count (39.5 × 10(6) /mL vs. 72.5 × 10(6) /mL, p = 0.036). Immunostaining of spermatozoa from infected samples and in vitro-infected cells detected CMV in sperm heads, tails and connecting pieces and revealed attachment to sperm membrane and intracellular localization. HHV-6 was the more common in fertile men with chronic IUTD than in the other groups combined (19% vs. 6.3%, p = 0.018) and had no effect on sperm parameters. The results suggest that both CMV and HHV-6 may contribute to the aetiology of IUTD and, moreover, CMV-associated IUTD can lead to male sterility.", "The testis is considered an immunologically privileged site where germ cell antigens are protected from autoimmune attack. Yet in response to infections, inflammatory diseases, or trauma, there is an influx of leukocytes to testicular interstitium. Interactions between endothelial cells (EC) and circulating leukocytes are implicated in the initiation and evolution of inflammatory processes. Chemokines are a family of chemoattractant cytokines characterized by their ability to both recruit and activate cells. Thus, we investigated the expression of CCL3, its receptors, and adhesion molecules CD31 and CD106 in an in vivo model of experimental autoimmune orchitis (EAO). In EAO, the highest content of CCL3 in testicular fluid coincides with onset of the disease. However, CCL3 released in vitro by testicular macrophages is higher during the immunization period. The specific chemokine receptors, CCR1 and CCR5, were expressed by testicular monocytes/macrophages and an increased number of CCR5+ cells was associated with the degree of testicular lesion. EC also play an essential role by facilitating leukocyte recruitment via their ability to express cell surface adhesion molecules that mediate interactions with leukocytes in the bloodstream. Rats with EAO showed a significant increase in the percentage of CD31+ EC that upregulate the expression of CD106. The percentage of leukocytes isolated from peripheral blood and lymph nodes expressing CD49d (CD106 ligand) also increases during orchitis. These data suggest that cell adhesion molecules, in conjunction with chemokines, contribute to the formation of a chemotactic gradient within the testis, causing the leukocyte infiltration characteristic of EAO histopathology.", "The incidence of mononuclear cell (MNC) infiltrates was studied in the testes of the following: 45 patients with carcinoma in situ (CIS) as the only lesion in the testis, CIS accompanied by an early invasion of CIS or CIS accompanied by invasive germ cell tumour (GCT) of the testis; 100 men and 146 boys who had died suddenly and unexpectedly, and 100 infertility patients. The results suggest that (1) the incidence of MNC infiltration increases with increasing severity of testicular malignant changes, (2) increased MNC infiltration is also evident in the contralateral testis where no malignant cells can be observed, (3) the incidence of MNC infiltration in men who died suddenly is not different from that in men who have had their testes biopsied because of infertility, and (4) the incidence of MNC infiltration is high in testes of boys younger than 1 year of age.", "Spermatogenic cells express cell-specific molecules with the potential to be seen as \"foreign\" by the immune system. Owing to the time difference between their appearance in puberty and the editing of the lymphocyte repertoire around birth, local adaptations of the immune system coined immune privilege are required to confer protection from autoattack. Testicular macrophages (TM) play an important role in maintaining testicular immune privilege and display reduced proinflammatory capacity compared with other macrophages. However, the molecular mechanism underlying this macrophage phenotype remained elusive. We demonstrate that TM have a lower constitutive expression of TLR pathway-specific genes compared with peritoneal macrophages. Moreover, in TM stimulated with LPS, the NF-κB signaling pathway is blocked due to lack of IκBα ubiquitination and, hence, degradation. Instead, challenge of TM with LPS or polyinosinic-polycytidylic acid induces MAPK, AP-1, and CREB signaling pathways, which leads to production of proinflammatory cytokines such as TNF-α, although at much lower levels than in peritoneal macrophages. Pretreatment of TM with inhibitors for MAPKs p38 and ERK1/2 suppresses activation of AP-1 and CREB signaling pathways and attenuates LPS-induced TNF-α and IL-10 secretion. High levels of IL-10 production and activation of STAT3 by LPS stimulation in TM indicate a regulatory macrophage phenotype. Our results suggest that TM maintain testicular immune privilege by inhibiting NF-κB signaling through impairment of IκBα ubiquitination and a general reduction of TLR cascade gene expression. However, TM do maintain some capacity for innate immune responses through AP-1 and CREB signaling pathways.", "Whether Ureaplasma spp. are a causative agent of male infertility remains controversial. Previous studies concerning Ureaplasma spp. and male infertility have been confined to the species level of Ureaplasma. Currently, an expanded multilocus sequence typing (eMLST) scheme has been established with high discriminatory power. The aim of this study was to use eMLST to explore the distribution of Ureaplasma spp. and to analyze its role in oligozoospermia and semen quality. A total of 480 semen samples were obtained from Chinese infertile males. The associations between Ureaplasma spp. with oligozoospermia and semen characteristics were further evaluated. Phylogenetic analysis revealed that 102 Ureaplasma spp. could be separated into two clusters and seven sub-groups. Within cluster I (U. parvum), eST16 and eST41 were the most frequent clones. For cluster II (U. urealyticum), eST82 and eST147 were the most prevalent clones. Sub-groups A and C belonging to cluster I and sub-group 1 belonging to cluster II showed an association with oligozoospermia, in contrast with the Ureaplasma spp. negative group (P < 0.05). Compared with the negative group, semen motility decreased in sub-group 2, especially for non-progressive motility (P < 0.05). These results indicated that sub-groups A and C belonging to cluster I (U. parvum) and sub-group 1 belonging to cluster II (U. urealyticum) were shown to be associated with oligozoospermia. Sub-group 2 belonging to cluster II may have the ability to impair semen motility, especially for non-progressive motility.", "The authors have investigated lymphocyte subpopulations and macrophages in normal human testes and the testes of patients under investigation and treatment for subfertility. Specific monoclonal antibodies were used in an indirect immunoperoxidase technique. In normal tissues, T lymphocytes (Leu 4-positive cells) were present in the rete testis with a preponderance of cells of the suppressor/cytotoxic phenotype. In contrast, no lymphocytes were detected within the peripheral portions of the testis. Cells reacting with the anti-Leu M3 monoclonal antibody, which defines monocytes/macrophages, were detected in appreciable numbers in peripheral testis with a specific location around the seminiferous tubules. HLA-DR-positive cells (human leukocyte antigens--class II [DR] determinants of the major histocompatibility complex) also were identified and showed a similar pattern of distribution to that of the Leu-M3 positive cells. While no lymphocytes were seen in the normal peripheral testis, T lymphocytes were detected in testicular biopsies from subfertile patients. Suppressor/cytotoxic T cells (Leu 2a-positive) predominated in patients with oligozoospermia and obstructive azoospermia while T cells of the helper/inducer phenotype predominated in patients with unilateral testicular obstruction and in postvasectomy patients. Sperm antibody measurements correlated with these findings.", "Experimental autoimmune epididymo-orchitis (EAEO) is a model of chronic inflammation, induced by immunisation with testicular antigens, which reproduces the pathology of some types of human infertility. Activins A and B regulate spermatogenesis and steroidogenesis, but are also pro-inflammatory, pro-fibrotic cytokines. Expression of the activins and their endogenous antagonists, inhibin and follistatin, was examined in murine EAEO. Adult untreated and adjuvant-treated control mice showed no pathology. All mice immunised with testis antigens developed EAEO by 50 days, characterised by loss of germ cells, immune cell infiltration and fibrosis in the testis, similar to biopsies from human inflamed testis. An increase of total CD45+ leukocytes, comprising CD3+ T cells, CD4 + CD8- and CD4 + CD25+ T cells, and a novel population of CD4 + CD8+ double positive T cells was also detected in EAEO testes. This was accompanied by increased expression of TNF, MCP-1 and IL-10. Activin A and B and follistatin protein levels were elevated in EAEO testes, with peak activin expression during the active phase of the disease, whereas mRNA expression of the inhibin B subunits (Inha and Inhbb) and activin receptor subunits (Acvr1b and Acvr2b) were downregulated. These data suggest that activin-follistatin regulation may play a role during the development of EAEO.", "There are currently no sensitive and specific assays for activin B that could be utilized to study human biological fluids. The aim of this project was to develop and validate a 'total' activin B ELISA for use with human biological fluids and establish concentrations of activin B in the circulation and fluids from the reproductive organs. The new ELISA was validated and then used to measure activin B levels in the circulation of healthy participants, IVF patients, pregnant women and in ovarian follicular fluid and seminal plasma. Healthy adult subjects (n = 143), subjects from an IVF clinic (n = 27) and pregnancy groups (n = 29) were sampled. The sensitivity of the assay was 0.019 ng/ml. Validation of the activin B ELISA showed good recovery (90.7 +/- 9.8%) and linearity in biological fluid and cell culture media and low cross-reactivity with related analytes (inhibin B = 0.077% and activin A = 0.0034%). There was a negative correlation between activin B concentration (r = -0.281, P < 0.011) and females with increasing age. Patients attending IVF clinics had significantly lower levels of activin B compared with gender-matched control subjects. Ovarian follicular fluid and seminal plasma had 50-80 fold higher levels of activin B (mean = 5.35 and 3.66 ng/ml respectively) than sera (mean = 0.071 ng/ml). This fully validated ELISA for activin B offers a tremendous utility for measuring this protein in a variety of normal physiological processes and in various clinical pathologies." ]
Trimetallic Dual-Atom Alloys: A New Class of Dual-Atom Catalysts	Single-atom catalysts have received significant attention for their ability to enable highly selective reactions. However, many reactions require more than one adjacent site to align reactants or break specific bonds. For example, breaking a C-O or O-H bond may be facilitated by a dual site containing an oxophilic element and a carbophilic or "hydrogenphilic" element that binds each molecular fragment. However, design of stable and well-defined dual-atom sites with desirable reactivity is difficult due to the complexity of multicomponent catalytic surfaces. Here, we describe a new type of dual-atom system, trimetallic dual-atom alloys, which were designed via computation of the alloying energetics. Through a broad computational screening we discovered that Pt-Cr dimers embedded in Ag(111) can be formed by virtue of the negative mixing enthalpy of Pt and Cr in Ag and the favorable interaction between Pt and Cr. These dual-atom alloy sites were then realized experimentally through surface science experiments that enabled the active sites to be imaged and their reactivity related to their atomic-scale structure. Specifically, Pt-Cr sites in Ag(111) can convert ethanol, whereas PtAg and CrAg are unreactive toward ethanol. Calculations show that the oxophilic Cr atom and the hydrogenphilic Pt atom act synergistically to break the O-H bond. Furthermore, ensembles with more than one Cr atom, present at higher dopant loadings, produce ethylene. Our calculations have identified many other thermodynamically favorable dual-atom alloy sites, and hence this work highlights a new class of materials that should offer new and useful chemical reactivity beyond the single-atom paradigm.	[ "Single-atom alloys can be effective catalysts and have been compared to supported single-atom catalysts. To rationally design single-atom alloys and other surfaces with localized ensembles, it is crucial to understand variations in reactivity when varying the dopant and the ensemble size. Here, we examined hydrogen adsorption on surfaces embedded with localized clusters and discovered general trends. Counterintuitively, increasing the amount of a more reactive metal sometimes makes a surface site less reactive. This behavior is due to the hybridization and splitting of narrow peaks in the electronic density of states of many of these surfaces, making them analogous to free-standing nanoclusters. When a single-atom alloy has a peak just below the Fermi energy, the corresponding two-dopant cluster often has weaker adsorption than the single-atom alloy due to splitting of this peak across the Fermi energy. Furthermore, single-atom alloys have qualitatively different behaviors than larger ensembles. Specifically, the adsorption energy is a U-shaped function of the dopant's group for single-atom alloys. Additionally, adsorption energies on single-atom alloys correlate more strongly with the dopant's p-band center than with the d-band center.", "For the large-scale sustainable implementation of polymer electrolyte membrane fuel cells in vehicles, high-performance electrocatalysts with low platinum consumption are desirable for use as cathode material during the oxygen reduction reaction in fuel cells. Here we report a carbon black-supported cost-effective, efficient and durable platinum single-atom electrocatalyst with carbon monoxide/methanol tolerance for the cathodic oxygen reduction reaction. The acidic single-cell with such a catalyst as cathode delivers high performance, with power density up to 680 mW cm-2 at 80 °C with a low platinum loading of 0.09 mgPt cm-2, corresponding to a platinum utilization of 0.13 gPt kW-1 in the fuel cell. Good fuel cell durability is also observed. Theoretical calculations reveal that the main effective sites on such platinum single-atom electrocatalysts are single-pyridinic-nitrogen-atom-anchored single-platinum-atom centres, which are tolerant to carbon monoxide/methanol, but highly active for the oxygen reduction reaction.", "Developing efficient catalysts for nitrogen fixation is becoming increasingly important but is still challenging due to the lack of robust design criteria for tackling the activity and selectivity problems, especially for electrochemical nitrogen reduction reaction (NRR). Herein, by means of large-scale density functional theory (DFT) computations, we reported a descriptor-based design principle to explore the large composition space of two-dimensional (2D) biatom catalysts (BACs), namely, metal dimers supported on 2D expanded phthalocyanine (M2-Pc or MM'-Pc), toward the NRR at the acid conditions. We sampled both homonuclear (M2-Pc) and heteronuclear (MM'-Pc) BACs and constructed the activity map of BACs by using N2H* adsorption energy as the activity descriptor, which reduces the number of promising catalyst candidates from over 900 to less than 100. This strategy allowed us to readily identify 3 homonuclear and 28 heteronuclear BACs, which could break the metal-based activity benchmark toward the efficient NRR. Particularly, using the free energy difference of H* and N2H* as a selectivity descriptor, we screened out five systems, including Ti2-Pc, V2-Pc, TiV-Pc, VCr-Pc, and VTa-Pc, which exhibit a strong capability of suppressing the competitive hydrogen evolution reaction (HER) with favorable limiting potential of -0.75, -0.39, -0.74, -0.85, and -0.47 V, respectively. This work not only broadens the possibility of discovering more efficient BACs toward N2 fixation but also provides a feasible strategy for rational design of NRR electrocatalysts and helps pave the way to fast screening and design of efficient BACs for the NRR and other electrochemical reactions.", "A combination of interpolation methods and local saddle-point search algorithms is probably the most efficient way of finding transition states in chemical reactions. Interpolation methods such as the growing-string method and the nudged-elastic band are able to find an approximation to the minimum-energy pathway and thereby provide a good initial guess for a transition state and imaginary mode connecting both reactant and product states. Since interpolation methods employ usually just a small number of configurations and converge slowly close to the minimum-energy pathway, local methods such as partitioned rational function optimization methods using either exact or approximate Hessians or minimum-mode-following methods such as the dimer or the Lanczos method have to be used to converge to the transition state. A modification to the original dimer method proposed by [Henkelman and Jonnson J. Chem. Phys. 111, 7010 (1999)] is presented, reducing the number of gradient calculations per cycle from six to four gradients or three gradients and one energy, and significantly improves the overall performance of the algorithm on quantum-chemical potential-energy surfaces, where forces are subject to numerical noise. A comparison is made between the dimer methods and the well-established partitioned rational function optimization methods for finding transition states after the use of interpolation methods. Results for 24 different small- to medium-sized chemical reactions covering a wide range of structural types demonstrate that the improved dimer method is an efficient alternative saddle-point search algorithm on medium-sized to large systems and is often even able to find transition states when partitioned rational function optimization methods fail to converge.", "Heterogeneous catalysts are workhorses in the industrial production of most commodity and specialty chemicals, and have widespread energy and environmental applications, with the annual market value of the catalysts themselves reaching almost $20 billion in 2018. These catalysts are complex, comprising multicomponent materials and multiple structures, making their rational design challenging, if not impossible. Furthermore, typical active metals like Pt, Pd, and Rh are expensive and can be susceptible to poisoning by CO, coking, and they are not always 100% selective. Efforts to use these elements sparingly and improve their selectivity has led to recent identification of single-atom heterogeneous catalysts in which individual transition metal atoms anchored on oxide or carbon-based supports are excellent catalysts for reactions like the CO oxidation, water-gas shift, alcohol dehydrogenation, and steam reforming. In this Account, we describe a new class of single-atom heterogeneous catalysts, namely, Single-Atom Alloys (SAAs) that comprise catalytically active elements like Pt, Pd, and Ni alloyed in more inert host metals at the single-atom limit. These materials evolved by complementary surface science and scanning probe studies using single crystals, and catalytic evaluation of the corresponding alloy nanoparticles with compositions informed by the surface science findings. The well-defined nature of the active sites in SAAs makes accurate modeling with theory relatively easy, enabling the rational design of SAA catalysts via a complementary three-prong approach, encompassing surface science model catalysts, theory, and real catalyst synthesis and testing under industrially relevant conditions. SAAs constitute one of just a few examples of when heterogeneous catalyst design has been guided by an understanding of fundamental surface processes. The Account starts by describing scanning tunneling microscopy studies of highly dilute alloys formed by doping small amounts of a catalytically active element into a more inert host metal. We first discuss hydrogenation reactions in which dissociation of H2 is often rate limiting. Results indicate how the SAA geometry allows the transition state and the binding site of the reaction intermediates to be decoupled, which enables both facile dissociation of reactants and weak binding of intermediates, two key factors for efficient and selective catalysis. These results were exploited to design the first PtCu SAA hydrogenation catalysts which showed high selectivity, stability and resistance to poisoning in industrially relevant hydrogenation reactions, such as the selective conversion of butadiene to butenes. Model studies also revealed spillover of hydrogen atoms from the Pt site where dissociation of H2 occurs to Cu sites where selective hydrogenation is facilitated in a bifunctional manner. We then discuss selective dehydrogenations on SAAs demonstrating that they enable efficient C-H activation, while being resistant to coking that plagues typical Pt catalysts. SAA PtCu nanoparticle catalysts showed excellent stability in butane dehydrogenation for days-on-stream at 400 °C. Another advantage of SAA catalysts is that on many alloy combinations CO, a common catalyst poison, binds more weakly to the alloy than the pure metal. We conclude by discussing recent theory results that predict the energetics of many key reaction steps on a wide range of SAAs and the exciting possibilities this reductionist approach to heterogeneous catalysis offers for the rational design of new catalysts.", "Single atom catalysts exhibit particularly high catalytic activities in contrast to regular nanomaterial-based catalysts. Until recently, research has been mostly focused on single atom catalysts, and it remains a great challenge to synthesize bimetallic dimer structures. Herein, we successfully prepare high-quality one-to-one A-B bimetallic dimer structures (Pt-Ru dimers) through an atomic layer deposition (ALD) process. The Pt-Ru dimers show much higher hydrogen evolution activity (more than 50 times) and excellent stability compared to commercial Pt/C catalysts. X-ray absorption spectroscopy indicates that the Pt-Ru dimers structure model contains one Pt-Ru bonding configuration. First principle calculations reveal that the Pt-Ru dimer generates a synergy effect by modulating the electronic structure, which results in the enhanced hydrogen evolution activity. This work paves the way for the rational design of bimetallic dimers with good activity and stability, which have a great potential to be applied in various catalytic reactions." ]
Obesity Paradox and Breast Cancer	Breast cancer is the most commonly occurring cancer among women. The relationship between the obesity paradox and breast cancer is still unclear. The goal of this study is to elucidate the association between high body mass index (BMI) and pathological findings by age.	[ "The root cause of non-inherited Alzheimer's disease (AD) remains unknown despite hundreds of research studies performed to attempt to solve this problem. Since proper prophylaxis remains the best strategy, many scientists have studied the risk factors that may affect AD development. There is robust evidence supporting the hypothesis that cardiovascular diseases (CVD) may contribute to AD progression, as the diseases often coexist. Therefore, a lack of well-defined diagnostic criteria makes studying the relationship between AD and CVD complicated. Additionally, inflammation accompanies the pathogenesis of AD and CVD, and is not only a consequence but also implicated as a significant contributor to the course of the diseases. Of note, АроЕε4 is found to be one of the major risk factors affecting both the cardiovascular and nervous systems. According to genome wide association and epidemiological studies, numerous common risk factors have been associated with the development of AD-related pathology. Furthermore, the risk of developing AD and CVDs appears to be increased by a wide range of conditions and lifestyle factors: hypertension, dyslipidemia, hypercholesterolemia, hyperhomocysteinemia, gut/oral microbiota, physical activity, and diet. This review summarizes the literature and provides possible mechanistic links between CVDs and AD.", "Many studies suggest increased body mass index (BMI) is associated with worse breast cancer outcomes, but few account for variability in screening, access to treatment, and tumor differences. We examined the association between BMI and risk of breast cancer recurrence, breast cancer-specific mortality, and all-cause mortality, and evaluated whether tumor characteristics differ by BMI among a mammographically screened population with access to treatment. Using a retrospective cohort study design, we followed 485 women aged ≥40 years diagnosed with stage I/II breast cancer within 24 months of a screening mammogram occurring between 1988 and 1993 for 10-year outcomes. BMI before diagnosis was categorized as normal (<25 kg/m(2)), overweight (25-29.9 kg/m(2)), and obese (≥30 kg/m(2)). Tumor marker expression was assessed via immunohistochemistry using tissue collected before adjuvant treatment. Medical records were abstracted to identify treatment, recurrence, and mortality. We used Cox proportional hazards to separately model the hazard ratios (HR) of our three outcomes by BMI while adjusting for age, stage, and tamoxifen use. Relative to normal-weight women, obese women experienced increased risk of recurrence (HR 2.43; 95 % CI 1.34-4.41) and breast cancer death (HR 2.41; 95 % CI 1.00-5.81) within 10 years of diagnosis. There was no association between BMI and all-cause mortality. Obese women had significantly faster growing tumors, as measured by Ki-67. Our findings add to the growing evidence that obesity may contribute to poorer breast cancer outcomes, and also suggest that increased tumor proliferation among obese women is a pathway that explains part of their excess risk of adverse outcomes.", "Molecular epidemiological studies have shown that gene polymorphisms of estrogen receptor alpha gene (ESR-α) are associated with breast cancer risk. However, previous results from many molecular studies have been inconsistent. In this study, we examined two polymorphisms (PvuII and XbaI RFLPs) of the ESR-α gene in 542 breast cancer cases and 1,016 controls from China. Associations between the polymorphisms and breast cancer risk were calculated with an unconditional logistic regression model. Linkage disequilibrium and haplotypes were analyzed with the SHEsis software. In addition, we also performed a systematic meta-analysis of 24 published studies evaluating the association. No significant associations were found between the PvuII polymorphism and breast cancer risk. However, a significantly decreased risk of breast cancer was observed among carriers of the XbaI 'G' allele (age-adjusted OR = 0.80; 95% CI = 0.66- 0.97) compared with carriers of the 'A' allele. Haplotype analysis showed significantly decreased cancer risk for carriers of the 'CG' haplotype (OR = 0.79; 95% CI = 0.66- 0.96). In the systematic meta-analysis, the XbaI 'G' allele was associated with an overall significantly decreased risk of breast cancer (OR = 0.90, 95% CI = 0.82- 1.00). In addition, the PvuII 'C' allele showed a 0.96- fold decreased disease risk (95% CI = 0.92- 0.99). In subgroup analysis, an association between the PvuII 'C' and XbaI 'G' alleles and breast cancer risk was significant in Asians ('C' vs. 'T': OR = 0.93, 95% CI = 0.85- 1.00; 'G' vs. 'A': OR = 0.82, 95% CI = 0.68- 0.98), but not in Euro-Americans. Thus, our results provide evidence that ESR-α polymorphisms are associated with susceptibility to breast cancer. These associations may largely depend on population characteristics and geographic location.", "The aim of this study was to describe and analyze the incidence and mortality of female breast cancer in Jiangsu Province of China. Incidence and mortality data for female breast cancer and corresponding population statistics from eligible cancer registries in Jiangsu from 2006 to 2010 were collected and analyzed. Crude rates, age-specific rates and age-standardized rates of incidence and mortality were calculated, and annual present changes (APCs) were estimated to describe the time trends. From 2006 to 2010, 11,013 new cases and 3,068 deaths of female breast cancer were identified in selected cancer registry areas of Jiangsu. The annual average crude incidence and age-standardized incidence by world population (ASW) were 25.2/ and 17.9/100,000 respectively. The annual average crude and ASW for mortality rates were 7.03/ and 4.81/100,000. The incidence was higher in urban areas than that in rural areas, and this was consistent in all age groups. No significant difference was observed in mortality between urban and rural areas. Two peaks were observed when looking at age-specific rates, one at 50-59 years and another at over 85 years. During the 5 years, incidence and mortality increased with APCs of 4.47% and 6.89%, respectively. Compared to the national level, Jiangsu is an area with relatively low risk of female breast cancer. Breast cancer has become a main public health problem among Chinese females. More prevention and control activities should be conducted to reduce the burden of this disease, even in relatively low risk areas like Jiangsu.", "We investigated the role of common genetic variation in immune-related genes on breast cancer disease-free survival (DFS) in Korean women. 107 breast cancer patients of the Seoul Breast Cancer Study (SEBCS) were selected for this study. A total of 2,432 tag single nucleotide polymorphisms (SNPs) in 283 immune-related genes were genotyped with the GoldenGate Oligonucleotide pool assay (OPA). A multivariate Cox-proportional hazard model and polygenic risk score model were used to estimate the effects of SNPs on breast cancer prognosis. Harrell's C index was calculated to estimate the predictive accuracy of polygenic risk score model. Subsequently, an extended gene set enrichment analysis (GSEA-SNP) was conducted to approximate the biological pathway. In addition, to confirm our results with current evidence, previous studies were systematically reviewed. Sixty-two SNPs were statistically significant at p-value less than 0.05. The most significant SNPs were rs1952438 in SOCS4 gene (hazard ratio (HR) = 11.99, 95% CI = 3.62-39.72, P = 4.84E-05), rs2289278 in TSLP gene (HR = 4.25, 95% CI = 2.10-8.62, P = 5.99E-05) and rs2074724 in HGF gene (HR = 4.63, 95% CI = 2.18-9.87, P = 7.04E-05). In the polygenic risk score model, the HR of women in the 3rd tertile was 6.78 (95% CI = 1.48-31.06) compared to patients in the 1st tertile of polygenic risk score. Harrell's C index was 0.813 with total patients and 0.924 in 4-fold cross validation. In the pathway analysis, 18 pathways were significantly associated with breast cancer prognosis (P<0.1). The IL-6R, IL-8, IL-10RB, IL-12A, and IL-12B was associated with the prognosis of cancer in data of both our study and a previous study. Therefore, our results suggest that genetic polymorphisms in immune-related genes have relevance to breast cancer prognosis among Korean women.", "Background. To evaluate whether insulin use was predictive for mortality from breast cancer in Taiwanese women with diabetes mellitus. Methods. A total of 48,880 diabetic women were followed up to determine the mortality from breast cancer during 1995-2006. Cox models were used, considering the following independent variables: age, sex, diabetes type, diabetes duration, body mass index, smoking, insulin use, and area of residence. Insulin use was also considered for its duration of use at cutoffs of 3 years and 5 years. Results. Age was a significant predictor in all analyses. The multivariable-adjusted hazard ratio (95% confidence interval, P value) for insulin use without considering the duration of use was not statistically significant (1.339 [0.782-2.293, P = 0.2878]). Compared with nonusers, insulin users showed the following adjusted hazard ratios for insulin use <3 years, ≥3 years, <5 years, and ≥5 years: 0.567 (0.179-1.791, P = 0.3333), 2.006 (1.102-3.653, P = 0.0228), 1.045 (0.505-2.162, P = 0.9048), and 1.899 (0.934-3.860, P = 0.0763). Conclusions. Insulin use (mainly human insulin) for ≥3 years may be associated with a higher risk of breast cancer mortality.", "Nitric oxide- (NO-) dependent oxidative stress results in mitochondrial ultrastructural alterations and DNA damage in cases of Alzheimer disease (AD). However, little is known about these pathways in human cancers, especially during the development as well as the progression of primary brain tumors and metastatic colorectal cancer. One of the key features of tumors is the deficiency in tissue energy that accompanies mitochondrial lesions and formation of the hypoxic smaller sized mitochondria with ultrastructural abnormalities. We speculate that mitochondrial involvement may play a significant role in the etiopathogenesis of cancer. Recent studies also demonstrate a potential link between AD and cancer, and anticancer drugs are being explored for the inhibition of AD-like pathology in transgenic mice. Severity of the cancer growth, metastasis, and brain pathology in AD (in animal models that mimic human AD) correlate with the degree of mitochondrial ultrastructural abnormalities. Recent advances in the cell-cycle reentry of the terminally differentiated neuronal cells indicate that NO-dependent mitochondrial abnormal activities and mitotic cell division are not the only important pathogenic factors in pathogenesis of cancer and AD, but open a new window for the development of novel treatment strategies for these devastating diseases.", "Overweight and obesity represent a rapidly growing threat to the health of populations in an increasing number of countries. Indeed they are now so common that they are replacing more traditional problems such as undernutrition and infectious diseases as the most significant causes of ill-health. Obesity comorbidities include coronary heart disease, hypertension and stroke, certain types of cancer, non-insulin-dependent diabetes mellitus, gallbladder disease, dyslipidaemia, osteoarthritis and gout, and pulmonary diseases, including sleep apnoea. In addition, the obese suffer from social bias, prejudice and discrimination, on the part not only of the general public but also of health professionals, and this may make them reluctant to seek medical assistance. WHO therefore convened a Consultation on obesity to review current epidemiological information, contributing factors and associated consequences, and this report presents its conclusions and recommendations. In particular, the Consultation considered the system for classifying overweight and obesity based on the body mass index, and concluded that a coherent system is now available and should be adopted internationally. The Consultation also concluded that the fundamental causes of the obesity epidemic are sedentary lifestyles and high-fat energy-dense diets, both resulting from the profound changes taking place in society and the behavioural patterns of communities as a consequence of increased urbanization and industrialization and the disappearance of traditional lifestyles. A reduction in fat intake to around 20-25% of energy is necessary to minimize energy imbalance and weight gain in sedentary individuals. While there is strong evidence that certain genes have an influence on body mass and body fat, most do not qualify as necessary genes, i.e. genes that cause obesity whenever two copies of the defective allele are present; it is likely to be many years before the results of genetic research can be applied to the problem. Methods for the treatment of obesity are described, including dietary management, physical activity and exercise, and antiobesity drugs, with gastrointestinal surgery being reserved for extreme cases.", "Large body size has been associated with decreased risk of breast cancer in premenopausal women but with increased risk in postmenopausal women. Limited information is available about African-American women and differences by estrogen and progesterone receptor status. We analyzed data from the Women's Contraceptive and Reproductive Experiences Study among 3,997 white and African-American breast cancer case patients diagnosed in 1994 to 1998 and 4,041 control participants ages 35 to 64 years. We calculated multivariate odds ratios (OR) as measures of relative risk of breast cancer associated with self-reported body mass index (BMI) at age 18 and 5 years before diagnosis (recent BMI). Risk tended to decrease with increasing BMI at age 18 years in all women [OR(BMI > or = 25 kg/m(2) versus < 20 kg/m(2)) = 0.76; 95% confidence interval (CI), 0.63-0.90; P(trend) = 0.005] and with recent BMI in premenopausal women (OR(BMI > or = 35 kg/m(2) versus < 25 kg/m(2)) = 0.81; 95% CI, 0.61-1.06; P(trend) = 0.05), unmodified by race. Among postmenopausal white but not African-American women, there was an inverse relation between recent BMI and risk. High recent BMI was associated with increased risk of estrogen receptor- and progesterone receptor-positive tumors among postmenopausal African-American women (OR(BMI > or = 35 kg/m(2) versus < 25 kg/m(2)) = 1.83; 95% CI, 1.08-3.09; P(trend) = 0.03). Among women at age 35 to 64 years, BMI at age 18 years is inversely associated with risk of breast cancer, but association with recent BMI varies by menopause status, race, and hormone receptor status. Our findings indicate that studies of BMI and breast cancer should consider breast cancer subtypes.", "Background: Obesity and cancer are recognized worldwide health threats. While there is no reported causal relationship, the increasing frequency of both conditions results in a higher incidence of obese patients who are being treated for cancer. Physiological data indicate that there is a relationship between obesity and susceptibility to pain; however, currently, there are no specific pharmacological interventions. Objective: To evaluate the self-reported intensity of postoperative pain in obese and nonobese lung cancer who receive either thoracotomy or video-assisted thoracic surgery (VATS) surgical therapy. Material and Methods: In 50 obese [mean body mass index (BMI) of 34.1 ± 3.2 kg/m2] and 62 nonobese (mean BMI of 24.9 ± 3 kg/m2) lung cancer patients, the intensity of pain was estimated every 4 h using a visual analog scale (VAS, 0 indicating no pain and 10 indicating \"worst imaginable pain\") beginning shortly after surgery (Day O) and continuing until the day of discharge (Day D). Results: The self-reported pain was more severe in obese than in nonobese patients, both at the time of the operation [Day O (4.5 ± 1.2 vs 3.4 ± 1.1; p < 0.0001)] and at the day of discharge [Day D (3.9 ± 1.4 vs 2.6 ± 0.9, p < 0.0001)]. This finding was consistent both in the patients after thoracotomy and after video-assisted thoracic surgery (VATS, p < 0.0001). The patients with severe pain shortly after surgery (VAS score >4) had significantly higher BMI (31.8 ± 5.6 kg/m2 vs 28.8 ± 5.2 kg/m2, p < 0.01) and were hospitalized longer than the remaining patients (13.0 ± 13.6 days vs 9.5 ± 3.6 days, p < 0.05). Conclusion: The reported perception of pain in obese lung cancer patients is greater than in nonobese patients undergoing the same thoracic surgery. In obese patients, severe pain persisted longer. Pain management is an important consideration in the postoperative care of lung cancer patients, even more so with obese patients." ]
-Aminobutyric Acid and Pancreatic Islets	The pancreatic islets are essential microorgans controlling the glucose level in the blood. The islets consist of different cell types which communicate with each other by means of auto- and paracrine interactions. One of the communication molecules produced by and released within the islets is γ-aminobutyric acid (GABA), a well-known inhibitor of neuronal excitability in the mammalian nervous system. Interestingly, GABA is also present in the blood in the nanomolar concentration range. Thus, GABA can affect not only islet function	[ "γ-Aminobutyric acid (GABA) administration has been shown to increase β-cell mass, leading to a reversal of type 1 diabetes in mice. Whether GABA has any effect on β cells of healthy and prediabetic/glucose-intolerant obese mice remains unknown. In the present study, we show that oral GABA administration ( ad libitum) to mice indeed increased pancreatic β-cell mass, which led to a modest enhancement in insulin secretion and glucose tolerance. However, GABA treatment did not further increase insulin-positive islet area in high fat diet-fed mice and was unable to prevent or reverse glucose intolerance and insulin resistance. Mechanistically, whether in vivo or in vitro, GABA treatment increased β-cell proliferation. In vitro, the effect was shown to be mediated via the GABAA receptor. Single-cell RNA sequencing analysis revealed that GABA preferentially up-regulated pathways linked to β-cell proliferation and simultaneously down-regulated those networks required for other processes, including insulin biosynthesis and metabolism. Interestingly, single-cell differential expression analysis revealed GABA treatment gave rise to a distinct subpopulation of β cells with a unique transcriptional signature, including urocortin 3 ( ucn3), wnt4, and hepacam2. Taken together, this study provides new mechanistic insight into the proliferative nature of GABA but suggests that β-cell compensation associated with prediabetes overlaps with, and negates, its proliferative effects.-Untereiner, A., Abdo, S., Bhattacharjee, A., Gohil, H., Pourasgari, F., Ibeh, N., Lai, M., Batchuluun, B., Wong, A., Khuu, N., Liu, Y., Al Rijjal, D., Winegarden, N., Virtanen, C., Orser, B. A., Cabrera, O., Varga, G., Rocheleau, J., Dai, F. F., Wheeler, M. B. GABA promotes β-cell proliferation, but does not overcome impaired glucose homeostasis associated with diet-induced obesity.", "Hormone-secreting cells within pancreatic islets of Langerhans play important roles in metabolic homeostasis and disease. However, their transcriptional characterization is still incomplete. Here, we sequenced the transcriptomes of thousands of human islet cells from healthy and type 2 diabetic donors. We could define specific genetic programs for each individual endocrine and exocrine cell type, even for rare δ, γ, ε, and stellate cells, and revealed subpopulations of α, β, and acinar cells. Intriguingly, δ cells expressed several important receptors, indicating an unrecognized importance of these cells in integrating paracrine and systemic metabolic signals. Genes previously associated with obesity or diabetes were found to correlate with BMI. Finally, comparing healthy and T2D transcriptomes in a cell-type resolved manner uncovered candidates for future functional studies. Altogether, our analyses demonstrate the utility of the generated single-cell gene expression resource.", "We sought to determine the mRNA transcriptome of all major human pancreatic endocrine and exocrine cell subtypes, including human alpha, beta, duct and acinar cells. In addition, we identified the cell type-specific distribution of transcription factors, signalling ligands and their receptors. Islet samples from healthy human donors were enzymatically dispersed to single cells and labelled with cell type-specific surface-reactive antibodies. Live endocrine and exocrine cell subpopulations were isolated by FACS and gene expression analyses were performed using microarray analysis and quantitative RT-PCR. Computational tools were used to evaluate receptor-ligand representation in these populations. Analysis of the transcriptomes of alpha, beta, large duct, small duct and acinar cells revealed previously unrecognised gene expression patterns in these cell types, including transcriptional regulators HOPX and HDAC9 in the human beta cell population. The abundance of some regulatory proteins was different from that reported in mouse tissue. For example, v-maf musculoaponeurotic fibrosarcoma oncogene homologue B (avian) (MAFB) was detected at equal levels in adult human alpha and beta cells, but is absent from adult mouse beta cells. Analysis of ligand-receptor interactions suggested that EPH receptor-ephrin communication between exocrine and endocrine cells contributes to pancreatic function. This is the first comprehensive analysis of the transcriptomes of human exocrine and endocrine pancreatic cell types-including beta cells-and provides a useful resource for diabetes research. In addition, paracrine signalling pathways within the pancreas are shown. These results will help guide efforts to specify human beta cell fate by embryonic stem cell or induced pluripotent stem cell differentiation or genetic reprogramming.", "Poorly controlled diabetes is characterised by a partial or complete loss of pancreatic islet β-cells, which deprives the remaining islet cells of important β-cell-derived soluble signals, such as insulin or GABA. We aimed to dissect the role of the two signals in the development of islet α-cells, focusing specifically on α-/β-cell transdifferentiation and using the stem cell differentiation factor nicotinamide as a comparator. Streptozotocin (STZ)-treated diabetic mice expressing a fluorescent reporter in pancreatic islet α-cells were injected with GABA (10 mg/kg once daily), nicotinamide (150 mg/kg once daily) or insulin (1U/kg three times daily) for 10 days. The impact of the treatment on metabolic status of the animals as well as the morphology, proliferative potential and transdifferentiation of pancreatic islet cells was assessed using biochemical methods and immunofluorescence. Metabolic status of STZ-diabetic mice was not dramatically altered by the treatment interventions, although GABA therapy did reduce circulating glucagon and augment pancreatic insulin stores. The effects of the exogenous agents on islet β-cells ranged from the attenuation of apoptosis (insulin, nicotinamide) to enhancement of proliferation (GABA). Furthermore, insulin and GABA but not nicotinamide enhanced the differentiation of α-cells into β-cells and increased relative number of 'bihormonal' cells, expressing both insulin and glucagon. Our data suggest a role for endogenous insulin and GABA signalling in α-cell plasticity, which is likely to bypass the common nicotinamide-sensitive stem cell differentiation pathway.", "Insulin-secreting β cells and glucagon-secreting α cells maintain physiological blood glucose levels, and their malfunction drives diabetes development. Using ChIP sequencing and RNA sequencing analysis, we determined the epigenetic and transcriptional landscape of human pancreatic α, β, and exocrine cells. We found that, compared with exocrine and β cells, differentiated α cells exhibited many more genes bivalently marked by the activating H3K4me3 and repressing H3K27me3 histone modifications. This was particularly true for β cell signature genes involved in transcriptional regulation. Remarkably, thousands of these genes were in a monovalent state in β cells, carrying only the activating or repressing mark. Our epigenomic findings suggested that α to β cell reprogramming could be promoted by manipulating the histone methylation signature of human pancreatic islets. Indeed, we show that treatment of cultured pancreatic islets with a histone methyltransferase inhibitor leads to colocalization of both glucagon and insulin and glucagon and insulin promoter factor 1 (PDX1) in human islets and colocalization of both glucagon and insulin in mouse islets. Thus, mammalian pancreatic islet cells display cell-type-specific epigenomic plasticity, suggesting that epigenomic manipulation could provide a path to cell reprogramming and novel cell replacement-based therapies for diabetes.", "The role of gamma-aminobutyric acid (GABA) and A-type GABA receptors (GABA(A)Rs) in modulating islet endocrine function has been actively investigated since the identification of GABA and GABA(A)Rs in the pancreatic islets. However, the reported effects of GABA(A)R activation on insulin secretion from islet beta cells have been controversial. This study examined the hypothesis that the effect of GABA on beta cell insulin secretion is dependent on glucose concentration. Perforated patch-clamp recordings in INS-1 cells demonstrated that GABA, at concentrations ranging from 1 to 1,000 micromol/l, induced a transmembrane current (I(GABA)) which was sensitive to the GABA(A)R antagonist bicuculline. The current-voltage relationship revealed that I(GABA) reversed at -42+/-2.2 mV, independently of glucose concentration. Nevertheless, the glucose concentration critically controlled the membrane potential (V (M)), i.e., at low glucose (0 or 2.8 mmol/l) the endogenous V (M) of INS-1 cells was below the I(GABA) reversal potential and at high glucose (16.7 or 28 mmol/l), the endogenous V (M) of INS-1 cells was above the I(GABA) reversal potential. Therefore, GABA dose-dependently induced membrane depolarisation at a low glucose concentration, but hyperpolarisation at a high glucose concentration. Consistent with electrophysiological findings, insulin secretion assays demonstrated that at 2.8 mmol/l glucose, GABA increased insulin secretion in a dose-dependent fashion (p<0.05, n=7). This enhancement was blocked by bicuculline (p<0.05, n=4). In contrast, in the presence of 28 mmol/l glucose, GABA suppressed the secretion of insulin (p<0.05, n=5). These findings indicate that activation of GABA(A)Rs in beta cells regulates insulin secretion in concert with changes in glucose levels.", "The different forms of diabetes mellitus differ in their pathogenesis but, ultimately, they are all characterized by progressive islet β-cell loss. Restoring the β-cell mass is therefore a major goal for future therapeutic approaches. The number of β-cells found at birth is determined by proliferation and differentiation of pancreatic progenitor cells, and it has been considered to remain mostly unchanged throughout adult life. Recent studies in mice have revealed an unexpected plasticity in islet endocrine cells in response to stress; under certain conditions, islet non-β-cells have the potential to reprogram into insulin producers, thus contributing to restore the β-cell mass. Here, we discuss the latest findings on pancreas and islet cell plasticity upon physiological, pathological and experimental conditions of stress. Understanding the mechanisms involved in cell reprogramming in these models will allow the development of new strategies for the treatment of diabetes, by exploiting the intrinsic regeneration capacity of the pancreas." ]
Implementation of a comprehensive cardiovascular disease prevention program in general practice	Our aim was to evaluate the implementation process of a comprehensive cardiovascular disease prevention program in general practice, to enhance understanding of influencing factors to implementation success and sustainability, and to learn how to overcome barriers.	[ "A major problem in the dissemination of most interventions found to be efficacious is that they are of limited or unknown generalizability. To document the \"robustness,\" or external validity, of a computer-assisted diabetes self-management program across different patient characteristics, healthcare settings (mixed payer vs health maintenance organization), intervention staff, and outcomes. A randomized controlled trial evaluating a computer-assisted behavior change program for adult patients with type 2 diabetes mellitus (n = 217) vs a computerized health risk assessment. Outcomes were identified using the RE-AIM framework and included program adoption among physicians, reach across patient groups, implementation, and behavioral (fat intake and physical activity) and biological (glycosylated hemoglobin and lipid levels) effectiveness measures. The program achieved 41% patient participation, variable adoption across healthcare settings (76% of health maintenance organization physicians vs 18% of non-health maintenance organization physicians participated), good implementation, and improvement in behavioral outcomes. There were few significant interactions between treatment condition and patient characteristics, type of healthcare setting, or interventionist experience on effectiveness measures. Patients and physicians were willing to participate in a computer-assisted dietary and physical activity goal-setting intervention, although participation varied by healthcare setting. Interventionists from different backgrounds successfully delivered the intervention, and the results appear robust across various patient and delivery characteristics.", "The DASH diet was designed for helping control of blood pressure but, fortunately, it can also be prescribed for many other chronic conditions. The current study intended to assess the potential effects of DASH diet on metabolic risk factors in patients with chronic disease. We carried out a systematic literature search for RCTs from inception until July 2020. A total of 54 clinical trials were included in the final analysis. Compared to control groups, a significant lower effect of the DASH diet was noted for body weight (-1.59 kg; p < 0.001), BMI (-0.64 kg/m2; p < 0.001), and WC (-1.93 cm; p < 0.001) as well as for SBP (-3.94 mmHg; p < 0.001) and DBP (-2.44 mmHg; P < 0.001). The DASH diet significantly decreased TC (-5.12 mg/dl; p = 0.008) and LDL-C levels (-3.53 mg/dl; p = 0.041), but not HDL-C (0.30 mg/dl; p = 0.510), TG (-4.22 mg/dl; p = 0.067), and VLDL-C (-2.16 mg/dl; p = 0.062). No significant effect of the DASH diet was noted for blood glucose (-0.38 mg/dl; p = 0.216), insulin (-0.03 μIU/mL; p = 0.817), HOMA-IR (-0.15; p = 0.132), and CRP (-0.33 mg/l; p = 0.173). The DASH diet is a feasible approach to weight loss and to control blood pressure and hypercholesterolemia.", "The number of older adults with different ethnic and socioeconomic background is steadily increasing. There is a need for community-based health promotion interventions for older adults that are responsive to ethnic and socioeconomic diversity among target populations. The aim of this study is to explore encounters between older adults living in disadvantaged areas and health care professionals in the context of community-based health promotion. Qualitative methods were used involving interviews and focus groups with older adults (n = 22) and municipal health care professionals (n = 8), and multiple observations were conducted. Data were analyzed thematically. Findings show a gap between health promotion services and older adults due to a perception of services as being neither accessible nor acceptable in the context of complex health and psychosocial needs. Health care professionals reported trust, proximity and presence as fundamental factors for improving acceptability and accessibility of health promotion services. There is a need to develop participatory approaches to engage older adults who live in disadvantaged areas in municipal health promotion services and to ensure that these services are relevant to these groups.", "Many interventions found to be effective in health services research studies fail to translate into meaningful patient care outcomes across multiple contexts. Health services researchers recognize the need to evaluate not only summative outcomes but also formative outcomes to assess the extent to which implementation is effective in a specific setting, prolongs sustainability, and promotes dissemination into other settings. Many implementation theories have been published to help promote effective implementation. However, they overlap considerably in the constructs included in individual theories, and a comparison of theories reveals that each is missing important constructs included in other theories. In addition, terminology and definitions are not consistent across theories. We describe the Consolidated Framework For Implementation Research (CFIR) that offers an overarching typology to promote implementation theory development and verification about what works where and why across multiple contexts. We used a snowball sampling approach to identify published theories that were evaluated to identify constructs based on strength of conceptual or empirical support for influence on implementation, consistency in definitions, alignment with our own findings, and potential for measurement. We combined constructs across published theories that had different labels but were redundant or overlapping in definition, and we parsed apart constructs that conflated underlying concepts. The CFIR is composed of five major domains: intervention characteristics, outer setting, inner setting, characteristics of the individuals involved, and the process of implementation. Eight constructs were identified related to the intervention (e.g., evidence strength and quality), four constructs were identified related to outer setting (e.g., patient needs and resources), 12 constructs were identified related to inner setting (e.g., culture, leadership engagement), five constructs were identified related to individual characteristics, and eight constructs were identified related to process (e.g., plan, evaluate, and reflect). We present explicit definitions for each construct. The CFIR provides a pragmatic structure for approaching complex, interacting, multi-level, and transient states of constructs in the real world by embracing, consolidating, and unifying key constructs from published implementation theories. It can be used to guide formative evaluations and build the implementation knowledge base across multiple studies and settings.", "To increase the likelihood of successful implementation of interventions and promote dissemination across real-world settings, it is essential to evaluate outcomes related to dimensions other than Effectiveness alone. Glasgow and colleagues' RE-AIM framework specifies four additional types of outcomes that are important to decision-makers: Reach, Adoption, Implementation (including cost), and Maintenance. To further strengthen RE-AIM, we propose integrating qualitative assessments in an expanded framework: RE-AIM Qualitative Evaluation for Systematic Translation (RE-AIM QuEST), a mixed methods framework. RE-AIM QuEST guides formative evaluation to identify real-time implementation barriers and explain how implementation context may influence translation to additional settings. RE-AIM QuEST was used to evaluate a pharmacist-led hypertension management intervention at 3 VA facilities in 2008-2009. We systematically reviewed each of the five RE-AIM dimensions and created open-ended companion questions to quantitative measures and identified qualitative and quantitative data sources, measures, and analyses. To illustrate use of the RE-AIM QuEST framework, we provide examples of real-time, coordinated use of quantitative process measures and qualitative methods to identify site-specific issues, and retrospective use of these data sources and analyses to understand variation across sites and explain outcomes. For example, in the Reach dimension, we conducted real-time measurement of enrollment across sites and used qualitative data to better understand and address barriers at a low-enrollment site. The RE-AIM QuEST framework may be a useful tool for improving interventions in real-time, for understanding retrospectively why an intervention did or did not work, and for enhancing its sustainability and translation to other settings.", "The aim of this study was to evaluate the association of pre-existing cardiovascular comorbidities, including hypertension and coronary heart disease, with coronavirus disease 2019 (COVID-19) severity and mortality. PubMed, ScienceDirect, and Scopus were searched between January 1, 2020, and July 18, 2020, to identify eligible studies. Random-effect models were used to estimate the pooled event rates of pre-existing cardiovascular disease comorbidities and odds ratio (OR) with 95% confidence intervals (95% CIs) of disease severity and mortality associated with the exposures of interest. A total of 34 studies involving 19,156 patients with COVID-19 infection met the inclusion criteria. The prevalence of pre-existing cardiovascular disease in the included studies was 14.0%. Pre-existing cardiovascular disease in COVID-19 patients was associated with severe outcomes (OR, 4.1; 95% CI, 2.9 to 5.7) and mortality (OR, 6.1; 95% CI, 2.9 to 12.7). Hypertension and coronary heart disease increased the risk of severe outcomes by 2.6 times (OR, 2.6; 95% CI, 1.9 to 3.6) and 2.5 times (OR, 2.5; 95% CI, 1.7 to 3.8), respectively. No significant publication bias was indicated. COVID-19 patients with pre-existing cardiovascular comorbidities have a higher risk of severe outcomes and mortality. Awareness of pre-existing cardiovascular comorbidity is important for the early management of COVID-19.", "Step Ahead was a randomized controlled trial testing ecologically based weight gain prevention interventions in the hospital workplace. The RE-AIM framework is used to assess the intervention's Reach, Effectiveness, Adoption, Implementation, and Maintenance. Some intervention components reached a large percentage of the workforce. Although the intervention was not effective in changing BMI on a population level, a dose response was observed, in which persons who used more of the intervention components and materials were more likely to prevent weight gain. Adoption of the intervention by sites invited was 100%. Implementation of healthy eating interventions in the hospital setting was especially challenging because close collaboration was necessary with hospital employees and contractors, and their mission and priorities often were at odds with the intervention goals. There are some notable instances of intervention maintenance at the institutional level: Farmers markets have been expanded at both sites since the end of the intervention period, and new wellness programs are being adopted and implemented. Implications for practice include the translation of this research into other workplace settings." ]
KCND3 Genetic Screening for Cardiac Disorders	Globally, cardiac channelopathies leading to electrical disorders are responsible for a significant number of sudden cardiac deaths without structural heart disease. Many genes encoding different ion channels in the heart were identified and their impairment was found to be associated with life-threatening cardiac abnormalities. KCND3, one of the genes expressed both in the heart and brain, is reported to have an association with Brugada syndrome, early-onset atrial fibrillation, early repolarization syndrome, and sudden unexplained death syndrome. KCND3 genetic screening could be a promising tool for functional studies for an understanding of the pathogenesis and genetic determinants of the above-mentioned electrical disorders.	[ "While a KCND3 V392I mutation uniquely displays a mixed electrophysiological phenotype of Kv4.3, only limited clinical information on the mutation carriers is available. We report two teenage siblings exhibiting both cardiac (early repolarization syndrome and paroxysmal atrial fibrillation) and cerebral phenotypes (epilepsy and intellectual disability), in whom we identified the KCND3 V392I mutation. We propose a link between the KCND3 mutation with a mixed electrophysiological phenotype and cardiocerebral phenotypes, which may be defined as a novel cardiocerebral channelopathy.", "The prevalence of atrial fibrillation (AF) is related to age. Anticoagulation is highly effective in preventing stroke in patients with AF, but the risk of hemorrhage may be increased in older patients. We reviewed the available epidemiologic data to define the age and sex distribution of people with AF. From four large recent population-based surveys, we estimated the overall age- and gender-specific prevalence of AF. These estimates were applied to the recent US census data to calculate the number of men and women with AF in each age group. There are an estimated 2.2 million people in the United States with AF, with a median age of about 75 years. The prevalence of AF is 2.3% in people older than 40 years and 5.9% in those older than 65 years. Approximately 70% of individuals with AF are between 65 and 85 years of age. The absolute number of men and women with AF is about equal. After age 75 years, about 60% of the people with AF are women. In contrast to people with AF in the general population, patients with AF in recent anticoagulation trials had a mean age of 69 years, and only 20% were older than 75 years. The risks and benefits of antithrombotic therapy in older individuals are important considerations in stroke prevention in AF.", "Early repolarization syndrome is rare in children. Mutation of genes encoding ion channels could display mixed electrophysiological phenotype of Kv4.3 including both cardiac phenotype (early repolarization syndrome, atrial fibrillation) and cerebral phenotype (epilepsy, intellectual disability). This situation is rare and was named as cardiocerebral channelopathy. Here, we report a case of an 11-year-old-girl with cardiocerebral channelopathy caused by KCND3 mutation, who was successfully treated with oral quinidine, metoprolol and implantable cardioverter-defibrillator. Clinicians should be vigilant on the risk of cardiogenic syncope and sudden cardiac death in a patient with epilepsy, intellectual disability and early repolarization pattern." ]
Retrospective comparative study	Retrospective comparative study.	[ "Retrospective review of anterior and posterior fusions for treatment of adolescent idiopathic thoracic scoliosis. To delineate the best factors determining final lumbar curve magnitude in patients with adolescent idiopathic scoliosis undergoing a selective thoracic anterior or posterior spinal fusion at or proximal to the first lumbar vertebra. Although spontaneous lumbar curve correction occurs consistently following a selective thoracic anterior or posterior spinal fusion, the degree of correction is somewhat unpredictable. One hundred consecutive patients with major thoracic-compensatory lumbar adolescent idiopathic scoliosis treated by a single surgeon with either selective posterior spinal fusion (n = 44) or anterior spinal fusion (n = 56) of the main thoracic region with an unfused lumbar spine with a lumbar B modifier (lumbar apex touching the center sacral vertical line) or lumbar C modifier (lumbar apex completely lateral to the center sacral vertical line) were retrospectively reviewed. Those patients who maintained excellent postoperative coronal balance, with spontaneous lumbar curve correction, had their thoracic Cobb corrected intraoperatively to a measurement very close to but not more than that of the preoperative thoracic push-prone Cobb. Stepwise multiple linear regression analysis was used to develop a formula to help predict lumbar response in those patients undergoing selective thoracic fusion. This is represented in the following formula: Final lumbar Cobb = 14.4 + 3.06 (lumbar modifier; 0 = B, 1 = C) + 0.30 (preoperative standing lumbar Cobb) - 0.18 (preoperative supine lower Cobb) + 0.81(preoperative push/prone lumbar Cobb) - 0.15(preoperative standing thoracic Cobb) - 0.16(% thoracic Cobb change from preoperative to immediate postoperative). Final model R2 = 0.72. Of the preoperative measurements examined, the preoperative push-prone is the best preoperative flexibility radiograph to predict the final lumbar curve measurement and, along with other factors, can be used to formulate a model that will help the treating surgeon more confidently predict the final lumbar curve response in patients undergoing a selective thoracic fusion.", "A prospective radiographic analysis of adolescent idiopathic scoliosis (AIS) patients managed with alternate-level pedicle screw fixation was performed. The objective of this study was to characterize segmental curve flexibility and to determine its predictive value in curve correction in AIS patients. Little is known regarding the distinct segmental curve characteristics and their ability to predict curve correction in patients with AIS. The segmental Cobb angle was measured on posteroanterior standing radiographs and on fulcrum bending radiographs. Radiographs were analyzed preoperatively and at 2 years postoperatively and the curve was divided into upper, mid, and lower segments based on predefined criteria. The segmental flexibility and the segmental fulcrum bending correction index (FBCI) were calculated. Eighty patients were included with mean age of 15 years. Preoperative mean segmental Cobb angles were 18, 31, and 17 degrees in the upper, mid, and lower segments, respectively. Segmental bending Cobb angles were 6, 13, and 4 degrees, respectively, corresponding to segmental flexibilities of 50%, 47%, and 83% in the upper, mid, and lower segments, respectively (P < 0.001). At 2-year follow up, the mean segmental FBCI were 155%, 131%, and 100% in the upper, mid, and lower segments, respectively (P < 0.001), which suggested that the lower segment of the curve was more flexible than the other segments and that higher correction was noted in the upper segments. A significant, positive correlation was noted between the segmental bending Cobb angle and the segmental FBCI (P < 0.05), whereby the strength of the correlation varied based on the curve segment. This is the first study to demonstrate the segmental variations in curve flexibility using the fulcrum bending radiograph in AIS patients. Curve flexibility is not uniform throughout the curve and different segments exhibit greater flexibility/correctibility than others. Segmental flexibility should be considered in assessing AIS patients and in the clinical decision-making strategy to optimize curve correction outcomes. 03.", "Coronal decompensation following correction of adolescent idiopathic scoliosis (AIS) has been reported to be due to the Cotrel-Dubousset rod derotation maneuver, or to a hypercorrection of the main thoracic curve. The treatment of such decompensation consists classically in observation, bracing, or extension of the instrumentation in the lumbar spine for a King 2 curve, or in the upper thoracic spine for a King 5 curve. As the postoperative decompensation is related to a hypercorrection of the main thoracic curve (relative to the compensatory curve), we hypothesized that if we were to \"let the spine go\" to some of its initial deformity, the balance of the patient would be improved. The purpose of the study was therefore to report on two cases where a postoperative imbalance following scoliosis surgery was successfully treated by decreasing the correction of the main thoracic curve. Two patients with AIS were found to have significant imbalance after scoliosis surgery. Both patients had been treated for a right thoracic curve (82 degrees and 85 degrees respectively) with an anterior release and posterior instrumentation. The revision surgery consisted for both patients in removing all the hooks between the end vertebrae of the main thoracic curve. This was done before the 3rd postoperative month for both patients. After revision surgery, the balance of both patients improved dramatically within a few weeks. The shoulders became almost level, and the trunk shift improved concomitantly. The Cobb angle increased by 8 degrees and 10 degrees, and the apical vertebra shifted to the right by 15 and 10 mm for the respective patients. These results were stable at 1-year follow-up. In the event of a persisting imbalance, we recommend, in selected cases, letting the spine go by removing all the implants located between the end vertebrae of the main thoracic curve. This adjustment or fine-tuning of the instrumentation should be done before the fusion takes place, and is best achieved with an instrumentation in which the hooks can be easily removed from the rod.", "Retrospective review of anterior and posterior fusions for treatment of adolescent idiopathic thoracic scoliosis. To evaluate both the instrumented thoracic and the spontaneous lumbar curve corrections after treatment of the primary thoracic scoliosis by either anterior or posterior fusion. Recent reports of thoracic scoliosis fusions have concentrated on the thoracic correction obtained by posterior segmental instrumentation systems. Coronal decompensation occurring because of curve progression with imbalance of the unfused lumbar spine has also been investigated. No report comparing spontaneous lumbar curve response after selective anterior versus posterior thoracic scoliosis fusions are available. One hundred twenty-three cases of primary thoracic-compensatory lumbar adolescent idiopathic scoliosis were treated by selective thoracic instrumentation and fusion with either an anterior (n = 70) or posterior (n = 53) single approach. Thoracic and lumbar Cobb measurements and lumbar apical translation parameters were assessed before surgery, 1 week after surgery, and 2 years after surgery on upright coronal radiographs. All patients had a minimum 2-year follow-up. At 2-year follow-up, the percentage of thoracic curve correction was superior for the anterior (58%) versus the posterior (38%) group (P < 0.05), whereas the spontaneous lumbar curve correction was also superior for the anterior (56%) group versus the posterior (37%) group for all curve types investigated (P < 0.05). Both treatment groups consistently improved lumbar apical positioning after the thoracic fusion procedure. Spontaneous lumbar curve correction occurs consistently after both selective anterior and posterior thoracic fusion implying intrinsic ability of the lumbar spine to follow thoracic spine correction. In the current study, using multisegmented hook-rod systems posteriorly with intentional limitation of posterior thoracic correction to avoid decompensation, instrumented thoracic and spontaneous lumbar curve correction was statistically better after anterior thoracic instrumentation and fusion, with the results most dramatic for lumbar curve Type C (true King II curves).", "Retrospective cohort study. The purpose of the study is to evaluate the role of supine radiographs in determining flexibility of thoracic and thoracolumbar curves. Ninety operative AIS patients with 2-year follow-up from a single institution were queried and classified into MT structural and TL structural groups. Equations were derived using linear regression to compute cut-off values for MT and TL curves. Thresholds were externally validated in a separate database of 60 AIS patients, and positive and negative predictive values were determined for each curve. MT supine values were highly predictive of MT side-bending values (TL group: 0.63, P < 0.001; MT group: 0.66, P = 0.006). Similarly, TL supine values were highly predictive of TL side-bending values (TL group: 0.56, P = 0.001 MT group: 0.68, P = 0.001). From our derived equations, MT and TL curves were considered structural on supine films if they were ≥ 30° and 35°, respectively. Contingency table analysis of external validity sample showed that supine films were highly predictive of structurality of MT curve (Sensitivity = 0.91, PPV = 0.95, NPV = 0.81) and TL curve (Sensitivity = 0.77, PPV = 0.81, NPV = 0.94). ROC analysis revealed that the area under curve for MT structurality from supine films was 0.931 (SEM: 0.03, CI: 0.86-0.99, P < 0.001) and TL structurality from supine films was 0.922 (SEM: 0.03, CI- 0.84-0.98, P < 0.001). A single preoperative supine radiograph is highly predictive of side-bending radiographs to assess curve flexibility in AIS. A cut-off of ≥ 30° for MT and ≥ 35° for TL curves in supine radiographs can determine curve structurality.", "Retrospective radiographic review. Our objectives were to (1) compare the ability of fulcrum bend radiographs and traction radiographs under general anesthesia to predict correction of adolescent idiopathic scoliosis (AIS) using pedicle screw only constructs and (2) compare the fulcrum bend correction index (FBCI) with a new measurement: the traction correction index (TCI). This is a retrospective radiographic review of 80 AIS patients (62 female and 18 male), who underwent scoliosis correction with pedicle screw only constructs. The mean age at surgery was 14 years (range 9-20 years). Radiographic analysis was carried out on the preoperative and immediate postoperative posteroanterior standing radiographs and the preoperative fulcrum bend radiographs and traction radiographs under general anesthesia. FBCI is calculated by dividing the correction rate by the fulcrum flexibility and TCI is calculated by dividing the correction rate by the traction flexibility. Preoperative mean Cobb angle of 63.9° was corrected to 25.8° postoperatively. The mean fulcrum bending Cobb angle was 37.6° and traction Cobb angle was 26.6°. The mean fulcrum flexibility was 41.1%, traction flexibility 58.4%, and correction rate 59.6%. The median FBCI was 137% and TCI was 104.3%. When comparing fulcrum bend and traction radiographs, we found the traction radiographs to be more predictive of curve correction in AIS using pedicle screw constructs. TCI takes into account the curve flexibility better than FBCI.", "To implement a clinically applicable, predictive model for the lumbar Cobb angle below a selective thoracic fusion in adolescent idiopathic scoliosis. A series of 146 adolescents with Lenke 1 or 2 idiopathic scoliosis, surgically treated with posterior selective fusion, and minimum follow-up of 5 years (average 7) was analyzed. The cohort was divided in 2 groups: if lumbar Cobb angle at last follow-up was, respectively, ≥ or < 10°. A logistic regression-based prediction model (PredictMed) was implemented to identify variables associated with the group ≥ 10°. The guidelines of the TRIPOD statement were followed. Mean Cobb angle of thoracic main curve was 56° preoperatively and 25° at last follow-up. Mean lumbar Cobb angle was 33° (20; 59) preoperatively and 11° (0; 35) at last follow-up. 53 patients were in group ≥ 10°. The 2 groups had similar demographics, flexibility of both main and lumbar curves, and magnitude of the preoperative main curve, p > 0.1. From univariate analysis, mean magnitude of preoperative lumbar curves (35° vs. 30°), mean correction of main curve (65% vs. 58%), mean ratio of main curve/distal curve (1.9 vs. 1.6) and distribution of lumbar modifiers were statistically different between groups (p < 0.05). PredictMed identified the following variables significantly associated with the group ≥ 10°: main curve % correction at last follow-up (p = 0.01) and distal curve angle (p = 0.04) with a prediction accuracy of 71%. The main modifiable factor influencing uninstrumented lumbar curve was the correction of main curve. The clinical model PredictMed showed an accuracy of 71% in prediction of lumbar Cobb angle ≥ 10° at last follow-up. Longitudinal comparative study." ]
Cell resolution characterization of cervical esophagus epithelium	Mouse esophagus is lined with a stratified epithelium, which is maintained by the constant renewal of unipotent progenitors. In this study, we profiled mouse esophagus by single-cell RNA sequencing and found taste buds specifically in the cervical segment of the esophagus. These taste buds have the same cellular composition as the ones from the tongue but express fewer taste receptor types. State-of-the-art transcriptional regulatory network analysis allowed the identification of specific transcription factors associated to the differentiation of immature progenitors into the three different taste bud cell types. Lineage tracing experiments revealed that esophageal taste buds arise from squamous bipotent progenitor, thus demonstrating that all esophageal progenitors are not unipotent. Our cell resolution characterization of cervical esophagus epithelium will enable a better understanding of esophageal progenitor potency and insights into the mechanisms involved in the development of taste buds.	[ "The esophageal lumen is lined by a stratified squamous epithelium comprised of proliferative basal cells that differentiate while migrating toward the luminal surface and eventually desquamate. Rapid epithelial renewal occurs, but the specific cell of origin that supports this high proliferative demand remains unknown. Herein, we have described a long-lived progenitor cell population in the mouse esophageal epithelium that is characterized by expression of keratin 15 (Krt15). Genetic in vivo lineage tracing revealed that the Krt15 promoter marks a long-lived basal cell population able to self-renew, proliferate, and generate differentiated cells, consistent with a progenitor/stem cell population. Transcriptional profiling demonstrated that Krt15+ basal cells are molecularly distinct from Krt15- basal cells. Depletion of Krt15-derived cells resulted in decreased proliferation, thereby leading to atrophy of the esophageal epithelium. Further, Krt15+ cells were radioresistant and contributed to esophageal epithelial regeneration following radiation-induced injury. These results establish the presence of a long-lived and indispensable Krt15+ progenitor cell population that provides additional perspective on esophageal epithelial biology and the widely prevalent diseases that afflict this epithelium.", "Epithelial functions are fundamentally determined by cytoskeletal keratin network organization. However, our understanding of keratin network plasticity is only based on analyses of cultured cells overexpressing fluorescently tagged keratins. In order to learn how keratin network organization is affected by various signals in functional epithelial tissues in vivo, we generated a knock-in mouse that produces fluorescence-tagged keratin 8. Homozygous keratin 8-YFP knock-in mice develop normally and show the expected expression of the fluorescent keratin network both in fixed and in vital tissues. In developing embryos, we observe for the first time de novo keratin network biogenesis in close proximity to desmosomal adhesion sites, keratin turnover in interphase cells and keratin rearrangements in dividing cells at subcellular resolution during formation of the first epithelial tissue. This mouse model will help to further dissect keratin network dynamics in its native tissue context during physiological and also pathological events.", "Adult taste buds are maintained by the lifelong proliferation of epithelial stem and progenitor cells, the identities of which have remained elusive. It has been proposed that these cells reside either within the taste bud (intragemmal) or in the surrounding epithelium (perigemmal). Here, we apply three different in vivo approaches enabling single-cell resolution of proliferative history to identify putative stem and progenitor cells associated with adult mouse taste buds. Experiments were performed across the circadian peak in oral epithelial proliferation (04:00 h), a time period in which mitotic activity in taste buds has not yet been detailed. Using double label pulse-chase experiments, we show that defined intragemmal cells (taste and basal) and perigemmal cells undergo rapid, sequential cell divisions and thus represent potential progenitor cells. Strikingly, mitotic activity was observed in taste cells previously thought to be postmitotic (labelled cells occur in 30% of palatal taste buds after 1 h of BrdU exposure). Basal cells showed expression of the transcription factor p63, required for maintaining the self-renewal potential of various epithelial stem cell types. Candidate taste stem cells were identified almost exclusively as basal cells using the label-retaining cell approach to localize slow-cycling cells (0.06 +/- 0.01 cells per taste bud; n = 436 taste buds). Together, these results indicate that both stem- and progenitor-like cells reside within the mammalian taste bud.", "Colchicine blocks mitotic division of the epithelial cells surrounding the taste bud of the rat tongue. Response to chemical stimulation decreases 50 per cent 3 hours after colchicine injection as measured by recording the electrical activity from the taste nerve bundle. Radioautography, using tritiated thymidine, shows that those epithelial cells surrounding the taste bud divide and that some of the daughter cells enter the taste bud and slowly move toward the center. The life span of the average cell is about 250 +/- 50 hours, although some cells have a much shorter and others a much longer life span. These studies suggest that the cells within the taste bud, as well as the nerves, undergo considerable change with time. Corresponding changes in function are considered.", "Single-cell transcriptomics has transformed our ability to characterize cell states, but deep biological understanding requires more than a taxonomic listing of clusters. As new methods arise to measure distinct cellular modalities, a key analytical challenge is to integrate these datasets to better understand cellular identity and function. Here, we develop a strategy to \"anchor\" diverse datasets together, enabling us to integrate single-cell measurements not only across scRNA-seq technologies, but also across different modalities. After demonstrating improvement over existing methods for integrating scRNA-seq data, we anchor scRNA-seq experiments with scATAC-seq to explore chromatin differences in closely related interneuron subsets and project protein expression measurements onto a bone marrow atlas to characterize lymphocyte populations. Lastly, we harmonize in situ gene expression and scRNA-seq datasets, allowing transcriptome-wide imputation of spatial gene expression patterns. Our work presents a strategy for the assembly of harmonized references and transfer of information across datasets.", "The electrical responses of the taste cell of the rat to chemical stimuli were studied by means of microelectrode techniques. Although large positive potential changes in the taste cell were usually elicited by taste stimuli, the response was a small negative potential change with respect to surrounding tissues if the microelectrode was thrust deeply into the taste bud. Both FeCl(3) and cocaine produced a positive change in the steady potential. If this new potential is larger than a certain equilibrium potential, reversal of the polarity of the potential change caused by a taste stimulus is observed. Gamma-aminobutyric acid and acetylcholine had no effect on the receptor steady potential nor on the receptor responses elicited by taste stimuli." ]
Role of ARID1A as tumor suppressor or oncogene in human cancers	ARID1A belongs to a class of chromatin regulatory proteins that function by maintaining accessibility at most promoters and enhancers, thereby regulating gene expression. The high frequency of ARID1A alterations in human cancers has highlighted its significance in tumorigenesis. The precise role of ARID1A in cancer is highly variable since ARID1A alterations can have a tumor suppressive or oncogenic role, depending on the tumor type and context. ARID1A is mutated in about 10% of all tumor types including endometrial, bladder, gastric, liver, biliopancreatic cancer, some ovarian cancer subtypes, and the extremely aggressive cancers of unknown primary. Its loss is generally associated with disease progression more often than onset. In some cancers, ARID1A loss is associated with worse prognostic features, thus supporting a major tumor suppressive role. However, some exceptions have been reported. Thus, the association of ARID1A genetic alterations with patient prognosis is controversial. However, ARID1A loss of function is considered conducive for the use of inhibitory drugs which are based on synthetic lethality mechanisms. In this review we summarize the current knowledge on the role of ARID1A as tumor suppressor or oncogene in different tumor types and discuss the strategies for treating ARID1A mutated cancers.	[ "Lung squamous cell carcinoma is a common type of lung cancer, causing approximately 400,000 deaths per year worldwide. Genomic alterations in squamous cell lung cancers have not been comprehensively characterized, and no molecularly targeted agents have been specifically developed for its treatment. As part of The Cancer Genome Atlas, here we profile 178 lung squamous cell carcinomas to provide a comprehensive landscape of genomic and epigenomic alterations. We show that the tumour type is characterized by complex genomic alterations, with a mean of 360 exonic mutations, 165 genomic rearrangements, and 323 segments of copy number alteration per tumour. We find statistically recurrent mutations in 11 genes, including mutation of TP53 in nearly all specimens. Previously unreported loss-of-function mutations are seen in the HLA-A class I major histocompatibility gene. Significantly altered pathways included NFE2L2 and KEAP1 in 34%, squamous differentiation genes in 44%, phosphatidylinositol-3-OH kinase pathway genes in 47%, and CDKN2A and RB1 in 72% of tumours. We identified a potential therapeutic target in most tumours, offering new avenues of investigation for the treatment of squamous cell lung cancers.", "ARID1A mutation is frequently found in clear cell ovarian cancer (CCC) and endometrioid ovarian cancer (EC). Anti-PD-1 monotherapy has been found to have limited efficacy in epithelial ovarian cancer; however, anti-PD-1 therapy showed significant clinical benefit in some CCC. We sought to define the relationship of ARID1A mutation/ARID1A expression to the immunogenic profile of different histologic subtypes of ovarian cancer. We performed next-generation sequencing of 160 cancer-related genes. Also, we analyzed the immunohistochemical status of ARID1A, PD-L1, and CD8 with survival in different histologic subtypes of ovarian cancer in a total of 103 cases. ARID1A mutation was found in 0% of the high-grade serous ovarian cancer (HGSC) (n = 36), 41.5% of the CCC (n = 41), 45.0% of the EC (n = 20), and 33.3% of the mucinous ovarian cancer (MC) (n = 6) cases. ARID1A loss was found in 19.4% of the HGSC, 75.6% of the CCC, 60.0% of the EC and 0% of the MC cases. ARID1A mutation was found to be associated with high PD-L1 (p < 0.001) or CD8 levels (p < 0.001) in CCC but not in other histologic subtypes. Meanwhile, ARID1A loss was associated with high PD-L1 or CD8 levels in CCC (p < 0.001) and HGSC (p < 0.001) but not in EC and MC. In addition, ARID1A mutation was associated with high tumor mutation burden in CCC (p = 0.006). ARID1A mutation/ARID1A expression is associated with immune microenvironmental factors in CCC but not in EC. ARID1A status can be a biomarker for selecting candidates for immune checkpoint blockade in CCC.", "SWI/SNF is a multisubunit chromatin-remodeling complex that performs fundamental roles in gene regulation, cell lineage specification, and organismal development. Mutations that inactivate SWI/SNF subunits are found in nearly 20% of human cancers, which indicates that the proper functioning of this complex is necessary to prevent tumor formation in diverse tissues. Recent studies show that SWI/SNF-mutant cancers depend on residual SWI/SNF complexes for their aberrant growth, thus revealing synthetic lethal interactions that could be exploited for therapeutic purposes. Other studies reveal that certain acute leukemias and small cell lung cancers, which lack SWI/SNF mutations, can be vulnerable to inhibition of the SWI/SNF ATPase subunit BRG1, whereas several normal and malignant cell types do not show this sensitivity. Here, we review the emerging evidence that implicates SWI/SNF as a tumor-dependency and candidate drug target in human cancer.", "Adenine-thymine-rich inactive domain-containing protein 1A (ARID1A) is a large subunit of the switch-sucrose nonfermenting (SWI-SNF) complex. ARID1A is considered to be a tumor suppressor in various cancers. We investigated the clinicopathological significance including prognosis of ARID1A expression in non-small cell lung cancer (NSCLC). ARID1A expression was studied by tissue microarray immunohistochemical analysis of 171 surgically resected NSCLC specimens including adenocarcinoma (ADC) and squamous cell carcinoma (SCC) on tissue microarray. Semiquantitative immunohistochemical score was obtained by multiplying the intensity and percentage scores. The overall score was further simplified by dichotomizing into either negative (score < 4) or positive (score ≥ 4) for each patient. The ARID1A-negative group revealed significantly higher correlations with male sex (p = 0.020), larger tumor size (p = 0.007), SCC than with ADC (p = 0.023) and smoking (p = 0.001). Univariate survival analysis showed that the ARID1A-negative group had a significantly shorter cancer specific survival than the ARID1A-positive group (p = 0.018). Multivariate survival analysis showed that ARID1A negativity (p = 0.022) were independent prognostic factors related with shorter cancer specific survival for NSCLC. In conclusion, Loss of ARID1A expression is a potential molecular marker to predictive of poor prognosis of NSCLC.", "Clear cell renal cell carcinoma (CCRCC) is the most common renal cell carcinoma. It has a relatively unfavorable prognosis compared to other common renal cell carcinomas. Recently, comprehensive molecular studies in CCRCC revealed important genetic alterations, including changes in the VHL, PBRM1, and ARID1A genes. The expression of ARID1A protein is associated with tumor progression and prognosis in many cancers. This study aimed to evaluate the nuclear expression of ARID1A in CCRCC and to assess its expression with the clinical prognosis. The nuclear expression of ARID1A was evaluated in 290 cases of CCRCC by immunohistochemistry. To determine the clinicopathological association with ARID1A, each of the cases was divided into 2 groups, low- and high-expression groups, according to the average proportion of nuclear staining. Decreased ARID1A expression was associated with the higher nuclear grade and higher pTNM stage (P < .001 and P = .013, respectively). The ARID1A low-expression group revealed significantly shorter cancer-specific and progression-free survival times (P = .001 and P < .001, respectively). Furthermore, Cox regression analysis showed that ARID1A expression was an independent prognostic factor for progression-free survival (P = .009). These results suggest that nuclear expression of ARID1A may serve as a new prognostic marker in CCRCC patients.", "Subunits of mammalian SWI/SNF (mSWI/SNF or BAF) complexes have recently been implicated as tumor suppressors in human malignancies. To understand the full extent of their involvement, we conducted a proteomic analysis of endogenous mSWI/SNF complexes, which identified several new dedicated, stable subunits not found in yeast SWI/SNF complexes, including BCL7A, BCL7B and BCL7C, BCL11A and BCL11B, BRD9 and SS18. Incorporating these new members, we determined mSWI/SNF subunit mutation frequency in exome and whole-genome sequencing studies of primary human tumors. Notably, mSWI/SNF subunits are mutated in 19.6% of all human tumors reported in 44 studies. Our analysis suggests that specific subunits protect against cancer in specific tissues. In addition, mutations affecting more than one subunit, defined here as compound heterozygosity, are prevalent in certain cancers. Our studies demonstrate that mSWI/SNF is the most frequently mutated chromatin-regulatory complex (CRC) in human cancer, exhibiting a broad mutation pattern, similar to that of TP53. Thus, proper functioning of polymorphic BAF complexes may constitute a major mechanism of tumor suppression.", "ARID1A is located in 1p36.11, a region frequently deleted in human cancers. Using a novel method to screen for tumorigenic cDNA sequences, we have identified ARID1A as a presumptive tumor suppressor gene. The transforming ARID1A sequence was an antisense cDNA, and was the product of a genomic rearrangement, as corroborated in the primary breast carcinoma from which the cDNA had been obtained. In further screening, we identified a lung adenocarcinoma cell line with a highly localized homozygous genomic deletion involving the 5' end of ARID1A. These studies provide strong evidence that ARID1A is a tumor suppressor gene. (c) 2007 Wiley-Liss, Inc." ]
Ataxia-telangiectasia mutated chromodomain helicase DNA binding protein 7 is involved in both morphogenetic and DSB-response functions	Following radiation exposure, unrepaired DNA double-strand breaks (DSBs) persist to some extent in a subset of cells as residual damage; they can exert adverse effects, including late-onset diseases. In search of the factor(s) that characterize(s) cells bearing such damage, we discovered ataxia-telangiectasia mutated (ATM)-dependent phosphorylation of the transcription factor chromodomain helicase DNA binding protein 7 (CHD7). CHD7 controls the morphogenesis of cell populations derived from neural crest cells during vertebrate early development. Indeed, malformations in various fetal bodies are attributable to CHD7 haploinsufficiency. Following radiation exposure, CHD7 becomes phosphorylated, ceases promoter/enhancer binding to target genes, and relocates to the DSB-repair protein complex, where it remains until the damage is repaired. Thus, ATM-dependent CHD7 phosphorylation appears to act as a functional switch. As such stress responses contribute to improved cell survival and canonical nonhomologous end joining, we conclude that CHD7 is involved in both morphogenetic and DSB-response functions. Thus, we propose that higher vertebrates have evolved intrinsic mechanisms underlying the morphogenesis-coupled DSB stress response. In fetal exposure, if the function of CHD7 becomes primarily shifted toward DNA repair, morphogenic activity is reduced, resulting in malformations.	[ "We show that DNA double-strand breaks (DSBs) induce complex subcompartmentalization of genome surveillance regulators. Chromatin marked by gamma-H2AX is occupied by ataxia telangiectasia-mutated (ATM) kinase, Mdc1, and 53BP1. In contrast, repair factors (Rad51, Rad52, BRCA2, and FANCD2), ATM and Rad-3-related (ATR) cascade (ATR, ATR interacting protein, and replication protein A), and the DNA clamp (Rad17 and -9) accumulate in subchromatin microcompartments delineated by single-stranded DNA (ssDNA). BRCA1 and the Mre11-Rad50-Nbs1 complex interact with both of these compartments. Importantly, some core DSB regulators do not form cytologically discernible foci. These are further subclassified to proteins that connect DSBs with the rest of the nucleus (Chk1 and -2), that assemble at unprocessed DSBs (DNA-PK/Ku70), and that exist on chromatin as preassembled complexes but become locally modified after DNA damage (Smc1/Smc3). Finally, checkpoint effectors such as p53 and Cdc25A do not accumulate at DSBs at all. We propose that subclassification of DSB regulators according to their residence sites provides a useful framework for understanding their involvement in diverse processes of genome surveillance.", "Neural crest cells are highly migratory pluripotent cells that give rise to diverse derivatives including cartilage, bone, smooth muscle, pigment, and endocrine cells as well as neurons and glia. Abnormalities in neural crest-derived tissues contribute to the etiology of CHARGE syndrome, a complex malformation disorder that encompasses clinical symptoms like coloboma, heart defects, atresia of the choanae, retarded growth and development, genital hypoplasia, ear anomalies, and deafness. Mutations in the chromodomain helicase DNA-binding protein 7 (CHD7) gene are causative of CHARGE syndrome and loss-of-function data in different model systems have firmly established a role of CHD7 in neural crest development. Here, we will summarize our current understanding of the function of CHD7 in neural crest development and discuss possible links of CHARGE syndrome to other developmental disorders.", "SignificanceOur work focuses on the critical longstanding question of the nontranscriptional role of p53 in tumor suppression. We demonstrate here that poly(ADP-ribose) polymerase (PARP)-dependent modification of p53 enables rapid recruitment of p53 to damage sites, where it in turn directs early repair pathway selection. Specifically, p53-mediated recruitment of 53BP1 at early time points promotes nonhomologous end joining over the more error-prone microhomology end-joining. Similarly, p53 directs nucleotide excision repair by mediating DDB1 recruitment. This property of p53 also correlates with tumor suppression in vivo. Our study provides mechanistic insight into how certain transcriptionally deficient p53 mutants may retain tumor-suppressive functions through regulating the DNA damage response.", "Doxorubicin (Adriamycin) is one of the most commonly used chemotherapeutic drugs and exhibits a wide spectrum of activity against solid tumors, lymphomas, and leukemias. Doxorubicin is classified as a topoisomerase II poison, although other mechanisms of action have been characterized. Here, we show that doxorubicin-DNA adducts (formed by the coadministration of doxorubicin with non-toxic doses of formaldehyde-releasing prodrugs) induce a more cytotoxic response in HL-60 cells than doxorubicin as a single agent. Doxorubicin-DNA adducts seem to be independent of classic topoisomerase II-mediated cellular responses (as observed by employing topoisomerase II catalytic inhibitors and HL-60/MX2 cells). Apoptosis induced by doxorubicin-DNA adducts initiates a caspase cascade that can be blocked by overexpressed Bcl-2, suggesting that adducts induce a classic mode of apoptosis. A reduction in the level of topoisomerase II-mediated double-strand-breaks was also observed with increasing levels of doxorubicin-DNA adducts and increased levels of apoptosis, further confirming that adducts exhibit a separate mechanism of action compared with the classic topoisomerase II poison mode of cell death by doxorubicin alone. Collectively, these results indicate that the presence of formaldehyde transfers doxorubicin from topoisomerase II-mediated cellular damage to the formation of doxorubicin-DNA adducts, and that these adducts are more cytotoxic than topoisomerase II-mediated lesions. These results also show that doxorubicin can induce apoptosis by a non-topoisomerase II-dependent mechanism, and this provides exciting new prospects for enhancing the clinical use of this agent and for the development of new derivatives and new tumor-targeted therapies." ]
Enrichment of adherent-invasive Escherichia coli in inflammatory bowel disease	Enrichment of adherent-invasive Escherichia coli (AIEC) has been consistently detected in subsets of inflammatory bowel disease (IBD) patients. Although some AIEC strains cause colitis in animal models, these studies did not systematically compare AIEC with non-AIEC strains, and causal links between AIEC and disease are still disputed. Specifically, it remains unclear whether AIEC shows enhanced pathogenicity compared to that of commensal E. coli found in the same ecological microhabitat and if the	[ "Adherent-invasive Escherichia coli (AIEC) are a leading candidate bacterial trigger for Crohn's disease (CD). The AIEC pathovar is defined by in vitro cell-line assays examining specific bacteria/cell interactions. No molecular marker exists for their identification. Our aim was to identify a molecular property common to the AIEC phenotype. 41 B2 phylogroup E. coli strains were isolated from 36 Australian subjects: 19 patients with IBD and 17 without. Adherence/invasion assays were conducted using the I-407 epithelial cell line and survival/replication assays using the THP-1 macrophage cell line. Cytokine secretion tumour necrosis factor ((TNF)-α, interleukin (IL) 6, IL-8 and IL-10) was measured using ELISA. The genomes were assembled and annotated, and cluster analysis performed using CD-HIT. The resulting matrices were analysed to identify genes unique/more frequent in AIEC strains compared with non-AIEC strains. Base composition differences and clustered regularly interspaced palindromic repeat (CRISPR) analyses were conducted. Of all B2 phylogroup strains assessed, 79% could survive and replicate in macrophages. Among them, 11/41 strains (5 CD, 2 UCs, 5 non-IBD) also adhere to and invade epithelial cells, a phenotype assigning them to the AIEC pathovar. The AIEC strains were phylogenetically heterogeneous. We did not identify a gene (or nucleic acid base composition differences) common to all, or the majority of, AIEC. Cytokine secretion and CRISPRs were not associated with the AIEC phenotype. Comparative genomic analysis of AIEC and non-AIEC strains did not identify a molecular property exclusive to the AIEC phenotype. We recommend a broader approach to the identification of the bacteria-host interactions that are important in the pathogenesis of Crohn's disease.", "Urinary tract infections (UTIs) caused by uropathogenic Escherichia coli (UPEC) affect 150 million people annually. Despite effective antibiotic therapy, 30-50% of patients experience recurrent UTIs. In addition, the growing prevalence of UPEC that are resistant to last-line antibiotic treatments, and more recently to carbapenems and colistin, make UTI a prime example of the antibiotic-resistance crisis and emphasize the need for new approaches to treat and prevent bacterial infections. UPEC strains establish reservoirs in the gut from which they are shed in the faeces, and can colonize the periurethral area or vagina and subsequently ascend through the urethra to the urinary tract, where they cause UTIs. UPEC isolates encode up to 16 distinct chaperone-usher pathway pili, and each pilus type may enable colonization of a habitat in the host or environment. For example, the type 1 pilus adhesin FimH binds mannose on the bladder surface, and mediates colonization of the bladder. However, little is known about the mechanisms underlying UPEC persistence in the gut. Here, using a mouse model, we show that F17-like and type 1 pili promote intestinal colonization and show distinct binding to epithelial cells distributed along colonic crypts. Phylogenomic and structural analyses reveal that F17-like pili are closely related to pilus types carried by intestinal pathogens, but are restricted to extra-intestinal pathogenic E. coli. Moreover, we show that targeting FimH with M4284, a high-affinity inhibitory mannoside, reduces intestinal colonization of genetically diverse UPEC isolates, while simultaneously treating UTI, without notably disrupting the structural configuration of the gut microbiota. By selectively depleting intestinal UPEC reservoirs, mannosides could markedly reduce the rate of UTIs and recurrent UTIs.", "An Escherichia coli (E. coli) pathotype with invasive properties, first reported by Darfeuille-Michaud and termed adherent-invasive E. coli (AIEC), was shown to be prevalent in up to half the individuals with Crohn's Disease (CD), suggesting that these bacteria could be involved in the pathophysiology of CD. Among the genes related to AIEC pathogenicity, fim has the potential to generate an inflammatory reaction from the intestinal epithelial cells and macrophages, as it interacts with TLR4, inducing the production of inflammatory cytokines independently of LPS. Therefore, targeting the bacterial adhesion of FimH-expressing bacteria seems a promising therapeutic approach, consisting of disarming bacteria without killing them, representing a selective strategy to suppress a potentially critical trigger of intestinal inflammation, without disturbing the intestinal microbiota. We analyzed the metagenomic composition of the gut microbiome of 358 patients with CD from two different cohorts and characterized the presence of FimH-expressing bacteria. To assess the pathogenic role of FimH, we used human intestinal explants and tested a specific FimH blocker to prevent bacterial adhesion and associated inflammation. We observed a significant and disease activity-dependent enrichment of Enterobacteriaceae in the gut microbiome of patients with CD. Bacterial FimH expression was functionally confirmed in ileal biopsies from 65% of the patients with CD. Using human intestinal explants, we further show that FimH is essential for adhesion and to trigger inflammation. Finally, a specific FimH-blocker, TAK-018, inhibits bacterial adhesion to the intestinal epithelium and prevents inflammation, thus preserving mucosal integrity. We propose that TAK-018, which is safe and well tolerated in humans, is a promising candidate for the treatment of CD and in particular in preventing its recurrence. Video abstract.", "Components of bacteria have been shown to induce innate antiviral immunity via Toll-like receptors (TLRs). We have recently shown that FimH, the adhesin portion of type 1 fimbria, can induce the innate immune system via TLR4. Here we report that FimH induces potent in vitro and in vivo innate antimicrobial responses. FimH induced an innate antiviral state in murine macrophage and primary MEFs which was correlated with IFN-beta production. Moreover, FimH induced the innate antiviral responses in cells from wild type, but not from MyD88(-/-), Trif(-/-), IFN-alpha/betaR(-/-) or IRF3(-/-) mice. Vaginal delivery of FimH, but not LPS, completely protected wild type, but not MyD88(-/-), IFN-alpha/betaR(-/-), IRF3(-/-) or TLR4(-/-) mice from subsequent genital HSV-2 challenge. The FimH-induced innate antiviral immunity correlated with the production of IFN-beta, but not IFN-alpha or IFN-gamma. To examine whether FimH plays a role in innate immune induction in the context of a natural infection, the innate immune responses to wild type uropathogenic E. coli (UPEC) and a FimH null mutant were examined in the urinary tract of C57Bl/6 (B6) mice and TLR4-deficient mice. While UPEC expressing FimH induced a robust polymorphonuclear response in B6, but not TLR4(-/-) mice, mutant bacteria lacking FimH did not. In addition, the presence of TLR4 was essential for innate control of and protection against UPEC. Our results demonstrate that FimH is a potent inducer of innate antimicrobial responses and signals differently, from that of LPS, via TLR4 at mucosal surfaces. Our studies suggest that FimH can potentially be used as an innate microbicide against mucosal pathogens.", "Intestinal bacteria are implicated increasingly as a pivotal factor in the development of Crohn's disease, but the specific components of the complex polymicrobial enteric environment driving the inflammatory response are unresolved. This study addresses the role of the ileal mucosa-associated microflora in Crohn's disease. A combination of culture-independent analysis of bacterial diversity (16S rDNA library analysis, quantitative PCR and fluorescence in situ hybridization) and molecular characterization of cultured bacteria was used to examine the ileal mucosa-associated flora of patients with Crohn's disease involving the ileum (13), Crohn's disease restricted to the colon (CCD) (8) and healthy individuals (7). Analysis of 16S rDNA libraries constructed from ileal mucosa yielded nine clades that segregated according to their origin (P<0.0001). 16S rDNA libraries of ileitis mucosa were enriched in sequences for Escherichia coli (P<0.001), but relatively depleted in a subset of Clostridiales (P<0.05). PCR of mucosal DNA was negative for Mycobacterium avium subspecies paratuberculosis, Shigella and Listeria. The number of E. coli in situ correlated with the severity of ileal disease (rho 0.621, P<0.001) and invasive E. coli was restricted to inflamed mucosa. E. coli strains isolated from the ileum were predominantly novel in phylogeny, displayed pathogen-like behavior in vitro and harbored chromosomal and episomal elements similar to those described in extraintestinal pathogenic E. coli and pathogenic Enterobacteriaceae. These data establish that dysbiosis of the ileal mucosa-associated flora correlates with an ileal Crohn's disease (ICD) phenotype, and raise the possibility that a selective increase in a novel group of invasive E. coli is involved in the etiopathogenesis to Crohn's disease involving the ileum.", "Crohn's disease (CD) is a life-long chronic disorder characterized by intestinal inflammation. Current treatments for CD are directed towards abnormal immune responses rather than the intestinal bacteria that trigger intestinal inflammation. Areas covered: Adherent-Invasive Escherichia coli (AIEC) bacteria abnormally colonize the ileal mucosa in a subgroup of CD patients. They can promote or perpetuate chronic inflammation and are therefore an interesting therapeutic target. Various strategies that target these E. coli strains have been developed to promote their intestinal clearance. Here, we review current AIEC-targeted strategies, especially anti-adhesive strategies, that are based on the development of FimH antagonists. We discuss their potential as personalized microbiota-targeted treatments for CD patients abnormally colonized by AIEC. Expert opinion: A large panel of mannose-derived FimH antagonists were tested for their ability to inhibit E. coli adhesion to host cells. Documented reports suggest that monovalent mannosides are promising candidates that could represent a complementary therapeutic strategy to prevent intestinal inflammation in the E. coli-colonized CD patient subgroup. Ongoing research continues to improve the pharmacokinetic properties of mannosides, and hopefully, clinical trials will be performed in CD patients in the near future.", "A major cause of enteric infection, Gram-negative pathogenic bacteria activate mucosal inflammation through lipopolysaccharide (LPS) binding to intestinal toll-like receptor 4 (TLR4). Breast feeding lowers risk of disease, and human milk modulates inflammation. This study tested whether human milk oligosaccharides (HMOSs) influence pathogenic Escherichia coli-induced interleukin (IL)-8 release by intestinal epithelial cells (IECs), identified specific proinflammatory signalling molecules modulated by HMOSs, specified the active HMOS and determined its mechanism of action. Models of inflammation were IECs invaded by type 1 pili enterotoxigenic E. coli (ETEC) in vitro: T84 modelled mature, and H4 modelled immature IECs. LPS-induced signalling molecules co-varying with IL-8 release in the presence or absence of HMOSs were identified. Knockdown and overexpression verified signalling mediators. The oligosaccharide responsible for altered signalling was identified. HMOSs attenuated LPS-dependent induction of IL-8 caused by ETEC, uropathogenic E. coli, and adherent-invasive E. coli (AIEC) infection, and suppressed CD14 transcription and translation. CD14 knockdown recapitulated HMOS-induced attenuation. Overexpression of CD14 increased the inflammatory response to ETEC and sensitivity to inhibition by HMOSs. 2'-fucosyllactose (2'-FL), at milk concentrations, displayed equivalent ability as total HMOSs to suppress CD14 expression, and protected AIEC-infected mice. HMOSs and 2'-FL directly inhibit LPS-mediated inflammation during ETEC invasion of T84 and H4 IECs through attenuation of CD14 induction. CD14 expression mediates LPS-TLR4 stimulation of portions of the 'macrophage migration inhibitory factors' inflammatory pathway via suppressors of cytokine signalling 2/signal transducer and activator of transcription 3/NF-κB. HMOS direct inhibition of inflammation supports its functioning as an innate immune system whereby the mother protects her vulnerable neonate through her milk. 2'-FL, a principal HMOS, quenches inflammatory signalling.", "Adherent-invasive Escherichia coli (AIEC) strains are genetically variable and virulence factors for AIEC are non-specific. FimH is the most studied pathogenicity-related protein, and there have been few studies on other proteins, such as Serine Protease Autotransporters of Enterobacteriacea (SPATEs). The goal of this study is to characterize E. coli strains isolated from patients with Crohn's disease (CD) in Chile and Spain, and identify genetic differences between strains associated with virulence markers and clonality. We characterized virulence factors and genetic variability by pulse field electrophoresis (PFGE) in 50 E. coli strains isolated from Chilean and Spanish patients with CD, and also determined which of these strains presented an AIEC phenotype. Twenty-six E. coli strains from control patients were also included. PFGE patterns were heterogeneous and we also observed a highly diverse profile of virulence genes among all E. coli strains obtained from patients with CD, including those strains defined as AIEC. Two iron transporter genes chuA, and irp2, were detected in various combinations in 68-84% of CD strains. We found that the most significant individual E. coli genetic marker associated with CD E. coli strains was chuA. In addition, patho-adaptative fimH mutations were absent in some of the highly adherent and invasive strains. The fimH adhesin, the iron transporter irp2, and Class-2 SPATEs did not show a significant association with CD strains. The V27A fimH mutation was detected in the most CD strains. This study highlights the genetic variability of E. coli CD strains from two distinct geographic origins, most of them affiliated with the B2 or D E. coli phylogroups and also reveals that nearly 40% of Chilean and Spanish CD patients are colonized with E.coli with a characteristic AIEC phenotype.", "Escherichia coli (E. coli), and particularly the adherent invasive E. coli (AIEC) pathotype, has been increasingly implicated in the ethiopathogenesis of Crohn's disease (CD). E. coli strains with similar pathogenic features to AIEC have been associated with other intestinal disorders such as ulcerative colitis, colorectal cancer, and coeliac disease, but AIEC prevalence in these diseases remains largely unexplored. Since AIEC was described one decade ago, substantial progress has been made in deciphering its mechanisms of pathogenicity. However, the molecular bases that characterize the phenotypic properties of this pathotype are still not well resolved. A review of studies focused on E. coli populations in inflammatory bowel disease (IBD) is presented here and we discuss about the putative role of this species on each IBD subtype. Given the relevance of AIEC in CD pathogenesis, we present the latest research findings concerning AIEC host-microbe interactions and pathogenicity. We also review the existing data regarding the prevalence and abundance of AIEC in CD and its association with other intestinal diseases from humans and animals, in order to discuss the AIEC disease- and host-specificity. Finally, we highlight the fact that dietary components frequently found in industrialized countries may enhance AIEC colonization in the gut, which merits further investigation and the implementation of preventative measures.", "Colonic mucus appears to consist of two glycoprotein fractions, one of which contains mannose, whereas the other is mannose-free. The mannose-containing fraction is significantly increased in ulcerative colitis." ]
Trapomyosin kinase receptor B signaling in the heart	Loss of brain-derived neurotrophic factor (BDNF)/TrkB (tropomyosin kinase receptor B) signaling accounts for brain and cardiac disorders. In neurons, β-adrenergic receptor stimulation enhances local BDNF expression. It is unclear if this occurs in a pathophysiological relevant manner in the heart, especially in the β-adrenergic receptor-desensitized postischemic myocardium. Nor is it fully understood whether and how TrkB agonists counter chronic postischemic left ventricle (LV) decompensation, a significant unmet clinical milestone.	[ "At least three distinct beta-adrenergic receptor (beta-AR) subtypes exist in mammals. These receptors modulate a wide variety of processes, from development and behavior, to cardiac function, metabolism, and smooth muscle tone. To understand the roles that individual beta-AR subtypes play in these processes, we have used the technique of gene targeting to create homozygous beta 1-AR null mutants (beta 1-AR -/-) in mice. The majority of beta 1-AR -/- mice die prenatally, and the penetrance of lethality shows strain dependence. Beta l-AR -/- mice that do survive to adulthood appear normal, but lack the chronotropic and inotropic responses seen in wild-type mice when beta-AR agonists such as isoproterenol are administered. Moreover, this lack of responsiveness is accompanied by markedly reduced stimulation of adenylate cyclase in cardiac membranes from beta 1-AR -/- mice. These findings occur despite persistent cardiac beta 2-AR expression, demonstrating the importance of beta 1-ARs for proper mouse development and cardiac function, while highlighting functional differences between beta-AR subtypes.", "Autonomic control of heart rate is mediated by cardioinhibitory parasympathetic cholinergic neurons located in the brainstem and stimulatory sympathetic noradrenergic neurons. During embryonic development the survival and cholinergic phenotype of brainstem autonomic neurons is promoted by brain-derived neurotrophic factor (BDNF). We now provide evidence that BDNF regulates heart rate by a mechanism involving increased brainstem cardioinhibitory parasympathetic activity. Mice with a BDNF haploinsufficiency exhibit elevated resting heart rate, and infusion of BDNF intracerebroventricularly reduces heart rate in both wild-type and BDNF+/- mice. The atropine-induced elevation of heart rate is diminished in BDNF+/- mice and is restored by BDNF infusion, whereas the atenolol-induced decrease in heart rate is unaffected by BDNF levels, suggesting that BDNF signaling enhances parasympathetic tone which is diminished with BDNF haploinsufficiency. Whole-cell recordings from pre-motor cholinergic cardioinhibitory vagal neurons in the nucleus ambiguus indicate that BDNF haploinsufficiency reduces cardioinhibitory vagal neuron activity by increased inhibitory GABAergic and diminished excitatory glutamatergic neurotransmission to these neurons. Our findings reveal a previously unknown role for BDNF in the control of heart rate by a mechanism involving increased activation of brainstem cholinergic parasympathetic neurons.", "Brain-derived neurotrophic factor (BDNF) is a neurotrophin essential for the development of normal respiratory rhythm and ventilatory control. Chronic exposure to Intermittent Hypercapnic Hypoxia (IHH) has been shown to alter ventilatory responses of piglets. This study investigated changes in BDNF distribution and expression in seven nuclei of the caudal medulla, from piglets exposed to IHH for 1, 2, 3, or 4 days before death, using non-radioactive in situ hybridisation (for mRNA) and immunohistochemistry (for protein). Compared to controls, BDNF mRNA was markedly increased across the entire medulla of the brainstem, after all durations of IHH (1-4 days). In contrast, BDNF protein expression increased after 1 day of exposure to IHH (p=0.003), but, thereafter, was not different to controls. Amongst individual nuclei, neurons of the dorsal motor nucleus of the vagus (DMNV) showed increased BDNF mRNA (p<0.01), but decreased protein expression (p=0.05) after all durations of IHH. In the ION, both mRNA and protein for BDNF were significantly increased after 1 day IHH (p<0.01 and p=0.001, respectively), but these increases were not sustained. This study is the first to investigate changes in BDNF expression in response to environmental challenges during postnatal development in the brainstem. Implications of the wide distribution of BDNF in the piglet caudal medulla and increased expression after IHH exposure are discussed, with particular reference to roles for BDNF-dependent neurons at this stage of development.", "The expression of vesicular acetylcholine transporter (VAChT), which transports ACh into synaptic vesicles, is coregulated with choline acetyltransferase (ChAT). Therefore, the effects of nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) on the levels of VAChT in cultured neurons from the septum of embryonic rats were investigated by immunocytochemistry. NGF and BDNF increased the number of VAChT-immunoreactive neurons by approximately 1.5-fold and enhanced the immunoreactivity in each positive cell. These results suggest that the neurotrophins enhance not only synthesis but also storage of ACh in septal neurons.", "beta-Adrenergic receptors (beta-ARs) are members of the superfamily of G-protein-coupled receptors that mediate the effects of catecholamines in the sympathetic nervous system. Three distinct beta-AR subtypes have been identified (beta1-AR, beta2-AR, and beta3-AR). In order to define further the role of the different beta-AR subtypes, we have used gene targeting to inactivate selectively the beta2-AR gene in mice. Based on intercrosses of heterozygous knockout (beta2-AR +/-) mice, there is no prenatal lethality associated with this mutation. Adult knockout mice (beta2-AR -/-) appear grossly normal and are fertile. Their resting heart rate and blood pressure are normal, and they have a normal chronotropic response to the beta-AR agonist isoproterenol. The hypotensive response to isoproterenol, however, is significantly blunted compared with wild type mice. Despite this defect in vasodilation, beta2-AR -/- mice can still exercise normally and actually have a greater total exercise capacity than wild type mice. At comparable workloads, beta2-AR -/- mice had a lower respiratory exchange ratio than wild type mice suggesting a difference in energy metabolism. beta2-AR -/- mice become hypertensive during exercise and exhibit a greater hypertensive response to epinephrine compared with wild type mice. In summary, the primary physiologic consequences of the beta2-AR gene disruption are observed only during the stress of exercise and are the result of alterations in both vascular tone and energy metabolism.", "The mTORC1 (mechanistic target of rapamycin complex-1) controls metabolism and protein homeostasis and is activated following ischemia reperfusion (IR) injury and by ischemic preconditioning (IPC). However, studies vary as to whether this activation is beneficial or detrimental, and its influence on metabolism after IR is little reported. A limitation of prior investigations is their use of broad gain/loss of mTORC1 function, mostly applied before ischemic stress. This can be circumvented by regulating one serine (S1365) on TSC2 (tuberous sclerosis complex) to achieve bidirectional mTORC1 modulation but only with TCS2-regulated costimulation. We tested the hypothesis that reduced TSC2 S1365 phosphorylation protects the myocardium against IR and is required for IPC by amplifying mTORC1 activity to favor glycolytic metabolism. Mice with either S1365A (TSC2SA; phospho-null) or S1365E (TSC2SE; phosphomimetic) knockin mutations were studied ex vivo and in vivo. In response to IR, hearts from TSC2SA mice had amplified mTORC1 activation and improved heart function compared with wild-type and TSC2SE hearts. The magnitude of protection matched IPC. IPC requited less S1365 phosphorylation, as TSC2SE hearts gained no benefit and failed to activate mTORC1 with IPC. IR metabolism was altered in TSC2SA, with increased mitochondrial oxygen consumption rate and glycolytic capacity (stressed/maximal extracellular acidification) after myocyte hypoxia-reperfusion. In whole heart, lactate increased and long-chain acylcarnitine levels declined during ischemia. The relative IR protection in TSC2SA was lost by lowering glucose in the perfusate by 36%. Adding fatty acid (palmitate) compensated for reduced glucose in wild type and TSC2SE but not TSC2SA which had the worst post-IR function under these conditions. TSC2-S1365 phosphorylation status regulates myocardial substrate utilization, and its decline activates mTORC1 biasing metabolism away from fatty acid oxidation to glycolysis to confer protection against IR. This pathway is also engaged and reduced TSC2 S1365 phosphorylation required for effective IPC. Graphic Abstract: A graphic abstract is available for this article.", "Chronic activation of brain-derived neurotrophic factor (BDNF) receptor TrkB is a potential method to prevent development of obesity, but the short half-life and nonbioavailable nature of BDNF hampers validation of the hypothesis. We report here that activation of muscular TrkB by the BDNF mimetic, 7,8-dihydroxyflavone (7,8-DHF), is sufficient to protect the development of diet-induced obesity in female mice. Using in vitro and in vivo models, we found that 7,8-DHF treatment enhanced the expression of uncoupling protein 1 (UCP1) and AMP-activated protein kinase (AMPK) activity in skeletal muscle, which resulted in increased systemic energy expenditure, reduced adiposity, and improved insulin sensitivity in female mice fed a high-fat diet. This antiobesity activity of 7,8-DHF is muscular TrkB-dependent as 7,8-DHF cannot mitigate diet-induced obesity in female muscle-specific TrkB knockout mice. Hence, our data reveal that chronic activation of muscular TrkB is useful in alleviating obesity and its complications." ]
Applications of molecular cytogenetics in domestic bovids	The discovery of the Robertsonian translocation (rob) involving cattle chromosomes 1 and 29 and the demonstration of its deleterious effects on fertility focused the interest of many scientific groups on using chromosome banding techniques to reveal chromosome abnormalities and verify their effects on fertility in domestic animals. At the same time, comparative banding studies among various species of domestic or wild animals were found useful for delineating chromosome evolution among species. The advent of molecular cytogenetics, particularly the use of fluorescence in situ hybridization (FISH), has allowed a deeper investigation of the chromosomes of domestic animals through: (a) the physical mapping of specific DNA sequences on chromosome regions; (b) the use of specific chromosome markers for the identification of the chromosomes or chromosome regions involved in chromosome abnormalities, especially when poor banding patterns are produced; (c) better anchoring of radiation hybrid and genetic maps to specific chromosome regions; (d) better comparisons of related and unrelated species by comparative FISH mapping and/or Zoo-FISH techniques; (e) the study of meiotic segregation, especially by sperm-FISH, in some chromosome abnormalities; (f) better demonstration of conserved or lost DNA sequences in chromosome abnormalities; (g) the use of informatic and genomic reconstructions, in addition to CGH arrays, to predict conserved or lost chromosome regions in related species; and (h) the study of some chromosome abnormalities and genomic stability using PCR applications. This review summarizes the most important applications of molecular cytogenetics in domestic bovids, with an emphasis on FISH mapping applications.	[ "Globally, cattle production has more than doubled since the 1960s, with widespread use of artificial insemination (AI) and an emphasis on a small pool of high genetic merit animals. Selecting AI bulls with optimal fertility is, therefore, vital, as impaired fertility reduces genetic gains and production, resulting in heavy financial and environmental losses. Chromosome translocations, particularly the 1;29 Robertsonian translocation, are a common cause of reduced fertility; however, reciprocal translocations are significantly underreported due to the difficulties inherent in analysing cattle chromosomes. Based on our porcine work, we have developed an approach for the unambiguous detection of Robertsonian and reciprocal translocations, using a multiple-hybridization probe detection strategy. We applied this method on the chromosomes of 39 bulls, detecting heterozygous and homozygous 1;29 translocations and a 12;23 reciprocal translocation in a total of seven animals. Previously, karyotype analysis was the only method of diagnosing chromosomal rearrangements in cattle, and was time-consuming and error-prone. With calving rates of only 50-60%, it is vital to reduce further fertility loss in order to maximise productivity. The approach developed here identifies abnormalities that DNA sequencing will not, and has the potential to lead to long-term gains, delivering meat and milk products in a more cost-effective and environmentally-responsible manner to a growing population.", "We report an extended river buffalo (Bubalus bubalis, 2n = 50; BBU) cytogenetic map including 388 loci, of which 68 have been FISH-mapped on autosomes in the present study. Ovine and caprine BAC clones containing both type I loci (known genes) and type II loci (simple sequence repeats (SRs), microsatellite marker, sequence-tagged sites (STSs)), previously assigned to sheep chromosomes, have been localized on R-banded river buffalo chromosomes (BBU), which expands the cytogenetic map of this important domestic species and increases our knowledge of the physical organization of its genome. The loci mapped in the present study correspond to loci already localized on homoeologous cattle (and sheep) chromosomes and chromosome bands, further confirming the high degree of chromosome homoeologies among bovids. The comparison of the integrated cytogenetic maps of BBU2p/BBU10 and BBU5p/BBU16 with those of human chromosomes (HSA) 6 and 11, respectively, identified, at least, nine conserved chromosome segments in each case and complex rearrangements differentiating river buffalo (and cattle) and human chromosomes.", "The development of a completely annotated sheep genome sequence is a key need for understanding the phylogenetic relationships and genetic diversity among the many different sheep breeds worldwide and for identifying genes controlling economically and physiologically important traits. The ovine genome sequence assembly will be crucial for developing optimized breeding programs based on highly productive, healthy sheep phenotypes that are adapted to modern breeding and production conditions. Scientists and breeders around the globe have been contributing to this goal by generating genomic and cDNA libraries, performing genome-wide and trait-associated analyses of polymorphism, expression analysis, genome sequencing, and by developing virtual and physical comparative maps. The International Sheep Genomics Consortium (ISGC), an informal network of sheep genomics researchers, is playing a major role in coordinating many of these activities. In addition to serving as an essential tool for monitoring chromosome abnormalities in specific sheep populations, ovine molecular cytogenetics provides physical anchors which link and order genome regions, such as sequence contigs, genes and polymorphic DNA markers to ovine chromosomes. Likewise, molecular cytogenetics can contribute to the process of defining evolutionary breakpoints between related species. The selective expansion of the sheep cytogenetic map, using loci to connect maps and identify chromosome bands, can substantially contribute to improving the quality of the annotated sheep genome sequence and will also accelerate its assembly. Furthermore, identifying major morphological chromosome anomalies and micro-rearrangements, such as gene duplications or deletions, that might occur between different sheep breeds and other Ovis species will also be important to understand the diversity of sheep chromosome structure and its implications for cross-breeding. To date, 566 loci have been assigned to specific chromosome regions in sheep and the new cytogenetic map is presented as part of this review. This review will also summarize the current cytogenomic status of the sheep genome, describe current activities in the sheep cytogenomics research sector, and will discuss the cytogenomics data in context with other major sheep genomics projects.", "Thirteen goat BAC clones containing coding sequences from HSA7, HSA12q, HSA4 and HSA6p were fluorescence in situ mapped to river buffalo (Bubalus bubalis, BBU) and sheep (Ovis aries, OAR) R-banded chromosomes. The following type I loci were mapped: BCP to BBU8q32 and OAR4q32, CLCN1 to BBU8q34 and OAR4q34, IGFBP3 to BBU8q24 and OAR4q27, KRT to BBU4q21 and OAR 3q21, IFNG to BBU4q23 and OAR3q23, IGF1 to BBU4q31 and OAR3q31, GNRHR to BBU7q32 and OAR6q32, MTP to BBU7q21 and OAR6q15, PDE6B to BBU7q36 and OAR6q36, BF to BBU2p22 and OAR20q22, EDN1 to BBU2p24 and OAR20q24, GSTA1 to BBU2p22 and OAR20q22, OLADRB (MHC) to BBU2p22 and OAR20q22. All mapped loci appeared to be located on homologous chromosomes and chromosome bands in both bovids. Comparison between gene orders in bovid (BBU and OAR) and human (HSA) chromosomes revealed complex rearrangements, especially between BBU7/OAR6 and HSA4, as well as between BBU2p/OAR20 and HSA6p.", "Aneuploidy is associated with spontaneous abortions, perinatal mortality, mental retardation and with embryonic and foetal mortality. Most of these abnormalities originate as a result of meiosis errors during gametogenesis. The main purpose of the study was to analyse frequency of aneuploidies of sex chromosomes and chromosome 6 by three-colour fluorescence in situ hybridization (FISH) in 47 young bulls, candidates for artificial insemination programme with cryopreserved semen and to investigate the influence of aneuploidies and disturbed sperm chromatin integrity on non-return rates, the frequencies of abortions, perinatal mortality and stillbirths. The average frequencies of spermatozoa with disomy for chromosomes X, Y, XY and 6 were 0.032, 0.005, 0.003 and 0.039% respectively. The incidence of XX66, YY66 and XY66 diploidy was 0.017, 0.006 and 0.015% respectively. Frequencies of meiotic II errors were significantly higher than meiotic I errors (p < 0.01). More X bearing spermatozoa than Y bearing spermatozoa were detected (5151 vs. 5022; p < 0.01). Sperm chromatin damage expressed by DNA fragmentation index (DFI) was 5.3 +/- 3.7 and percentage of cells with defective chromatin condensation (HDS) was 1.4 +/- 0.8. No correlation was found between sperm aneuploidy and basic sperm analysis. The relationship was found between non-return rate and total aneuploidy (r = -0.310; p = 0.036). Significant correlation was found between sex disomy, total aneuploidy (disomy of chromosomes 6, X, Y and XY spermatozoa and diploidy) and stillbirths (r = 0.390; p = 0.013; and r = 0.331; p = 0.037). Chromosome 6 disomy correlated with perinatal mortality (r = 0.317; p = 0.047). HDS correlated significantly with total aneuploidy (r = 0.449; p = 0.002). Our study indicated that aneuploidy frequencies in young fertile bull spermatozoa are relatively low. Nevertheless, there exists a variability in aneuploidy frequencies amongst bulls, which appears to be able to have an influence on the fertility of these animals.", "A genetic map of bovine Chromosome (Chr) 11 (BTA11, synteny group U16) has been constructed from 330 animals belonging to 21 families, which constitute the international bovine reference panel (IBRP). This map is based on 13 polymorphic microsatellite markers, two of which were chosen in previously published maps. Three markers have been isolated from cosmids. Two of the three cosmids have been physically localized by fluorescence in situ hybridization (FISH), to anchor the genetic map on the chromosome. In addition, a biallelic polymorphism in the beta-lactoglobulin gene (LGB) has been genetically positioned relative to the microsatellite markers. The most probable order of the markers is: cen-INRA044-BM716-INRA177-(TGLA327, INRA198, INRA131)-INRA111-INRABERN169-(INRA115, INRA032)-INRA108-INRABERN162-INRA195-LGB. The total linkage group spans 126 cM, which probably corresponds to most of the chromosome length. The average intermarker distance is about 10.5 cM, allowing the potential detection of a genetic linkage with any Economic Trait Loci (ETL) of this chromosome. Seven of these markers have been previously published by Vaiman and coworkers (1994), two will be published as part of a set of markers (Eggen et al. in preparation), two are described in this paper, and two (BM716, TGLA327) were chosen from the published maps of BTA11 in order to integrate our data with existing maps. All these markers were assigned to synteny group U16 by use of a previously characterized panel of hamster/bovine somatic hybrid cell lines (Guérin et al. 1994).(ABSTRACT TRUNCATED AT 250 WORDS)", "Thirteen male river buffaloes, 119 females with reproductive problems (which had reached reproductive age but had failed to become pregnant in the presence of bulls) and two male co-twins underwent both clinical and cytogenetic investigation. Clinical analyses performed by veterinary practitioners revealed normal body conformation and external genitalia for most females. However, some subjects showed some slight male traits such as large base horn circumference, prominent withers and tight pelvis. Rectal palpation revealed damage to internal sex adducts varying between atrophy of Mullerian ducts to complete lack of internal sex adducts (with closed vagina). All bulls had normal karyotypes at high resolution banding, while 25 animals (23 females and 2 male co-twins) (20.7%) with reproductive problems were found to carry the following sex chromosome abnormalities: X monosomy (2 females); X trisomy (1 female); sex reversal syndrome (2 females); and free-martinism (18 females and 2 males). All female carriers were sterile.", "A G- and R-banding comparison of cattle (Bos taurus, 2n = 60), goat (Capra hircus, 2n = 60) and sheep (Ovis aries, 2n = 54) chromosomes at the 450 band level was made. The study revealed a large number of banding homologies among the autosomes of the three species and resolved some ambiguities in arranging some of their small disputed acrocentrics by direct and indirect comparisons with some bovid marker chromosomes. A loss of the subcentromeric G-positive band in sheep chromosome 2g was observed when the G-banding patterns of sheep 2q and homologous cattle and goat chromosome 2 were compared. The chromosomal divergences among cattle, goat and river buffalo (Bubalus bubalis, 2n = 50) sex chromosomes are shown to have occurred by pericentric and paracentric inversions with a loss (or acquisition of constitutive heterochromatin.", "To identify physical and genetic anchor loci on bovine chromosomes, 13 cosmids, obtained after the screening of partial bovine cosmid libraries with the (CA)n microsatellite motif, were mapped by fluorescence in situ hybridization (FISH). Eleven cosmid probes yielded a specific signal on one of the bovine chromosomes and identified the following loci: D5S2, D5S3, D6S3, D8S1, D11S5, D13S1, D16S5, D17S2, D19S2, D19S3, D21S8. Two cosmids produced centromeric signals on many chromosomes. The microsatellite-containing regions were subcloned and sequenced. The sequence information revealed that the two centromeric cosmids were derived from bovine satellites 1.723 and 1.709, respectively. A cosmid located in the subtelomeric region of Chromosome (Chr) 17 (D17S2) had features of a chromosome-specific satellite. Primers were designed for eight of the nonsatellite cosmids, and seven of these microsatellites were polymorphic with between three and eight alleles on a set of outbred reference families. The polymorphic and chromosomally mapped loci can now be used to physically anchor other bovine polymorphic markers by linkage analysis. The microsatellite primers were also applied to DNA samples of a previously characterized panel of somatic hybrid cell lines, allowing the assignment of seven microsatellite loci to defined syntenic groups. These assignments confirmed earlier mapping results, revealed a probable case of false synteny, and placed two formerly unassigned syntenic groups on specific chromosomes.", "We report the most extensive genetic linkage map for a livestock species produced to date. We have linked 376 microsatellite (MS) loci with seven restriction fragment length polymorphic loci in a backcross reference population. The 383 markers were placed into 24 linkage groups which span 1997 cM. Seven additional MS did not fall into a linkage group. Linkage groups are assigned to 13 autosomes and the X chromosome (haploid n = 19). This map provides the basis for genetic analysis of quantitative inheritance of phenotypic and physiologic traits in swine." ]
Pathogenesis of diabetic foot ulcers.	Diabetic foot ulcers (DFUs) are associated with complex pathogenic factors and are considered a serious complication of diabetes. The potential mechanisms underlying DFUs have been increasingly investigated. Previous studies have focused on the three aspects of diabetic peripheral vascular disease, neuropathy, and wound infections. With advances in technology, researchers have been gradually conducting studies using immune cells, endothelial cells, keratinocytes, and fibroblasts, as they are involved in wound healing. It has been reported that the upregulation or downregulation of molecular signaling pathways is essential for the healing of DFUs. With a recent increase in the awareness of epigenetics, its regulatory role in wound healing has become a much sought-after trend in the treatment of DFUs. This review focuses on four aspects involved in the pathogenesis of DFUs: physiological and pathological mechanisms, cellular mechanisms, molecular signaling pathway mechanisms, and epigenetics. Given the challenge in the treatment of DFUs, we are hopeful that our review will provide new ideas for peers.	[ "Diabetic foot ulcer (DFU) caused by impaired wound healing is a common vascular complication of diabetes. The current study revealed that plasma levels of pigment epithelium-derived factor (PEDF) were elevated in type 2 diabetic patients with DFU and in db/db mice. To test whether elevated PEDF levels contribute to skin wound-healing delay in diabetes, endogenous PEDF was neutralized with an anti-PEDF antibody in db/db mice. Our results showed that neutralization of PEDF accelerated wound healing, increased angiogenesis in the wound skin, and improved the functions and numbers of endothelial progenitor cells (EPCs) in the diabetic mice. Further, PEDF-deficient mice showed higher baseline blood flow in the skin, higher density of cutaneous microvessels, increased skin thickness, improved numbers and functions of circulating EPCs, and accelerated wound healing compared with wild-type mice. Overexpression of PEDF suppressed the Wnt signaling pathway in the wound skin. Lithium chloride-induced Wnt signaling activation downstream of the PEDF interaction site attenuated the inhibitory effect of PEDF on EPCs and rescued the wound-healing deficiency in diabetic mice. Taken together, these results suggest that elevated circulating PEDF levels contribute to impaired wound healing in the process of angiogenesis and vasculogenesis through the inhibition of Wnt/β-catenin signaling.", "The developmental pathway toward diabetic foot ulceration is best described as multifactorial. A critical triad of neuropathy, minor foot trauma, and foot deformity has been identified in greater than 63% of foot ulcers. In addition to these risk factors, increased peak plantar pressures, autonomic and motor neuropathy, limited joint mobility, and impaired wound healing contribute to the formation of diabetic foot ulcers. It is essential to understand the pathogenesis of foot ulcers so that appropriate treatments may be developed.", "Transplantation of human cord blood-derived endothelial progenitor cells (EPCs) is reported to contribute to neovascularization in various ischemic diseases. However, the possible beneficial role and underlying mechanisms in diabetes-impaired wound healing have been less well characterized. In this study, EPC transplantation stimulated keratinocyte and fibroblast proliferation substantially as early as 3 days after injury, leading to significantly accelerated wound closure in streptozotocin-induced diabetic nude mice, compared to PBS control. RT-PCR analysis showed that EPCs secreted various wound healing-related growth factors. Among them, keratinocyte growth factor and platelet-derived growth factor were highly expressed in the EPCs and were present at substantial levels in the EPC-injected dermal tissue. Using EPC-conditioned medium (CM), we found that paracrine factors from EPCs directly exerted mitogenic and chemotactic effects on keratinocytes and fibroblasts. Moreover, injection of EPC-CM alone into the same diabetic wound mice promoted wound healing and increased neovascularization to a similar extent as achieved with EPC transplantation. These results indicate that the beneficial effect of EPC transplantation on diabetic wounds was mainly achieved by their direct paracrine action on keratinocytes, fibroblasts, and endothelial cells, rather than through their physical engraftment into host tissues (vasculogenesis). In addition, EPC-CM was shown to be therapeutically equivalent to EPCs, at least for the treatment of diabetic dermal wounds, suggesting that conditioned medium may serve as a novel therapeutic option that is free from allograft-associated immune rejection concern.", "Diabetic foot ulcers (DFUs) are a life-threatening disease that often result in lower limb amputations and a shortened lifespan. However, molecular mechanisms contributing to the pathogenesis of DFUs remain poorly understood. We use next-generation sequencing to generate a human dataset of pathogenic DFUs to compare to transcriptional profiles of human skin and oral acute wounds, oral as a model of \"ideal\" adult tissue repair due to accelerated closure without scarring. Here we identify major transcriptional networks deregulated in DFUs that result in decreased neutrophils and macrophages recruitment and overall poorly controlled inflammatory response. Transcription factors FOXM1 and STAT3, which function to activate and promote survival of immune cells, are inhibited in DFUs. Moreover, inhibition of FOXM1 in diabetic mouse models (STZ-induced and db/db) results in delayed wound healing and decreased neutrophil and macrophage recruitment in diabetic wounds in vivo. Our data underscore the role of a perturbed, ineffective inflammatory response as a major contributor to the pathogenesis of DFUs, which is facilitated by FOXM1-mediated deregulation of recruitment of neutrophils and macrophages, revealing a potential therapeutic strategy.", "The endothelium, a cellular monolayer lining the blood vessel wall, plays a critical role in maintaining multiorgan health and homeostasis. Endothelial functions in health include dynamic maintenance of vascular tone, angiogenesis, hemostasis, and the provision of an antioxidant, anti-inflammatory, and antithrombotic interface. Dysfunction of the vascular endothelium presents with impaired endothelium-dependent vasodilation, heightened oxidative stress, chronic inflammation, leukocyte adhesion and hyperpermeability, and endothelial cell senescence. Recent studies have implicated altered endothelial cell metabolism and endothelial-to-mesenchymal transition as new features of endothelial dysfunction. Endothelial dysfunction is regarded as a hallmark of many diverse human panvascular diseases, including atherosclerosis, hypertension, and diabetes. Endothelial dysfunction has also been implicated in severe coronavirus disease 2019. Many clinically used pharmacotherapies, ranging from traditional lipid-lowering drugs, antihypertensive drugs, and antidiabetic drugs to proprotein convertase subtilisin/kexin type 9 inhibitors and interleukin 1β monoclonal antibodies, counter endothelial dysfunction as part of their clinical benefits. The regulation of endothelial dysfunction by noncoding RNAs has provided novel insights into these newly described regulators of endothelial dysfunction, thus yielding potential new therapeutic approaches. Altogether, a better understanding of the versatile (dys)functions of endothelial cells will not only deepen our comprehension of human diseases but also accelerate effective therapeutic drug discovery. In this review, we provide a timely overview of the multiple layers of endothelial function, describe the consequences and mechanisms of endothelial dysfunction, and identify pathways to effective targeted therapies. SIGNIFICANCE STATEMENT: The endothelium was initially considered to be a semipermeable biomechanical barrier and gatekeeper of vascular health. In recent decades, a deepened understanding of the biological functions of the endothelium has led to its recognition as a ubiquitous tissue regulating vascular tone, cell behavior, innate immunity, cell-cell interactions, and cell metabolism in the vessel wall. Endothelial dysfunction is the hallmark of cardiovascular, metabolic, and emerging infectious diseases. Pharmacotherapies targeting endothelial dysfunction have potential for treatment of cardiovascular and many other diseases.", "Diabetes is primarily characterized by hyperglycemia, and its high incidence is often very costly to patients, their families, and national economies. Unsurprisingly, the number and function of endothelial progenitor cells (EPCs) decrease in patients resulting in diabetic wound non-healing. As precursors of endothelial cells (ECs), these cells were discovered in 1997 and found to play an essential role in wound healing. Their function, number, and role in wound healing has been widely investigated. Hitherto, a lot of complex molecular mechanisms have been discovered. In this review, we summarize the mechanisms of how hyperglycemia affects the function and number of EPCs and how the affected cells impact wound healing. We aim to provide a complete summary of the relationship between diabetic hyperglycosemia, EPCs, and wound healing, as well as a better comprehensive platform for subsequent related research.", "Macrophages are critical for the initiation and resolution of the inflammatory phase of wound repair. In diabetes, macrophages display a prolonged inflammatory phenotype in late wound healing. Mixed-lineage leukemia-1 (MLL1) has been shown to direct gene expression by regulating nuclear factor-κB (NF-κB)-mediated inflammatory gene transcription. Thus, we hypothesized that MLL1 influences macrophage-mediated inflammation in wound repair. We used a myeloid-specific Mll1 knockout (Mll1 ) to determine the function of MLL1 in wound healing. Mll1 mice display delayed wound healing and decreased wound macrophage inflammatory cytokine production compared with control animals. Furthermore, wound macrophages from Mll1 mice demonstrated decreased histone H3 lysine 4 trimethylation (H3K4me3) (activation mark) at NF-κB binding sites on inflammatory gene promoters. Of note, early wound macrophages from prediabetic mice displayed similarly decreased MLL1, H3K4me3 at inflammatory gene promoters, and inflammatory cytokines compared with controls. Late wound macrophages from prediabetic mice demonstrated an increase in MLL1, H3K4me3 at inflammatory gene promoters, and inflammatory cytokines. Prediabetic macrophages treated with an MLL1 inhibitor demonstrated reduced inflammation. Finally, monocytes from patients with type 2 diabetes had increased Mll1 compared with control subjects without diabetes. These results define an important role for MLL1 in regulating macrophage-mediated inflammation in wound repair and identify a potential target for the treatment of chronic inflammation in diabetic wounds." ]
Impact of high-level trauma centers on population access and injury mortality	Timely access to high-level (I/II) trauma centers (HLTCs) is essential to minimize mortality after injury. Over the last 15 years, there has been a proliferation of HLTC nationally. The current study evaluates the impact of additional HLTC on population access and injury mortality.	[ "This observational study assesses the association of a new trauma center with transport times for trauma patients as a measure of prompt access to care and specifically examines changes in racial, ethnic, and income disparities in transport times.", "Previous studies have reported that the number and distribution of trauma centers are uneven across states, suggesting large differences in access to trauma center care. To estimate the proportion of US residents having access to trauma centers within 45 and 60 minutes. Cross-sectional study using data from 2 national databases as part of the Trauma Resource Allocation Model for Ambulances and Hospitals (TRAMAH) project. Trauma centers, base helipads, and block group population were counted for all 50 states and the District of Columbia as of January 2005. Percentages of national, regional, and state populations having access to all 703 level I, II, and III trauma centers in the United States by either ground ambulance or helicopter within 45 and 60 minutes. An estimated 69.2% and 84.1% of all US residents had access to a level I or II trauma center within 45 and 60 minutes, respectively. The 46.7 million Americans who had no access within an hour lived mostly in rural areas, whereas the 42.8 million Americans who had access to 20 or more level I or II trauma centers within an hour lived mostly in urban areas. Within 45 and 60 minutes, respectively, 26.7% and 27.7% of US residents had access to level I or II trauma centers by helicopter only and 1.9% and 3.1% of US residents had access to level I or II centers only from trauma centers or base helipads outside their home states. Selecting trauma centers based on geographic need, appropriately locating medical helicopter bases, and establishing formal agreements for sharing trauma care resources across states should be considered to improve access to trauma care in the United States.", "The 'golden hour' is a well-known concept, suggesting that shortening time from injury to definitive care is critically important for better outcome of trauma patients. However, there was no established evidence to support it. We aimed to validate the association between time to definitive care and mortality in hemodynamically unstable patients for the current trauma care settings. The data were collected from the Japan Trauma Data Bank between 2006 and 2015. The inclusion criteria were patients with systolic blood pressure (SBP) <90 mm Hg and heart rate (HR) >110 beats/min or SBP <70 mm Hg who underwent definitive care within 4 hours from the onset of injury and survived for more than 4 hours. The outcome measure was in-hospital mortality. We evaluated the relationship between time to definitive care and mortality using the generalized additive model (GAM). Subgroup analysis was also conducted using GAM after dividing the patients into the severe (SBP <70 mm Hg) and moderate (SBP ≥70 mm Hg and <90 mm Hg, and HR >110 beats/min) shock group. 1169 patients were enrolled in this study. Of these, 386 (33.0%) died. Median time from injury to definitive care was 137 min. Only 61 patients (5.2%) received definitive care within 60 min. The GAM models demonstrated that mortality remained stable for the early phase, followed by a decrease over time. The severe shock group presented with a paradoxical decline of mortality with time, whereas the moderate shock group had a time-dependent increase in mortality. We did not observe the association of shorter time to definitive care with a decrease in mortality. However, this was likely an offset result of severe and moderate shock groups. The result indicated that early definitive care could have a positive impact on survival outcome of patients with moderate shock. Level Ⅳ, prognostic study.", "This study uses data from the American Trauma Society’s Trauma Information Exchange Program to evaluate trends in nationwide 60-minute access to American College of Surgeons Committee on Trauma–verified level I-IV trauma centers between 2013 and 2019.", "Rural, low-income, and historically underrepresented minority communities face substantial barriers to trauma care and experience high injury incidence and mortality rates. Characteristics of injury incident locations may contribute to poor injury outcomes. To examine the association of injury scene characteristics with injury mortality. In this cross-sectional study, data from trauma center and emergency medical services provided by emergency medical services companies and designated trauma centers in the state of Maryland from January 1, 2015, to December 31, 2015, were geocoded by injury incident locations and linked with injury scene characteristics. Participants included adults who experienced traumatic injury in Maryland and were transported to a designated trauma center or died while in emergency medical services care at the incident scene or in transit. The primary exposures of interest were geographic characteristics of injury incident locations, including distance to the nearest trauma center, designation level and ownership status of the nearest trauma center, and land use, as well as community-level characteristics such as median age and per capita income. Odds of death were estimated with multilevel logistic regression, controlling for individual demographic measures and measures of injury and health. Of the 16 082 patients included in this study, 8716 (52.4%) were white, and 5838 (36.3%) were African American. Most patients were male (10 582; 65.8%) and younger than 65 years (12 383; 77.0%). Odds of death increased by 8.0% for every 5-mile increase in distance to the nearest trauma center (OR, 1.08; 95% CI, 1.01-1.15; P = .03). Compared with privately owned level 1 or 2 centers, odds of death increased by 49.9% when the nearest trauma center was level 3 (OR, 1.50; 95% CI, 1.06-2.11; P = .02), and by 80.7% when the nearest trauma center was publicly owned (OR, 1.81; 95% CI, 1.39-2.34; P < .001). At the zip code tabulation area level, odds of death increased by 16.0% for every 5-year increase in median age (OR, 1.16; 95% CI, 1.03-1.30; P = .02), and decreased by 26.6% when the per capita income was greater than $25 000 (OR, 0.73; 95% CI, 0.54-0.99; P = .05). Injury scene characteristics are associated with injury mortality. Odds of death are highest for patients injured in communities with higher median age or lower per capita income and at locations farthest from level 1 or 2 trauma centers.", "Although death rates from injuries are higher in rural areas compared with large metropolitan areas, little is known about how non-fatal injury rates vary by rurality. Data from the 1997-2001 US National Health Interview Surveys were used to explore associations between rurality and non-fatal injury. A nationally representative survey. The annual injury rates per 1000 adults and 95% CIs were computed for medically attended injuries. Counties of residence were coded according to urban influence codes into four categories: large urban, small urban, suburban and rural. A linear-by-linear trend test was used to determine whether injury rates increase monotonically with county rurality. Logistic regression was used to control potential confounders. Compared with large urban counties, small urban counties experienced 8% higher injury odds (95% CI 1% to 15%); suburban counties 20% higher injury odds (95% CI 10% to 31%); and rural counties 30% higher injury odds (95% CI 17% to 43%) after adjusting for age, gender, marital status, education and health insurance. Rural residents had higher non-fatal injury rates than urban and suburban residents. Exploring this discrepancy can further contribute to new hypotheses regarding rural injury risk and ultimately lead to better suited interventions for rural residents.", "The purpose of this study is to evaluate the current evidence regarding the impact of the ACA on trauma outcomes and the financial impact on trauma patients and trauma systems. Traumatic injuries are the leading cause of death and disability between the ages of 1 to 47 years. Uninsured status has been associated with worse outcomes and higher financial strain. The Affordable Care Act (ACA) was signed into law with the aim of increasing health insurance coverage. Despite improvements in insured status, it is unclear how the ACA has affected trauma. We conducted a literature search using PubMed and Google Scholar for peer-reviewed studies investigating the impact of the ACA on trauma published between January 2017 and April 2021. Our search identified 20 studies that evaluated the impact of ACA implementation on trauma. The evidence suggests ACA implementation has been associated with increased postacute care access but not significant changes in trauma mortality. ACA implementation has been associated with a decreased likelihood of catastrophic health expenditures for trauma patients. ACA was also associated with an increase in overall reimbursement and amount billed for trauma visits, but a decrease in Medicaid reimbursement. Some improvements on the financial impact of ACA implementation on trauma patients and trauma systems have been shown, but studies are limited by methods of calculating costs and by inconsistent pre-/post-ACA timeframes. Further studies on cost-effectiveness and cost-benefit analysis will need to be conducted to definitively determine the impact of ACA on trauma." ]
How do I create a query that returns a list of values from a list?	A series of	[ "A series of nitric oxide donating acridone derivatives are synthesized and evaluated for in vitro cytotoxic activity against different sensitive and resistant cancer cell lines MCF7/Wt, MCF7/Mr (BCRP overexpression) and MCF7/Dx (P-gp expression). The results showed that NO-donating acridones are potent against both the sensitive and resistant cells. Structure activity relationship indicate that the nitric oxide donating moiety connected through a butyl chain at N(10) position as well as morpholino moiety linkage through an amide bridge on the acridone ring system at C-2 position, are required to exert a good cytotoxic effect. Further, good correlations were observed when cytotoxic properties were compared with in vitro nitric oxide release rate, nitric oxide donating group potentiated the cytotoxic effect of the acridone derivatives. Exogenous release of nitric oxide by NO donating acridones enhanced the accumulation of doxorubicin in MCF7/Dx cell lines when it was coadministered with doxorubicin, which inhibited the efflux process of doxorubicin. In summary, a nitric oxide donating group can potentiate the anti-MDR property of acridones.", "Mast cells, neutrophils, and macrophages are important inflammatory cells that have been implicated in the pathogenesis of acute and chronic inflammatory diseases. To explore a novel antiinflammatory agent, we have synthesized two types of acridines, 9-anilinoacridine and 9-phenoxyacridine derivatives, for evaluation on the grounds that acridine is a versatile heterocycle possessing a wide variety of biological properties. The title compounds were synthesized by reaction of 9-chloroacridine with appropriate Ar-NH(2) and Ar-OH, and their antiinflammatory activities on inhibitory effects on the activation of mast cells, neutrophils, and macrophages were studied. Three acridine derivatives 4, 10, and 11 were proved to be more potent than the reference inhibitor mepacrine for the inhibition of rat peritoneal mast cell degranulation with similar IC(50) values (16-21 microM). Compound 3 also showed potent inhibitory activity (IC(50) = 8.2 and 4.4 microM, respectively) for the secretion of lysosomal enzyme and beta-glucuronidase from neutrophils. Moreover, compounds 5 and 9 were shown to be efficacious inhibitors of TNF-alpha production in macrophage-like cell lines RAW 264.7. Compounds 2 and 12 were the potent inhibitors of TNF-alpha production in murine microglial cell lines N9. To further explore the cytotoxic properties of these acridine derivatives, (E)-12 was selected for NCI's in vitro disease-oriented tumor cells screen. The results indicated that this compound had no significant cytotoxicity with a mean GI(50) of 58.0 microM. These results indicated that the antiinflammatory effects of acridine derivatives were mediated, at least in part, through the suppression of chemical mediators released from mast cells, neutrophils, and macrophages and that these compounds have the potential to be novel antiinflammatory agents with no significant cytotoxicity.", "Despite the worldwide public health impact of malaria, neither the mechanism by which the Plasmodium parasite detoxifies and sequesters haem, nor the action of current antimalarial drugs is well understood. The haem groups released from the digestion of the haemoglobin of infected red blood cells are aggregated into an insoluble material called haemozoin or malaria pigment. Synthetic beta-haematin (FeIII-protoporphyrin-IX)2 is chemically, spectroscopically and crystallographically identical to haemozoin and is believed to consist of strands of FeIII-porphyrin units, linked into a polymer by propionate oxygen-iron bonds. Here we report the crystal structure of beta-haematin determined using simulated annealing techniques to analyse powder diffraction data obtained with synchrotron radiation. The molecules are linked into dimers through reciprocal iron-carboxylate bonds to one of the propionic side chains of each porphyrin, and the dimers form chains linked by hydrogen bonds in the crystal. This result has implications for understanding the action of current antimalarial drugs and possibly for the design of new therapeutic agents.", "Radioisotopic assays involve expense, multistep protocols, equipment, and radioactivity safety requirements which are problematic in high-throughput drug testing. This study reports an alternative, simple, robust, inexpensive, one-step fluorescence assay for use in antimalarial drug screening. Parasite growth is determined by using SYBR Green I, a dye with marked fluorescence enhancement upon contact with Plasmodium DNA. A side-by-side comparison of this fluorescence assay and a standard radioisotopic method was performed by testing known antimalarial agents against Plasmodium falciparum strain D6. Both assay methods were used to determine the effective concentration of drug that resulted in a 50% reduction in the observed counts (EC(50)) after 48 h of parasite growth in the presence of each drug. The EC(50)s of chloroquine, quinine, mefloquine, artemisinin, and 3,6-bis-epsilon-(N,N-diethylamino)-amyloxyxanthone were similar or identical by both techniques. The results obtained with this new fluorescence assay suggest that it may be an ideal method for high-throughput antimalarial drug screening.", "Sorafenib was the only small-molecule drug approved by FDA for treatment of the advanced hepatocellular carcinoma (HCC). Recent study indicated that YM155 was a promising agent for HCC cells with high survivin expression, however, the antitumor activity needs to be further improved. Based on molecular docking and rational design method, a series of multi-substituted benzyl acridone derivatives were designed and synthesized. MTT assay indicated that some of the synthesized compounds displayed better antiproliferative activity against HepG2 cells than YM155. Later study indicated that the representive compound 8u may directly interact with survivin protein and induce HepG2 cells apoptosis, which is different from YM155. In addition, ADME property was predicted in silico, and it performed well. Moreover, in vivo preliminary experiments showed that 8u may be a good lead compound in the treatment of HCC.", "Preventing and delaying the emergence of drug resistance is an essential goal of antimalarial drug development. Monotherapy and highly mutable drug targets have each facilitated resistance, and both are undesirable in effective long-term strategies against multi-drug-resistant malaria. Haem remains an immutable and vulnerable target, because it is not parasite-encoded and its detoxification during haemoglobin degradation, critical to parasite survival, can be subverted by drug-haem interaction as in the case of quinolines and many other drugs. Here we describe a new antimalarial chemotype that combines the haem-targeting character of acridones, together with a chemosensitizing component that counteracts resistance to quinoline antimalarial drugs. Beyond the essential intrinsic characteristics common to deserving candidate antimalarials (high potency in vitro against pan-sensitive and multi-drug-resistant Plasmodium falciparum, efficacy and safety in vivo after oral administration, inexpensive synthesis and favourable physicochemical properties), our initial lead, T3.5 (3-chloro-6-(2-diethylamino-ethoxy)-10-(2-diethylamino-ethyl)-acridone), demonstrates unique synergistic properties. In addition to 'verapamil-like' chemosensitization to chloroquine and amodiaquine against quinoline-resistant parasites, T3.5 also results in an apparently mechanistically distinct synergism with quinine and with piperaquine. This synergy, evident in both quinoline-sensitive and quinoline-resistant parasites, has been demonstrated both in vitro and in vivo. In summary, this innovative acridone design merges intrinsic potency and resistance-counteracting functions in one molecule, and represents a new strategy to expand, enhance and sustain effective antimalarial drug combinations." ]
Correlation between the MAPK7 gene and colorectal cancer risk in the Turkish population	Mitogen-activated protein kinases (MAPKs) are important regulatory molecules, which have essential roles in physiology and pathology. In the present study, we examined the possible correlation between the MAPK7 gene and colorectal cancer risk in the Turkish population.	[ "The occurrence of colorectal cancer is related to the interaction that takes place at several levels between hereditary factors, environmental and individual ones. Understanding the molecular basis is important because it can identify factors that contribute to the initiation of development, maintenance of progression but also determine the response or resistance to antitumor agents. Understanding colorectal cancer at the molecular level has provided data used for genetic tests of family forms, it defined predictive markers used to select patients susceptible to certain forms of therapy and also for the development of molecular diagnostic tests to detect early non-invasive cancers.", "Hereditary forms of colorectal cancer, as is the case with virtually all forms of hereditary cancer, show extensive phenotypic and genotypic heterogeneity, a phenomenon discussed throughout this special issue of Familial Cancer. Clearly, the family physician, oncology specialist, genetic counselor, and cancer geneticist must know fully the complexity of hereditary cancer syndromes, their differential diagnosis, in order to establish a diagnosis, direct highly-targeted surveillance and management, and then be able to communicate effectively with the molecular geneticist so that an at-risk patient's DNA can be tested in accord with the syndrome of concern. Thus, a family with features of the Lynch syndrome will merit microsatellite instability testing, consideration for immunohistochemistry evaluation, and mismatch repair gene testing, while, in contrast, a patient with FAP will require APC testing. However, other germline mutations, yet to be identified, may be important should testing for these mutations prove to be absent and, therein, unrewarding to the patient. Nevertheless, our position is that if the patient's family history is consistent with one of these syndromes, but a mutation is not found in the family, we still recommend the same surveillance and management strategies for patients from families with an established cancer-causing germline mutation. Our purpose in this paper is to provide a concise coverage of the major hereditary colorectal cancer syndromes, to discuss genetic counseling, molecular genetic evaluation, highly targeted surveillance and management, so that cancer control can be maximized for these high hereditary cancer risk patients.", "Wnts are a large family of proteins that participate in an array of cellular biological processes such as embryogenesis, organogenesis and tumor formation. These proteins bind to membrane receptor complexes comprised of a frizzled (Fz) G-protein-coupled receptor and other membrane co-receptors forming molecular groups that initiate, at least, three different intracellular signaling cascades leading to nuclear generation of transcription factors which regulate various cellular events. These events result in selective cellular differentiation, reduction or inhibition of the apoptotic mechanisms or changes in the biologic behavior of various cell lines. During the last decade, canonical Wnt signaling has been shown to play a significant role in the control of osteoblastogenesis and bone formation. In several clinical cases, mutations have been found in the Wnt receptor complexes that are associated with changes in bone mineral density and fractures. Loss-of-function mutations in LRP5 receptors cause, osteoporosis-pseudoglioma syndrome, while gain-of-function mutations in the same group lead to high bone mass phenotypes. Furthermore, osteocytes secrete proteins such as sclerostin, which blocks the membrane complex activation by Wnt, resulting in inhibition of bone formation. Studies of knockout and transgenic mouse models for Wnt pathway components have demonstrated that canonical signaling regulates most aspects of osteoblast physiology including commitment, differentiation, bone matrix formation/mineralization and apoptosis as well as coupling to osteoclastogenesis and bone resorption. Future studies in this rapidly growing area of research can possibly lead to the identification of targets of pharmacological intervention useful in the management of osteoporosis. In the present review we summarized the current knowledge related to the various components of the Wnt signaling pathway, the ways they cooperate in inducing and directing transcriptional functions as well as the interacting points with the TGFbeta superfamily. We also outlined a probable working integrating model of the mechanism of bone formation.", "Ionizing radiation, like a variety of other cellular stress factors, can activate or down-regulate multiple signaling pathways, leading to either increased cell death or increased cell proliferation. Modulation of the signaling process, however, depends on the cell type, radiation dose, and culture conditions. The mitogen-activated protein kinase (MAPK) pathway transduces signals from the cell membrane to the nucleus in response to a variety of different stimuli and participates in various intracellular signaling pathways that control a wide spectrum of cellular processes, including growth, differentiation, and stress responses, and is known to have a key role in cancer progression. Multiple signal transduction pathways stimulated by ionizing radiation are mediated by the MAPK superfamily including the extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 MAPK. The ERK pathway, activated by mitogenic stimuli such as growth factors, cytokines, and phorbol esters, plays a major role in regulating cell growth, survival, and differentiation. In contrast, JNK and p38 MAPK are weakly activated by growth factors but respond strongly to stress signals including tumor necrosis factor (TNF), interleukin-1, ionizing and ultraviolet radiation, hyperosmotic stress, and chemotherapeutic drugs. Activation of JNK and p38 MAPK by stress stimuli is strongly associated with apoptotic cell death. MAPK signaling is also known to potentially influence tumor cell radiosensitivity because of their activity associated with radiation-induced DNA damage response. This review will discuss the MAPK signaling pathways and their roles in cellular radiation responses.", "ERK5 is a protein kinase that also contains a nuclear localisation signal and a transcriptional transactivation domain. Inhibition of ERK5 has therapeutic potential in cancer and inflammation and this has prompted the development of ERK5 kinase inhibitors (ERK5i). However, few ERK5i programmes have taken account of the ERK5 transactivation domain. We have recently shown that the binding of small molecule ERK5i to the ERK5 kinase domain stimulates nuclear localisation and paradoxical activation of its transactivation domain. Other kinase inhibitors paradoxically activate their intended kinase target, in some cases leading to severe physiological consequences highlighting the importance of mitigating these effects. Here, we review the assays used to monitor ERK5 activities (kinase and transcriptional) in cells, the challenges faced in development of small molecule inhibitors to the ERK5 pathway, and classify the molecular mechanisms of paradoxical activation of protein kinases by kinase inhibitors.", "Micro-RNAs have been reported to play crucial roles in a diversity of cellular processes such as cell proliferation, differentiation and development by regulating the expression of specific genes. They have also been shown to play vital roles in several diseases such as cancer. In the present study, we investigated the role of miR-143-3p in breast cancer. Our results showed that the expression of miR-143-3p is significantly downregulated in breast cancer cells. Upregulation of miR-143-3p inhibited the proliferation and migration of breast cancer cells. Conversely, inhibition of miR-143-3p promoted the proliferation of cancer cells. Bioinformatics analysis and other several experiments revealed MAPK7 as the potential target of miR-143-3p. Quantitative RT-PCR showed that the expression of MAPK7 correlated well with the expression of miR-143. Moreover, the inhibition of MAPK 7 in breast cancer cells abrogated the effects of miR-143 indicating that miR-143-3p-exerted effects on breast cancer are mediated by MAPK7. Takentogether, these results provide strong clues about the therapeutic potential of miR-143-3p in the treatment of breast cancer.", "Microsatellite instability (MSI) is intrinsic to most colorectal carcinomas (CRC) from patients with hereditary nonpolyposis colorectal cancer (HNPCC), reflecting germline mutations in the mismatch-repair (MMR) genes. Its occurrence and chronological sequence of development in sporadic CRC appears less well defined. To explore the time sequence in acquisition of MSI, and the role it plays during tumor progression in sporadic CRC, we compared the incidence of MSI in tissue samples from 40 Dukes'-B and 30 Dukes'-D CRC patients with liver metastases, at 4 different microsatellite loci, representing sites on the APC, DCC and p53 genes respectively as well as the D2S123 site. Among the 30 patients with hepatic metastases, MSI was found in 9 (30%) of the primary, and 13 (43.3%) of the metastatic tumors. In comparison, among the 40 Dukes'-B CRC, MSI was found in only 8 cases (20%). CRC with MSI were more frequently located in the right colon, less frequently on the left side, and seldom in the rectum. Tumor ploidy analysis shows that 46.2% of Dukes'-D primary tumors with MSI are diploid (chi2 = 4.46, p = 0.035). With a mean follow-up time of 4.2 years for the Dukes'-B CRC, there were no recurrences in the 8 patients with MSI, whilst 6 (18.8%) relapses occurred amongst the 32 patients without MSI, average time to recurrence being 15 months. In Dukes'-D CRC, mean survival time for patients with MSI was 37 months (95% CI, 24 to 51 months), for those without MSI 26 months (95% CI, 18 to 35 months), although this was not statistically significant. Our data suggest that tumor progression may involve increased genetic instability." ]
Positive-roll bending of poly-dC:dG mononucleotide repeats	In recent decades, study of DNA structure has largely been focused on the interrelationships between nucleotides at the level of nearest neighbours. A little-utilized approach to probing structure on a larger scale is non-denaturing bisulfite modification of genomic DNA in conjunction with high-throughput sequencing. This technique revealed a marked gradient in reactivity increasing towards the 5' end of poly-dC:dG mononucleotide repeats as short as two base pairs, suggesting that access of the anion may be greater at these points due to positive-roll bending not predicted by existing models. Consistent with this, the 5' ends of these repeats are strikingly enriched at positions relative to the nucleosome dyad that bend towards the major groove, while their 3' ends tend to sit outside these areas. Mutation rates are also higher at the 5' ends of poly-dC:dG when CpG dinucleotides are excluded. These findings shed light on the mechanisms underlying bending/flexibility of the DNA double helix as well as the sequences that facilitate DNA packaging.	[ "For many aspects of DNA-protein interaction, it is vital to know how DNA bending rigidity (or persistence length, a) depends on its sequence. We addressed this problem using the method based on cyclization of short DNA fragments, which allows very accurate determination of a. Our approach was based on assigning specific values of a to each of 10 distinct dinucleotide steps. We prepared DNA fragments, each about 200 bp in length, with various quasi-periodic sequences, measured their cyclization efficiencies (j factors), and fitted the data by the theoretical equation to obtain the values of a for each fragment. From these data, we obtained a set of a for the dinucleotide steps. To test this set, we used it to design DNA sequences that should correspond to very low and very high values of a, prepared the corresponding fragments, and determined their values of a experimentally. The measured and calculated values of a were very close to one another, confirming that we have found the correct solution to this long-standing problem. The same experimental data also allowed us to determine the sequence dependence of DNA helical repeat.", "This study investigates the effect of Mg(2+) bound to the DNA major groove on DNA structure and dynamics. The analysis of a comprehensive dataset of B-DNA crystallographic structures shows that divalent cations are preferentially located in the DNA major groove where they interact with successive bases of (A/G)pG and the phosphate group of 5'-CpA or TpG. Based on this knowledge, molecular dynamics simulations were carried out on a DNA oligomer without or with Mg(2+) close to an ApG step. These simulations showed that the hydrated Mg(2+) forms a stable intra-strand cross-link between the two purines in solution. ApG generates an electrostatic potential in the major groove that is particularly attractive for cations; its intrinsic conformation is well-adapted to the formation of water-mediated hydrogen bonds with Mg(2+). The binding of Mg(2+) modulates the behavior of the 5'-neighboring step by increasing the BII (ε-ζ>0°) population of its phosphate group. Additional electrostatic interactions between the 5'-phosphate group and Mg(2+) strengthen both the DNA-cation binding and the BII character of the 5'-step. Cation binding in the major groove may therefore locally influence the DNA conformational landscape, suggesting a possible avenue for better understanding how strong DNA distortions can be stabilized in protein-DNA complexes.", "Mechanical deformations of DNA such as bending are ubiquitous and have been implicated in diverse cellular functions1. However, the lack of high-throughput tools to measure the mechanical properties of DNA has limited our understanding of how DNA mechanics influence chromatin transactions across the genome. Here we develop 'loop-seq'-a high-throughput assay to measure the propensity for DNA looping-and determine the intrinsic cyclizabilities of 270,806 50-base-pair DNA fragments that span Saccharomyces cerevisiae chromosome V, other genomic regions, and random sequences. We found sequence-encoded regions of unusually low bendability within nucleosome-depleted regions upstream of transcription start sites (TSSs). Low bendability of linker DNA inhibits nucleosome sliding into the linker by the chromatin remodeller INO80, which explains how INO80 can define nucleosome-depleted regions in the absence of other factors2. Chromosome-wide, nucleosomes were characterized by high DNA bendability near dyads and low bendability near linkers. This contrast increases for deeper gene-body nucleosomes but disappears after random substitution of synonymous codons, which suggests that the evolution of codon choice has been influenced by DNA mechanics around gene-body nucleosomes. Furthermore, we show that local DNA mechanics affect transcription through TSS-proximal nucleosomes. Overall, this genome-scale map of DNA mechanics indicates a 'mechanical code' with broad functional implications.", "Bowtie is an ultrafast, memory-efficient alignment program for aligning short DNA sequence reads to large genomes. For the human genome, Burrows-Wheeler indexing allows Bowtie to align more than 25 million reads per CPU hour with a memory footprint of approximately 1.3 gigabytes. Bowtie extends previous Burrows-Wheeler techniques with a novel quality-aware backtracking algorithm that permits mismatches. Multiple processor cores can be used simultaneously to achieve even greater alignment speeds. Bowtie is open source (http://bowtie.cbcb.umd.edu).", "DNA helical twist imposes geometric constraints on the location of histone-DNA interaction sites along nucleosomal DNA. Certain 10.5-bp periodic nucleotides in phase with these geometric constraints have been suggested to facilitate nucleosome positioning. However, the extent of nucleotide periodicity in nucleosomal DNA and its significance in directing nucleosome positioning still remain unclear. We clarify these issues by applying categorical spectral analysis to high-resolution nucleosome maps in two yeast species. We find that only a small fraction of nucleosomal sequences contain significant 10.5-bp periodicity. We further develop a spectral decomposition method to show that the previously observed periodicity in aligned nucleosomal sequences mainly results from proper phasing among nucleosomal sequences, and not from a preponderant occurrence of periodicity within individual sequences. Importantly, we show that this phasing may arise from the histones' proclivity for putting preferred nucleotides at some of the evenly spaced histone-DNA contact points with respect to the dyad axis. We demonstrate that 10.5-bp periodicity, when present, significantly facilitates rotational, but not translational, nucleosome positioning. Finally, although periodicity only moderately affects nucleosome occupancy genome wide, reduced periodicity is an evolutionarily conserved signature of nucleosome-depleted regions around transcription start/termination sites.", "DNA fragments which are intrinsically bent or curved migrate anomalously during electrophoresis through polyacrylamide gels. Starting with an initial population of approximately 10(12) unique DNA sequences, DNA which exhibited the kind of anomalous mobility associated with DNA bending was selected and enriched using a variation of the SELEX procedure. After seven rounds of selection and amplification, the vast majority of the remaining population of DNA fragments migrated as bent DNA. Cloning and sequencing of 30 individual sequences from this population has yielded information regarding the relationship between DNA sequence and bending. Some of the previous conclusions on DNA bending have been confirmed while others have been modified, by the results presented here. In addition, the dinucleotide base step CA/TG, which had not been thought to be a major factor in DNA bending, appears to be important.", "Cyclization of DNA with sticky ends is commonly used to measure DNA bendability as a function of length and sequence, but how its kinetics depend on the rotational positioning of the sticky ends around the helical axis is less clear. Here, we measured cyclization (looping) and decyclization (unlooping) rates (kloop and kunloop) of DNA with sticky ends over three helical periods (100-130 bp) using single-molecule fluorescence resonance energy transfer (FRET). kloop showed a nontrivial undulation as a function of DNA length whereas kunloop showed a clear oscillation with a period close to the helical turn of DNA (∼10.5 bp). The oscillation of kunloop was almost completely suppressed in the presence of gaps around the sticky ends. We explain these findings by modeling double-helical DNA as a twisted wormlike chain with a finite width, intrinsic curvature, and stacking interaction between the end base pairs. We also discuss technical issues for converting the FRET-based cyclization/decyclization rates to an equilibrium quantity known as the J factor that is widely used to characterize DNA bending mechanics.", "Nucleosome organization has been suggested to affect local mutation rates in the genome. However, the lack of de novo mutation and high-resolution nucleosome data has limited the investigation of this hypothesis. Additionally, analyses using indirect mutation rate measurements have yielded contradictory and potentially confounding results. Here, we combine data on >300,000 human de novo mutations with high-resolution nucleosome maps and find substantially elevated mutation rates around translationally stable ('strong') nucleosomes. We show that the mutational mechanisms affected by strong nucleosomes are low-fidelity replication, insufficient mismatch repair and increased double-strand breaks. Strong nucleosomes preferentially locate within young SINE/LINE transposons, suggesting that when subject to increased mutation rates, transposons are then more rapidly inactivated. Depletion of strong nucleosomes in older transposons suggests frequent positioning changes during evolution. The findings have important implications for human genetics and genome evolution.", "Recent studies of genome-wide nucleosomal organization suggest that the DNA sequence is one of the major determinants of nucleosome positioning. Although the search for underlying patterns encoded in nucleosomal DNA has been going on for about 30 years, our knowledge of these patterns still remains limited. Based on our evaluations of DNA deformation energy, we developed new scoring functions to predict nucleosome positioning. There are three principal differences between our approach and earlier studies: (i) we assume that the length of nucleosomal DNA varies from 146 to 147 bp; (ii) we consider the anisotropic flexibility of pyrimidine-purine (YR) dimeric steps in the context of their neighbors (e.g., YYRR versus RYRY); (iii) we postulate that alternating AT-rich and GC-rich motifs reflect sequence-dependent interactions between histone arginines and DNA in the minor groove. Using these functions, we analyzed 20 nucleosome positions mapped in vitro at single nucleotide resolution (including clones 601, 603, 605, the pGUB plasmid, chicken beta-globin and three 5S rDNA genes). We predicted 15 of the 20 positions with 1-bp precision, and two positions with 2-bp precision. The predicted position of the '601' nucleosome (i.e., the optimum of the computed score) deviates from the experimentally determined unique position by no more than 1 bp - an accuracy exceeding that of earlier predictions. Our analysis reveals a clear heterogeneity of the nucleosomal sequences which can be divided into two groups based on the positioning 'rules' they follow. The sequences of one group are enriched by highly deformable YR/YYRR motifs at the minor-groove bending sites SHL+/- 3.5 and +/- 5.5, which is similar to the alpha-satellite sequence used in most crystallized nucleosomes. Apparently, the positioning of these nucleosomes is determined by the interactions between histones H2A/H2B and the terminal parts of nucleosomal DNA. In the other group (that includes the '601' clone) the same YR/YYRR motifs occur predominantly at the sites SHL +/- 1.5. The interaction between the H3/H4 tetramer and the central part of the nucleosomal DNA is likely to be responsible for the positioning of nucleosomes of this group, and the DNA trajectory in these nucleosomes may differ in detail from the published structures. Thus, from the stereochemical perspective, the in vitro nucleosomes studied here follow either an X-ray-like pattern (with strong deformations in the terminal parts of nucleosomal DNA), or an alternative pattern (with the deformations occurring predominantly in the central part of the nucleosomal DNA). The results presented here may be useful for genome-wide classification of nucleosomes, linking together structural and thermodynamic characteristics of nucleosomes with the underlying DNA sequence patterns guiding their positions.", "We have used the emerging rules for the sequence dependence of DNA bendability to design and test a series of DNA molecules that incorporate strongly into nucleosomes. Competitive reconstitution experiments showed the superiority in histone octamer binding of DNA molecules in which segments consisting exclusively of A and T or G and C, separated by 2 base pairs (bp), are repeated with a 10-bp period. These repeated (A/T)3NN(G/C)3NN motifs are superior in nucleosome formation to natural positioning sequences and to other repeated motifs such as AANNNTTNNN and GGNNNCCNNN. Studies of different lengths of repetitive anisotropically flexible DNA showed that a segment of approximately 40 bp embedded in a 160-bp fragment is sufficient to generate nucleosome binding equivalent to that of natural nucleosome positioning sequences from 5S RNA genes. Bending requirements along the surface of the nucleosome seem to be quite constant, with no large jumps in binding free energy attributable to protein-induced kinks. The most favorable sequences incorporate into nucleosomes more strongly by 100-fold than bulk nucleosomal DNA, but differential bending free energies are small when normalized to the number of bends: a free energy difference of only about 100 cal/mol per bend (1 cal = 4.184 J) distinguishes the best bending sequences and bulk DNA. We infer that the distortion energy of DNA bending in the nucleosome is only weakly dependent on DNA sequence." ]
Towards a FAIR Framework for the Life Sciences	The life sciences are one of the biggest suppliers of scientific data. Reusing and connecting these data can uncover hidden insights and lead to new concepts. Efficient reuse of these datasets is strongly promoted when they are interlinked with a sufficient amount of machine-actionable metadata. While the FAIR (Findable, Accessible, Interoperable, Reusable) guiding principles have been accepted by all stakeholders, in practice, there are only a limited number of easy-to-adopt implementations available that fulfill the needs of data producers.	[ "The Center for Expanded Data Annotation and Retrieval (CEDAR) aims to revolutionize the way that metadata describing scientific experiments are authored. The software we have developed-the CEDAR Workbench-is a suite of Web-based tools and REST APIs that allows users to construct metadata templates, to fill in templates to generate high-quality metadata, and to share and manage these resources. The CEDAR Workbench provides a versatile, REST-based environment for authoring metadata that are enriched with terms from ontologies. The metadata are available as JSON, JSON-LD, or RDF for easy integration in scientific applications and reusability on the Web. Users can leverage our APIs for validating and submitting metadata to external repositories. The CEDAR Workbench is freely available and open-source.", "Biomedical ontologies provide essential domain knowledge to drive data integration, information retrieval, data annotation, natural-language processing and decision support. BioPortal (http://bioportal.bioontology.org) is an open repository of biomedical ontologies that provides access via Web services and Web browsers to ontologies developed in OWL, RDF, OBO format and Protégé frames. BioPortal functionality includes the ability to browse, search and visualize ontologies. The Web interface also facilitates community-based participation in the evaluation and evolution of ontology content by providing features to add notes to ontology terms, mappings between terms and ontology reviews based on criteria such as usability, domain coverage, quality of content, and documentation and support. BioPortal also enables integrated search of biomedical data resources such as the Gene Expression Omnibus (GEO), ClinicalTrials.gov, and ArrayExpress, through the annotation and indexing of these resources with ontologies in BioPortal. Thus, BioPortal not only provides investigators, clinicians, and developers 'one-stop shopping' to programmatically access biomedical ontologies, but also provides support to integrate data from a variety of biomedical resources.", "There is an urgent need to improve the infrastructure supporting the reuse of scholarly data. A diverse set of stakeholders-representing academia, industry, funding agencies, and scholarly publishers-have come together to design and jointly endorse a concise and measureable set of principles that we refer to as the FAIR Data Principles. The intent is that these may act as a guideline for those wishing to enhance the reusability of their data holdings. Distinct from peer initiatives that focus on the human scholar, the FAIR Principles put specific emphasis on enhancing the ability of machines to automatically find and use the data, in addition to supporting its reuse by individuals. This Comment is the first formal publication of the FAIR Principles, and includes the rationale behind them, and some exemplar implementations in the community." ]
HIV Incidence Estimates for Ending the HIV Epidemic	HIV incidence estimates are published each year for all Ending the HIV Epidemic (EHE) counties, but they are not stratified by the demographic variables highly associated with risk of infection. Regularly updated estimates of HIV incident diagnoses available at local levels are required to monitor the epidemic in the United States over time and could contribute to background incidence rate estimates for alternative clinical trial designs for new HIV prevention products.	[ "Despite the recent success of antiretrovirals for HIV prevention, additional, more effective, or more acceptable biomedical interventions will ultimately be needed to end the HIV epidemic. Designing clinical trials to evaluate the efficacy of new products that reduce HIV infection risk is challenging because of the existence of highly effective interventions to prevent HIV. However, the implementation of these interventions is uneven, and the fact that multiple HIV prevention efficacy trials are currently evaluating new products means the field confronts uncertainty in the emerging standard of prevention. In this Viewpoint, we take stock of the current state of HIV prevention, and subsequently discuss the key challenges in designing future trials to evaluate the next generation of HIV prevention products. We also highlight gaps in the knowledge base that need to be addressed to advance the design of research. Future trials are tenable, even in the context of existing and effective interventions, and should involve careful statistical approaches and multidisciplinary collaborative design.", "AIDSVu is a public resource for visualizing HIV surveillance data and other population-based information relevant to HIV prevention, care, policy, and impact assessment. The site, AIDSVu.org, aims to make data about the US HIV epidemic widely available, easily accessible, and locally relevant to inform public health decision making. AIDSVu develops visualizations, maps, and downloadable datasets using results from HIV surveillance systems, other population-based sources of information (eg, US Census and national probability surveys), and other data developed specifically for display and dissemination through the website (eg, pre-exposure prophylaxis [PrEP] prescriptions). Other types of content are developed to translate surveillance data into summarized content for diverse audiences using infographic panels, interactive maps, local and state fact sheets, and narrative blog posts. Over 10 years, AIDSVu.org has used an expanded number of data sources and has progressively provided HIV surveillance and related data at finer geographic levels, with current data resources providing HIV prevalence data down to the census tract level in many of the largest US cities. Data are available at the county level in 48 US states and at the ZIP Code level in more than 50 US cities. In 2019, over 500,000 unique users consumed AIDSVu data and resources, and HIV-related data and insights were disseminated through nearly 4,000,000 social media posts. Since AIDSVu's inception, at least 249 peer-reviewed publications have used AIDSVu data for analyses or referenced AIDSVu resources. Data uses have included targeting of HIV testing programs, identifying areas with inequitable PrEP uptake, including maps and data in academic and community grant applications, and strategically selecting locations for new HIV treatment and care facilities to serve high-need areas. Surveillance data should be actively used to guide and evaluate public health programs; AIDSVu translates high-quality, population-based data about the US HIV epidemic and makes that information available in formats that are not consistently available in surveillance reports. Bringing public health surveillance data to an online resource is a democratization of data, and presenting information about the HIV epidemic in more visual formats allows diverse stakeholders to engage with, understand, and use these important public health data to inform public health decision making.", "Trials of candidate agents for HIV pre-exposure prophylaxis (PrEP) might randomly assign participants to be given a new PrEP agent or oral coformulated tenofovir disoproxil fumarate plus emtricitabine. This design presents unique challenges in interpretation. First, with two active arms, HIV incidence might be low. Second, the effectiveness of tenofovir disoproxil fumarate plus emtricitabine varies across populations; thus, similar HIV incidence between groups could be consistent with a wide range of effectiveness for the new PrEP. We propose a two-part approach to trial results. First, we use Bayesian methods to incorporate assumptions about the background incidence of HIV in the trial in the absence of PrEP, possibly augmented by external data. On the basis of the estimated background incidence, we estimate and compare the number of averted (or prevented) HIV infections in each of the two trial groups, calculating the averted infections ratio. We apply these methods to a completed trial of tenofovir alafenamide plus emtricitabine for PrEP. Our framework shows that leveraging external information to estimate averted infections and the averted infections ratio enhances the efficiency and interpretation of active-controlled PrEP trials.", "Stan is a probabilistic programming language for specifying statistical models. A Stan program imperatively defines a log probability function over parameters conditioned on specified data and constants. As of version 2.14.0, Stan provides full Bayesian inference for continuous-variable models through Markov chain Monte Carlo methods such as the No-U-Turn sampler, an adaptive form of Hamiltonian Monte Carlo sampling. Penalized maximum likelihood estimates are calculated using optimization methods such as the limited memory Broyden-Fletcher-Goldfarb-Shanno algorithm. Stan is also a platform for computing log densities and their gradients and Hessians, which can be used in alternative algorithms such as variational Bayes, expectation propagation, and marginal inference using approximate integration. To this end, Stan is set up so that the densities, gradients, and Hessians, along with intermediate quantities of the algorithm such as acceptance probabilities, are easily accessible. Stan can be called from the command line using the cmdstan package, through R using the rstan package, and through Python using the pystan package. All three interfaces support sampling and optimization-based inference with diagnostics and posterior analysis. rstan and pystan also provide access to log probabilities, gradients, Hessians, parameter transforms, and specialized plotting." ]
Acute myeloid leukemia measurable residual disease	Preventing relapse for adults with acute myeloid leukemia (AML) in first remission is the most common indication for allogeneic hematopoietic cell transplant. The presence of AML measurable residual disease (MRD) has been associated with higher relapse rates, but testing is not standardized.	[ "Patients with acute myeloid leukemia (AML) and a FLT3 mutation have poor outcomes. We conducted a phase 3 trial to determine whether the addition of midostaurin - an oral multitargeted kinase inhibitor that is active in patients with a FLT3 mutation - to standard chemotherapy would prolong overall survival in this population. We screened 3277 patients, 18 to 59 years of age, who had newly diagnosed AML for FLT3 mutations. Patients were randomly assigned to receive standard chemotherapy (induction therapy with daunorubicin and cytarabine and consolidation therapy with high-dose cytarabine) plus either midostaurin or placebo; those who were in remission after consolidation therapy entered a maintenance phase in which they received either midostaurin or placebo. Randomization was stratified according to subtype of FLT3 mutation: point mutation in the tyrosine kinase domain (TKD) or internal tandem duplication (ITD) mutation with either a high ratio (>0.7) or a low ratio (0.05 to 0.7) of mutant to wild-type alleles (ITD [high] and ITD [low], respectively). Allogeneic transplantation was allowed. The primary end point was overall survival. A total of 717 patients underwent randomization; 360 were assigned to the midostaurin group, and 357 to the placebo group. The FLT3 subtype was ITD (high) in 214 patients, ITD (low) in 341 patients, and TKD in 162 patients. The treatment groups were well balanced with respect to age, race, FLT3 subtype, cytogenetic risk, and blood counts but not with respect to sex (51.7% in the midostaurin group vs. 59.4% in the placebo group were women, P=0.04). Overall survival was significantly longer in the midostaurin group than in the placebo group (hazard ratio for death, 0.78; one-sided P=0.009), as was event-free survival (hazard ratio for event or death, 0.78; one-sided P=0.002). In both the primary analysis and an analysis in which data for patients who underwent transplantation were censored, the benefit of midostaurin was consistent across all FLT3 subtypes. The rate of severe adverse events was similar in the two groups. The addition of the multitargeted kinase inhibitor midostaurin to standard chemotherapy significantly prolonged overall and event-free survival among patients with AML and a FLT3 mutation. (Funded by the National Cancer Institute and Novartis; ClinicalTrials.gov number, NCT00651261 .).", "Measurable residual disease (MRD; previously termed minimal residual disease) is an independent, postdiagnosis, prognostic indicator in acute myeloid leukemia (AML) that is important for risk stratification and treatment planning, in conjunction with other well-established clinical, cytogenetic, and molecular data assessed at diagnosis. MRD can be evaluated using a variety of multiparameter flow cytometry and molecular protocols, but, to date, these approaches have not been qualitatively or quantitatively standardized, making their use in clinical practice challenging. The objective of this work was to identify key clinical and scientific issues in the measurement and application of MRD in AML, to achieve consensus on these issues, and to provide guidelines for the current and future use of MRD in clinical practice. The work was accomplished over 2 years, during 4 meetings by a specially designated MRD Working Party of the European LeukemiaNet. The group included 24 faculty with expertise in AML hematopathology, molecular diagnostics, clinical trials, and clinical medicine, from 19 institutions in Europe and the United States.", "Genomic analysis is essential for risk stratification in patients with acute myeloid leukemia (AML) or myelodysplastic syndromes (MDS). Whole-genome sequencing is a potential replacement for conventional cytogenetic and sequencing approaches, but its accuracy, feasibility, and clinical utility have not been demonstrated. We used a streamlined whole-genome sequencing approach to obtain genomic profiles for 263 patients with myeloid cancers, including 235 patients who had undergone successful cytogenetic analysis. We adapted sample preparation, sequencing, and analysis to detect mutations for risk stratification using existing European Leukemia Network (ELN) guidelines and to minimize turnaround time. We analyzed the performance of whole-genome sequencing by comparing our results with findings from cytogenetic analysis and targeted sequencing. Whole-genome sequencing detected all 40 recurrent translocations and 91 copy-number alterations that had been identified by cytogenetic analysis. In addition, we identified new clinically reportable genomic events in 40 of 235 patients (17.0%). Prospective sequencing of samples obtained from 117 consecutive patients was performed in a median of 5 days and provided new genetic information in 29 patients (24.8%), which changed the risk category for 19 patients (16.2%). Standard AML risk groups, as defined by sequencing results instead of cytogenetic analysis, correlated with clinical outcomes. Whole-genome sequencing was also used to stratify patients who had inconclusive results by cytogenetic analysis into risk groups in which clinical outcomes were measurably different. In our study, we found that whole-genome sequencing provided rapid and accurate genomic profiling in patients with AML or MDS. Such sequencing also provided a greater diagnostic yield than conventional cytogenetic analysis and more efficient risk stratification on the basis of standard risk categories. (Funded by the Siteman Cancer Research Fund and others.).", "The role of decentralized assessment of measurable residual disease (MRD) for risk stratification in acute myeloid leukemia (AML) remains largely unknown, and so it does which methodological aspects are critical to empower the evaluation of MRD with prognostic significance, particularly if using multiparameter flow cytometry (MFC). We analyzed 1076 AML patients in first remission after induction chemotherapy, in whom MRD was evaluated by MFC in local laboratories of 60 Hospitals participating in the PETHEMA registry. We also conducted a survey on technical aspects of MRD testing to determine the impact of methodological heterogeneity in the prognostic value of MFC. Our results confirmed the recommended cutoff of 0.1% to discriminate patients with significantly different cumulative-incidence of relapse (-CIR- HR:0.71, P < 0.001) and overall survival (HR: 0.73, P = 0.001), but uncovered the limited prognostic value of MFC based MRD in multivariate and recursive partitioning models including other clinical, genetic and treatment related factors. Virtually all aspects related with methodological, interpretation, and reporting of MFC based MRD testing impacted in its ability to discriminate patients with different CIR. Thus, this study demonstrated that \"real-world\" assessment of MRD using MFC is prognostic in patients at first remission, and urges greater standardization for improved risk-stratification toward clinical decisions in AML.", "This phase 1b trial investigated several doses and schedules of midostaurin in combination with daunorubicin and cytarabine induction and high-dose cytarabine post-remission therapy in newly diagnosed patients with acute myeloid leukemia (AML). The discontinuation rate on the 50-mg twice-daily dose schedule was lower than 100 mg twice daily, and no grade 3/4 nausea or vomiting was seen. The complete remission rate for the midostaurin 50-mg twice-daily dose schedule was 80% (FMS-like tyrosine kinase 3 receptor (FLT3)-wild-type: 20 of 27 (74%), FLT3-mutant: 12 of 13 (92%)). Overall survival (OS) probabilities of patients with FLT3-mutant AML at 1 and 2 years (0.85 and 0.62, respectively) were similar to the FLT3-wild-type population (0.78 and 0.52, respectively). Midostaurin in combination with standard chemotherapy demonstrated high complete response and OS rates in newly diagnosed younger adults with AML, and was generally well tolerated at 50 mg twice daily for 14 days. A phase III prospective trial is ongoing (CALGB 10603, NCT00651261)." ]
Endoscopic mucosal resection with SOUTEN (PEMR-S) for colorectal lesions	SOUTEN (KANEKA Co., Tokyo, Japan) is a unique snare with a disk tip. We analyzed the efficacy of precutting endoscopic mucosal resection with SOUTEN (PEMR-S) for colorectal lesions.	[ "A proportion of neoplastic polyps are incompletely resected, resulting in local recurrence, especially after resection of large polyps or piecemeal resection. Local recurrences that develop after endoscopic resection of intramucosal neoplasms that lacked risk factors for lymph node metastasis or positive vertical margins are usually treated endoscopically. Endoscopic submucosal dissection (ESD) is indicated for local residual or recurrent early carcinomas after endoscopic resection. However, ESD for such recurrent lesions is technically difficult and is typically a lengthy procedure. Underwater endoscopic mucosal resection (UEMR), which was developed in 2012, is suitable for recurrent or residual lesions and reportedly achieves superior en bloc resection rates and endoscopic complete resection rates than conventional EMR. However, a large recurrent lesion is a negative independent predictor of successful en bloc resection and of complete endoscopic removal. We therefore perform UEMR for relatively small (≤ 10-15 mm) recurrent lesions and ESD for larger lesions.", "We analyzed the efficacy of precutting endoscopic mucosal resection (EMR), which is a method of making a full or partial circumferential mucosal incision around a tumor with a snare tip for en bloc resection. We reviewed cases from 2011 to 2018 in which precutting EMR (n = 167) and standard EMR (n = 557) were performed for lesions of 10 - 30 mm. Precutting EMR was indicated for benign lesions of 20 - 30 mm or lesions of < 20 mm for which standard EMR was difficult. Through propensity score matching of the two groups, the therapeutic outcomes for 35 lesions of ≥ 20 mm and 98 lesions of < 20 mm in each group were analyzed. In the two sizes of lesion, there were significant differences between the precutting and standard groups in the en bloc resection rate (≥ 20 mm 88.6 % vs. 48.5 %, P < 0.001; < 20 mm 98.0 % vs. 85.7 %, P = 0.004) and the histological complete resection rate (≥ 20 mm 71.4 % vs. 42.9 %, P = 0.02; < 20 mm 87.8 % vs. 67.3 %, P < 0.001). Precutting EMR enabled high en bloc resection rates in cases involving difficult lesions.", "Endoscopic mucosal resection (EMR) is currently the most used technique for resection of large distal colorectal polyps. However, in large lesions EMR can often only be performed in a piecemeal fashion resulting in relatively low radical (R0)-resection rates and high recurrence rates. Endoscopic submucosal dissection (ESD) is a newer procedure that is more difficult resulting in a longer procedural time, but is promising due to the high en-bloc resection rates and the very low recurrence rates. We aim to evaluate the (cost-)effectiveness of ESD against EMR on both short (i.e. 6 months) and long-term (i.e. 36 months). We hypothesize that in the short-run ESD is more time consuming resulting in higher healthcare costs, but is (cost-) effective on the long-term due to lower patients burden, a higher number of R0-resections and lower recurrence rates with less need for repeated procedures. This is a multicenter randomized clinical trial in patients with a non-pedunculated polyp larger than 20 mm in the rectum, sigmoid, or descending colon suspected to be an adenoma by means of endoscopic assessment. Primary endpoint is recurrence rate at follow-up colonoscopy at 6 months. Secondary endpoints are R0-resection rate, perceived burden and quality of life, healthcare resources utilization and costs, surgical referral rate, complication rate and recurrence rate at 36 months. Quality-adjusted-life-year (QALY) will be estimated taking an area under the curve approach and using EQ-5D-indexes. Healthcare costs will be calculated by multiplying used healthcare services with unit prices. The cost-effectiveness of ESD against EMR will be expressed as incremental cost-effectiveness ratios (ICER) showing additional costs per recurrence free patient and as ICER showing additional costs per QALY. If this trial confirms ESD to be favorable on the long-term, the burden of extra colonoscopies and repeated procedures can be prevented for future patients. NCT02657044 (Clinicaltrials.gov), registered January 8, 2016.", "Many clinical studies on narrow-band imaging (NBI) magnifying endoscopy classifications advocated so far in Japan (Sano, Hiroshima, Showa, and Jikei classifications) have reported the usefulness of NBI magnifying endoscopy for qualitative and quantitative diagnosis of colorectal lesions. However, discussions at professional meetings have raised issues such as: (i) the presence of multiple terms for the same or similar findings; (ii) the necessity of including surface patterns in magnifying endoscopic classifications; and (iii) differences in the NBI findings in elevated and superficial lesions. To resolve these problems, the Japan NBI Expert Team (JNET) was constituted with the aim of establishing a universal NBI magnifying endoscopic classification for colorectal tumors (JNET classification) in 2011. Consensus was reached on this classification using the modified Delphi method, and this classification was proposed in June 2014. The JNET classification consists of four categories of vessel and surface pattern (i.e. Types 1, 2A, 2B, and 3). Types 1, 2A, 2B, and 3 are correlated with the histopathological findings of hyperplastic polyp/sessile serrated polyp (SSP), low-grade intramucosal neoplasia, high-grade intramucosal neoplasia/shallow submucosal invasive cancer, and deep submucosal invasive cancer, respectively.", "Endoscopic mucosal resection (EMR) is effective for large laterally spreading flat and sessile lesions (LSLs). Sessile serrated adenomas/polyps (SSA/Ps) are linked to the relative failure of colonoscopy to prevent proximal colorectal cancer. We aimed to examine the technical success, adverse events and recurrence following EMR for large SSA/Ps in comparison with large conventional adenomas. Over 74 months till August 2014, prospective multicentre data of LSLs ≥20 mm were analysed. A standardised dye-based conventional EMR technique followed by scheduled surveillance colonoscopy was used. From a total of 2000 lesions, 323 SSA/Ps in 246 patients and 1527 adenomas in 1425 patients were included for analysis. Technical success for EMR was superior in SSA/Ps compared with adenomas (99.1% vs 94.5%, p<0.001). Significant bleeding and perforation were similar in both cohorts. The cumulative recurrence rates for adenomas after 6, 12, 18 and 24 months were 16.1%, 20.4%, 23.4% and 28.4%, respectively. For SSA/Ps, they were 6.3% at 6 months and 7.0% from 12 months onwards (p<0.001). Following multivariable adjustment, the HR of recurrence for adenomas versus SSA/Ps was 1.7 (95% CI 0.9 to 3.0, p=0.097). Subgroup analysis by lesion size revealed an eightfold increased risk of recurrence for 20-25 mm adenomas versus SSA/Ps, but no significantly different risk between lesion types in larger lesion groups. Recurrence after EMR of 20-25 mm LSLs is significantly less frequent in SSA/Ps compared with adenomatous lesions. SSA/Ps can be more effectively removed than adenomatous LSLs with equivalent safety. Ensuring complete initial resection is imperative for avoiding recurrence. ClinicalTrials.gov NCT01368289." ]
Prognostic gene model and competing endogenous RNA network in pancreatic adenocarcinoma.	Recent studies have indicated that pyroptosis may participate in the regulation of tumorigenesis and immune microenvironment. However, the role of pyroptosis-related genes (PRGs) in pancreatic adenocarcinoma (PAAD) remains unclear. Through multiple bioinformatics analysis, we constructed a prognostic gene model and competing endogenous RNA network. The correlation between PRGs and prognosis, immune infiltration, immune checkpoints, and tumor mutational burden was analyzed by Kaplan-Meier curve, univariate Cox, multivariate regression, and Spearman's analysis in PAAD patients. The qRT-PCR, Western blotting, CCK-8, Wound healing, and Transwell assay were applied to examine the role of CASP6 in PANC-1 cell. Thirty-one PRGs were upregulated in PAAD. Functional enrichment analysis revealed that the PRGs were mainly involved in pyroptosis, NOD-like receptor signaling pathway, and response to bacteria. We established a novel 4-gene signature related to PRGs for evaluating the prognosis of PAAD patients. Patients with PAAD in the low-risk group had a better prognosis than those in the high-risk group. The nomogram suggested that the 1-, 3-, and 5-years survival probability exhibited robust predictive performance. Significant correlation was observed between prognostic PRGs and immune infiltration, immune checkpoints, and tumor mutational burden. We first identified the potential competing endogenous RNA regulatory axis in PAAD: lncRNA PVT1/hsa-miR-16-5p/CASP6/CASP8. Moreover, knockdown of CASP6 dramatically inhibited the proliferation, migration, and invasion ability of PANC-1 cell in vitro. In conclusion, CASP6 could be a potential biomarker, promoting the occurrence and progression in PAAD. The lncRNA PVT1/hsa-miR-16-5p/CASP6/CASP8 regulatory axis plays an vital role in regulating the anti-tumor immune responses for PAAD.	[ "Hepatocellular carcinoma (HCC) is the second most lethal malignant tumor because of its significant heterogeneity and complicated molecular pathogenesis. Novel prognostic biomarkers are urgently needed because no effective and reliable prognostic biomarkers currently exist for HCC patients. Increasing evidence has revealed that pyroptosis plays a role in the occurrence and progression of malignant tumors. However, the relationship between pyroptosis-related genes (PRGs) and HCC patient prognosis remains unclear. In this study, 57 PRGs were obtained from previous studies and GeneCards. The gene expression profiles and clinical data of HCC patients were acquired from public data portals. Least absolute shrinkage and selection operator (LASSO) Cox regression analysis was performed to establish a risk model using TCGA data. Additionally, the risk model was further validated in an independent ICGC dataset. Our results showed that 39 PRGs were significantly differentially expressed between tumor and normal liver tissues in the TCGA cohort. Functional analysis confirmed that these PRGs were enriched in pyroptosis-related pathways. According to univariate Cox regression analysis, 14 differentially expressed PRGs were correlated with the prognosis of HCC patients in the TCGA cohort. A risk model integrating two PRGs was constructed to classify the patients into different risk groups. Poor overall survival was observed in the high-risk group of both TCGA (p < 0.001) and ICGC (p < 0.001) patients. Receiver operating characteristic curves demonstrated the accuracy of the model. Furthermore, the risk score was confirmed as an independent prognostic indicator via multivariate Cox regression analysis (TCGA cohort: HR = 3.346, p < 0.001; ICGC cohort: HR = 3.699, p < 0.001). Moreover, the single-sample gene set enrichment analysis revealed different immune statuses between high- and low-risk groups. In conclusion, our new pyroptosis-related risk model has potential application in predicting the prognosis of HCC patients.", "Programmed cell death (PCD) refers to the way in which cells die depending on specific genes encoding signals or activities. Apoptosis, autophagy, and pyroptosis are all mechanisms of PCD. Among these mechanisms, pyroptosis is mediated by the gasdermin family, accompanied by inflammatory and immune responses. The relationship between pyroptosis and cancer is complex, and the effects of pyroptosis on cancer vary in different tissues and genetic backgrounds. On one hand, pyroptosis can inhibit the occurrence and development of tumors; on the other hand, as a type of proinflammatory death, pyroptosis can form a suitable microenvironment for tumor cell growth and thus promote tumor growth. In addition, the induction of tumor pyroptosis is also considered a potential cancer treatment strategy. Studies have shown that DFNA5 (nonsyndromic hearing impairment protein 5)/GSDME (Gasdermin-E) mRNA methylation results in lower expression levels of DFNA5/GSDME in most tumor cells than in normal cells, making it difficult to activate the pyroptosis in most tumor cells. During the treatment of malignant tumors, appropriate chemotherapeutic drugs can be selected according to the expression levels of DFNA5/GSDME, which can be upregulated in tumor cells, thereby increasing the sensitivity to chemotherapeutic drugs and reducing drug resistance. Therefore, induced pyroptosis may play a predominant role in the treatment of cancer. Here, we review the latest research on the anti- and protumor effects of pyroptosis and its potential applications in cancer treatment.", "Inflammasome activation leads to caspase-1 activation, which causes the maturation and secretion of pro-IL-1β and pro-IL-18 among other substrates. A subgroup of the NLR (nucleotide-binding domain, leucine-rich repeat containing) proteins are key mediators of the inflammasome. Studies of gene-deficient mice and cells have implicated NLR inflammasomes in a host of responses to a wide range of microbial pathogens, inflammatory diseases, cancer, and metabolic and autoimmune disorders. Determining exactly how the inflammasome is activated in these diseases and disease models remains a challenge. This review presents and integrates recent progress in the field.", "Gasdermin is a recently identified family of pore-forming proteins consisting of Gasdermin A (GSDMA), Gasdermin B (GSDMB), Gasdermin C (GSDMC), Gasdermin D (GSDMD), Gasdermin E (GSDME), and DFNB59. Gasdermin D (GSDMD) is a downstream effector of inflammasomes, which are supramolecular complexes that activate inflammatory caspases (-1, -4, and -5 in human and -1 and -11 in mouse). GSDMD contains a functionally important N-terminal domain (GSDMD-N), a C-terminal domain, and a linker in between that is recognized and cleaved by the activated inflammatory caspases. Upon cleavage, the GSDMD-N fragments translocate on the membrane and oligomerize to form membrane-embedded pores after specifically binding to acidic lipids such as phosphatidylinositol phosphates (PIPs), phosphatidic acid (PA), phosphatidylserine (PS), and cardiolipin. The pore exhibits strong membrane-disrupting cytotoxicity in mammalian cells by disrupting the osmotic potential and also serves as a gate for extracellular release of mature IL-1β and IL-18 during pyroptosis. In this chapter, we review our current understanding of GSDM proteins in physiological and pathological cell death, with more focused discussions on its structural basis for GSDM activation and pore formation.", "A hallmark of Entamoeba histolytica (Eh) invasion in the gut is acute inflammation dominated by the secretion of pro-inflammatory cytokines TNF-α and IL-1β. This is initiated when Eh in contact with macrophages in the lamina propria activates caspase-1 by recruiting the NLRP3 inflammasome complex in a Gal-lectin and EhCP-A5-dependent manner resulting in the maturation and secretion of IL-1β and IL-18. Here, we interrogated the requirements and mechanisms for Eh-induced caspase-4/1 activation in the cleavage of gasdermin D (GSDMD) to regulate bioactive IL-1β release in the absence of cell death in human macrophages. Unlike caspase-1, caspase-4 activation occurred as early as 10 min that was dependent on Eh Gal-lectin and EhCP-A5 binding to macrophages. By utilizing CRISPR-Cas9 gene edited CASP4/1, NLRP3 KO and ASC-def cells, caspase-4 activation was found to be independent of the canonical NLRP3 inflammasomes. In CRISPR-Cas9 gene edited CASP1 macrophages, caspase-4 activation was significantly up regulated that enhanced the enzymatic cleavage of GSDMD at the same cleavage site as caspase-1 to induce GSDMD pore formation and sustained bioactive IL-1β secretion. Eh-induced IL-1β secretion was independent of pyroptosis as revealed by pharmacological blockade of GSDMD pore formation and in CRISPR-Cas9 gene edited GSDMD KO macrophages. This was in marked contrast to the potent positive control, lipopolysaccharide + Nigericin that induced high expression of predominantly caspase-1 that efficiently cleaved GSDMD with high IL-1β secretion/release associated with massive cell pyroptosis. These results reveal that Eh triggered \"hyperactivated macrophages\" allowed caspase-4 dependent cleavage of GSDMD and IL-1β secretion to occur in the absence of pyroptosis that may play an important role in disease pathogenesis.", "Chronic inflammation is a known risk factor for tumorigenesis, yet the precise mechanism of this association is currently unknown. The inflammasome, a multiprotein complex formed by NOD-like receptor (NLR) family members, has recently been shown to orchestrate multiple innate and adaptive immune responses, yet its potential role in inflammation-induced cancer has been little studied. Using the azoxymethane and dextran sodium sulfate colitis-associated colorectal cancer model, we show that caspase-1-deficient (Casp1(-/-)) mice have enhanced tumor formation. Surprisingly, the role of caspase-1 in tumorigenesis was not through regulation of colonic inflammation, but rather through regulation of colonic epithelial cell proliferation and apoptosis. Consequently, caspase-1-deficient mice demonstrate increased colonic epithelial cell proliferation in early stages of injury-induced tumor formation and reduced apoptosis in advanced tumors. We suggest a model in which the NLRC4 inflammasome is central to colonic inflammation-induced tumor formation through regulation of epithelial cell response to injury.", "Skin cutaneous melanoma (SKCM) is a chronically malignant tumor with a high mortality rate. Pyroptosis, a kind of pro-inflammatory programmed cell death, has been linked to cancer in recent studies. However, the value of pyroptosis in the diagnosis and prognosis of SKCM is not clear. In this study, it was discovered that 20 pyroptosis-related genes (PRGs) differed in expression between SKCM and normal tissues, which were related to diagnosis and prognosis. Firstly, based on these genes, nine machine-learning algorithms were shown to perform well in constructing diagnostic classifiers, including K-Nearest Neighbor (KNN), logistic regression, Support Vector Machine (SVM), Artificial Neural Network (ANN), decision tree, random forest, XGBoost, LightGBM, and CatBoost. Secondly, the least absolute shrinkage and selection operator (LASSO) Cox regression analysis was applied and the prognostic model was constructed based on 9 PRGs. Subgroups in low and high risks determined by the prognostic model were shown to have different survival. Thirdly, functional enrichment analyses were performed by applying the gene set enrichment analysis (GSEA), and results suggested that the risk was related to immune response. In conclusion, the expression signatures of pyroptosis-related genes are effective and robust in the diagnosis and prognosis of SKCM, which is related to immunity.", "Pancreatic adenocarcinoma (PAAD) has become one of the deadliest malignancies in the world. Since necroptosis plays a crucial role in regulating the immune system, it is necessary to develop novel prognostic biomarkers associated with necroptosis and explore its potential role in PAAD. The transcriptome RNA-seq data of PAAD were downloaded from the TCGA and GTEx databases. A prognostic signature was constructed by the least absolute shrinkage and selection operator (LASSO) Cox regression, and its prognostic value was evaluated by nomogram and validated in an independent GEO cohort. We identified a total of 24 differentially expressed NRGs in PAAD, and constructed a prognostic signature with 5 NRGs, which showed good performance in predicting the prognosis of PAAD patients. The ROC curves for 1-, 3-, and 5-year survival rate were 0.652, 0.778, and 0.817, respectively. This prognostic signature showed consistent prognosis prediction in an independent patient cohort. Furthermore, the correlations between 5-NRGs signature and TMB, MSI, histopathological classification, immune infiltration, immune types, and immunomodulators were all significant. Notably, the expression profiles of the five NRGs in exosomes of serum were consistent with their expression in tumor tissues. These data suggested that the 5-NRGs signature is a promising biomarker for predicting the prognosis of PAAD.", "γδ T cells are considered to be innate lymphocytes that play an important role in host defense against tumors and infections. We recently reported that IL-18 markedly amplified γδ T cell responses to zoledronate (ZOL)/IL-2. In an extension of this finding, we analyzed the mechanism underlying the IL-18-mediated expansion of γδ T cells. After incubation of PBMCs with ZOL/IL-2/IL-18, the majority of the cells expressed γδ TCR, and the rest mostly exhibited CD56(bright)CD11c(+) under the conditions used in this study. CD56(bright)CD11c(+) cells were derived from a culture of CD56(int)CD11c(+) cells and CD14(+) cells in the presence of IL-2 and IL-18 without the addition of ZOL. They expressed IL-18Rs, HLA-DR, CD25, CD80, CD83, CD86, and CD11a/CD18. In addition, they produced IFN-γ, TNF-α, but not IL-12, when treated with IL-2/IL-18, and they exerted cytotoxicity against K562 cells, thus exhibiting characteristics of both NK cells and dendritic cells. Incubation of purified γδ T cells with CD56(bright)CD11c(+) cells in the presence of ZOL/IL-2/IL-18 resulted in the formation of massive cell clusters and led to the marked expansion of γδ T cells. However, both conventional CD56(-/int)CD11c(high) dendritic cells induced by GM-CSF/IL-4 and CD56(+)CD11c(-) NK cells failed to support the expansion of γδ T cells. These results strongly suggest that CD56(bright)CD11c(+) cells play a key role in the IL-18-mediated proliferation of γδ T cells.", "When pathogenic stresses are recognized by innate immune cells, inflammasomes are assembled and caspase-1 is activated, resulting in the conversion of pro-IL-18 into mature IL-18. Because natural killer (NK) cells express IL-18 receptors, IL-18 may play roles in immune functions of NK cells. In the present study, we examined the effect of IL-18 on NK cells derived from lung cancer patients and healthy adult volunteers. When peripheral blood NK cells were stimulated with IL-2, the cells formed clusters beginning on day 5-6 and proliferated thereafter, in which the number of NK cells increased by 10-fold in 10 days. When IL-18 was added, cell clusters were observed as early as on day 4 and NK cells proliferated vigorously. On day 10, the expansion rate was 56-fold on average, showing that IL-18 promoted the expansion of NK cells. It was also notable that IL-18 enhanced the expression of CD80, CD86, HLA-DR and HLA-DQ on NK cells, suggesting that IL-18 conferred NK cells an APC-like phenotype. When cellular cytotoxicity was determined, APC-like NK cells efficiently killed tumor cells and anti-tumor activity was augmented by the addition of tumor antigen-specific mAbs. In addition, IFN-γ was produced by APC-like NK cells in response to tumor cells, and the cytokine production was further enhanced by mAbs. Taken together, IL-18 not only promoted the expansion of NK cells, but also changed the phenotype of NK cells. IL-2/IL-18-induced NK cells might, therefore, serve as a bridge between innate immunity and adaptive immunity and be useful for cancer immunotherapy." ]
Prognostic Significance of Mitochondrial Function-Related Long Noncoding RNAs and Their Link to Immunological Microenvironment in Breast Cancer	Mitochondrial function, as the core of the cell's energy metabolism, is firmly connected to cancer metabolism and growth. However, the involvement of long noncoding RNAs (lncRNAs) related to mitochondrial function in breast cancer (BRCA) has not been thoroughly investigated. As a result, the objective of this research was to dissect the prognostic implication of mitochondrial function-related lncRNAs and their link to the immunological microenvironment in BRCA. The Cancer Genome Atlas (TCGA) database was used to acquire clinicopathological and transcriptome information for BRCA samples. Mitochondrial function-related lncRNAs were recognized by coexpression analysis of 944 mitochondrial function-related mRNAs obtained from the MitoMiner 4.0 database. A novel prognostic signature was built in the training cohort using integrated analysis of mitochondrial function-related lncRNA and the corresponding clinical information through univariate analysis, lasso regression, and stepwise multivariate Cox regression analysis. The prognostic worth was judged in the training cohort and validated in the test cohort. In addition, functional enrichment and immune microenvironment analyses were performed to explore the risk score on the basis of the prognostic signature. An 8-mitochondrial function-related lncRNA signature was generated by integrated analysis. Individuals within the higher-risk category had a worse overall survival rate (OS) (training cohort: P < 0.001; validation cohort: P < 0.001; whole cohort: P < 0.001). The risk score was identified as an independent risk factor by multivariate Cox regression analysis (training cohort: HR 1.441, 95% CI 1.229-1.689, P < 0.001; validation cohort: HR 1.343, 95% CI 1.166-1.548, P < 0.001; whole cohort: HR 1.241, 95% CI 1.156-1.333, P < 0.001). Following that, the predictive accuracy of the model was confirmed by the ROC curves. In addition, nomograms were generated, and the calibration curves revealed that the model had excellent prediction accuracy for 3- and 5-year OS. Besides, the higher-risk BRCA individuals have relatively decreased amounts of infiltration of tumor-killing immune cells, lower levels of immune checkpoint molecules, and immune function. We constructed and verified a novel mitochondrial function-related lncRNA signature that might accurately predict the outcome of BRCA, play an essential role in immunotherapy, and might be exploited as a therapeutic target for precise BRCA therapy.	[ "Accumulating preclinical and clinical evidence indicates that the success of several anticancer agents-including some conventional chemotherapeutics, targeted anticancer agents as well as specific forms of radiotherapy-depends (at least in part) on their ability to stimulate anticancer immune responses. Such immunostimulatory effects can be \"on-target,\" i.e., they originate within cancer cells, or \"off-target,\" i.e., they develop from a heretofore unsuspected interaction between cancer therapy and the immune system. Here, we briefly discuss the immunologic mechanisms that underlie the efficacy of some forms of cancer therapy, as we highlight the rationale for combining these treatment modalities with immunotherapy to achieve superior therapeutic effects. Cancer Immunol Res; 4(11); 895-902. ©2016 AACR.", "Upon stimulation, small numbers of naive CD8+ T cells proliferate and differentiate into a variety of memory and effector cell types. CD8+ T cells can persist for years and kill tumour cells and virally infected cells. The functional and phenotypic changes that occur during CD8+ T cell differentiation are well characterized, but the epigenetic states that underlie these changes are incompletely understood. Here, we review the epigenetic processes that direct CD8+ T cell differentiation and function. We focus on epigenetic modification of DNA and associated histones at genes and their regulatory elements. We also describe structural changes in chromatin organization that affect gene expression. Finally, we examine the translational potential of epigenetic interventions to improve CD8+ T cell function in individuals with chronic infections and cancer.", "Contrary to conventional wisdom, functional mitochondria are essential for the cancer cell. Although mutations in mitochondrial genes are common in cancer cells, they do not inactivate mitochondrial energy metabolism but rather alter the mitochondrial bioenergetic and biosynthetic state. These states communicate with the nucleus through mitochondrial 'retrograde signalling' to modulate signal transduction pathways, transcriptional circuits and chromatin structure to meet the perceived mitochondrial and nuclear requirements of the cancer cell. Cancer cells then reprogramme adjacent stromal cells to optimize the cancer cell environment. These alterations activate out-of-context programmes that are important in development, stress response, wound healing and nutritional status.", "Immuno-oncology is an emerging field that has revolutionized cancer treatment. Most immunomodulatory strategies focus on enhancing T cell responses, but there has been a recent surge of interest in harnessing the relatively underexplored natural killer (NK) cell compartment for therapeutic interventions. NK cells show cytotoxic activity against diverse tumour cell types, and some of the clinical approaches originally developed to increase T cell cytotoxicity may also activate NK cells. Moreover, increasing numbers of studies have identified novel methods for increasing NK cell antitumour immunity and expanding NK cell populations ex vivo, thereby paving the way for a new generation of anticancer immunotherapies. The role of other innate lymphoid cells (group 1 innate lymphoid cell (ILC1), ILC2 and ILC3 subsets) in tumours is also being actively explored. This Review provides an overview of the field and summarizes current immunotherapeutic approaches for solid tumours and haematological malignancies.", "Natural killer (NK) cells are innate cytotoxic lymphocytes involved in the surveillance and elimination of cancer. As we have learned more and more about the mechanisms NK cells employ to recognize and eliminate tumor cells, and how, in turn, cancer evades NK cell responses, we have gained a clear appreciation that NK cells can be harnessed in cancer immunotherapy. Here, we review the evidence for NK cells' critical role in combating transformed and malignant cells, and how cancer immunotherapies potentiate NK cell responses for therapeutic purposes. We highlight cutting-edge immunotherapeutic strategies in preclinical and clinical development such as adoptive NK cell transfer, chimeric antigen receptor-expressing NK cells (CAR-NKs), bispecific and trispecific killer cell engagers (BiKEs and TriKEs), checkpoint blockade, and oncolytic virotherapy. Further, we describe the challenges that NK cells face (e.g., postsurgical dysfunction) that must be overcome by these therapeutic modalities to achieve cancer clearance.", "Checkpoint blockade immunotherapy targeting the PD-1/PD-L1 inhibitory axis has produced remarkable results in the treatment of several types of cancer. Whereas cytotoxic T cells are known to provide important antitumor effects during checkpoint blockade, certain cancers with low MHC expression are responsive to therapy, suggesting that other immune cell types may also play a role. Here, we employed several mouse models of cancer to investigate the effect of PD-1/PD-L1 blockade on NK cells, a population of cytotoxic innate lymphocytes that also mediate antitumor immunity. We discovered that PD-1 and PD-L1 blockade elicited a strong NK cell response that was indispensable for the full therapeutic effect of immunotherapy. PD-1 was expressed on NK cells within transplantable, spontaneous, and genetically induced mouse tumor models, and PD-L1 expression in cancer cells resulted in reduced NK cell responses and generation of more aggressive tumors in vivo. PD-1 expression was more abundant on NK cells with an activated and more responsive phenotype and did not mark NK cells with an exhausted phenotype. These results demonstrate the importance of the PD-1/PD-L1 axis in inhibiting NK cell responses in vivo and reveal that NK cells, in addition to T cells, mediate the effect of PD-1/PD-L1 blockade immunotherapy.", "We have identified a novel human gene of the Ig superfamily, designated LAG-3. Expression of this gene is undetectable in resting PBL, while it is found (a 2-kb message) in activated T and NK cells. The LAG-3 gene includes eight exons; the corresponding cDNA encodes a 498-amino acid membrane protein with four extracellular IgSF domains. The first one belongs to the V-SET; it is particular since it includes an extra loop in the middle of the domain and an unusual intrachain disulphide bridge. The three other domains belong to the C2-SET. Strong internal homologies are found in the LAG-3 molecule between domains 1 and 3, as well as between domains 2 and 4. It is therefore likely that LAG-3 has evolved by duplication of a pre-existing gene encoding a two IgSF-domain structure. The compared analysis of LAG-3 and CD4, with respect to both their peptidic sequence as well as their exon/intron organization, indicated that the two molecules are closely related. This point is strengthened by the finding that both genes are located on the distal part of the short arm of chromosome 12." ]
Well-TEMP-seq: a high-throughput, cost-effective, accurate, and efficient method for profiling the dynamics of single-cell gene expression	Single-cell RNA sequencing (scRNA-seq) reveals the transcriptional heterogeneity of cells, but the static snapshots fail to reveal the time-resolved dynamics of transcription. Herein, we develop Well-TEMP-seq, a high-throughput, cost-effective, accurate, and efficient method for massively parallel profiling the temporal dynamics of single-cell gene expression. Well-TEMP-seq combines metabolic RNA labeling with scRNA-seq method Well-paired-seq to distinguish newly transcribed RNAs marked by T-to-C substitutions from pre-existing RNAs in each of thousands of single cells. The Well-paired-seq chip ensures a high single cell/barcoded bead pairing rate (~80%) and the improved alkylation chemistry on beads greatly alleviates chemical conversion-induced cell loss (~67.5% recovery). We further apply Well-TEMP-seq to profile the transcriptional dynamics of colorectal cancer cells exposed to 5-AZA-CdR, a DNA-demethylating drug. Well-TEMP-seq unbiasedly captures the RNA dynamics and outperforms the splicing-based RNA velocity method. We anticipate that Well-TEMP-seq will be broadly applicable to unveil the dynamics of single-cell gene expression in diverse biological processes.	[ "Hairy/enhancer-of-split related with YRPW motif-like (HEYL) protein was first identified as a transcriptional repressor. It is a downstream gene of the Notch and transforming growth factor-β pathways. Little is known about its role in the pathogenesis of hepatocellular carcinoma (HCC). Eighty surgically resected paired HCC and adjacent non-cancerous tissues were analyzed for HEYL expression by reverse transcription quantitative polymerase chain reaction (RT-qPCR) and immunohistochemistry (IHC). HCC cells were transfected with pHEYL-EGFP vector to overexpress the HEYL gene or infected with specific shHEYL lentiviral vector to silence HEYL gene expression. HEYL expressional analysis and functional characterization were assessed by 3-(4 5-dimethylthiazol-2-yl)-2 5-diphenyltetrazolium bromide assays, flow cytometry, RT-qPCR, western blotting and methylation-specific PCR. We determined that HEYL expression was inactivated in more than 75% of HCC. In addition, overexpression of HEYL in SK-Hep 1 cells caused apoptosis by the cleavage of caspase 3 and poly (ADP-ribose) polymerase. We discovered that HEYL apoptosis was preceded by serine 15 phosphorylation and accumulation of P53. Molecular analysis revealed that HEYL overexpression led to increased p16, p19, p21, p27 and Bad protein expression, and reduced c-Myc, Bcl-2 and Cyclin B1 expression. Epigenetic silencing of HEYL expression by DNA hypermethylation in HCC directly correlated with loss of HEYL expression in HCC. HEYL is frequently downregulated by promoter methylation in HCC. HEYL may be a tumor suppressor of liver carcinogenesis through upregulation of P53 gene expression and activation of P53-mediated apoptosis.", "It is now established that the transcription factors E2A, EBF1 and Foxo1 have critical roles in B cell development. Here we show that E2A and EBF1 bound regulatory elements present in the Foxo1 locus. E2A and EBF1, as well as E2A and Foxo1, in turn, were wired together by a vast spectrum of cis-regulatory sequences. These associations were dynamic during developmental progression. Occupancy by the E2A isoform E47 directly resulted in greater abundance, as well as a pattern of monomethylation of histone H3 at lysine 4 (H3K4) across putative enhancer regions. Finally, we divided the pro-B cell epigenome into clusters of loci with occupancy by E2A, EBF and Foxo1. From this analysis we constructed a global network consisting of transcriptional regulators, signaling and survival factors that we propose orchestrates B cell fate.", "Cellular RNA levels are determined by the interplay of RNA production, processing and degradation. However, because most studies of RNA regulation do not distinguish the separate contributions of these processes, little is known about how they are temporally integrated. Here we combine metabolic labeling of RNA at high temporal resolution with advanced RNA quantification and computational modeling to estimate RNA transcription and degradation rates during the response of mouse dendritic cells to lipopolysaccharide. We find that changes in transcription rates determine the majority of temporal changes in RNA levels, but that changes in degradation rates are important for shaping sharp 'peaked' responses. We used sequencing of the newly transcribed RNA population to estimate temporally constant RNA processing and degradation rates genome wide. Degradation rates vary significantly between genes and contribute to the observed differences in the dynamic response. Certain transcripts, including those encoding cytokines and transcription factors, mature faster. Our study provides a quantitative approach to study the integrative process of RNA regulation.", "The T-box transcription factors T-bet and Eomesodermin (Eomes) have been well defined as key drivers of immune cell development and cytolytic function. While the majority of studies have defined the roles of these factors in the context of murine T-cells, recent results have revealed that T-bet, and possibly Eomes, are expressed in other immune cell subsets. To date, the expression patterns of these factors in subsets of human peripheral blood mononuclear cells beyond T-cells remain relatively uncharacterized. In this study, we used multiparametric flow cytometry to characterize T-bet and Eomes expression in major human blood cell subsets, including total CD4(+) and CD8(+) T-cells, γδ T-cells, invariant NKT cells, natural killer cells, B-cells, and dendritic cells. Our studies identified novel cell subsets that express T-bet and Eomes and raise implications for their possible functions in the context of other human immune cell subsets besides their well-known roles in T-cells.", "This protocol explains how to perform a fast SCENIC analysis alongside standard best practices steps on single-cell RNA-sequencing data using software containers and Nextflow pipelines. SCENIC reconstructs regulons (i.e., transcription factors and their target genes) assesses the activity of these discovered regulons in individual cells and uses these cellular activity patterns to find meaningful clusters of cells. Here we present an improved version of SCENIC with several advances. SCENIC has been refactored and reimplemented in Python (pySCENIC), resulting in a tenfold increase in speed, and has been packaged into containers for ease of use. It is now also possible to use epigenomic track databases, as well as motifs, to refine regulons. In this protocol, we explain the different steps of SCENIC: the workflow starts from the count matrix depicting the gene abundances for all cells and consists of three stages. First, coexpression modules are inferred using a regression per-target approach (GRNBoost2). Next, the indirect targets are pruned from these modules using cis-regulatory motif discovery (cisTarget). Lastly, the activity of these regulons is quantified via an enrichment score for the regulon's target genes (AUCell). Nonlinear projection methods can be used to display visual groupings of cells based on the cellular activity patterns of these regulons. The results can be exported as a loom file and visualized in the SCope web application. This protocol is illustrated on two use cases: a peripheral blood mononuclear cell data set and a panel of single-cell RNA-sequencing cancer experiments. For a data set of 10,000 genes and 50,000 cells, the pipeline runs in <2 h.", "Single-cell RNA sequencing (scRNA-seq) has highlighted the important role of intercellular heterogeneity in phenotype variability in both health and disease1. However, current scRNA-seq approaches provide only a snapshot of gene expression and convey little information on the true temporal dynamics and stochastic nature of transcription. A further key limitation of scRNA-seq analysis is that the RNA profile of each individual cell can be analysed only once. Here we introduce single-cell, thiol-(SH)-linked alkylation of RNA for metabolic labelling sequencing (scSLAM-seq), which integrates metabolic RNA labelling2, biochemical nucleoside conversion3 and scRNA-seq to record transcriptional activity directly by differentiating between new and old RNA for thousands of genes per single cell. We use scSLAM-seq to study the onset of infection with lytic cytomegalovirus in single mouse fibroblasts. The cell-cycle state and dose of infection deduced from old RNA enable dose-response analysis based on new RNA. scSLAM-seq thereby both visualizes and explains differences in transcriptional activity at the single-cell level. Furthermore, it depicts 'on-off' switches and transcriptional burst kinetics in host gene expression with extensive gene-specific differences that correlate with promoter-intrinsic features (TBP-TATA-box interactions and DNA methylation). Thus, gene-specific, and not cell-specific, features explain the heterogeneity in transcriptomes between individual cells and the transcriptional response to perturbations.", "Large parts of mammalian genomes are transcriptionally inactive and enriched with various classes of interspersed and tandem repeats. Here we show that the tumor suppressor protein p53 cooperates with DNA methylation to maintain silencing of a large portion of the mouse genome. Massive transcription of major classes of short, interspersed nuclear elements (SINEs) B1 and B2, both strands of near-centromeric satellite DNAs consisting of tandem repeats, and multiple species of noncoding RNAs was observed in p53-deficient but not in p53 wild-type mouse fibroblasts treated with the DNA demethylating agent 5-aza-2'-deoxycytidine. The abundance of these transcripts exceeded the level of β-actin mRNA by more than 150-fold. Accumulation of these transcripts, which are capable of forming double-stranded RNA (dsRNA), was accompanied by a strong, endogenous, apoptosis-inducing type I IFN response. This phenomenon, which we named \"TRAIN\" (for \"transcription of repeats activates interferon\"), was observed in spontaneous tumors in two models of cancer-prone mice, presumably reflecting naturally occurring DNA hypomethylation and p53 inactivation in cancer. These observations suggest that p53 and IFN cooperate to prevent accumulation of cells with activated repeats and provide a plausible explanation for the deregulation of IFN function frequently seen in tumors. Overall, this work reveals roles for p53 and IFN that are key for genetic stability and therefore relevant to both tumorigenesis and the evolution of species." ]
Osteonecrosis of the jaw: a rare but serious adverse drug reaction.	Osteonecrosis of the jaw (ONJ) is a rare but serious adverse drug reaction (ADR) commonly associated with bisphosphonate and denosumab therapy. Prior research utilized an online, public FDA Adverse Event Reporting System (FAERS) Database to explore this ADR. This data identified and described several novel medications associated with ONJ. Our study aims to build upon the prior findings, reporting trends of medication induced ONJ over time and identifying newly described medications.	[ "Social media is becoming increasingly popular as a platform for sharing personal health-related information. This information can be utilized for public health monitoring tasks, particularly for pharmacovigilance, via the use of natural language processing (NLP) techniques. However, the language in social media is highly informal, and user-expressed medical concepts are often nontechnical, descriptive, and challenging to extract. There has been limited progress in addressing these challenges, and thus far, advanced machine learning-based NLP techniques have been underutilized. Our objective is to design a machine learning-based approach to extract mentions of adverse drug reactions (ADRs) from highly informal text in social media. We introduce ADRMine, a machine learning-based concept extraction system that uses conditional random fields (CRFs). ADRMine utilizes a variety of features, including a novel feature for modeling words' semantic similarities. The similarities are modeled by clustering words based on unsupervised, pretrained word representation vectors (embeddings) generated from unlabeled user posts in social media using a deep learning technique. ADRMine outperforms several strong baseline systems in the ADR extraction task by achieving an F-measure of 0.82. Feature analysis demonstrates that the proposed word cluster features significantly improve extraction performance. It is possible to extract complex medical concepts, with relatively high performance, from informal, user-generated content. Our approach is particularly scalable, suitable for social media mining, as it relies on large volumes of unlabeled data, thus diminishing the need for large, annotated training data sets.", "Adverse drug events cause substantial morbidity and mortality and are often discovered after a drug comes to market. We hypothesized that Internet users may provide early clues about adverse drug events via their online information-seeking. We conducted a large-scale study of Web search log data gathered during 2010. We pay particular attention to the specific drug pairing of paroxetine and pravastatin, whose interaction was reported to cause hyperglycemia after the time period of the online logs used in the analysis. We also examine sets of drug pairs known to be associated with hyperglycemia and those not associated with hyperglycemia. We find that anonymized signals on drug interactions can be mined from search logs. Compared to analyses of other sources such as electronic health records (EHR), logs are inexpensive to collect and mine. The results demonstrate that logs of the search activities of populations of computer users can contribute to drug safety surveillance.", "Identification of adverse drug reactions (ADRs) during the post-marketing phase is one of the most important goals of drug safety surveillance. Spontaneous reporting systems (SRS) data, which are the mainstay of traditional drug safety surveillance, are used for hypothesis generation and to validate the newer approaches. The publicly available US Food and Drug Administration (FDA) Adverse Event Reporting System (FAERS) data requires substantial curation before they can be used appropriately, and applying different strategies for data cleaning and normalization can have material impact on analysis results. We provide a curated and standardized version of FAERS removing duplicate case records, applying standardized vocabularies with drug names mapped to RxNorm concepts and outcomes mapped to SNOMED-CT concepts, and pre-computed summary statistics about drug-outcome relationships for general consumption. This publicly available resource, along with the source code, will accelerate drug safety research by reducing the amount of time spent performing data management on the source FAERS reports, improving the quality of the underlying data, and enabling standardized analyses using common vocabularies.", "Adverse drug reactions (ADRs) represent an important risk for patients and have a significant economic impact on health systems. ADRs are the fifth most common cause of hospital death, with a burden estimated at 197,000 deaths per year in the EU. This has a societal cost of <euro>79 billion per year. Because of this strong impact in public health, regulatory authorities (RAs) worldwide are implementing new pharmacovigilance legislation to promote and protect public health by reducing the burden of ADRs through the detection of safety signals. Although, traditionally, signal detection activities have mainly been performed based on spontaneous reporting from healthcare professionals and national health RAs, the new pharmacovigilance legislation underlines the relevance of other sources of information (such as scientific literature) for the evaluation of the benefit-risk balance of a certain product. This review aims to highlight the relevance of periodic scientific literature screening in the safety signal detection process. The authors present four practical examples where a safety signal that was detected from a literature report had an impact on the lifecycle of a drug. In addition, based on practical experience of the screening of medical and scientific literature for safety purposes, this article analyses the requirements of the new pharmacovigilance guidelines on literature screening and highlights the need for the implementation of a literature review procedure and the main challenges encountered when performing literature screening for safety aspects." ]
The miR-200b/429 is a critical regulator of cervical cancer.	The miR-200b/429 located at 1p36 is a highly conserved miRNA cluster emerging as a critical regulator of cervical cancer. Using publicly available miRNA expression data from TCGA and GEO followed by independent validation, we aimed to identify the association between miR-200b/429 expression and cervical cancer. miR-200b/429 cluster was significantly overexpressed in cancer samples compared to normal samples. miR-200b/429 expression did not correlate with patient survival; however, its overexpression correlated with histological type. Protein-protein interaction analysis of 90 target genes of miR-200b/429 identified EZH2, FLT1, IGF2, IRS1, JUN, KDR, SOX2, MYB, ZEB1, and TIMP2 as the top ten hub genes. PI3K-AKT and MAPK signaling pathways emerged as major target pathways of miR-200b/429 and their hub genes. Kaplan-Meier survival analysis showed the expression of seven miR-200b/429 target genes (EZH2, FLT1, IGF2, IRS1, JUN, SOX2, and TIMP2) to influence the overall survival of patients. The miR-200a-3p and miR-200b-5p could help predict cervical cancer with metastatic potential. The cancer hallmark enrichment analysis showed hub genes to promote growth, sustained proliferation, resistance to apoptosis, induction of angiogenesis, activation of invasion, and metastasis, enabling replicative immortality, evading immune destruction, and tumor-promoting inflammation. The drug-gene interaction analysis identified 182 potential drugs to interact with 27 target genes of miR-200b/429 with paclitaxel, doxorubicin, dabrafenib, bortezomib, docetaxel, ABT-199, eribulin, vorinostat, etoposide, and mitoxantrone emerging as the top ten best candidate drugs. Taken together, miR-200b/429 and associated hub genes can be helpful for prognostic application and clinical management of cervical cancer.	[ "The miR-15a/16-1 cluster is abnormally expressed in cervical cancer (CC) tissues and plays a vital role in cervical carcinogenesis. We aimed to evaluate the miR-15a/16-1 expression in healthy and cancerous cervical tissues, identify the associated networks, and to test its prognostic significance. miR-15a/16-1-MC expressions were analyzed in TCGA-CESC datasets by UALCAN, GEPIA2, and Datasetviewer. miR-15a/16-1 validated targets were extracted from mirTarBase and in silico functional analysis of the target genes were performed using WebGestalt. The interaction networks were constructed by the miRNet, STRING, and NetworkAnalyst tools. The prognostic significance and metastatic potential of the target genes were predicted using UALCAN and HCMDB. The FDA approved drugs to target miR-15a/16-1 and target gene network in CC were performed using DGIdb, STITCH and PanDrugs. TCGA-CESC and GEO data analysis suggested significant overexpression of miR-15a/16-1 in CC samples. The Kaplan-Meier survival analysis showed that miR-15a and its four target genes (BCL2, CCNE1, NUP50, and RBPJ) influence the overall survival of CC patients. Among the 66 differentially expressed target genes, 12 of them are linked to head, neck, or lung metastasis. Functional enrichment analysis predicted the association of this cluster with p53 signaling, human papillomavirus infection, PI3-AKT signaling pathway, and pathways in cancer. Drug-gene interaction analysis showed 52 potential FDA approved drugs to interact with the miR-15a/16-1 target genes. Nine of the 52 drugs are currently used as a chemotherapeutic agent for the treatment of CC patients. The present study shows that miR-15a/16-1 expression can be used as a clinical marker and target for therapy in CC.", "Many patients with locally advanced cervical cancer experience recurrence within the radiation field after chemoradiotherapy. Biomarkers of tumor radioresistance are required to identify patients in need of intensified treatment. Here, the biomarker potential of miR-200 family members was investigated in this disease. Also, involvement of tumor hypoxia in the radioresistance mechanism was determined, using a previously defined 6-gene hypoxia classifier. miR-200 expression was measured in pretreatment tumor biopsies of an explorative cohort (n = 90) and validation cohort 1 (n = 110) by RNA sequencing. Publicly available miR-200 data of 79 patients were included for the validation of prognostic significance. A score based on expression of the miR-200a/b/-429 (miR-200a, miR-200b, and miR-429) cluster showed prognostic significance in all cohorts. The score was significant in multivariate analysis of central pelvic recurrence. No association with distant recurrence or hypoxia status was found. Potential miRNA target genes were identified from gene expression profiles and showed enrichment of genes in extracellular matrix organization and cell adhesion. miR-200a/b/-429 overexpression had a pronounced radiosensitizing effect in tumor xenografts, whereas the effect was minor in vitro. In conclusion, miR-200a/b/-429 downregulation is a candidate biomarker of central pelvic recurrence and seems to predict cell adhesion-mediated tumor radioresistance independent of clinical markers and hypoxia.", "Diffusion-weighted magnetic resonance imaging (DWI) is increasingly being used to assess tumour response to a variety of anticancer treatments. The technique is quick to perform without the need for administration of exogenous contrast medium, and enables the apparent diffusion coefficient (ADC) of tissues to be quantified. Studies have shown that ADC increases in response to a variety of treatments including chemotherapy, radiotherapy, minimally invasive therapies and novel therapeutics. In this article, we review the rationale of applying DWI for tumour assessment, the evidence for ADC measurements in relation to specific treatments and some of the practical considerations for using ADC to evaluate treatment response.", "Two mature miRNA species may be generated from the 5' and 3' arms of a pre-miRNA precursor. In most cases, only one species remains while the complementary species is degraded. However, co-existence of miRNA-5p and -3p species is increasingly being reported. In this work, we aimed to systematically investigate co-expression of miRNA-5p/3p in colon cancer cells in a genome-wide analysis, and to examine cross-targeting of the dysregulated miRNAs and 5p/3p species. Four colon cancer cell lines were examined relative to two normal colon tissues. Of the 1,190 miRNAs analyzed, 92 and 36 were found to be up- or down-regulated, respectively, in cancer cells. Nineteen co-expressed miRNA-5p/3p pairs were further identified suggesting frequent 5p/3p co-accumulation in colon cancer cells. Of these, 14 pairs were co-up-regulated and 3 pairs were co-down-regulated indicating concerted 5p/3p dysregulation. Nine dysregulated miRNA pairs fell into three miRNA gene families, namely let-7, mir-8/200 and mir-17, which showed frequent cross-targeting in the metastasis process. Focusing on the let-7d-5p/3p pair, the respectively targeted IGF1R and KRAS were shown to be in a reverse relationship with expression of the respective miRNA, which was confirmed in transient transfection assays using let-7d mimic or inhibitor. Targeting of KRAS by let-7d was previous reported; targeting of IGF1R by let-7d-5p was confirmed in luciferase assays in this study. The findings of let-7d-5p/3p and multiple other miRNAs targeting IGF1R, KRAS and other metastasis-related factors suggest that 5p/3p miRNAs contribute to cross-targeting of multiple cancer-associated factors and processes possibly to evade functional abolishment when any one of the crucial factors are inactivated. miRNA-5p/3p species are frequently co-expressed and are coordinately regulated in colon cancer cells. In cancer cells, multiple cross-targeting by the miRNAs, including the co-existing 5p/3p species, frequently occurs in an apparent safe-proof scheme of miRNA regulation of important tumorigenesis processes. Further systematic analysis of co-existing miRNA-5p/3p pairs in clinical tissues is important in elucidating 5p/3p contributions to cancer pathogenesis.", "The many functional partnerships and interactions that occur between proteins are at the core of cellular processing and their systematic characterization helps to provide context in molecular systems biology. However, known and predicted interactions are scattered over multiple resources, and the available data exhibit notable differences in terms of quality and completeness. The STRING database (http://string-db.org) aims to provide a critical assessment and integration of protein-protein interactions, including direct (physical) as well as indirect (functional) associations. The new version 10.0 of STRING covers more than 2000 organisms, which has necessitated novel, scalable algorithms for transferring interaction information between organisms. For this purpose, we have introduced hierarchical and self-consistent orthology annotations for all interacting proteins, grouping the proteins into families at various levels of phylogenetic resolution. Further improvements in version 10.0 include a completely redesigned prediction pipeline for inferring protein-protein associations from co-expression data, an API interface for the R computing environment and improved statistical analysis for enrichment tests in user-provided networks.", "Chemoradiation, either alone or in combination with surgery or induction chemotherapy, is the current standard of care for most locally advanced solid tumors. Though chemoradiation is usually performed at the maximum tolerated doses of both chemotherapy and radiation, current cure rates are not satisfactory for many tumor entities, since tumor heterogeneity and plasticity result in chemo- and radioresistance. Advances in the understanding of tumor biology, a rapidly growing number of molecular targeting agents and novel technologies enabling the in-depth characterization of individual tumors, have fuelled the hope of entering an era of precision oncology, where each tumor will be treated according to its individual characteristics and weaknesses. At present though, molecular targeting approaches in combination with radiotherapy or chemoradiation have not yet proven to be beneficial over standard chemoradiation treatment in the clinical setting. A promising approach to improve efficacy is the combined usage of two targeting agents in order to inhibit backup pathways or achieve a more complete pathway inhibition. Here we review preclinical attempts to utilize such dual targeting strategies for future tumor radiosensitization.", "Cytoscape is an open source software project for integrating biomolecular interaction networks with high-throughput expression data and other molecular states into a unified conceptual framework. Although applicable to any system of molecular components and interactions, Cytoscape is most powerful when used in conjunction with large databases of protein-protein, protein-DNA, and genetic interactions that are increasingly available for humans and model organisms. Cytoscape's software Core provides basic functionality to layout and query the network; to visually integrate the network with expression profiles, phenotypes, and other molecular states; and to link the network to databases of functional annotations. The Core is extensible through a straightforward plug-in architecture, allowing rapid development of additional computational analyses and features. Several case studies of Cytoscape plug-ins are surveyed, including a search for interaction pathways correlating with changes in gene expression, a study of protein complexes involved in cellular recovery to DNA damage, inference of a combined physical/functional interaction network for Halobacterium, and an interface to detailed stochastic/kinetic gene regulatory models." ]
Leadership behaviour of nurse managers and nurses' levels of job satisfaction and compassion fatigue during the COVID-19 pandemic	To reveal the relationships between the leadership behaviour of nurse managers and nurses' levels of job satisfaction and compassion fatigue during the COVID-19 pandemic.	[ "Pre-COVID-19 research highlighted the nursing profession worldwide as being at high risk from symptoms of burnout, post-traumatic stress disorder (PTSD) and suicide. The World Health Organization declared a pandemic on 11th March 2020 due to the sustained risk of further global spread of COVID-19. The high healthcare burden associated with COVID-19 has increased nurses' trauma and workload, thereby exacerbating pressure on an already strained workforce and causing additional psychological distress for staff. The Impact of COVID-19 on Nurses (ICON) interview study examined the impacts of the pandemic on frontline nursing staff's psychosocial and emotional wellbeing. Longitudinal qualitative interview study. Nurses who had completed time 1 and 2 of the ICON survey were sampled to include a range of UK work settings including acute, primary and community care and care homes. Interviewees were purposively sampled for maximum variation to cover a broad range of personal and professional factors, and experiences during the COVID-19 pandemic, including redeployment. Nurses participated in qualitative in-depth narrative interviews after the first wave of COVID-19 in July 2020 (n = 27) and again at the beginning of the second wave in December 2020 (n = 25) via video and audio platform software. Rigorous qualitative narrative analysis was undertaken both cross-sectionally (within wave) and longitudinally (cross wave) to explore issues of consistency and change. The terms moral distress, compassion fatigue, burnout and PTSD describe the emotional states reported by the majority of interviewees leading many to consider leaving the profession. Causes of this identified included care delivery challenges; insufficient staff and training; PPE challenges and frustrations. Four themes were identified: (1) 'Deathscapes' and impoverished care (2) Systemic challenges and self-preservation (3) Emotional exhaustion and (4) (Un)helpful support. Nurses have been deeply affected by what they have experienced and report being forever altered with the impacts of COVID-19 persisting and deeply felt. There is an urgent need to tackle stigma to create a psychologically safe working environment and for a national COVID-19 nursing workforce recovery strategy to help restore nurse's well-being and demonstrate a valuing of the nursing workforce and therefore support retention.", "To examine the experience of Italian nurses posted on social media and discover changes, if any, over the waves. A mixed methods study reported according to the Good Reporting of a Mixed Methods Study criteria. All narratives (texts, letters and interviews) posted by Italian nurses from February 2020 to May 2020 (first wave) and from October 2020 to May 2021 (second/third wave) on the five most famous Italian professional social media platforms. The data were analysed qualitatively (first wave) and then quantitatively (second/third wave). A total of 380 narratives (202,626 words, 2510 quotes) were posted in the first wave, and 161 (68,388 words, 835 quotes) in the second/third wave. In the first wave, the following five themes emerged: (a) 'sharing what is happening within myself' (891; 35.5%); (b) 'experiencing unprecedented working conditions' (749; 29.8%); (c) 'failing to rehabilitate the image of nurses in society' (376; 15%); (d) 'experiencing a deep change' (253; 10.1%) and (e) 'do not abandon us' (241; 9.6%). The same themes and subthemes also emerged in the second/third wave with some significant differences, indicating changes in the lived experience of nurses. Moreover, in the second/third wave, a new theme emerged: 'experiencing the mixed emotions towards jabs'. By analysing their posts, Italian nurses continue to face challenges during the COVID-19 pandemic, with changes in their lived experiences across the waves. Governments, nursing associations and health care organizations should consider these changes to design policies to prevent the further loss of nurses.", "To evaluate the correlation between burnout, self-esteem and quality of life among nurses. Analytical, cross-sectional study. Sakarya Training and Research Hospital, Turkey, in 2013. The sample was made-up of 131 volunteering nurses after obtainnig informed written consent from the participants, ethical committee, and corresponding institutions. Data were collected by personal information form (21 questions), Maslach Burnout Inventory (MBI-22 items), Rosenberg Self-Esteem Scale (RSES-10 items), Professional Quality Of Life Scale(ProQOL-30 items). The scales were analysed in terms of internal consistency. Cronbach Alpha coefficients were determined as reliable for our sample. MBI 3 subscale total scores of the participant nurses were low for emotional exhaustion and depersonalization, high for personal accomplishment. Total score from RSES was 15.32 ±3.70. Total scores from 3 subscales of ProQOL were 29.78 ±9.02 for compassion satisfaction, 24.65 ±5.75 for burnout, and 15.12 ±6.54 for compassion fatigue. In this study, it was detected that burnout in nurses affected compassion satisfaction and individual success negatively.", "Qualitative content analysis and thematic analysis are two commonly used approaches in data analysis of nursing research, but boundaries between the two have not been clearly specified. In other words, they are being used interchangeably and it seems difficult for the researcher to choose between them. In this respect, this paper describes and discusses the boundaries between qualitative content analysis and thematic analysis and presents implications to improve the consistency between the purpose of related studies and the method of data analyses. This is a discussion paper, comprising an analytical overview and discussion of the definitions, aims, philosophical background, data gathering, and analysis of content analysis and thematic analysis, and addressing their methodological subtleties. It is concluded that in spite of many similarities between the approaches, including cutting across data and searching for patterns and themes, their main difference lies in the opportunity for quantification of data. It means that measuring the frequency of different categories and themes is possible in content analysis with caution as a proxy for significance.", "Mixed methods research is the use of quantitative and qualitative methods in a single study or series of studies. It is an emergent methodology which is increasingly used by health researchers, especially within health services research. There is a growing literature on the theory, design and critical appraisal of mixed methods research. However, there are few papers that summarize this methodological approach for health practitioners who wish to conduct or critically engage with mixed methods studies. The objective of this paper is to provide an accessible introduction to mixed methods for clinicians and researchers unfamiliar with this approach. We present a synthesis of key methodological literature on mixed methods research, with examples from our own work and that of others, to illustrate the practical applications of this approach within health research. We summarize definitions of mixed methods research, the value of this approach, key aspects of study design and analysis, and discuss the potential challenges of combining quantitative and qualitative methods and data. One of the key challenges within mixed methods research is the successful integration of quantitative and qualitative data during analysis and interpretation. However, the integration of different types of data can generate insights into a research question, resulting in enriched understanding of complex health research problems.", "Nurses' burnout and psychological well-being are a significant concern during the pandemic. The aim of this study is to (i) examine the level of burnout, anxiety, depression, perceived stress and self-rated health for nurses at two time-points, 2020 and 2021, and (ii) examine the socio-demographic characteristics, work-related factors and perceived workplace support factors in relation to the level of burnout. This is a cross-sectional study with a longitudinal approach. A convenience sample of registered nurses who worked in an acute care tertiary hospital in Singapore were surveyed during two time-points. Participants' health, socio-demographic characteristics, work-related factors and perceived workplace support factors were collected. Among the 179 nurses, there was a significant increase in burnout level, poorer self-rated health and reduced job dedication. A decrease in the percentage of nurses who felt appreciated at work was reported in 2021 (p = 0.04). Nurses who felt their team was not working well together were 3.30 times more likely to experience burnout (95% CI 1.12 to 9.69; p = 0.03). Nurses who reported that they never felt appreciated by their department/hospital were 8.84 times more likely to experience burnout (95% CI 2.67 to 29.21; p < 0.001). Nurses with poorer self-rated health were more likely to report burnout (95% CI: 1.32-6.03; p = 0.008). Nurses had an increased experience of burnout, reduced job dedication and poorer self-rated health after the outbreak. Interventions at the departmental and organizational levels are needed to improve the workplace support. Strategies to support nurses' psychological well-being during the aftermath of COVID-19 are vital to managing nurses' burnout and improving job dedication and self-rated health.", "Quoted words and phrases from research participants are a common feature of qualitative research reports. Quoting is a process that requires the achievement of the proper balance between the obligations of scientific reporting and the taking of artistic license. Quotes are used to support researcher claims, illustrate ideas, illuminate experience, evoke emotion, and provoke response. Quoting involves researchers in acts of choosing that lie in the domains of aesthetics and ethics." ]
Phytochemical and pharmacological properties of Thai herbs and spices	Seeking cures for chronic inflammation-associated diseases and infectious diseases caused by critical human pathogens is challenging and time-consuming. Even as the research community searches for novel bioactive agents, consuming a healthy diet with functional ability might be an effective way to delay and prevent the progression of severe health conditions. Many plant ingredients in Thai food are considered medicinal, and these vegetables, herbs, and spices collectively possess multiple biological and pharmacological activities, such as anti-inflammatory, antimicrobial, antidiabetic, antipyretic, anticancer, hepatoprotective, and cardioprotective effects.	[ "Two phenylbutenoids, (E)-4-(3',4'-dimethoxyphenyl)but-3-enyl acetate and (E)-4-(3',4'-dimethoxyphenyl)but-1,3-diene, were separated from the rhizomes of Zingiber Cassumunar using a preparative upright counter-current chromatography (CCC). With a two-phase solvent system composed of light petroleum (b.p. 60-90 degrees C)-ethanol-diethyl ether-water (5:4:2:1, v/v), 150 mg of (E)-4-(3',4'-dimethoxyphenyl)but-3-enyl acetate and 175 mg of (E)-4-(3','-dimethoxyphenyl)but- 1,3-diene with the purity of 98.7 and 95.1%, respectively, were obtained from 600 mg of the crude sample of Z. Cassumunar in a single-step separation. Structures of these two compounds were identified by ESI-MS, 1H NMR and 13C NMR.", "Zingiber cassumunar Roxb. (Zingiberaceae), is an important medicinal plant known as \"Plai (Phlai)\" in Thailand, \"Bangle\" in Indonesia, and \"Bulei\" in China. Traditionally, this plant has been used to treat inflammation, pain, and respiratory problems. The rhizomes are the primary part of the plant that has been used for medicinal purposes due to their constituents with therapeutic properties, including phenylbutenoids, curcuminoids, and essential oils. Since the 1970s, many studies have been conducted on the phytochemicals and bioactivities of Z. cassumunar to establish fundamental scientific evidence that supports its use in traditional medicine. The accumulated biological studies on the extracts, solvent fractions, and constituents of Z. cassumunar have described their diverse medicinal properties, including antioxidant, anti-inflammatory, anticancer, neuroprotective/neurotrophic, cosmeceutical, and antifungal/antimicrobial bioactivities. In this review, we summarize information on the phytochemicals of Z. cassumunar and the bioactivities of its extracts and constituents.", "The Curcuma genus has been extensively used for therapeutic purposes in traditional or folk medicine worldwide, including for its anti-inflammatory activity. Curcuma spp.'s active constituents, such as alkaloids, flavonoids, and terpenoids, can act on various targets in the signaling pathway, restrain pro-inflammatory enzymes, lower the production of inflammatory cytokines and chemokines, and reduce oxidative stress, which subsequently suppresses inflammatory processes. Preclinical and clinical studies have reported the predominant anti-inflammatory activity of several Curcuma species. This review provides an overview of the anti-inflammatory effects of different extracts, preparations, and bioactive components in this genus. This analysis may provide a scientific basis for developing new and alternative methods for the isolation of a single entity from this genus to attenuate inflammatory conditions. The Curcuma genus is waiting for researchers interested in developing safe and efficient anti-inflammatory agents for further investigation.", "The objective of this study was to investigate the bioactivity of twenty-nine known isolated compounds from Cratoxylum species including three anthraquinones, four triterpenes, and twenty-two xanthones. All isolated compounds were subjected to antibacterial, anti-inflammatory and anti-oxidant activities. Cytotoxicity evaluations were performed by MTT assay. The anti-oxidatant activity was performed using DPPH assay. The anti-inflammatory activity was evaluated from the production of cytokines TNF-α and IL1-β using ELISA assay. Human gingival fibroblasts and monocytes could tolerate both anthraquinones and triterpenes. All isolated anthraquinones showed moderate-to-high antibacterial efficacy while compound A3 also demonstrated moderate anti-inflammatory effect. None of the isolated triterpenes, except for T1, inhibited the expression of TNF-α. A number of isolated xanthones was toxic to HGFs and monocytes. Compound X5, X14 and a 1:1 mixture of X5 and X6 showed comparative anti-inflammatory activity to dexamethasone. Several triterpene and xanthone compounds also expressed antibacterial effect against P. gingivalis. Some isolated xanthones exerted anti-oxidant activity comparable to ascorbic acid. Accordingly, selected pure compounds from plants of Cratoxylum genus might be of benefit in developing medications that are important in treating periodontal diseases.", "Acacia pennata subsp insuavis, or Cha-om in Thai, is a common vegetable found in Thailand. It has been used as a medicinal herb for a long time. From the literature, antinociceptive, anti-inflammatory, antimicrobial, and anti-helminthic activities were reported. In this study, we investigated two new actions of this plant: larvicide and pupicide. The crude ethanolic and fractionated extracts of A. pennata shoot tips were tested against aquatic stages of the dengue virus vector, Aedes aegypti mosquito. The 1st-4th instar larvae and pupae of Ae. aegypti were subjected for bioassays by following the standard protocol of WHO. The larval and pupal mortalities were observed after 24- and 48-hour exposure times. The bioassays demonstrated that stronger efficacy was found from the fractionated extracts than the crude extracts. The LC50 values against the 3rd instar larvae were 39.45-50.75 mg/l (fractionated extracts) and 244.50 mg/l (crude extracts). It also effects the pupae with the LC50 values of 44.10-53.73 mg/l and 87.27 mg/l for the fractionnated and the crude extracts, respectively. The bioassays demonstrated the effective mosquito larvicide and pupicide of A. pennata extracts. It could be an alternative candidate for the development of phytotoxin for controlling mosquito vectors.", "Coronavirus disease-19 (COVID-19), a devastating respiratory illness caused by SARS-associated coronavirus-2 (SARS-CoV-2), has already affected over 64 million people and caused 1.48 million deaths, just 12 months from the first diagnosis. COVID-19 patients develop serious complications, including severe pneumonia, acute respiratory distress syndrome (ARDS), and or multiorgan failure due to exaggerated host immune response following infection. Currently, drugs that were effective against SARS-CoV are being repurposed for SARS-CoV-2. During this public health emergency, food nutraceuticals could be promising prophylactic therapeutics for COVID-19. Curcumin, a bioactive compound in turmeric, exerts diverse pharmacological activities and is widely used in foods and traditional medicines. This review presents several lines of evidence, which suggest curcumin as a promising prophylactic, therapeutic candidate for COVID-19. First, curcumin exerts antiviral activity against many types of enveloped viruses, including SARS-CoV-2, by multiple mechanisms: direct interaction with viral membrane proteins; disruption of the viral envelope; inhibition of viral proteases; induce host antiviral responses. Second, curcumin protects from lethal pneumonia and ARDS via targeting NF-κB, inflammasome, IL-6 trans signal, and HMGB1 pathways. Third, curcumin is safe and well-tolerated in both healthy and diseased human subjects. In conclusion, accumulated evidence indicates that curcumin may be a potential prophylactic therapeutic for COVID-19 in the clinic and public health settings.", "High-speed counter-current chromatography (HSCCC) was utilized as an effective procedure for isolation of targeted three anthocyanins di-glucosides from Syzygium cumini (SC) pulp by using an optimized biphasic successful combination of different solvents. The resulted fractions described by HPLC/ESI-MS to be delphinidin 3,5-diglucoside (DDG), petunidin 3,5-diglucoside (PDG), and malvidin 3,5-diglucoside (MDG). A weight of 150 mg of sample yielded 7.53, 22.68, and 39.09 mg for DDG, PDG, and MDG, respectively. It was stated that the target three anthocyanins possessed strong antioxidant activities. Furthermore, MDG exhibited definite advantages for inhibition of nitric oxide release and pro-inflammatory mediators like mouse interleukin 6 (IL-6), mouse interleukin (IL-1β) and mouse tumor necrosis factor (TNF-α) in lipopolysaccharide (LPS)-induced RAW264.7 macrophages. The results propose that HSCCC can be utilized to separate highly antioxidative and anti-inflammatory natural components from SC pulp. PRACTICAL APPLICATIONS: Anthocyanins are water-soluble pigments and considered one of the groups of bioactive compounds, which generally concentrate in the skin, and often also the flesh of some fruits and vegetables as glycosides like acylglycosides and aglycones of anthocyanidins. The fully ripe fruits of SC contain anthocyanins, like as delphinidin, cyanidin, and petunidin, which supply them a distinctive color and good antioxidant characteristics. HSCCC considers a system of liquid-liquid extraction with separating large quantities of materials, using a minimum of solvent. The findings of the study would pave a separation strategy for potential large-scale preparation of anthocyanins di-glucosides standards for compounds detection and reduce the inflammation symptoms through declining the induction of pro-inflammatory cytokines such as IL-1β, TNF-α and IL-6, which will also enhance the future notification on the structure-activity correlations of anthocyanins di-glucosides." ]
CD47 expression is upregulated following DNA damage in cancer cells	CD47 is a cell surface ligand expressed on all nucleated cells. It is a unique immune checkpoint protein acting as "don't eat me" signal to prevent phagocytosis and is constitutively overexpressed in many tumors. However, the underlying mechanism(s) for CD47 overexpression is not clear. Here, we show that irradiation (IR) as well as various other genotoxic agents induce elevated expression of CD47. This upregulation correlates with the extent of residual double-strand breaks (DSBs) as determined by γH2AX staining. Interestingly, cells lacking mre-11, a component of the MRE11-RAD50-NBS1 (MRN) complex that plays a central role in DSB repair, or cells treated with the mre-11 inhibitor, mirin, fail to elevate the expression of CD47 upon DNA damage. On the other hand, both p53 and NF-κB pathways or cell-cycle arrest do not play a role in CD47 upregualtion upon DNA damage. We further show that CD47 expression is upregulated in livers harvested from mice treated with the DNA-damage inducing agent Diethylnitrosamine (DEN) and in cisplatin-treated mesothelioma tumors. Hence, our results indicate that CD47 is upregulated following DNA damage in a mre-11-dependent manner. Chronic DNA damage response in cancer cells might contribute to constitutive elevated expression of CD47 and promote immune evasion.	[ "Knockdown or gene disruption of the ubiquitously expressed cell surface receptor CD47 protects non-malignant cells from genotoxic stress caused by ionizing radiation or cytotoxic chemotherapy but sensitizes tumors in an immune competent host to genotoxic stress. The selective radioprotection of non-malignant cells is mediated in part by enhanced autophagy and protection of anabolic metabolism pathways, but differential H2AX activation kinetics suggested that the DNA damage response is also CD47-dependent. A high throughput screen of drug sensitivities indicated that CD47 expression selectively sensitizes Jurkat T cells to inhibitors of topoisomerases, which are known targets of Schlafen-11 (SLFN11). CD47 mRNA expression positively correlated with schlafen-11 mRNA expression in a subset of human cancers but not the corresponding non-malignant tissues. CD47 mRNA expression was also negatively correlated with SLFN11 promoter methylation in some cancers. CD47 knockdown, gene disruption, or treatment with a CD47 function-blocking antibody decreased SLFN11 expression in Jurkat cells. The CD47 signaling ligand thrombospondin-1 also suppressed schlafen-11 expression in wild type but not CD47-deficient T cells. Re-expressing SLFN11 restored radiosensitivity to a CD47-deficient Jurkat cells. Disruption of CD47 in PC3 prostate cancer cells similarly decreased schlafen-11 expression and was associated with a CD47-dependent decrease in acetylation and increased methylation of histone H3 in the SLFN11 promoter region. The ability of histone deacetylase or topoisomerase inhibitors to induce SLFN11 expression in PC3 cells was lost when CD47 was targeted in these cells. Disrupting CD47 in PC3 cells increased resistance to etoposide but, in contrast to Jurkat cells, not to ionizing radiation. These data identify CD47 as a context-dependent regulator of SLFN11 expression and suggest an approach to improve radiotherapy and chemotherapy responses by combining with CD47-targeted therapeutics.", "The maintenance of genome stability depends on the DNA damage response (DDR), which is a functional network comprising signal transduction, cell cycle regulation and DNA repair. The metabolism of DNA double-strand breaks governed by the DDR is important for preventing genomic alterations and sporadic cancers, and hereditary defects in this response cause debilitating human pathologies, including developmental defects and cancer. The MRE11 complex, composed of the meiotic recombination 11 (MRE11), RAD50 and Nijmegen breakage syndrome 1 (NBS1; also known as nibrin) proteins is central to the DDR, and recent insights into its structure and function have been gained from in vitro structural analysis and studies of animal models in which the DDR response is deficient.", "The Mre11/Rad50/NBS1 (MRN) complex maintains genomic stability by bridging DNA ends and initiating DNA damage signaling through activation of the ATM kinase. Mre11 possesses DNA nuclease activities that are highly conserved in evolution but play unknown roles in mammals. To define the functions of Mre11, we engineered targeted mouse alleles that either abrogate nuclease activities or inactivate the entire MRN complex. Mre11 nuclease deficiency causes a striking array of phenotypes indistinguishable from the absence of MRN, including early embryonic lethality and dramatic genomic instability. We identify a crucial role for the nuclease activities in homology-directed double-strand-break repair and a contributing role in activating the ATR kinase. However, the nuclease activities are not required to activate ATM after DNA damage or telomere deprotection. Therefore, nucleolytic processing by Mre11 is an essential function of fundamental importance in DNA repair, distinct from MRN control of ATM signaling.", "Failing to properly repair damaged DNA drives the ageing process. Furthermore, age-related inflammation contributes to the manifestation of ageing. Recently, we demonstrated that the efficiency of repair of diethylnitrosamine (DEN)-induced double-strand breaks (DSBs) rapidly declines with age. We therefore hypothesised that with age, the decline in DNA damage repair stems from age-related inflammation. We used DEN-induced DNA damage in mouse livers and compared the efficiency of their resolution in different ages and following various permutations aimed at manipulating the liver age-related inflammation. We found that age-related deregulation of innate immunity was linked to altered gut microbiota. Consequently, antibiotic treatment, MyD88 ablation or germ-free mice had reduced cytokine expression and improved DSBs rejoining in 6-month-old mice. In contrast, feeding young mice with a high-fat diet enhanced inflammation and facilitated the decline in DSBs repair. This latter effect was reversed by antibiotic treatment. Kupffer cell replenishment or their inactivation with gadolinium chloride reduced proinflammatory cytokine expression and reversed the decline in DSBs repair. The addition of proinflammatory cytokines ablated DSBs rejoining mediated by macrophage-derived heparin-binding epidermal growth factor-like growth factor. Taken together, our results reveal a previously unrecognised link between commensal bacteria-induced inflammation that results in age-dependent decline in DNA damage repair. Importantly, the present study support the notion of a cell non-autonomous mechanism for age-related decline in DNA damage repair that is based on the presence of 'inflamm-ageing' cytokines in the tissue microenvironment, rather than an intrinsic cellular deficiency in the DNA repair machinery.", "Mre11 forms the core of the multifunctional Mre11-Rad50-Nbs1 (MRN) complex that detects DNA double-strand breaks (DSBs), activates the ATM checkpoint kinase, and initiates homologous recombination (HR) repair of DSBs. To define the roles of Mre11 in both DNA bridging and nucleolytic processing during initiation of DSB repair, we combined small-angle X-ray scattering (SAXS) and crystal structures of Pyrococcus furiosus Mre11 dimers bound to DNA with mutational analyses of fission yeast Mre11. The Mre11 dimer adopts a four-lobed U-shaped structure that is critical for proper MRN complex assembly and for binding and aligning DNA ends. Further, mutations blocking Mre11 endonuclease activity impair cell survival after DSB induction without compromising MRN complex assembly or Mre11-dependant recruitment of Ctp1, an HR factor, to DSBs. These results show how Mre11 dimerization and nuclease activities initiate repair of DSBs and collapsed replication forks, as well as provide a molecular foundation for understanding cancer-causing Mre11 mutations in ataxia telangiectasia-like disorder (ATLD).", "A key cellular response to DNA double-strand breaks (DSBs) is 5'-to-3' DSB resection by nucleases to generate regions of ssDNA that then trigger cell cycle checkpoint signaling and DSB repair by homologous recombination (HR). Here, we reveal that in the absence of exonuclease Exo1 activity, deletion or mutation of the Saccharomyces cerevisiae RecQ-family helicase, Sgs1, causes pronounced hypersensitivity to DSB-inducing agents. Moreover, we establish that this reflects severely compromised DSB resection, deficient DNA damage signaling, and strongly impaired HR-mediated repair. Furthermore, we show that the mammalian Sgs1 ortholog, BLM--whose deficiency causes cancer predisposition and infertility in people--also functions in parallel with Exo1 to promote DSB resection, DSB signaling and resistance to DSB-generating agents. Collectively, these data establish evolutionarily conserved roles for the BLM and Sgs1 helicases in DSB processing, signaling, and repair.", "Malignant Mesothelioma is a highly aggressive cancer, which is difficult to diagnose and treat. Here we describe the molecular, cellular and morphological characterization of a syngeneic system consisting of murine AB1, AB12 and AB22 mesothelioma cells injected in immunocompetent BALB/c mice, which allows the study of the interplay of tumor cells with the immune system. Murine mesothelioma cells, like human ones, respond to exogenous High Mobility Group Box 1 protein, a Damage-Associated Molecular Pattern that acts as a chemoattractant for leukocytes and as a proinflammatory mediator. The tumors derived from AB cells are morphologically and histologically similar to human MM tumors, and respond to treatments used for MM patients. Our system largely recapitulates human mesothelioma, and we advocate its use for the study of MM development and treatment." ]
Palm leaf impressions from the latest Maastrichtian (late Cretaceous to early Danian) sediments	Ten palm leaf impressions are documented from the latest Maastrichtian (late Cretaceous) to early Danian (earliest Paleocene) sediments (K-Pg,	[ "The climates of Asia are affected significantly by the extent and height of the Himalayan mountains and the Tibetan plateau. Uplift of this region began about 50 Myr ago, and further significant increases in altitude of the Tibetan plateau are thought to have occurred about 10-8 Myr ago, or more recently. However, the climatic consequences of this uplift remain unclear. Here we use records of aeolian sediments from China and marine sediments from the Indian and North Pacific oceans to identify three stages of evolution of Asian climates: first, enhanced aridity in the Asian interior and onset of the Indian and east Asian monsoons, about 9-8 Myr ago; next, continued intensification of the east Asian summer and winter monsoons, together with increased dust transport to the North Pacific Ocean, about 3.6-2.6 Myr ago; and last, increased variability and possible weakening of the Indian and east Asian summer monsoons and continued strengthening of the east Asian winter monsoon since about 2.6 Myr ago. The results of a numerical climate-model experiment, using idealized stepwise increases of mountain-plateau elevation, support the argument that the stages in evolution of Asian monsoons are linked to phases of Himalaya-Tibetan plateau uplift and to Northern Hemisphere glaciation.", "The often-used phrase 'the uplift of the Tibetan Plateau' implies a flat-surfaced Tibet rose as a coherent entity, and that uplift was driven entirely by the collision and northward movement of India. Here, we argue that these are misconceptions derived in large part from simplistic geodynamic and climate modeling, as well as proxy misinterpretation. The growth of Tibet was a complex process involving mostly Mesozoic collisions of several Gondwanan terranes with Asia, thickening the crust and generating complex relief before the arrival of India. In this review, Earth system modeling, paleoaltimetry proxies and fossil finds contribute to a new synthetic view of the topographic evolution of Tibet. A notable feature overlooked in previous models of plateau formation was the persistence through much of the Cenozoic of a wide east-west orientated deep central valley, and the formation of a plateau occurred only in the late Neogene through compression and internal sedimentation.", "Knowledge of the topographic evolution of the Tibetan Plateau is essential for understanding its construction and its influences on climate, environment, and biodiversity. Previous elevations estimated from stable isotope records from the Lunpola Basin in central Tibet, which indicate a high plateau since at least 35 Ma, are challenged by recent discoveries of low-elevation tropical fossils apparently deposited at 25.5 Ma. Here, we use magnetostratigraphic and radiochronologic dating to revise the chronology of elevation estimates from the Lunpola Basin. The updated ages reconcile previous results and indicate that the elevations of central Tibet were generally low (<2.3 km) at 39.5 Ma and high (3.5 to 4.5 km) at ~26 Ma. This supports the existence in the Eocene of low-elevation longitudinally oriented narrow regions until their uplift in the early Miocene, with potential implications for the growth mechanisms of the Tibetan Plateau, Asian atmospheric circulation, surface processes, and biotic evolution.", "The Lhasa terrane is a key region for understanding the paleoelevation of the southern Tibetan Plateau after India-Asia collision. The Gerze Basin, located in the northern part of the Lhasa terrane, is a shortening-related basin. We discovered Lagena laevis (Bandy) fossils in upper Eocene strata of the Gerze Basin. This type of foraminifera is associated with lagoon and estuarine environments, indicating that the northern part of the Lhasa terrane was near sea level during the late Eocene. We speculate that these foraminifera were transported inland by storm surges to low elevation freshwater lakes during times of marine transgressions. This inference is consistent with the relatively positive δ(18)O values in carbonate from the same deposits that indicate low palaeoelevations close to sea level. Considering the palaeoelevation results from the nearby Oligocene basins at a similar latitude and the volcanic history of the Lhasa terrane, we infer that large-magnitude surface uplift of the northern Lhasa terrane occurred between late Eocene and late Oligocene time." ]
X. tropicalis motile cilia transduce Wnt-Gsk3-Ppp1r11-Pp1 signaling	It is widely thought that Wnt/Lrp6 signaling proceeds through the cytoplasm and that motile cilia are signaling-inert nanomotors. Contrasting both views, we here show in the mucociliary epidermis of X. tropicalis embryos that motile cilia transduce a ciliary Wnt signal that is distinct from canonical β-catenin signaling. Instead, it engages a Wnt-Gsk3-Ppp1r11-Pp1 signaling axis. Mucociliary Wnt signaling is essential for ciliogenesis and it engages Lrp6 co-receptors that localize to cilia via a VxP ciliary targeting sequence. Live-cell imaging using a ciliary Gsk3 biosensor reveals an immediate response of motile cilia to Wnt ligand. Wnt treatment stimulates ciliary beating in X. tropicalis embryos and primary human airway mucociliary epithelia. Moreover, Wnt treatment improves ciliary function in X. tropicalis ciliopathy models of male infertility and primary ciliary dyskinesia (ccdc108, gas2l2). We conclude that X. tropicalis motile cilia are Wnt signaling organelles that transduce a distinct Wnt-Pp1 response.	[ "Primary ciliary dyskinesia is an inherited, currently incurable condition. In the respiratory system, primary ciliary dyskinesia causes impaired functioning of the mucociliary escalator, leading to nasal congestion, cough, and recurrent otitis media, and commonly progresses to cause more serious and permanent damage, including hearing deficits, chronic sinusitis, and bronchiectasis. New treatment options for the condition are thus necessary. In characterizing an immortalized human bronchial epithelial cell line (BCi-NS1.1) grown at an air-liquid interface to permit differentiation, we have identified that these cells have dyskinetic motile cilia. The cells had a normal male karyotype, and phenotypic markers of epithelial cell differentiation emerged, as previously shown. Ciliary beat frequency (CBF) as assessed by high-speed videomicroscopy was lower than normal (4.4 Hz). Although changes in CBF induced by known modulators were as expected, the cilia displayed a dyskinetic, circular beat pattern characteristic of central microtubular agenesis with outer doublet transposition. This ultrastructural defect was confirmed by electron microscopy. We propose that the BCi-NS1.1 cell line is a useful model system for examination of modulators of CBF and more specifically could be used to screen for novel drugs with the ability to enhance CBF and perhaps repair a dyskinetic ciliary beat pattern.", "Epithelial tubules consist of multiple cell types that are specialized for specific aspects of organ function. In the zebrafish pronephros, multiciliated cells (MCCs) are specialized for fluid propulsion, whereas transporting epithelial cells recover filtered-blood solutes. These cell types are distributed in a ;salt-and-pepper' fashion in the pronephros, suggesting that a lateral inhibition mechanism may play a role in their differentiation. We find that the Notch ligand Jagged 2 is expressed in MCCs and that notch3 is expressed in pronephric epithelial cells. Morpholino knockdown of either jagged 2 or notch3, or mutation in mind bomb (in which Notch signaling is impaired), dramatically expands ciliogenic gene expression, whereas ion transporter expression is lost, indicating that pronephric cells are transfated to MCCs. Conversely, ectopic expression of the Notch1a intracellular domain represses MCC differentiation. Gamma-secretase inhibition using DAPT demonstrated a requirement for Notch signaling early in pronephric development, before the pattern of MCC differentiation is apparent. Strikingly, we find that jagged 2 knockdown generates extra cilia and is sufficient to rescue the kidney cilia mutant double bubble. Our results indicate that Jagged 2/Notch signaling modulates the number of multiciliated versus transporting epithelial cells in the pronephros by way of a genetic pathway involving repression of rfx2, a key transcriptional regulator of the ciliogenesis program.", "The canonical Wnt signaling pathway is of paramount importance in development and disease. An emergent question is whether the upstream cascade of the canonical Wnt pathway has physiologically relevant roles beyond β-catenin-mediated transcription, which is difficult to study due to the pervasive role of this protein. Here, we show that transcriptionally silent spermatozoa respond to Wnt signals released from the epididymis and that mice mutant for the Wnt regulator Cyclin Y-like 1 are male sterile due to immotile and malformed spermatozoa. Post-transcriptional Wnt signaling impacts spermatozoa through GSK3 by (1) reducing global protein poly-ubiquitination to maintain protein homeostasis; (2) inhibiting septin 4 phosphorylation to establish a membrane diffusion barrier in the sperm tail; and (3) inhibiting protein phosphatase 1 to initiate sperm motility. The results indicate that Wnt signaling orchestrates a rich post-transcriptional sperm maturation program and invite revisiting transcription-independent Wnt signaling in somatic cells as well.", "The recent finding of the most common mutation (DF508) in cystic fibrosis in white populations has led to the publication of numerous data regarding its distribution and frequency. A review of the available data shows that there is a gradient in both the incidence of cystic fibrosis and the frequency of the DF508 mutation in Europe, the highest values being found in western Europe. It is postulated that the DF508 mutation was present in a western European population before the arrival of the Indo-Europeans from the Middle East; the mutation then spread into these migrating populations.", "Airway secretions are cleared by mucociliary clearance (MCC), in addition to other mechanisms such as cough, peristalsis, two-phase gas-liquid flow and alveolar clearance. MCC comprises the cephalad movement of mucus caused by the cilia lining the conducting airways until it can be swallowed or expectorated. MCC is a very complex process in which many variables are involved, all of which may modify the final outcome. The structure, number, movement and co-ordination of the cilia present in the airways as well as the amount, composition and rheological properties of the periciliary and mucus layers are determinants of MCC. Physiological factors such as age, sex, posture, sleep and exercise are reported to influence MCC due to a change in the cilia, the mucus or the periciliary layer, or a combination of these. Environmental pollution is suspected to have a depressant effect on MCC dependent on different factors such as pollutant concentration and the duration of exposure. Most studies focus on sulphur dioxide, sulphuric acid, nitrogen dioxide and ozone. Tobacco smoke and hairspray have been noted to have a negative influence on MCC. Some diseases are known to affect MCC, mostly negatively. The underlying mechanism differs from one illness to another. Immotile cilia syndrome, asthma, bronchiectasis, chronic bronchitis, cystic fibrosis and some acute respiratory tract infections are among the most frequently reported. The present paper reviews normal mucociliary clearance and the effects of diseases on this process.", "Specialised epithelia such as mucociliary, secretory and transporting epithelia line all major organs, including the lung, gut and kidney. Malfunction of these epithelia is associated with many human diseases. The frog embryonic epidermis possesses mucus-secreting and multiciliated cells, and has served as an excellent model system for the biogenesis of cilia. However, ionic regulation is important for the function of all specialised epithelia and it is not clear how this is achieved in the embryonic frog epidermis. Here, we show that a third cell type develops alongside ciliated and mucus-secreting cells in the tadpole skin. These cells express high levels of ion channels and transporters; therefore, we suggest that they are analogous to ionocytes found in transporting epithelia such as the mammalian kidney. We show that frog ionocytes express the transcription factor foxi1e, which is required for the development of these cells. Depletion of ionocytes by foxi1e knockdown has detrimental effects on the development of multiciliated cells, which show fewer and aberrantly beating cilia. These results reveal a newly identified role for ionocytes and suggest that the frog embryonic skin is a model system that is particularly suited to studying the interactions of different cell types in mucociliary, as well as in secretory and transporting, epithelia.", "The catalytic subunit of mammalian protein phosphatase-1 (PP1) is known to bind to a number of regulatory subunits, whose functions include the targeting of the catalytic subunit to the molecular proximity of its substrate proteins. In addition, PP1 is potently inhibited by several inhibitory polypeptides that include inhibitor-1 and inhibitor-2. In this study the yeast two-hybrid system was used to screen a human cDNA library for putative PP1-binding proteins. Ten putative positive clones were identified, one of which was found to be a partial cDNA of the hemochromatosis candidate gene V (HCG V) whose function was previously unknown. The full-length protein of 126 amino acid residues was expressed in Escherichia coli as a glutathione S-transferase fusion protein and also as a nonfusion protein. The recombinant protein inhibited recombinant and rabbit muscle protein phosphatase-1 with IC50s of ca. 1 nM, but did not inhibit PP2A. The term inhibitor-3 is proposed for this novel inhibitor. It is extremely hydrophilic, is heat stable, and behaves anomalously on SDS-PAGE with an apparent molecular mass of 23 kDa and on gel filtration with a relative molecular weight of 55 000, in contrast to its calculated molecular mass of 14 kDa. These characteristics are shared by the previously described protein phosphatase-1 inhibitor-2 and inhibitor-1 proteins." ]
is atrial fibrillation a risk factor?	Physical inactivity is a known risk factor for atrial fibrillation (AF). Wearable devices, such as smartwatches, present an opportunity to investigate the relation between daily step count and AF risk.	[ "The effects of inactivity and exercise training on established and novel cardiovascular risk factors are relatively modest and do not account for the impact of inactivity and exercise on vascular risk. We examine evidence that inactivity and exercise have direct effects on both vasculature function and structure in humans. Physical deconditioning is associated with enhanced vasoconstrictor tone and has profound and rapid effects on arterial remodelling in both large and smaller arteries. Evidence for an effect of deconditioning on vasodilator function is less consistent. Studies of the impact of exercise training suggest that both functional and structural remodelling adaptations occur and that the magnitude and time-course of these changes depends upon training duration and intensity and the vessel beds involved. Inactivity and exercise have direct \"vascular deconditioning and conditioning\" effects which likely modify cardiovascular risk.", "Self-reported and accelerometer-measured physical activity levels generally exhibit low correlation and agreement. The objective of this study is to compare estimates of physical activity among adults from a newly developed Canadian questionnaire with those obtained objectively by accelerometry. Data for 18- to 79-year-olds (N = 2,372) were collected in 2014 and 2015 as part of the Canadian Health Measures Survey (CHMS). Moderate-to-vigorous physical activity (MVPA) was reported on the household questionnaire by domain (transportation, recreation, and occupational or household) as part of the new Physical Activity Adult Questionnaire (PAAQ) and measured objectively using the Actical accelerometer. Correlation and mean difference analyses were used to assess the relationships between measured and reported physical activity variables. Linear regression was used to test the association between measured and reported physical activity and measures of obesity. On average, Canadian adults reported more physical activity than they accumulated on an accelerometer (49 minutes versus 23 minutes per day). The highest correlation observed was between accelerometer-measured MVPA and the sum of self-reported recreation and transportation activity (R = 0.36, p ⟨ 0.0001). The sum of activity from all domains (recreation + transportation + occupational or household) exhibited a lower correlation with measured variables because the occupational or household domain was negatively correlated with MVPA (R = -0.04). The occupational or household domain was positively correlated with light-intensity physical activity (R = 0.20, p ⟨ 0.0001). Respondents in the least active quintile were more likely than those in the most active quintile to report more activity than was measured by the accelerometer. On average, the most active quintile reported less activity than was measured by the accelerometer. The newly developed Canadian physical activity questionnaire exhibited modest correlation and agreement with accelerometer-measured physical activity among adults. Accelerometers and questionnaires provide complementary information, about different aspects of physical activity (actual movement versus perceived time). Consequently, one should exercise caution in using estimates derived from these methods interchangeably.", "The benefits for elderly individuals of regular participation in both cardiovascular and resistance-training programmes are great. Health benefits include a significant reduction in risk of coronary heart disease, diabetes mellitus and insulin resistance, hypertension and obesity as well as improvements in bone density, muscle mass, arterial compliance and energy metabolism. Additionally, increases in cardiovascular fitness (maximal oxygen consumption and endurance), muscle strength and overall functional capacity are forthcoming allowing elderly individuals to maintain their independence, increase levels of spontaneous physical activity and freely participate in activities associated with daily living. Taken together, these benefits associated with involvement in regular exercise can significantly improve the quality of life in elderly populations. It is noteworthy that the quality and quantity of exercise necessary to elicit important health benefits will differ from that needed to produce significant gains in fitness. This review describes the current recommendations for exercise prescriptions for the elderly for both cardiovascular and strength/resistance-training programmes. However, it must be noted that the benefits described are of little value if elderly individuals do not become involved in regular exercise regimens. Consequently, the major challenges facing healthcare professionals today concern: (i) the implementation of educational programmes designed to inform elderly individuals of the health and functional benefits associated with regular physical activity as well as how safe and effective such programmes can be; and (ii) design interventions that will both increase involvement in regular exercise as well as improve adherence and compliance to such programmes.", "The purpose of this study was to examine the effects of walking on resting systolic and diastolic blood pressure in adults. A total of 24 primary outcomes from 16 studies and 650 subjects (410 exercise, 240 control) met the criteria for inclusion: (1) randomized and nonrandomized controlled trials, (2) walking as the only intervention, (3) subjects apparently sedentary, (4) adult humans > or =18 years of age, (5) English-language studies published between January 1966 and December 1998, (6) resting blood pressure assessed, (7) training studies > or =4 weeks. Using a random effects model, statistically significant decreases of approximately 2% were found for both resting systolic and diastolic blood pressure (systolic, mean +/- SEM = -3 +/- 1 mm Hg, 95% confidence interval: -5 to -2 mm Hg; diastolic, mean +/- SEM = -2 +/- 1 mm Hg, 95% confidence interval: -3 to -1 mm Hg). Walking exercise programs reduce resting blood pressure in adults.", "N-terminal pro-brain natriuretic peptide (NT-proBNP) and high sensitivity Troponin T (hsTnT) are markers of cardiac injury used in diagnosis of heart failure and myocardial infarction respectively, and associated with increased risk of cardiovascular disease. Since physical activity is protective against cardiovascular disease and heart failure, we investigated whether higher levels of physical activity, and less sedentary behaviour were associated with lower NT-proBNP and hsTnT. Cross sectional study of 1130 men, age 70-91years, from the British Regional Heart Study, measured in 2010-2012. Fasting blood samples were analysed for NT-proBNP and hsTnT. Physical activity and sedentary behaviour were measured using ActiGraph GT3X accelerometers. Relationships between activity and NT-proBNP or hsTnT were non-linear; biomarker levels were lower with higher total activity, steps, moderate/vigorous activity and light activity only at low to moderate levels of activity. For example, for each additional 10min of moderate/vigorous activity, NT-proBNP was lower by 35.7% (95% CI -47.9, -23.6) and hsTnT by 8.4% (95% CI -11.1, -5.6), in men who undertook <25 or 50min of moderate/vigorous activity per day respectively. Biomarker levels increased linearly with increasing sedentary behaviour, but not independently of moderate/vigorous activity. Associations between biomarkers and moderate/vigorous activity (and between hsTnT and light activity) were independent of sedentary behaviour, suggesting activity is driving the relationships. In these older men with concomitantly low levels of physical activity, activity may be more important in protecting against cardiac health deterioration in less active individuals, although reverse causality might be operating.", "Poor glucose control increases the risk of vascular complications and cardiovascular mortality in patients with diabetes mellitus (DM). Our aim was to evaluate the efficacy of a long-term exercise training program on metabolic control and arterial stiffness in patients with type 2 DM. Fifty men with DM (age 52.3 +/- 5.6 years) were randomly assigned to the exercise training (E) or standard treatment for DM (control [C]) group for 24 months. Supervised exercise training included both endurance and muscle strength training 4 times/week. All exercise sessions were controlled by heart rate and intensity. Glycated hemoglobin A1c, insulin, leptin, blood lipids, blood pressure, maximal oxygen consumption in spiroergometry, and muscle strength were measured every 6 months. Arterial stiffness was assessed by measuring pulse wave velocity. Maximal oxygen consumption in spiroergometry (E 31.9 to 34.8 vs C 32.6 to 31.8 ml/kg/min; p = 0.003), muscle strength (sit-up test, E 12.7 to 20.8 vs C 14.6 to 13.1 times; p <0.001), hemoglobin A1c (E 8.2% to 7.6% vs C 8.0% to 8.3%; p = 0.006), and leptin (E 7.4 to 6.7 vs C 7.4 to 7.9 microg/L; p = 0.013) improved significantly in the E group, but no change or worsening in these variables occurred in the C group. Body weight was not different between groups at 2 years. However, pulse wave velocity increased in both groups (E +0.600 vs C +1.300 m/s; p = 0.27). In conclusion, long-term endurance and strength training was effective and resulted in improved metabolic control of DM compared with standard treatment. Despite significant cardiovascular risk reduction, conduit arterial elasticity did not improve.", "Arterial stiffness is an important risk factor for cardiovascular disease. Carotid-femoral pulse wave velocity (cfPWV) is the most recognized and established index of arterial stiffness. An emerging automatic measure of PWV primarily used in the Asian countries is brachial-ankle PWV (baPWV). To systematically compare these two methodologies, we conducted a multicenter study involving a total of 2287 patients. There was a significant positive relation between baPWV and cfPWV (r = 0.73). Average baPWV was approximately 20% higher than cfPWV. Both cfPWV and baPWV were significantly and positively associated with age (r = 0.56 and 0.64), systolic blood pressure (r = 0.49 and 0.61), and the Framingham risk score (r = 0.48 and 0.63). The areas under the receiver operating curves (ROCs) of PWV to predict the presence of both stroke and coronary artery disease were comparable between cfPWV and baPWV. Collectively, these results indicate that cfPWV and baPWV are indices of arterial stiffness that exhibit similar extent of associations with cardiovascular disease risk factors and clinical events.", "Regular physical activity appears to attenuate or even reverse age-related arterial stiffening. Yet, it is not clear if the reduced stiffening associated with habitual physical activity is also observed in community-dwelling older adults. Among 3893 older adults in a prospective cohort study, we associated physical activity with measures of central arterial stiffness (via carotid-femoral pulse wave velocity or cfPWV) and pressure pulsatility (via central pulse pressure or cPP). We also examined the association of long-term habitual physical activity, measured as persistence in physical activity levels from mid-life to late-life, with cfPWV and cPP among 1747 participants. The adjusted mean difference in cfPWV was lower, reflecting less arterial stiffness, for those with moderate (ß = -0.30 m/s) or high (ß = -0.38 m/s) physical activity compared with no physical activity. The adjusted mean difference in cPP was also lower for those with high (ß = -2.49 mmHg) physical activity, relative to no physical activity. Stronger effect estimates were observed among those with persistent physical activity from mid-life to late-life. Higher physical activity in late-life, and habitual physical activity from mid-life to late-life, is associated with lower central arterial stiffness and pressure pulsatility in a large population-based sample of community-dwelling older adults.", "In recent years, great emphasis has been placed on the role of arterial stiffness in the development of cardiovascular diseases. Indeed, the assessment of arterial stiffness is increasingly used in the clinical assessment of patients. Although several papers have previously addressed the methodological issues concerning the various indices of arterial stiffness currently available, and their clinical applications, clinicians and researchers still report difficulties in selecting the most appropriate methodology for their specific use. This paper summarizes the proceedings of several meetings of the European Network for Non-invasive Investigation of Large Arteries and is aimed at providing an updated and practical overview of the most relevant methodological aspects and clinical applications in this area.", "Carotid-femoral PWV (cfPWV) is a well-established measure of central arterial stiffness, while brachial-ankle PWV (baPWV) is being used more frequently in East Asian countries. Few studies have simultaneously characterized the distributions and correlates of segment-specific PWV measures and their associations with cardiovascular risk factors. We evaluated segment-specific PWV (cfPWV, baPWV, and femoral-ankle (faPWV)) in 4,974 older-aged African American and Caucasian adults in the community-based Atherosclerosis Risk in Communities (ARIC) Study using a standardized protocol and the OMRON VP-1000 Plus system. We examined the distribution and multivariable-adjusted correlates of PWV measures by race and sex. Mean age ranged from 74 ± 5 to 76 ± 5 years across race-sex groups. In all race-sex groups, cfPWV correlated with baPWV but not with faPWV, and cfPWV and baPWV were higher with age, whereas faPWV was not. Heart rate and systolic blood pressure (SBP) were positively associated and weight was negatively associated with all PWV measures; however, the associations with age, glycated hemoglobin, triglycerides, and high-density lipoprotein (HDL) cholesterol varied by segment and race-sex group. Our findings indicate that cfPWV and faPWV reflect distinct aspects of segment-specific vascular stiffness and their associated profile of cardiovascular risk factors. Even among older adults, age is associated with higher cfPWV and baPWV, but not with faPWV. Understanding factors that ostensibly play a role in increasing arterial stiffness in different arterial territories can inform opportunities for cardiovascular disease (CVD) prevention and risk management." ]
MUC1-C is a target for the treatment of BRAF(V600E) colorectal cancers.	Colorectal cancers (CRCs) harboring the BRAF(V600E) mutation are associated with aggressive disease and resistance to BRAF inhibitors by feedback activation of the receptor tyrosine kinase (RTK)→RAS→MAPK pathway. The oncogenic MUC1-C protein promotes progression of colitis to CRC; whereas there is no known involvement of MUC1-C in BRAF(V600E) CRCs. The present work demonstrates that MUC1 expression is significantly upregulated in BRAF(V600E) vs wild-type CRCs. We show that BRAF(V600E) CRC cells are dependent on MUC1-C for proliferation and BRAF inhibitor (BRAFi) resistance. Mechanistically, MUC1-C integrates induction of MYC in driving cell cycle progression with activation of the SHP2 phosphotyrosine phosphatase, which enhances RTK-mediated RAS→ERK signaling. We demonstrate that targeting MUC1-C genetically and pharmacologically suppresses (i) activation of MYC, (ii) induction of the NOTCH1 stemness factor, and (iii) the capacity for self-renewal. We also show that MUC1-C associates with SHP2 and is required for SHP2 activation in driving BRAFi-induced feedback of ERK signaling. In this way, targeting MUC1-C in BRAFi-resistant BRAF(V600E) CRC tumors inhibits growth and sensitizes to BRAF inhibition. These findings demonstrate that MUC1-C is a target for the treatment of BRAF(V600E) CRCs and for reversing their resistance to BRAF inhibitors by suppressing the feedback MAPK pathway.	[ "Mucin 1 C-terminal subunit (MUC1-C) has been introduced as a key regulator for acquiring drug resistance in various cancers, but the functional role of MUC1-C in urothelial carcinoma (UC) cells remains unknown. We aimed to elucidate the molecular mechanisms underlying the acquisition of cisplatin (CDDP) resistance through MUC1-C oncoprotein in UC cells. MUC1-C expression was examined immunohistochemically in tumor specimens of 159 UC patients who received CDDP-based perioperative chemotherapy. As a result, moderate to high MUC1-C expression was independently associated with poor survival in UC patients. Using human bladder cancer cell lines and CDDP-resistant (CR) cell lines, we compared the expression levels of MUC1-C, multiple drug resistance 1 (MDR1), the PI3K-AKT-mTOR pathway, and x-cystine/glutamate transporter (xCT) to elucidate the biological mechanisms contributing to the acquisition of chemoresistance. MUC1-C was strongly expressed in CR cell lines, followed with MDR1 expression via activation of the PI3K-AKT-mTOR pathway. MUC1-C also stabilized the expression of xCT, which enhanced antioxidant defenses by increasing intracellular glutathione (GSH) levels. MUC1 down-regulation showed MDR1 inhibition along with PI3K-AKT-mTOR pathway suppression. Moreover, it inhibited xCT stabilization and resulted in significant decreases in intracellular GSH levels and increased reactive oxygen species (ROS) generation. The MUC1-C inhibitor restored sensitivity to CDDP in CR cells and UC murine xenograft models. In conclusion, we found that MUC1-C plays a pivotal role in the acquisition of CDDP resistance in UC cells, and therefore the combined treatment of CDDP with a MUC1-C inhibitor may become a novel therapeutic option in CR UC patients.", "Rational use of DNA-damaging chemotherapy, with new combinations to heighten DNA replication stress, could improve outcomes in small cell lung cancer.", "The Brg/Brahma-associated factor (BAF, mSWI/SNF) chromatin remodeling complex is of importance in development and has been linked to prostate oncogenesis. The oncogenic MUC1-C protein promotes lineage plasticity in the progression of neuroendocrine prostate cancer (NEPC), however, there is no known association between MUC1-C and BAF. We report here that MUC1-C binds directly to the E2F1 transcription factor and that the MUC1-C→E2F1 pathway induces expression of embryonic stem cell-specific BAF (esBAF) components BRG1, ARID1A, BAF60a, BAF155, and BAF170 in castrate-resistant prostate cancer (CRPC) and NEPC cells. In concert with this previously unrecognized pathway, MUC1 was associated with increased expression of E2F1 and esBAF components in NEPC tumors as compared with CRPC, supporting involvement of MUC1-C in activating the E2F1→esBAF pathway with progression to NEPC. MUC1-C formed a nuclear complex with BAF and activated cancer stem cell (CSC) gene signatures and the core pluripotency factor gene network. The MUC1-C→E2F1→BAF pathway was necessary for induction of both the NOTCH1 effector of CSC function and the NANOG pluripotency factor, and collectively, this network drove CSC self-renewal. These findings indicate that MUC1-C promotes NEPC progression by integrating activation of E2F1 and esBAF with induction of NOTCH1, NANOG, and stemness. SIGNIFICANCE: These findings show that MUC1-C, which promotes prostate cancer progression, activates a novel pathway that drives the BAF remodeling complex, induces NOTCH1 and NANOG, and promotes self-renewal of prostate cancer stem cells.", "Small cell lung cancer (SCLC) is a recalcitrant malignancy defined by subtypes on the basis of differential expression of the ASCL1, NEUROD1, and POU2F3 transcription factors. The MUC1-C protein is activated in pulmonary epithelial cells by exposure to environmental carcinogens and promotes oncogenesis; however, there is no known association between MUC1-C and SCLC. We report that MUC1-C is expressed in classic neuroendocrine (NE) SCLC-A, variant NE SCLC-N and non-NE SCLC-P cells and activates the MYC pathway in these subtypes. In SCLC cells characterized by NE differentiation and DNA replication stress, we show that MUC1-C activates the MYC pathway in association with induction of E2F target genes and dysregulation of mitotic progression. Our studies further demonstrate that the MUC1-C→MYC pathway is necessary for induction of (i) NOTCH2, a marker of pulmonary NE stem cells that are the proposed cell of SCLC origin, and (ii) ASCL1 and NEUROD1. We also show that the MUC1-C→MYC→NOTCH2 network is necessary for self-renewal capacity and tumorigenicity of NE and non-NE SCLC cells. Analyses of datasets from SCLC tumors confirmed that MUC1 expression in single SCLC cells significantly associates with activation of the MYC pathway. These findings demonstrate that SCLC cells are addicted to MUC1-C and identify a potential new target for SCLC treatment. This work uncovers addiction of SCLC cells to MUC1-C, which is a druggable target that could provide new opportunities for advancing SCLC treatment.", "The oncogenic MUC1-C protein promotes dedifferentiation of castrate-resistant prostate cancer (CRPC) and triple-negative breast cancer (TNBC) cells. Chromatin remodeling is critical for the cancer stem cell (CSC) state; however, there is no definitive evidence that MUC1-C regulates chromatin accessibility and thereby expression of stemness-associated genes. We demonstrate that MUC1-C drives global changes in chromatin architecture in the dedifferentiation of CRPC and TNBC cells. Our results show that MUC1-C induces differentially accessible regions (DAR) across their genomes, which are significantly associated with differentially expressed genes (DEG). Motif and cistrome analysis further demonstrated MUC1-C-induced DARs align with genes regulated by the JUN/AP-1 family of transcription factors. MUC1-C activates the BAF chromatin remodeling complex, which is recruited by JUN in enhancer selection. In studies of the NOTCH1 gene, which is required for CRPC and TNBC cell self-renewal, we demonstrate that MUC1-C is necessary for (i) occupancy of JUN and ARID1A/BAF, (ii) increases in H3K27ac and H3K4me3 signals, and (iii) opening of chromatin accessibility on a proximal enhancer-like signature. Studies of the EGR1 and LY6E stemness-associated genes further demonstrate that MUC1-C-induced JUN/ARID1A complexes regulate chromatin accessibility on proximal and distal enhancer-like signatures. These findings uncover a role for MUC1-C in chromatin remodeling that is mediated at least in part by JUN/AP-1 and ARID1A/BAF in association with driving the CSC state. These findings show that MUC1-C, which is necessary for the CRPC and TNBC CSC state, activates a novel pathway involving JUN/AP-1 and ARID1A/BAF that regulates chromatin accessibility of stemness-associated gene enhancers." ]
Program death-1 inhibitor cemiplimab-rwlc	Program death-1 inhibitors, a class of immune-checkpoint inhibitors, are now the standard of care in a variety of cancer settings, including cutaneous malignancies, such as melanomas, Merkel cell, and cutaneous squamous cell carcinomas (cSCCs). The clinical trials that led to the approval of the programmed death-1 inhibitor cemiplimab-rwlc (Libtayo	[ "Immune checkpoint inhibitors (CPIs), monoclonal antibodies that target inhibitory receptors expressed on T cells, represent an emerging class of immunotherapy used in treating solid organ and hematologic malignancies. We describe the clinical and histologic features of 13 patients with CPI-induced acute kidney injury (AKI) who underwent kidney biopsy. Median time from initiation of a CPI to AKI was 91 (range, 21 to 245) days. Pyuria was present in 8 patients, and the median urine protein to creatinine ratio was 0.48 (range, 0.12 to 0.98) g/g. An extrarenal immune-related adverse event occurred prior to the onset of AKI in 7 patients. Median peak serum creatinine was 4.5 (interquartile range, 3.6-7.3) mg/dl with 4 patients requiring hemodialysis. The prevalent pathologic lesion was acute tubulointerstitial nephritis in 12 patients, with 3 having granulomatous features, and 1 thrombotic microangiopathy. Among the 12 patients with acute tubulointerstitial nephritis, 10 received treatment with glucocorticoids, resulting in complete or partial improvement in renal function in 2 and 7 patients, respectively. However, the 2 patients with acute tubulointerstitial nephritis not given glucocorticoids had no improvement in renal function. Thus, CPI-induced AKI is a new entity that presents with clinical and histologic features similar to other causes of drug-induced acute tubulointerstitial nephritis, though with a longer latency period. Glucocorticoids appear to be a potentially effective treatment strategy. Hence, AKI due to CPIs may be caused by a unique mechanism of action linked to reprogramming of the immune system, leading to loss of tolerance.", "Tumor cells from malignancies of any type-carcinoma, sarcoma, lymphoma, leukemia-may cause systemic arteriolar and capillary obstructions. The high shear rates of blood passing through these obstructions result in fragmentation of the red cells and can cause severe anemia, described as microangiopathic hemolytic anemia (MAHA).The thrombi caused by these obstructions consume platelets and can lead to severe thrombocytopenia. MAHA (defined by fragmented red cells on the peripheral blood smear and evidence of hemolysis) and thrombocytopenia are the clinical features of syndromes described as thrombotic microangiopathies (TMAs). If a malignancy is not recognized as the cause of TMA, the diagnosis of thrombotic thrombocytopenic purpura (TTP) may be considered and plasma exchange, the essential treatment for TTP, may be initiated-a critical decision because this treatment carries a high risk of serious complications. This review describes the clinical features that should suggest a search for systemic malignancy as the cause of unexpected MAHA and thrombocytopenia. Recognition of a systemic malignancy is critical to the initiation of appropriate chemotherapy and avoidance of inappropriate use of plasma exchange treatment." ]
Graph convolutional network and self-attention for DNA storage encoding	DNA Encoding, as a key step in DNA storage, plays an important role in reading and writing accuracy and the storage error rate. However, currently, the encoding efficiency is not high enough and the encoding speed is not fast enough, which limits the performance of DNA storage systems. In this work, a DNA storage encoding system with a graph convolutional network and self-attention (GCNSA) is proposed. The experimental results show that DNA storage code constructed by GCNSA increases by 14.4% on average under the basic constraints, and by 5%-40% under other constraints. The increase of DNA storage codes effectively improves the storage density of 0.7-2.2% in the DNA storage system. The GCNSA predicted more DNA storage codes in less time while ensuring the quality of codes, which lays a foundation for higher read and write efficiency in DNA storage.	[ "Rapid advances in nanopore technologies for sequencing single long DNA and RNA molecules have led to substantial improvements in accuracy, read length and throughput. These breakthroughs have required extensive development of experimental and bioinformatics methods to fully exploit nanopore long reads for investigations of genomes, transcriptomes, epigenomes and epitranscriptomes. Nanopore sequencing is being applied in genome assembly, full-length transcript detection and base modification detection and in more specialized areas, such as rapid clinical diagnoses and outbreak surveillance. Many opportunities remain for improving data quality and analytical approaches through the development of new nanopores, base-calling methods and experimental protocols tailored to particular applications.", "Digital production, transmission and storage have revolutionized how we access and use information but have also made archiving an increasingly complex task that requires active, continuing maintenance of digital media. This challenge has focused some interest on DNA as an attractive target for information storage because of its capacity for high-density information encoding, longevity under easily achieved conditions and proven track record as an information bearer. Previous DNA-based information storage approaches have encoded only trivial amounts of information or were not amenable to scaling-up, and used no robust error-correction and lacked examination of their cost-efficiency for large-scale information archival. Here we describe a scalable method that can reliably store more information than has been handled before. We encoded computer files totalling 739 kilobytes of hard-disk storage and with an estimated Shannon information of 5.2 × 10(6) bits into a DNA code, synthesized this DNA, sequenced it and reconstructed the original files with 100% accuracy. Theoretical analysis indicates that our DNA-based storage scheme could be scaled far beyond current global information volumes and offers a realistic technology for large-scale, long-term and infrequently accessed digital archiving. In fact, current trends in technological advances are reducing DNA synthesis costs at a pace that should make our scheme cost-effective for sub-50-year archiving within a decade.", "DNA is an attractive medium to store digital information. Here we report a storage strategy, called DNA Fountain, that is highly robust and approaches the information capacity per nucleotide. Using our approach, we stored a full computer operating system, movie, and other files with a total of 2.14 × 106 bytes in DNA oligonucleotides and perfectly retrieved the information from a sequencing coverage equivalent to a single tile of Illumina sequencing. We also tested a process that can allow 2.18 × 1015 retrievals using the original DNA sample and were able to perfectly decode the data. Finally, we explored the limit of our architecture in terms of bytes per molecule and obtained a perfect retrieval from a density of 215 petabytes per gram of DNA, orders of magnitude higher than previous reports.", "DNA is a promising next-generation data storage medium, but challenges remain with synthesis costs and recording latency. Here, we describe a prototype of a DNA data storage system that uses an extended molecular alphabet combining natural and chemically modified nucleotides. Our results show that MspA nanopores can discriminate different combinations and ordered sequences of natural and chemically modified nucleotides in custom-designed oligomers. We further demonstrate single-molecule sequencing of the extended alphabet using a neural network architecture that classifies raw current signals generated by Oxford Nanopore sequencers with an average accuracy exceeding 60% (39× larger than random guessing). Molecular dynamics simulations show that the majority of modified nucleotides lead to only minor perturbations of the DNA double helix. Overall, the extended molecular alphabet may potentially offer a nearly 2-fold increase in storage density and potentially the same order of reduction in the recording latency, thereby enabling new implementations of molecular recorders.", "The density and long-term stability of DNA make it an appealing storage medium, particularly for long-term data archiving. Existing DNA storage technologies involve the synthesis and sequencing of multiple nominally identical molecules in parallel, resulting in information redundancy. We report the development of encoding and decoding methods that exploit this redundancy using composite DNA letters. A composite DNA letter is a representation of a position in a sequence that consists of a mixture of all four DNA nucleotides in a predetermined ratio. Our methods encode data using fewer synthesis cycles. We encode 6.4 MB into composite DNA, with distinguishable composition medians, using 20% fewer synthesis cycles per unit of data, as compared to previous reports. We also simulate encoding with larger composite alphabets, with distinguishable composition deciles, to show that 75% fewer synthesis cycles are potentially sufficient. We describe applicable error-correcting codes and inference methods, and investigate error patterns in the context of composite DNA letters.", "The cytochrome P450 (CYP) gene family catalyzes drug metabolism and bioactivation and is therefore relevant to drug development. We determined potency values for 17,143 compounds against five recombinant CYP isozymes (1A2, 2C9, 2C19, 2D6 and 3A4) using an in vitro bioluminescent assay. The compounds included libraries of US Food and Drug Administration (FDA)-approved drugs and screening libraries. We observed cross-library isozyme inhibition (30-78%) with important differences between libraries. Whereas only 7% of the typical screening library was inactive against all five isozymes, 33% of FDA-approved drugs were inactive, reflecting the optimized pharmacological properties of the latter. Our results suggest that low CYP 2C isozyme activity is a common property of drugs, whereas other isozymes, such as CYP 2D6, show little discrimination between drugs and unoptimized compounds found in screening libraries. We also identified chemical substructures that differentiated between the five isozymes. The pharmacological compendium described here should further the understanding of CYP isozymes.", "DNA has recently emerged as an attractive medium for archival data storage. Recent work has demonstrated proof-of-principle prototype systems; however, very uneven (biased) sequencing coverage has been reported, which indicates inefficiencies in the storage process. Deviations from the average coverage in the sequence copy distribution can either cause wasteful provisioning in sequencing or excessive number of missing sequences. Here, we use millions of unique sequences from a DNA-based digital data archival system to study the oligonucleotide copy unevenness problem and show that the two paramount sources of bias are the synthesis and amplification (PCR) processes. Based on these findings, we develop a statistical model for each molecular process as well as the overall process. We further use our model to explore the trade-offs between synthesis bias, storage physical density, logical redundancy, and sequencing redundancy, providing insights for engineering efficient, robust DNA data storage systems.", "Deeply understanding the properties (e.g., chemical or biological characteristics) of small molecules plays an essential role in drug development. A large number of molecular property datasets have been rapidly accumulated in recent years. However, most of these datasets contain only a limited amount of data, which hinders deep learning methods from making accurate predictions of the corresponding molecular properties. In this work, we propose a transfer learning strategy to alleviate such a data scarcity problem by exploiting the similarity between molecular property prediction tasks. We introduce an effective and interpretable computational framework, named MoTSE (Molecular Tasks Similarity Estimator), to provide an accurate estimation of task similarity. Comprehensive tests demonstrated that the task similarity derived from MoTSE can serve as useful guidance to improve the prediction performance of transfer learning on molecular properties. We also showed that MoTSE can capture the intrinsic relationships between molecular properties and provide meaningful interpretability for the derived similarity." ]
Atrial fibrillation, anticoagulation, and racial, ethnic, and socioeconomic disparities	Atrial fibrillation (AF) is the most prevalent arrhythmia in the United States and is responsible for 1 in 7 ischemic strokes. While anticoagulation is effective at preventing strokes, prior work has highlighted significant disparities in anticoagulation prescribing. Furthermore, racial, ethnic, sex, and socioeconomic disparities in AF outcomes have been described. As such, we aimed to review recent data on disparities with respect to anticoagulation for AF published between January 2018 and February 2021. The search string consisted of 7 phrases that combined AF, anticoagulation, and disparities involving sex, race, ethnicity, income, socioeconomic status (SES), and access to care and identified 13 relevant articles. The aggregate data demonstrated that Black patients were less likely to be prescribed anticoagulation than patients of other racial/ethnic groups. Additionally, Black patients were more likely to be prescribed warfarin instead of direct oral anticoagulants (DOACs) despite evidence that DOACs are safer and better tolerated. Lower-income patients and patients with less education were also less likely to receive DOACs. Some studies found that women were less likely to be anticoagulated than men even when their estimated stroke risk was higher, although other studies did not show sex-based differences. Building upon prior work, our study demonstrates that racial and ethnic disparities have persisted in the management of AF. Additionally, we our work highlights that there are significant disparities in anticoagulation management for AF associated with sex, income, and education. More work is needed to identify mechanisms for these disparities and identify potential solutions to achieve pharmacoequity.	[ "To examine sex differences in thromboprophylaxis in patients with atrial fibrillation before and after the introduction of non-vitamin K oral anticoagulants, we performed a cross-sectional registry study based on anonymized individual-level patient data of all individuals with a diagnosis of nonvalvular atrial fibrillation (International Classification of Diseases, Tenth Revision code I48) in the region of Stockholm, Sweden (2.2 million inhabitants), in 2011 and 2015, respectively. Thromboprophylaxis improved considerably during the period. During 2007 to 2011, 23,198 men and 18,504 women had an atrial fibrillation diagnosis. In 2011, more men than women (53% men vs 48% women) received oral anticoagulants (almost exclusively warfarin) and more women received aspirin only (35% women vs 30% men), whereas there was no sex difference for no thromboprophylaxis (17%). During 2011 to 2015, 27,237 men and 20,461 women had a diagnosis of atrial fibrillation. Compared with the earlier time period, a higher proportion used oral anticoagulants (71% women vs 70% men), but fewer women ≥80 years received anticoagulants (67% women vs 72% men), more women received aspirin (15% women vs 13% men), and fewer women had no thromboprophylaxis (15% women vs 17% men). Patients with co-morbidities potentially complicating oral anticoagulant use used more oral anticoagulant in 2015 compared with 2011. The sex differences observed in 2011 with fewer women using oral anticoagulants had disappeared in 2015 except in women 80 years and older and in patients with complicated co-morbidity.", "Although it is well established that atrial fibrillation (AF) causes ischemic stroke, the relationship between AF and cognitive impairment is unclear. The aim of this systematic review is to investigate whether AF is associated with cognitive impairment or dementia. An electronic search of Medline, Embase, Psychlit, Cinahl and the Cochrane library was performed in March 2000 to identify studies in which the primary aim was to investigate the relationship between AF and cognitive impairment or dementia. Studies with relevant data on both cognitive function and AF (even if that was not the primary aim of the study) were also identified. Further references were identified from these sources. Ten studies were identified, of which 4 were cross-sectional, 5 were case-control, and 1 was a prospective cohort study. The methodology and measures of cognition varied substantially, so it was not valid to apply formal meta-analysis techniques to the results. However, the methodology in all the studies was flawed in at least 1 aspect, including the reporting of results, external validity, and internal validity. Seven studies found an association between AF and at least 1 measure of cognition whereas 3 studies did not find an association. The evidence that AF is associated with cognitive impairment is inconclusive. Further studies are required to establish whether there is a relationship between AF and cognitive impairment, and if so, whether the relationship is causal.", "It is important to identify candidates who warrant extended cardiac monitoring after ischemic stroke. We investigated the predictive performance of the CHADS2 and CHA2DS2-VASc scores for previously unknown atrial fibrillation during in-hospital electrocardiographic monitoring. Patients were selected from a prospective trial in China. The clinical prediction of the scores was examined using the C statistic. Multivariate logistic regressions were performed to analyze the relevant risk factors. Among 1315 patients enrolled in study, previously unknown atrial fibrillation was detected in 110 (8.4%). Age, heart failure, NIHSS on admission, creatinine, and triglycerides were independently associated with newly detected atrial fibrillation. For newly detected atrial fibrillation, the C statistic value was 0.55 (OR 1.14, 95% CI: 0.97-1.33) for CHADS2 and 0.62 (OR 1.26, 95% CI: 1.12-1.42) for CHA2DS2-VASc; adding newly identified risk factors to these two scores, the value of C statistic was improved to 0.74 and 0.75, respectively. Age, heart failure, NIHSS on admission, creatinine and triglycerides were independent predictors of previously unknown atrial fibrillation. The CHADS2 and CHA2DS2-VASc scores are useful but not optimal for atrial fibrillation prediction. Addition of newly identified risk factors to these two scores resulted in significant improvement of the predictive performance.", "Atrial fibrillation (AF) is a common arrhythmia that poses a significant risk of stroke. Cross-sectional and case-control studies have shown evidence of associations between AF and breast or colorectal cancer, but there have been no longitudinal studies in which this has been assessed. We prospectively examined a cohort of 93,676 postmenopausal women enrolled in the Women's Health Initiative from 1994 to 1998 to determine whether there are relationships between baseline AF and the development of invasive breast or colorectal cancer. The prevalence of self-reported physician diagnosis of AF at baseline was 5.1%. Over approximately 15 years of follow-up, the incidence of invasive breast cancer was 5.7%, and the incidence of colorectal cancer was 1.6%. Adjusted hazard ratios and 95% confidence intervals were obtained using Cox proportional hazards models. We found no significant association between AF and incident colorectal cancer, but we did see a 19% excess risk of invasive breast cancer among those with AF (adjusted hazard ratio (HR) = 1.19, 95% confidence interval (CI): 1.03, 1.38). Additional adjustment for baseline use of cardiac glycosides attenuated the association between AF and invasive breast cancer (HR = 1.01, 95% CI: 0.85, 1.20). Cardiac glycoside use was strongly associated with incident invasive breast cancer (HR = 1.68, 95% CI: 1.33, 2.12) independent of AF and other confounders. Mechanisms of the associations among breast cancer, AF, and cardiac glycosides need further investigation.", "Hybrid ablation (HABL) of atrial fibrillation combining endoscopic, minimally invasive, closed chest epicardial ablation with endocardial CARTO-guided accuracy was introduced to overcome the limitations of current therapeutic options for patients with persistent (PSAF) and longstanding persistent atrial fibrillation (LSPAF). The purpose of this study was to evaluate the procedural safety and feasibility as well as effectiveness of HABL in patients with PSAF and LSPAF 1 year after the procedure. The study is a single-center, prospective clinical registry. From 07/2009 to 12.2014, 90 patients with PSAF (n = 39) and LSPAF (n = 51), at the mean age of 54.8 ±9.8, in mean EHRA class 2.6, underwent HABL. 64.4% of patients had a history of prior cardioversion or catheter ablation. Thirteen patients had LVEF less than 35%. Mean AF duration was 4.5 ±3.7 years. Patients were scheduled for 3-, 6- and 12-month follow-up with 7-day Holter monitoring. At 6 months after the procedure 78% (54/69) of patients were in SR. At 12 months after the procedure 86% (59/69) were in SR and 62.3% (43/69) in SR and off class I/III antiarrhythmic drugs (AADs). Only 1% (1/69) of patients required a repeat ablation for atrial flutter. A significant decrease in LA dimension and an increase in LVEF were noted. A combination of epicardial and endocardial RF ablation should be considered as a treatment option for patients with persistent and long-standing persistent atrial fibrillation as it is safe and effective in restoring sinus rhythm.", "Atrial fibrillation (AF) has been associated with cognitive decline independent of stroke, suggesting additional effects of AF on the brain. We aimed to assess the association between AF and brain function and structure in a general elderly population. This is a cross-sectional analysis of 4251 nondemented participants (mean age, 76 ± 5 years) in the population-based Age, Gene/Environment Susceptibility-Reykjavik Study. Medical record data were collected for the presence, subtype, and time from first diagnosis of AF; 330 participants had AF. Brain volume measurements, adjusted for intracranial volume, and presence of cerebral infarcts were determined with magnetic resonance imaging. Memory, speed of processing, and executive function composites were calculated from a cognitive test battery. In a multivariable linear regression model, adjustments were made for demographic factors, cardiovascular risk factors, and cerebral infarcts. Participants with AF had lower total brain volume compared with those without AF (P<0.001). The association was stronger with persistent/permanent than paroxysmal AF and with increased time from the first diagnosis of the disease. Of the brain tissue volumes, AF was associated with lower volume of gray and white matter hyperintensities (P<0.001 and P = 0.008, respectively), but not of white matter hyperintensities (P = 0.49). Participants with AF scored lower on tests of memory. AF is associated with smaller brain volume, and the association is stronger with increasing burden of the arrhythmia. These findings suggest that AF has a cumulative negative effect on the brain independent of cerebral infarcts.", "Atrial fibrillation (AF) remains a growing problem in the United States and worldwide, imposing a high individual and health system burden, including increased resource consumption due to repeated hospitalizations, stroke, dementia, heart failure, and death. This comprehensive review summarizes the most recent data on sex-related differences in risks associated with AF. Women with AF have increased risk of stroke and death compared to men, and possible reasons for this disparity are explored. Women also continue to have worse symptoms and quality of life, and poorer outcomes with stroke prevention, as well as with rate and rhythm control management strategies. Many current rhythm control treatment strategies for AF, including cardioversion and ablation, are used less frequently in women as compared to men, whereas women are more likely to be treated with rate control strategies or antiarrhythmic drugs. Sex differences should be considered in treating women with AF to improve outcomes and women and men should be offered the same interventions for AF. We need to improve the evidence base to understand if variation in utilization of rate and rhythm control management between men and women represents health inequities or appropriate clinical judgement.", "Worldwide, there is growing evidence that quality of international normalized ratio (INR) control in atrial fibrillation patients treated with Vitamin K Antagonists (VKA) is suboptimal. However, sex disparities in population-based real-world settings have been scarcely studied, as well as patterns of switching to second-line Non-VKA oral anticoagulants (NOAC). We aimed to assess the quality of INR control in atrial fibrillation patients treated with VKA in the region of Valencia, Spain, for the whole population and differencing by sex, and to identify factors associated with poor control. We also quantified switching to Non-VKA oral anticoagulants (NOAC) and we identified factors associated to switching. This is a cross-sectional, population-based study. Information was obtained through linking different regional electronic databases. Outcome measures were Time in Therapeutic Range (TTR) and percentage of INR determinations in range (PINRR) in 2015, and percentage of switching to NOAC in 2016, for the whole population and stratified by sex. We included 22,629 patients, 50.4% were women. Mean TTR was 62.3% for women and 63.7% for men, and PINNR was 58.3% for women and 60.1% for men (p<0.001). Considering the TTR<65% threshold, 53% of women and 49.3% of men had poor anticoagulation control (p<0.001). Women, long-term users antiplatelet users, and patients with comorbidities, visits to Emergency Department and use of alcohol were more likely to present poor INR control. 5.4% of poorly controlled patients during 2015 switched to a NOAC throughout 2016, with no sex differences. The quality of INR control of all AF patients treated with VKA in 2015 in our Southern European region was suboptimal, and women were at a higher risk of poor INR control. This reflects sex disparities in care, and programs for improving the quality of oral anticoagulation should incorporate the gender perspective. Clinical inertia may be lying behind the observed low rates of switching in patient with poor INR control.", "Atrial fibrillation (AF) is a growing health problem worldwide. While the disease plagues both men and women, this arrhythmia does not affect both sexes equally. Women are more likely to have major adverse outcomes such as stroke and its sequela; however, recent data on stroke prevention show improving outcomes. The purpose of this review of the recent literature is to summarize important updates on risk scores and management of patients with AF. It has been well known that women have a higher risk of strokes than men when untreated or when treated with warfarin. Current risk scores emphasizing new risk factors such as the higher risk of strokes in women have been incorporated into clinical guidelines. However, with the use of direct oral anticoagulants, this sex disparity on stroke is no longer seen and women have less major bleeding than men. The use of cardiac glycosides is associated with increased incidence of breast cancer, and this medication is used more in women. Procedural complications for the management of AF are higher in women. The study of the pathophysiology of AF and its management is a rapidly evolving area of cardiovascular medicine. Sex-specific data is necessary to achieve advances in the field and improve the outcomes in both men and women.", "The increasing availability of human cardiac tissues for study are critically important in increasing our understanding of the impact of gender, age, and other parameters, such as medications and cardiac disease, on arrhythmia susceptibility. In this study, we aimed to compare the mRNA expression of 89 ion channel subunits, calcium handling proteins, and transcription factors important in cardiac conduction and arrhythmogenesis in the left atria (LA) and ventricles (LV) of failing and nonfailing human hearts of both genders. Total RNA samples, prepared from failing male (n = 9) and female (n = 7), and from nonfailing male (n = 9) and female (n = 9) hearts, were probed using custom-designed Taqman gene arrays. Analyses were performed to explore the relationships between gender, failure state, and chamber expression. Hierarchical cluster analysis revealed chamber specific expression patterns, but failed to identify disease- or gender-dependent clustering. Gender-specific analysis showed lower expression levels in transcripts encoding for K(v)4.3, KChIP2, K(v)1.5, and K(ir)3.1 in the failing female as compared with the male LA. Analysis of LV transcripts, however, did not reveal significant differences based on gender. Overall, our data highlight the differential expression and transcriptional remodeling of ion channel subunits in the human heart as a function of gender and cardiac disease. Furthermore, the availability of such data sets will allow for the development of disease-, gender-, and, most importantly, patient-specific cardiac models, with the ability to utilize such information as mRNA expression to predict cardiac phenotype." ]
Multiple forms of stigmatization among caregivers of children with autism with an ethnic minority background	Multiple forms of stigmatization are experienced by caregivers of children with autism among ethnic minority (EM) groups in various countries. Such forms of stigmatization can lead to delayed assessment and services for mental health among children and caregivers. This review identified the research literature on the types of stigmatization experienced by caregivers of children with autism with an EM background. A total of 19 studies published after 2010 (i.e., 12 from the USA, 2 from the UK, 1 from Canada, and 1 from New Zealand) of caregivers of 20 ethnicities were identified and reviewed, and their reporting qualities systematically also assessed. Four main themes: (1) self-stigma, (2) social stigma, (3) stigma towards EM parents of children on the autism spectrum, and (4) service utilization stigma, and nine sub-themes were identified. The discrimination experienced by caregivers were extracted, synthesized, and further discussed. While the reporting quality of the studies included is good, the depth of the understanding of this under-researched yet important phenomenon is very limited. The multiple forms of stigmatization experiences are complex, and it may be difficult to disentangle whether the causes of stigmatization were autism and/or EM related, and the types of stigmatization can vary enormously among different ethnic groups in different societies. More quantitative studies are needed to quantify the impacts of multiple forms of stigmatization on families of children with autism in EM groups so that more socially inclusive support for caregivers with an EM background in host countries can be developed.	[ "Early identification of children with autism spectrum disorder (ASD) facilitates timely access to intervention services. Yet, few population-based data exist on ASD identification among preschool-aged children. The authors aimed to describe ASD prevalence and characteristics among 4-year-old children in 5 of 11 sites participating in the 2010 Autism and Developmental Disabilities Monitoring Network. Children with ASD were identified through screening of health and education records for ASD indicators, data abstraction and compilation for each child, and clinician review of records. ASD prevalence estimates, ages at first evaluation and ASD diagnosis, cognitive test scores, and demographics were compared for 4-year-old children and 8-year-old children living in the same areas. Among 58,467 children in these 5 sites, 4-year-old ASD prevalence was 13.4 per 1000, which was 30% lower than 8-year-old ASD prevalence. Prevalence of ASD without cognitive impairment was 40% lower among 4-year-olds compared with 8-year-olds, but prevalence of ASD with cognitive impairment was 20% higher among 4-year-olds compared with 8-year-olds. Among 4-year-olds with ASD, female and non-Hispanic white children were more likely to receive their first comprehensive evaluation by age 36 months compared with male and non-Hispanic black children, respectively. Among children diagnosed with ASD by age 48 months, median age at first comprehensive evaluation was 27 months for 4-year-olds compared with 32 months for 8-year-olds. Population-based ASD surveillance among 4-year-old children provides valuable information about the early identification of children with ASD and suggests progression toward lowering the age of first ASD evaluation within participating Autism and Developmental Disabilities Monitoring communities.", "This evidence base update examines the level of empirical support for interventions for children with autism spectrum disorder (ASD) younger than 5 years old. It focuses on research published since a previous review in this journal (Rogers & Vismara, 2008 ). We identified psychological or behavioral interventions that had been manualized and evaluated in either (a) experimental or quasi-experimental group studies or (b) systematic reviews of single-subject studies. We extracted data from all studies that met these criteria and were published after the previous review. Interventions were categorized across two dimensions. First, primary theoretical principles included applied behavior analysis (ABA), developmental social-pragmatic (DSP), or both. Second, practice elements included scope (comprehensive or focused), modality (individual intervention with the child, parent training, or classrooms), and intervention targets (e.g., spoken language or alternative and augmentative communication). We classified two interventions as well-established (individual, comprehensive ABA and teacher-implemented, focused ABA + DSP), 3 as probably efficacious (individual, focused ABA for augmentative and alternative communication; individual, focused ABA + DSP; and focused DSP parent training), and 5 as possibly efficacious (individual, comprehensive ABA + DSP; comprehensive ABA classrooms; focused ABA for spoken communication; focused ABA parent training; and teacher-implemented, focused DSP). The evidence base for ASD interventions has grown substantially since 2008. An increasing number of interventions have some empirical support; others are emerging as potentially efficacious. Priorities for future research include improving outcome measures, developing interventions for understudied ASD symptoms (e.g., repetitive behaviors), pinpointing mechanisms of action in interventions, and adapting interventions for implementation with fidelity by community providers.", "In this article, we discuss findings of a hermeneutic phenomenological study that sought to describe the experiences of parents who have a child with autism. Qualitative interviews were conducted with parents from 16 families of children with autism residing in a western Canadian province. \"Living in a world of our own\" emerged as the essence of the parents' experiences. In \"living in a world of our own,\" parents described a world of isolation. Three themes representing the essential challenging elements of the parents' experiences included vigilant parenting, sustaining the self and family, and fighting all the way. Although much is known about the fundamental importance of support to parents of children with chronic conditions and/or disabilities, findings from this study indicate that knowledge has not been adequately transferred to the care of children with autism.", "Parents of children with autism spectrum disorder (ASD) report high levels of stress that can interfere with important child and family treatments. Limited past research considers how the caregiving experience and social determinants of health may contribute to treatment engagement and outcomes, particularly in under-represented families, who already experience service and health disparities. We aimed to assess the experiences of caring for an individual with ASD, with specific emphasis on perceptions of stress. Three key informant interviews were conducted with parents (n=1) and providers (n=2) of children with ASD to refine interview guide questions. Once questions were refined, four focus groups (n=17) and one key informant interview were conducted with parents of children with ASD who were (a) non-white, (b) Spanish speakers, (c) of limited financial resources, and/or (d) living in rural counties. All participants lived in Western New York, with the majority residing in Rochester. Content analysis by two independent coders was used to identify and refine themes. Themes included: (a) caregiving for an individual with ASD can cause interference with family functioning, (b) misperceptions of ASD contribute to caregiver stress, (c) culture contributes to stressors for parents, and (d) service navigation difficulties are a significant source of stress. Suggestions for interventions to address parents stress included: modular and integrative treatments for multiple content areas, addressing cultural barriers to treatment engagement, and education on ASD to the community. Parent-focused interventions for caregivers of children with ASD should specifically explore and address service and health disparities for parents, especially those predicated on race, ethnicity, rurality, and language of origin. Interventions should also be individualized to parent characteristics and experiences. In future research on parent training, the unique contributions of caregiver stress and other characteristics (e.g., race-related stress, geographic location) should be included as potential modifiers of treatment.", "The aim of this article is to briefly review the literature on stigmatization and more generally identity threats, to focus more specifically of the way people appraise and cope with those threatening situations. Based on the transactional model of stress and coping of Lazarus and Folkman (1984), we propose a model of coping with identity threats that takes into accounts the principle characteristic of stigma, its devaluing aspect. We present a model with specific antecedents, a refined appraisal phase and a new classification of coping strategies based on the motives that may be elicited by the threatening situation, those of protecting and/or enhancing the personal and/or social identity.", "Latinos comprise the largest racial/ethnic group in the United States and have 2-3 times the prevalence of type 2 diabetes mellitus as Caucasians. The Lawrence Latino Diabetes Prevention Project (LLDPP) is a community-based translational research study which aims to reduce the risk of diabetes among Latinos who have a >/= 30% probability of developing diabetes in the next 7.5 years per a predictive equation. The project was conducted in Lawrence, Massachusetts, a predominantly Caribbean-origin urban Latino community. Individuals were identified primarily from a community health center's patient panel, screened for study eligibility, randomized to either a usual care or a lifestyle intervention condition, and followed for one year. Like the efficacious Diabetes Prevention Program (DPP), the LLDPP intervention targeted weight loss through dietary change and increased physical activity. However, unlike the DPP, the LLDPP intervention was less intensive, tailored to literacy needs and cultural preferences, and delivered in Spanish. The group format of the intervention (13 group sessions over 1 year) was complemented by 3 individual home visits and was implemented by individuals from the community with training and supervision by a clinical research nutritionist and a behavioral psychologist. Study measures included demographics, Stern predictive equation components (age, gender, ethnicity, fasting glucose, systolic blood pressure, HDL-cholesterol, body mass index, and family history of diabetes), glycosylated hemoglobin, dietary intake, physical activity, depressive symptoms, social support, quality of life, and medication use. Body weight was measured at baseline, 6-months, and one-year; all other measures were assessed at baseline and one-year. All surveys were orally administered in Spanish. A community-academic partnership enabled the successful recruitment, intervention, and assessment of Latinos at risk of diabetes with a one-year study retention rate of 93%. NCT00810290.", "This meta-analytic study synthesized findings from 108 independent data sets across 22 cultures to investigate whether the stigma internalization model (the internalization of experienced stigma and perceived stigma to self-stigma) is associated with well-being and recovery of people with mental illness. We also examined the moderating role of collectivism in the internalization process. Results of the meta-analytic structural equation modeling suggested that self-stigma is a significant mediator in the relationships between experienced stigma and perceived stigma with well-being and recovery variables (indirect effects = 0.02 to -0.16). Experienced and perceived stigma had significant direct effects on well-being and recovery variables (Bs = 0.07 to -0.21, p < 0.05), suggesting that both external (e.g., public stigma) and internal (i.e., self-stigma) influences of stigma work concurrently to affect recovery and well-being of people with mental illness. The results of the mixed effect three-level meta-analytic models showed that collectivism significantly moderated the relationship between experienced and perceived stigma with self-stigma (Bs = 0.06 to 0.11, p < 0.05). This implied that the more collectivistic a culture is, the stronger the correlation between experienced and perceived stigma with self-stigma. Implications to stigma reduction approaches were discussed." ]
Immunogenicity of Mayaro virus-like particles from insect cells and mammalian cells	Mayaro virus (MAYV) is a mosquito-transmitted alphavirus that causes often debilitating rheumatic disease in tropical Central and South America. There are currently no licensed vaccines or antiviral drugs available for MAYV disease. Here, we generated Mayaro virus-like particles (VLPs) using the scalable baculovirus-insect cell expression system. High-level secretion of MAYV VLPs in the culture fluid of Sf9 insect cells was achieved, and particles with a diameter of 64 to 70 nm were obtained after purification. We characterize a C57BL/6J adult wild-type mouse model of MAYV infection and disease and used this model to compare the immunogenicity of VLPs from insect cells with that of VLPs produced in mammalian cells. Mice received two intramuscular immunizations with 1 μg of nonadjuvanted MAYV VLPs. Potent neutralizing antibody responses were generated against the vaccine strain, BeH407, with comparable activity seen against a contemporary 2018 isolate from Brazil (BR-18), whereas neutralizing activity against chikungunya virus was marginal. Sequencing of BR-18 illustrated that this virus segregates with genotype D isolates, whereas MAYV BeH407 belongs to genotype L. The mammalian cell-derived VLPs induced higher mean neutralizing antibody titers than those produced in insect cells. Both VLP vaccines completely protected adult wild-type mice against viremia, myositis, tendonitis, and joint inflammation after MAYV challenge.	[ "Mayaro fever is an acute, self-limited, febrile, mosquito-borne viral disease manifested by fever, chills, headache, myalgias, and arthralgias. The virus belongs to the family Togaviridae and the genus Alphavirus. Five other mosquito-borne viruses have been described as causing a similar dengue-like illness. The virus was first isolated in 1954, and the first epidemics were described in 1955 in Brazil and Bolivia. Other cases have been reported in Suriname, Brazil, Peru, French Guiana, and Trinidad. Up to 10 to 15% of febrile illnesses in endemic areas have been attributed to Mayaro virus. The exact pathogenesis and pathophysiology among humans is unknown. Animal models have demonstrated necrosis of skeletal muscle, periosteum, perichondrial tissues, and evidence of meningitis and encephalitis. All previous cases of Mayaro fever describe a self-limited illness. No reports of recurrent symptoms exist in the literature. This report describes a case of recurrent arthralgias in a military service member presenting to the emergency department.", "Adaptation to mosquito vectors suited for transmission in urban settings is a major driver in the emergence of arboviruses. To better anticipate future emergence events, it is crucial to assess their potential to adapt to new vector hosts. In this work, we used two different experimental evolution approaches to study the adaptation process of an emerging alphavirus, Mayaro virus (MAYV), to Ae. aegypti, an urban mosquito vector of many other arboviruses. We identified E2-T179N as a key mutation increasing MAYV replication in insect cells and enhancing transmission after escaping the midgut of live Ae. aegypti. In contrast, this mutation decreased viral replication and binding in human fibroblasts, a primary cellular target of MAYV in humans. We also showed that MAYV E2-T179N generates reduced viremia and displays less severe tissue pathology in vivo in a mouse model. We found evidence in mouse fibroblasts that MAYV E2-T179N is less dependent on the Mxra8 receptor for replication than WT MAYV. Similarly, exogenous expression of human apolipoprotein receptor 2 and Mxra8 enhanced WT MAYV replication compared to MAYV E2-T179N. When this mutation was introduced in the closely related chikungunya virus, which has caused major outbreaks globally in the past two decades, we observed increased replication in both human and insect cells, suggesting E2 position 179 is an important determinant of alphavirus host-adaptation, although in a virus-specific manner. Collectively, these results indicate that adaptation at the T179 residue in MAYV E2 may result in increased vector competence-but coming at the cost of optimal replication in humans-and may represent a first step towards a future emergence event.", "A serologic survey for Mayaro virus (Alphavirus, Togaviridae) in 28 wild nonflying forest mammal species in French Guiana showed a prevalence ranging from 0% to 52% and increasing with age. Species active during the day and those who spent time in trees were significantly more infected, results consistent with transmission implicating diurnal mosquitoes and continuous infectious pressure.", "Mayaro virus (MAYV) is an arbovirus that circulates in Latin America and is emerging as a potential threat to public health. Infected individuals develop Mayaro fever, a severe inflammatory disease characterized by high fever, rash, arthralgia, myalgia and headache. The disease is often associated with a prolonged arthralgia mediated by a chronic inflammation that can last months. Although the immune response against other arboviruses, such as chikungunya virus (CHIKV), dengue virus (DENV) and Zika virus (ZIKV), has been extensively studied, little is known about the pathogenesis of MAYV infection. In this study, we established models of MAYV infection in macrophages and in mice and found that MAYV can replicate in bone marrow-derived macrophages and robustly induce expression of inflammasome proteins, such as NLRP3, ASC, AIM2, and Caspase-1 (CASP1). Infection performed in macrophages derived from Nlrp3-/-, Aim2-/-, Asc-/-and Casp1/11-/-mice indicate that the NLRP3, but not AIM2 inflammasome is essential for production of inflammatory cytokines, such as IL-1β. We also determined that MAYV triggers NLRP3 inflammasome activation by inducing reactive oxygen species (ROS) and potassium efflux. In vivo infections performed in inflammasome-deficient mice indicate that NLRP3 is involved with footpad swelling, inflammation and pain, establishing a role of the NLRP3 inflammasome in the MAYV pathogenesis. Accordingly, we detected higher levels of caspase1-p20, IL-1β and IL-18 in the serum of MAYV-infected patients as compared to healthy individuals, supporting the participation of the NLRP3-inflammasome during MAYV infection in humans.", "A cluster of protracted migratory polyarthritis involving four adult family members occurred in January 2000 after a brief overnight outing in a rural area of Venezuela. Laboratory testing demonstrated Mayaro virus as the cause of the cluster. These results documented the first human cases of Mayaro virus in Venezuela.", "Mayaro virus (MAYV), isolated for the first time in Trinidad and Tobago, has captured the attention of public health authorities worldwide following recent outbreaks in the Americas. It has a propensity to be exported outside its original geographical range, because of the vast distribution of its vectors. Moreover, most of the world population is immunologically naïve with respect to infection with MAYV which makes this virus a true threat. The recent invasion of several countries by Aedesalbopictus underscores the risk of potential urban transmission of MAYV in both tropical and temperate regions. In humans, the clinical manifestations of MAYV disease range from mild fever, rash, and joint pain to arthralgia. In the absence of a licensed vaccine and clinically proven therapeutics against Mayaro fever, prevention focuses mainly on household mosquito control. However, as demonstrated for other arboviruses, mosquito control is rather inefficient for outbreak management and alternative approaches to contain the spread of MAYV are therefore necessary. Despite its strong epidemic potential, little is currently known about MAYV. This review addresses various aspects of MAYV, including its epidemiology, vector biology, mode of transmission, and clinical complications, as well as the latest developments in MAYV diagnosis.", "Chikungunya virus (CHIKV), a mosquito-borne alphavirus of increasing public health significance, has caused large epidemics in Africa and the Indian Ocean basin; now it is spreading throughout the Americas. The primary vectors of CHIKV are Aedes (Ae.) aegypti and, after the introduction of a mutation in the E1 envelope protein gene, the highly anthropophilic and geographically widespread Ae. albopictus mosquito. We review here research efforts to characterize the viral genetic basis of mosquito-vector interactions, the use of RNA interference and other strategies for the control of CHIKV in mosquitoes, and the potentiation of CHIKV infection by mosquito saliva. Over the past decade, CHIKV has emerged on a truly global scale. Since 2013, CHIKV transmission has been reported throughout the Caribbean region, in North America, and in Central and South American countries, including Brazil, Columbia, Costa Rica, El Salvador, French Guiana, Guatemala, Guyana, Nicaragua, Panama, Suriname, and Venezuela. Closing the gaps in our knowledge of driving factors behind the rapid geographic expansion of CHIKV should be considered a research priority. The abundance of multiple primate species in many of these countries, together with species of mosquito that have never been exposed to CHIKV, may provide opportunities for this highly adaptable virus to establish sylvatic cycles that to date have not been seen outside of Africa. The short-term and long-term ecological consequences of such transmission cycles, including the impact on wildlife and people living in these areas, are completely unknown.", "Mayaro virus (MAYV), causative agent of Mayaro Fever, is an arbovirus transmitted by Haemagogus mosquitoes. Despite recent attention due to the identification of several cases in South and Central America and the Caribbean, limited information on MAYV evolution and epidemiology exists and represents a barrier to prevention of further spread. We present a thorough spatiotemporal evolutionary study of MAYV full-genome sequences collected over the last sixty years within South America and Haiti, revealing recent recombination events and adaptation to a broad host and vector range, including Aedes mosquito species. We employed a Bayesian phylogeography approach to characterize the emergence of recombinants in Brazil and Haiti and report evidence in favor of the putative role of human mobility in facilitating recombination among MAYV strains from geographically distinct regions. Spatiotemporal characteristics of recombination events and the emergence of this previously neglected virus in Haiti, a known hub for pathogen spread to the Americas, warrants close monitoring of MAYV infection in the immediate future.", "Emergence of arboviruses is a rising problem in several areas in the world. Here we report a case of Mayaro virus infection that was diagnosed in a French citizen presenting a dengue-like syndrome with prolonged arthralgia following a travel in French Guiana. Diagnosis was based on serological testing, a newly developed specific RT-PCR and sequencing. The real incidence of this viral infection among travelers is poorly known but this case is the first reported in a European area where Aedes albopictus mosquitoes are established, which underscores the necessity to determine the vector competence of the European strain of this mosquito species for Mayaro virus.", "A laboratory worker developed clinical signs of infection with Mayaro virus (Togaviridae), an arbovirus of South and Central America, 6 days after preparation of Mayaro viral antigen and 10 days after a trip to a rain forest. There was no evidence of skin lesions during the antigen preparation, and level 3 containment safety measures were followed. Therefore, molecular characterization of the virus was undertaken to identify the source of infection. RT-PCR and DNA sequence comparisons proved the infection was with the laboratory strain. Airborne Mayaro virus contamination is thus a hazard to laboratory personnel." ]
Detection of Structural Variations in Humans	Many studies have shown that structural variations (SVs) strongly impact human disease. As a common type of SV, insertions are usually associated with genetic diseases. Therefore, accurately detecting insertions is of great significance. Although many methods for detecting insertions have been proposed, these methods often generate some errors and miss some variants. Hence, accurately detecting insertions remains a challenging task.	[ "The fundamental challenge of multi-sample structural variant (SV) analysis such as merging and benchmarking is identifying when two SVs are the same. Common approaches for comparing SVs were developed alongside technologies which produce ill-defined boundaries. As SV detection becomes more exact, algorithms to preserve this refined signal are needed. Here, we present Truvari-an SV comparison, annotation, and analysis toolkit-and demonstrate the effect of SV comparison choices by building population-level VCFs from 36 haplotype-resolved long-read assemblies. We observe over-merging from other SV merging approaches which cause up to a 2.2× inflation of allele frequency, relative to Truvari.", "During the last quarter of the twentieth century, our knowledge about human genetic variation was limited mainly to the heterochromatin polymorphisms, large enough to be visible in the light microscope, and the single nucleotide polymorphisms (SNPs) identified by traditional PCR-based DNA sequencing. In the past five years, the rapid development and expanded use of microarray technologies, including oligonucleotide array comparative genomic hybridization and SNP genotyping arrays, as well as next-generation sequencing with \"paired-end\" methods, has enabled a whole-genome analysis with essentially unlimited resolution. The discovery of submicroscopic copy-number variations (CNVs) present in our genomes has changed dramatically our perspective on DNA structural variation and disease. It is now thought that CNVs encompass more total nucleotides and arise more frequently than SNPs. CNVs, to a larger extent than SNPs, have been shown to be responsible for human evolution, genetic diversity between individuals, and a rapidly increasing number of traits or susceptibility to traits; such conditions have been referred to as genomic disorders. In addition to well-known sporadic chromosomal microdeletion syndromes and Mendelian diseases, many common complex traits including autism and schizophrenia can result from CNVs. Both recombination- and replication-based mechanisms for CNV formation have been described.", "A high-confidence, comprehensive human variant set is critical in assessing accuracy of sequencing algorithms, which are crucial in precision medicine based on high-throughput sequencing. Although recent works have attempted to provide such a resource, they still do not encompass all major types of variants including structural variants (SVs). Thus, we leveraged the massive high-quality Sanger sequences from the HuRef genome to construct by far the most comprehensive gold set of a single individual, which was cross validated with deep Illumina sequencing, population datasets, and well-established algorithms. It was a necessary effort to completely reanalyze the HuRef genome as its previously published variants were mostly reported five years ago, suffering from compatibility, organization, and accuracy issues that prevent their direct use in benchmarking. Our extensive analysis and validation resulted in a gold set with high specificity and sensitivity. In contrast to the current gold sets of the NA12878 or HS1011 genomes, our gold set is the first that includes small variants, deletion SVs and insertion SVs up to a hundred thousand base-pairs. We demonstrate the utility of our HuRef gold set to benchmark several published SV detection tools.", "New technologies and analysis methods are enabling genomic structural variants (SVs) to be detected with ever-increasing accuracy, resolution and comprehensiveness. To help translate these methods to routine research and clinical practice, we developed a sequence-resolved benchmark set for identification of both false-negative and false-positive germline large insertions and deletions. To create this benchmark for a broadly consented son in a Personal Genome Project trio with broadly available cells and DNA, the Genome in a Bottle Consortium integrated 19 sequence-resolved variant calling methods from diverse technologies. The final benchmark set contains 12,745 isolated, sequence-resolved insertion (7,281) and deletion (5,464) calls ≥50 base pairs (bp). The Tier 1 benchmark regions, for which any extra calls are putative false positives, cover 2.51 Gbp and 5,262 insertions and 4,095 deletions supported by ≥1 diploid assembly. We demonstrate that the benchmark set reliably identifies false negatives and false positives in high-quality SV callsets from short-, linked- and long-read sequencing and optical mapping.", "The 1000 Genomes Project set out to provide a comprehensive description of common human genetic variation by applying whole-genome sequencing to a diverse set of individuals from multiple populations. Here we report completion of the project, having reconstructed the genomes of 2,504 individuals from 26 populations using a combination of low-coverage whole-genome sequencing, deep exome sequencing, and dense microarray genotyping. We characterized a broad spectrum of genetic variation, in total over 88 million variants (84.7 million single nucleotide polymorphisms (SNPs), 3.6 million short insertions/deletions (indels), and 60,000 structural variants), all phased onto high-quality haplotypes. This resource includes >99% of SNP variants with a frequency of >1% for a variety of ancestries. We describe the distribution of genetic variation across the global sample, and discuss the implications for common disease studies.", "The completion of the human genome reference sequence ushered in a new era for the study and discovery of human transposable elements. It now is undeniable that transposable elements, historically dismissed as junk DNA, have had an instrumental role in sculpting the structure and function of our genomes. In particular, long interspersed element-1 (LINE-1 or L1) and short interspersed elements (SINEs) continue to affect our genome, and their movement can lead to sporadic cases of disease. Here, we briefly review the types of transposable elements present in the human genome and their mechanisms of mobility. We next highlight how advances in DNA sequencing and genomic technologies have enabled the discovery of novel retrotransposons in individual genomes. Finally, we discuss how L1-mediated retrotransposition events impact human genomes.", "Polar bears (PBs) are superbly adapted to the extreme Arctic environment and have become emblematic of the threat to biodiversity from global climate change. Their divergence from the lower-latitude brown bear provides a textbook example of rapid evolution of distinct phenotypes. However, limited mitochondrial and nuclear DNA evidence conflicts in the timing of PB origin as well as placement of the species within versus sister to the brown bear lineage. We gathered extensive genomic sequence data from contemporary polar, brown, and American black bear samples, in addition to a 130,000- to 110,000-y old PB, to examine this problem from a genome-wide perspective. Nuclear DNA markers reflect a species tree consistent with expectation, showing polar and brown bears to be sister species. However, for the enigmatic brown bears native to Alaska's Alexander Archipelago, we estimate that not only their mitochondrial genome, but also 5-10% of their nuclear genome, is most closely related to PBs, indicating ancient admixture between the two species. Explicit admixture analyses are consistent with ancient splits among PBs, brown bears and black bears that were later followed by occasional admixture. We also provide paleodemographic estimates that suggest bear evolution has tracked key climate events, and that PB in particular experienced a prolonged and dramatic decline in its effective population size during the last ca. 500,000 years. We demonstrate that brown bears and PBs have had sufficiently independent evolutionary histories over the last 4-5 million years to leave imprints in the PB nuclear genome that likely are associated with ecological adaptation to the Arctic environment." ]
Genome-wide association studies of resting-state networks with resting-state networks	Functional connectivity within resting-state networks (RSN-FC) is vital for cognitive functioning. RSN-FC is heritable and partially translates to the anatomic architecture of white matter, but the genetic component of structural connections of RSNs (RSN-SC) and their potential genetic overlap with RSN-FC remain unknown. Here, we perform genome-wide association studies (	[ "Several recent reports in large, independent samples have demonstrated the influence of motion artifact on resting-state functional connectivity MRI (rsfc-MRI). Standard rsfc-MRI preprocessing typically includes regression of confounding signals and band-pass filtering. However, substantial heterogeneity exists in how these techniques are implemented across studies, and no prior study has examined the effect of differing approaches for the control of motion-induced artifacts. To better understand how in-scanner head motion affects rsfc-MRI data, we describe the spatial, temporal, and spectral characteristics of motion artifacts in a sample of 348 adolescents. Analyses utilize a novel approach for describing head motion on a voxelwise basis. Next, we systematically evaluate the efficacy of a range of confound regression and filtering techniques for the control of motion-induced artifacts. Results reveal that the effectiveness of preprocessing procedures on the control of motion is heterogeneous, and that improved preprocessing provides a substantial benefit beyond typical procedures. These results demonstrate that the effect of motion on rsfc-MRI can be substantially attenuated through improved preprocessing procedures, but not completely removed.", "Genome-wide association studies are carried out to identify unknown genes for a complex trait. Polymorphisms showing the most statistically significant associations are reported and followed up in subsequent confirmatory studies. In addition to the test of association, the statistical analysis provides point estimates of the relationship between the genotype and phenotype at each polymorphism, typically an odds ratio in case-control association studies. The statistical significance of the test and the estimator of the odds ratio are completely correlated. Selecting the most extreme statistics is equivalent to selecting the most extreme odds ratios. The value of the estimator, given the value of the statistical significance depends on the standard error of the estimator and the power of the study. This report shows that when power is low, estimates of the odds ratio from a genome-wide association study, or any large-scale association study, will be upwardly biased. Genome-wide association studies are often underpowered given the low alpha levels required to declare statistical significance and the small individual genetic effects known to characterize complex traits. Factors such as low allele frequency, inadequate sample size and weak genetic effects contribute to large standard errors in the odds ratio estimates, low power and upwardly biased odds ratios. Studies that have high power to detect an association with the true odds ratio will have little or no bias, regardless of the statistical significance threshold. The results have implications for the interpretation of genome-wide association analysis and the planning of subsequent confirmatory stages.", "Microscopic features (that is, microstructure) of axons affect neural circuit activity through characteristics such as conduction speed. To what extent axonal microstructure in white matter relates to functional connectivity (synchrony) between brain regions is largely unknown. Using MRI data in 11,354 subjects, we constructed multivariate models that predict functional connectivity of pairs of brain regions from the microstructural signature of white matter pathways that connect them. Microstructure-derived models provided predictions of functional connectivity that explained 3.5% of cross-subject variance on average (ranging from 1-13%, or r = 0.1-0.36) and reached statistical significance in 90% of the brain regions considered. The microstructure-function relationships were associated with genetic variants, co-located with genes DAAM1 and LPAR1, that have previously been linked to neural development. Our results demonstrate that variation in white matter microstructure predicts a fraction of functional connectivity across individuals, and that this relationship is underpinned by genetic variability in certain brain areas.", "Multiple studies have confirmed the contribution of rare de novo copy number variations to the risk for autism spectrum disorders. But whereas de novo single nucleotide variants have been identified in affected individuals, their contribution to risk has yet to be clarified. Specifically, the frequency and distribution of these mutations have not been well characterized in matched unaffected controls, and such data are vital to the interpretation of de novo coding mutations observed in probands. Here we show, using whole-exome sequencing of 928 individuals, including 200 phenotypically discordant sibling pairs, that highly disruptive (nonsense and splice-site) de novo mutations in brain-expressed genes are associated with autism spectrum disorders and carry large effects. On the basis of mutation rates in unaffected individuals, we demonstrate that multiple independent de novo single nucleotide variants in the same gene among unrelated probands reliably identifies risk alleles, providing a clear path forward for gene discovery. Among a total of 279 identified de novo coding mutations, there is a single instance in probands, and none in siblings, in which two independent nonsense variants disrupt the same gene, SCN2A (sodium channel, voltage-gated, type II, α subunit), a result that is highly unlikely by chance.", "Structural neuroimaging studies have provided some of the most consistent evidence for brain abnormalities in schizophrenia. Since the initial computed tomography study by Johnstone and co-workers, which reported lateral ventricular enlargement in schizophrenia, advances in brain imaging technology have enabled further and more refined characterization of abnormal brain structure in schizophrenia in vivo. This selective review discusses the major issues and findings in structural neuroimaging studies of schizophrenia. Among these are evidence for generalized and regional brain volume abnormalities, the specificity of anatomic findings to schizophrenia and to men versus women with schizophrenia, the contribution of genetic influences, and the timing of neuroanatomic pathology in schizophrenia. The second section reviews new approaches for examining brain structure in schizophrenia and their applications to studies on the pathophysiology of schizophrenia.", "Schizophrenia is a psychotic disorder characterized by functional dysconnectivity or abnormal integration between distant brain regions. Recent functional imaging studies have implicated large-scale thalamo-cortical connectivity as being disrupted in patients. However, observed connectivity differences in schizophrenia have been inconsistent between studies, with reports of hyperconnectivity and hypoconnectivity between the same brain regions. Using resting state eyes-closed functional imaging and independent component analysis on a multi-site data that included 151 schizophrenia patients and 163 age- and gender matched healthy controls, we decomposed the functional brain data into 100 components and identified 47 as functionally relevant intrinsic connectivity networks. We subsequently evaluated group differences in functional network connectivity, both in a static sense, computed as the pairwise Pearson correlations between the full network time courses (5.4 minutes in length), and a dynamic sense, computed using sliding windows (44 s in length) and k-means clustering to characterize five discrete functional connectivity states. Static connectivity analysis revealed that compared to healthy controls, patients show significantly stronger connectivity, i.e., hyperconnectivity, between the thalamus and sensory networks (auditory, motor and visual), as well as reduced connectivity (hypoconnectivity) between sensory networks from all modalities. Dynamic analysis suggests that (1), on average, schizophrenia patients spend much less time than healthy controls in states typified by strong, large-scale connectivity, and (2), that abnormal connectivity patterns are more pronounced during these connectivity states. In particular, states exhibiting cortical-subcortical antagonism (anti-correlations) and strong positive connectivity between sensory networks are those that show the group differences of thalamic hyperconnectivity and sensory hypoconnectivity. Group differences are weak or absent during other connectivity states. Dynamic analysis also revealed hypoconnectivity between the putamen and sensory networks during the same states of thalamic hyperconnectivity; notably, this finding cannot be observed in the static connectivity analysis. Finally, in post-hoc analyses we observed that the relationships between sub-cortical low frequency power and connectivity with sensory networks is altered in patients, suggesting different functional interactions between sub-cortical nuclei and sensorimotor cortex during specific connectivity states. While important differences between patients with schizophrenia and healthy controls have been identified, one should interpret the results with caution given the history of medication in patients. Taken together, our results support and expand current knowledge regarding dysconnectivity in schizophrenia, and strongly advocate the use of dynamic analyses to better account for and understand functional connectivity differences.", "This protocol describes how to appropriately design a genetic association case-control study, either focusing on a candidate gene (CG) or region or implementing a genome-wide approach. The steps described involve: (i) defining the case phenotype in adequate detail; (ii) checking the heritability of the disease in question; (iii) considering whether a population-based study is the appropriate design for the research question; (iv) the appropriate selection of controls; (v) sample size calculations and (vi) giving due consideration to whether it is a de novo or replication study. General guidelines are given, as well as specific examples of a CG and a genome-wide association study into type 2 diabetes. Software and websites used in this protocol include the International HapMap Consortium website, Genetic Power Calculator, CaT, and SNPSpD. Running each of the programs takes only a few seconds; the rate-limiting steps involve thinking through the designs and parameters in the disease models.", "In modern whole-genome scans, the use of stringent thresholds to control the genome-wide testing error distorts the estimation process, producing estimated effect sizes that may be on average far greater in magnitude than the true effect sizes. We introduce a method, based on the estimate of genetic effect and its standard error as reported by standard statistical software, to correct for this bias in case-control association studies. Our approach is widely applicable, is far easier to implement than competing approaches, and may often be applied to published studies without access to the original data. We evaluate the performance of our approach via extensive simulations for a range of genetic models, minor allele frequencies, and genetic effect sizes. Compared to the naive estimation procedure, our approach reduces the bias and the mean squared error, especially for modest effect sizes. We also develop a principled method to construct confidence intervals for the genetic effect that acknowledges the conditioning on statistical significance. Our approach is described in the specific context of odds ratios and logistic modeling but is more widely applicable. Application to recently published data sets demonstrates the relevance of our approach to modern genome scans.", "We examined the status of the neural network mediating the default mode of brain function, which typically exhibits greater activation during rest than during task, in patients in the early phase of schizophrenia and in young first-degree relatives of persons with schizophrenia. During functional MRI, patients, relatives, and controls alternated between rest and performance of working memory (WM) tasks. As expected, controls exhibited task-related suppression of activation in the default network, including medial prefrontal cortex (MPFC) and posterior cingulate cortex/precuneus. Patients and relatives exhibited significantly reduced task-related suppression in MPFC, and these reductions remained after controlling for performance. Increased task-related MPFC suppression correlated with better WM performance in patients and relatives and with less psychopathology in all 3 groups. For WM task performance, patients and relatives had greater activation in right dorsolateral prefrontal cortex (DLPFC) than controls. During rest and task, patients and relatives exhibited abnormally high functional connectivity within the default network. The magnitudes of default network connectivity during rest and task correlated with psychopathology in the patients. Further, during both rest and task, patients exhibited reduced anticorrelations between MPFC and DLPFC, a region that was hyperactivated by patients and relatives during WM performance. Among patients, the magnitude of MPFC task suppression negatively correlated with default connectivity, suggesting an association between the hyperactivation and hyperconnectivity in schizophrenia. Hyperactivation (reduced task-related suppression) of default regions and hyperconnectivity of the default network may contribute to disturbances of thought in schizophrenia and risk for the illness.", "Current methods for annotating and interpreting human genetic variation tend to exploit a single information type (for example, conservation) and/or are restricted in scope (for example, to missense changes). Here we describe Combined Annotation-Dependent Depletion (CADD), a method for objectively integrating many diverse annotations into a single measure (C score) for each variant. We implement CADD as a support vector machine trained to differentiate 14.7 million high-frequency human-derived alleles from 14.7 million simulated variants. We precompute C scores for all 8.6 billion possible human single-nucleotide variants and enable scoring of short insertions-deletions. C scores correlate with allelic diversity, annotations of functionality, pathogenicity, disease severity, experimentally measured regulatory effects and complex trait associations, and they highly rank known pathogenic variants within individual genomes. The ability of CADD to prioritize functional, deleterious and pathogenic variants across many functional categories, effect sizes and genetic architectures is unmatched by any current single-annotation method." ]
Collisionally dissociating intact protein ions in a trapped ion mobility spectrometry device	Mass spectrometry (MS)-based proteomics workflows of intact protein ions have increasingly been utilized to study biological systems. These workflows, however, frequently result in convoluted and difficult to analyze mass spectra. Ion mobility spectrometry (IMS) is a promising tool to overcome these limitations by separating ions by their mass- and size-to-charge ratios. In this work, we further characterize a newly developed method to collisionally dissociate intact protein ions in a trapped ion mobility spectrometry (TIMS) device. Dissociation occurs prior to ion mobility separation and thus, all product ions are distributed throughout the mobility dimension, enabling facile assignment of near isobaric product ions. We demonstrate that collisional activation within a TIMS device is capable of dissociating protein ions up to 66 kDa. We also demonstrate that the ion population size within the TIMS device significantly influences the efficiency of fragmentation. Lastly, we compare CIDtims to the other modes of collisional activation available on the Bruker timsTOF and demonstrate that the mobility resolution in CIDtims enables the annotation of overlapping fragment ions and improves sequence coverage.	[ "The presence of disulfide linkages in multiply charged polypeptide ions tends to inhibit the formation of structurally informative product ions under conventional quadrupole ion trap collisional activation conditions. In particular, fragmentation that requires two cleavages (i.e., cleavage of a disulfide linkage and a peptide linkage) is strongly suppressed. Reduction of the disulfide linkage(s) by use of dithiothreitol yields parent ions upon electrospray without this complication. Far richer structural information is revealed by ion trap collisional activation of the disulfide-reduced species than from the native species. These observations are illustrated with doubly protonated native and reduced somatosin, the [M + 5H](5+) ion of native bovine insulin and the [M + 4H](4+) and [M + 3H](3+) ions of the B-chain of bovine insulin produced by reduction of the disulfide linkages in insulin, and the [M + 11H](11+) ion of native chicken lysozyme and the [M + 11H](11+) and [M + 14H](14+) ions of reduced lysozyme. In each case, the product ions produced by ion trap collisional activation were subjected to ion/ion proton transfer reactions to facilitate interpretation of the product ion spectra. These studies clearly suggest that the identification of polypeptides with one or more disulfide linkages via application of ion trap collisional activation to the multiply charged parent ions formed directly by electrospray could be problematic. Means for cleaving the disulfide linkage, such as reduction by dithiothreitol prior to electrospray, are therefore desirable in these cases.", "The effect of the basic residue on the energetics, dynamics, and mechanisms of backbone fragmentation of protonated peptides was investigated. Time-resolved and collision energy-resolved surface-induced dissociation (SID) of singly protonated peptides with the N-terminal arginine residue and their analogues, in which arginine is replaced with less basic lysine and histidine residues, was examined using a specially configured Fourier transform ion cyclotron resonance mass spectrometer (FTICR-MS). SID experiments demonstrated different kinetics of formation of several primary product ions of peptides with and without arginine residue. The energetics and dynamics of these pathways were determined from Rice-Ramsperger-Kassel-Marcus (RRKM) modeling of the experimental data. Comparison between the kinetics and energetics of fragmentation of arginine-containing peptides and the corresponding methyl ester derivatives provides important information on the effect of dissociation pathways involving salt bridge (SB) intermediates on the observed fragmentation behavior. Because pathways involving SB intermediates are characterized by low threshold energies, they efficiently compete with classical oxazolone and imine/enol pathways of arginine-containing peptides on a long time scale of the FTICR instrument. In contrast, fragmentation of histidine- and lysine-containing peptides is largely determined by canonical pathways. Because SB pathways are characterized by negative activation entropies, fragmentation of arginine-containing peptides is kinetically hindered and observed at higher collision energies as compared to their lysine- and histidine-containing analogues.", "It has previously been reported that disulfide and backbone bonds of native intact proteins can be concurrently cleaved using electrospray ionization (ESI) and collision-induced dissociation (CID) tandem mass spectrometry (MS/MS). However, the cleavages of disulfide bonds result in different cysteine modifications in product ions, making it difficult to identify the disulfide-bonded proteins via database search. To solve this identification problem, we have developed a pseudo MS(3) approach by combining nozzle-skimmer dissociation (NSD) and CID on a quadrupole time-of-flight (Q-TOF) mass spectrometer using chicken lysozyme as a model. Although many of the product ions were similar to those typically seen in MS/MS spectra of enzymatically derived peptides, additional uncommon product ions were detected including c(i-1) ions (the i(th) residue being aspartic acid, arginine, lysine and dehydroalanine) as well as those from a scrambled sequence. The formation of these uncommon types of product ions, likely caused by the lack of mobile protons, were proposed to involve bond rearrangements via a six-membered ring transition state and/or salt bridge(s). A search of 20 pseudo MS(3) spectra against the Gallus gallus (chicken) database using Batch-Tag, a program originally designed for bottom up MS/MS analysis, identified chicken lysozyme as the only hit with the expectation values less than 0.02 for 12 of the spectra. The pseudo MS(3) approach may help to identify disulfide-bonded proteins and determine the associated post-translational modifications (PTMs); the confidence in the identification may be improved by incorporating the fragmentation characteristics into currently available search programs.", "There is now strong evidence for the existence of macrocyclic isomers of b(n)(+) ions, the formation and subsequent opening of which can lead to loss of sequence information from protonated peptides in multiple-stage tandem mass spectrometry experiments. In this study, the fragmentation patterns of protonated YARFLG and permuted isomers of the model peptide were investigated by collision-induced dissociation. Of interest was the potential influence of the arginine residue, and its position in the peptide sequence, on formation of the presumed macrocyclic b(5) ion isomer and potential loss of sequence information. We find that regardless of the sequence position (either internal or at the N- or C-terminus), only direct sequence ions or ions directly related to fragmentation of the arginine side chain are observed.", "Shotgun proteomics entails the identification of as many peptides as possible from complex mixtures. Here we investigate how many peptides are detectable by high resolution MS in standard LC runs of cell lysate and how many of them are accessible to data-dependent MS/MS. Isotope clusters were determined by MaxQuant and stringently filtered for charge states and retention times typical of peptides. This resulted in more than 100,000 likely peptide features, of which only about 16% had been targeted for MS/MS. Three instrumental attributes determine the proportion of additional peptides that can be identified: sequencing speed, sensitivity, and precursor ion isolation. In our data, an MS/MS scan rate of 25/s would be necessary to target all peptide features, but this drops to less than 17/s for reasonably abundant peptides. Sensitivity is a greater challenge, with many peptide features requiring long MS/MS injection times (>250 ms). The greatest limitation, however, is the generally low proportion of the target peptide ion intensity in the MS/MS selection window (the \"precursor ion fraction\" or PIF). Median PIF is only 0.14, making the peptides difficult to identify by standard MS/MS methods. Our results aid in developing strategies to further increase coverage in shotgun proteomics." ]
Microbiota-gut-brain axis in HIV-associated neurocognitive disorders	Although the advent of combination anti-retroviral therapy (cART) has transformed HIV into a manageable chronic disease, an estimated 30-50% of people living with HIV (PLWH) exhibit cognitive and motor deficits collectively known as HIV-associated neurocognitive disorders (HAND). A key driver of HAND neuropathology is chronic neuroinflammation, where proinflammatory mediators produced by activated microglia and macrophages are thought to inflict neuronal injury and loss. Moreover, the dysregulation of the microbiota-gut-brain axis (MGBA) in PLWH, consequent to gastrointestinal dysfunction and dysbiosis, can lead to neuroinflammation and persistent cognitive impairment, which underscores the need for new interventions.	[ "Enteric glia activation has been reported to amplify intestinal inflammation via the enteroglial-specific S100B protein. This neurotrophin promotes macrophage recruitment in the mucosa, amplify colonic inflammation and interacts with toll-like receptors (TLR). Molecules inhibiting S100B-driven enteric activation might mitigate the course of ulcerative colitis (UC). This study aims to investigate the effects of palmitoylethanolammide (PEA), a drug able to counteract astroglial activation in the central nervous system, on intestinal inflammation, in humans and mice. Mouse models of dextran sodium sulphate (DSS)-induced colitis, colonic biopsies deriving from UC patients and primary cultures of mouse and human enteric glial cells (EGC), have been used to assess the effects of PEA, alone or in the presence of specific PPARα or PPARγ antagonists, on: macroscopic signs of UC (DAI score, colon length, spleen weight, macrophages/neutrophils infiltration); the expression and release of proinflammatory markers typical of UC; TLR pathway in EGCs. PEA treatment improves all macroscopic signs of UC and decreases the expression and release of all the proinflammatory markers tested. PEA anti-inflammatory effects are mediated by the selective targeting of the S100B/TLR4 axis on ECG, causing a downstream inhibition of nuclear factor kappa B (NF-kB)-dependent inflammation. Antagonists at PPARα, but not PPARγ, abolished PEA effects, in mice and in humans. Because of its lack of toxicity, its ability in reducing inflammation and its selective PPARα action, PEA might be an innovative molecule to broaden pharmacological strategies against UC.", "Perivascular macrophages are located in the perivascular space of cerebral microvessels and thus uniquely situated at the intersection between the brain parenchyma and blood. Connections between the nervous and immune systems are mediated in part through these cells that are ideally located to sense perturbations in the periphery and turnover by cells entering the central nervous system (CNS) from the circulation. It has become clear that unique subsets of brain macrophages exist in normal and SIV- or HIV-infected brains, and perivascular macrophages and similar cells in the meninges and choroid plexus play a central role in lentiviral neuropathogenesis. Common to all these cell populations is their likely replacement within the CNS by monocytes. Studies of SIV-infected non-human primates and HIV-infected humans underscore the importance of virus-infected and activated monocytes, which traffic to the CNS from blood to become perivascular macrophages, potentially drive the blood-brain barrier damage and cause neuronal injury. This review summarizes what we know about SIV- and HIV-induced neuropathogenesis focusing on brain perivascular macrophages and their precursors in blood that may mediate HIV CNS infection and injury.", "Despite effective antiretroviral therapy (ART), people living with HIV (PLWH) still present persistent chronic immune activation and inflammation. This condition is the result of several factors including thymic dysfunction, persistent antigen stimulation due to low residual viremia, microbial translocation and dysbiosis, caused by the disruption of the gut mucosa, co-infections, and cumulative ART toxicity. All of these factors can create a vicious cycle that does not allow the full control of immune activation and inflammation, leading to an increased risk of developing non-AIDS co-morbidities such as metabolic syndrome and cardiovascular diseases. This review aims to provide an overview of the most recent data about HIV-associated inflammation and chronic immune exhaustion in PLWH under effective ART. Furthermore, we discuss new therapy approaches that are currently being tested to reduce the risk of developing inflammation, ART toxicity, and non-AIDS co-morbidities.", "Persistent gastrointestinal inflammation, a hallmark of progressive HIV/SIV infection, causes disruption of the gastrointestinal epithelial barrier, microbial translocation, and generalized immune activation/inflammation driving AIDS progression. Apart from protein regulators, recent studies strongly suggest critical roles for microRNAs (miRNAs) in regulating and managing certain aspects of the inflammatory process. To examine their immunoregulatory role, we profiled miRNA expression in the colon from 12 chronic SIV-infected and 4 control macaques. After applying multiple comparisons correction, 10 (3 upregulated and 7 downregulated) miRNAs showed differential expression. Most notably, miR-34a showed significant upregulation in both epithelial and lamina propria leukocyte (LPL) compartments. Intense γH2A.X expression in colonic epithelium and LPLs confirmed the contribution of DNA damage response in driving miR-34a upregulation. SIRT1 mRNA and protein decreased significantly in both colonic epithelium and LPLs. Luciferase reporter assays validated rhesus macaque SIRT1 as a direct miR-34a target. Decreased SIRT1 expression was associated with constitutively enhanced expression of the transcriptionally active form of the p65 (acetylated on lysine 310) subunit of NF-κB exclusively in the LPL compartment. The intensity and number of acetylated p65(+) cells was markedly elevated in LPLs of chronically SIV-infected macaques compared with uninfected controls and localized to increased numbers of IgA(+) and IgG(+) plasma cells. These findings provide new insights into the potential role of the miR-34a-SIRT1-p65 axis in causing hyperactivation of the intestinal B cell system. Our results point to a possible mechanism where the normal immunosuppressive function of SIRT1 is inhibited by elevated miR-34a expression resulting in constitutive activation of acetylated p65 (lysine 310).", "Chronic stress is associated with visceral hyperalgesia in functional gastrointestinal disorders. We investigated whether corticosterone plays a role in chronic psychological stress-induced visceral hyperalgesia. Male rats were subjected to 1-hour water avoidance (WA) stress or subcutaneous corticosterone injection daily for 10 consecutive days in the presence or absence of corticoid-receptor antagonist RU-486 and cannabinoid-receptor agonist WIN55,212-2. The visceromotor response to colorectal distension was measured. Receptor protein levels were measured and whole-cell patch-clamp recordings were used to assess transient receptor potential vanilloid type 1 (TRPV1) currents in L6-S2 dorsal root ganglion (DRG) neurons. Mass spectrometry was used to measure endocannabinoid anandamide content. Chronic WA stress was associated with visceral hyperalgesia in response to colorectal distension, increased stool output and reciprocal changes in cannabinoid receptor 1 (CB1) (decreased) and TRPV1 (increased) receptor expression and function. Treatment of WA stressed rats with RU-486 prevented these changes. Control rats treated with serial injections of corticosterone in situ showed a significant increase in serum corticosterone associated with visceral hyperalgesia, enhanced anandamide content, increased TRPV1, and decreased CB1 receptor protein levels, which were prevented by co-treatment with RU-486. Exposure of isolated control L6-S2 DRGs in vitro to corticosterone reproduced the changes in CB1 and TRPV1 receptors observed in situ, which was prevented by co-treatment with RU-486 or WIN55,212-2. These results support a novel role for corticosterone to modulate CB1 and TRPV1-receptor pathways in L6-S2 DRGs in the chronic WA stressed rat, which contributes to visceral hyperalgesia observed in this model.", "In 1964, the psychoactive ingredient of Cannabis sativa, Δ(9)-tetrahydrocannabinol (THC), was isolated. Nearly 30 years later the endogenous counterparts of THC, collectively termed endocannabinoids (eCBs), were discovered: N-arachidonoylethanolamine (anandamide) (AEA) in 1992 and 2-arachidonoylglycerol (2-AG) in 1995. Since then, considerable research has shed light on the impact of eCBs on human health and disease, identifying an ensemble of proteins that bind, synthesize, and degrade them and that together form the eCB system (ECS). eCBs control basic biological processes including cell choice between survival and death and progenitor/stem cell proliferation and differentiation. Unsurprisingly, in the past two decades eCBs have been recognized as key mediators of several aspects of human pathophysiology and thus have emerged to be among the most widespread and versatile signaling molecules ever discovered. Here some of the pioneers of this research field review the state of the art of critical eCB functions in peripheral organs. Our community effort is aimed at establishing consensus views on the relevance of the peripheral ECS for human health and disease pathogenesis, as well as highlighting emerging challenges and therapeutic hopes.", "Activation of cannabinoid receptors (CBs) by endocannabinoids impacts on a number of gastrointestinal functions. Recent data indicate that CB1 agonists improve 2,4-dinitrobenzene sulfonic acid-induced colitis in mice, thus suggesting a role for the endocannabinoid agonist anandamide (AEA) in protecting the gut against inflammation. We here examined the gut endocannabinoid system in inflammatory bowel disease (IBD) patients, and investigated the ex vivo and in vitro effects of the non-hydrolysable AEA analog methanandamide (MAEA) on the mucosal proinflammatory response. The content of AEA, but not of 2-arachidonoyl-glycerol and N-palmitoylethanolamine, was significantly lower in inflamed than uninflamed IBD mucosa, and this was paralleled by lower activity of the AEA-synthesizing enzyme N-acyl-phosphatidylethanolamine-specific phospholipase D and higher activity of the AEA-degrading enzyme fatty acid amide hydrolase. MAEA significantly downregulated interferon-γ and tumor necrosis factor-α secretion by both organ culture biopsies and lamina propria mononuclear cells. Although these results are promising, further studies are needed to determine the role of cannabinoid pathways in gut inflammation.", "Neurodegeneration leading to Parkinson's disease (PD) and Alzheimer's disease (AD) has become a major health burden globally. Current treatments mainly target controlling symptoms and there are no therapeutics available in clinical practice to preventing the neurodegeneration or inducing neuronal repairing. Thus, the demand of novel research for the two disorders is imperative. This literature review aims to provide a collection of published work on PD and AD and current uses of endocannabinoid system (ECS) as a potential drug target for neurodegeneration. PD is frequently treated with L-DOPA and deep brain stimulation. Recent gene modification and remodelling techniques, such as CRISPR through human embryonic stem cells and induced pluripotent stem cells, have shown promising strategy for personalised medicine. AD characterised by extracellular deposits of amyloid β-senile plaques and neurofibrillary tangles of tau protein commonly uses choline acetyltransferase enhancers as therapeutics. The ECS is currently being studied as PD and AD drug targets where overexpression of ECS receptors exerted neuroprotection against PD and reduced neuroinflammation in AD. The delta-9-tetrahydrocannabinoid (Δ9-THC) and cannabidiol (CBD) cannabinoids of plant Cannabis sativa have shown neuroprotection upon PD and AD animal models yet triggered toxic effects on patients when administered directly. Therefore, understanding the precise molecular cascade following cannabinoid treatment is suggested, focusing especially on gene expression to identify drug targets for preventing and repairing neurodegeneration.", "The actions of cannabis are mediated by receptors that are part of an endogenous cannabinoid system. The endocannabinoid system (ECS) consists of the naturally occurring ligands N-arachidonoylethanolamine (anandamide) and 2-arachidonoylglycerol (2-AG), their biosynthetic and degradative enzymes, and the cannabinoid (CB) receptors CB1 and CB2. The ECS is a widely distributed transmitter system that controls gut functions peripherally and centrally. It is an important physiologic regulator of gastrointestinal motility. Polymorphisms in the gene encoding CB1 (CNR1) have been associated with some forms of irritable bowel syndrome. The ECS is involved in the control of nausea and vomiting and visceral sensation. The homeostatic role of the ECS also extends to the control of intestinal inflammation. We review the mechanisms by which the ECS links stress and visceral pain. CB1 in sensory ganglia controls visceral sensation, and transcription of CNR1 is modified through epigenetic processes under conditions of chronic stress. These processes might link stress with abdominal pain. The ECS is also involved centrally in the manifestation of stress, and endocannabinoid signaling reduces the activity of hypothalamic-pituitary-adrenal pathways via actions in specific brain regions, notably the prefrontal cortex, amygdala, and hypothalamus. Agents that modulate the ECS are in early stages of development for treatment of gastrointestinal diseases. Increasing our understanding of the ECS will greatly advance our knowledge of interactions between the brain and gut and could lead to new treatments for gastrointestinal disorders.", "Sphingosine kinase (SPHK) is a key enzyme producing important messenger sphingosine 1-phosphate and is implicated in cell proliferation and suppression of apoptosis. Because the extent of agonist-induced activation of SPHK is modest, signaling via SPHK may be regulated through its localization at specific intracellular sites. Although the SPHK1 isoform has been extensively studied and characterized, the regulation of expression and function of the other isoform, SPHK2, remain largely unexplored. Here we describe an important post-translational modification, namely, phosphorylation of SPHK2 catalyzed by protein kinase D (PKD), which regulates its localization. Upon stimulation of HeLa cells by tumor promoter phorbol 12-myristate 13-acetate, a serine residue in a novel and putative nuclear export signal, identified for the first time, in SPHK2 was phosphorylated followed by SPHK2 export from the nucleus. Constitutively active PKD phosphorylated this serine residue in the nuclear export signal both in vivo and in vitro. Moreover, down-regulation of PKDs through RNA interference resulted in the attenuation of both basal and phorbol 12-myristate 13-acetate-induced phosphorylation, which was followed by the accumulation of SPHK2 in the nucleus in a manner rescued by PKD over-expression. These results indicate that PKD is a physiologically relevant enzyme for SPHK2 phosphorylation, which leads to its nuclear export for subsequent cellular signaling." ]
Estimating the Individuals' Willingness to Pay for a COVID-19 Vaccine	Understanding the individuals' willingness to pay (WTP) for the COVID-19 vaccine could help design policy interventions to control the COVID-19 pandemic. This study aimed to estimate the individuals' willingness to pay (WTP) for a COVID-19 vaccine and to identify its associated determinants.	[ "The rise in dengue fever cases and the absence of dengue vaccines will likely cause governments to consider various types of effective means for controlling the disease. Given strong public interests in potential dengue vaccines, it is essential to understand the private economic benefits of dengue vaccines for accelerated introduction of vaccines into the public sector program and private markets of high-risk countries. A contingent valuation study for a hypothetical dengue vaccine was administered to 400 households in a multi-country setting: Vietnam, Thailand, and Colombia. All respondents received a description of the hypothetical dengue vaccine scenarios of 70% or 95% effectiveness for 10 or 30 years with a three dose series. Five price points were determined after pilot tests in order to reflect different local situations such as household income levels and general perceptions towards dengue fever. We adopted either Poisson or negative binomial regression models to calculate average willingness-to-pay (WTP), as well as median WTP. We found that there is a significant demand for dengue vaccines. The parametric median WTP is $26.4 ($8.8 per dose) in Vietnam, $70.3 ($23.4 per dose) in Thailand, and $23 ($7.7 per dose) in Colombia. Our study also suggests that respondents place more value on vaccinating young children than school age children and adults. Knowing that dengue vaccines are not yet available, our study provides critical information to both public and private sectors. The study results can be used to ensure broad coverage with an affordable price and incorporated into cost benefit analyses, which can inform prioritization of alternative health interventions at the national level.", "The new coronavirus pandemic has appreciably impacted morbidity and mortality, as well as having an economic impact worldwide. New vaccines are a potential way forward to reduce transmission rates and subsequent infection. In Brazil, vaccines are being distributed via the public sector; however, in the future, they will be available in the private market. Information about consumers' willingness to pay (WTP) for a hypothetical vaccine against SARS CoV-2 can help future price setting discussions. A cross-sectional study was conducted with consumers in the five regions of Brazil regarding the WTP for a hypothetical vaccine against SARS CoV-2 with a 50% efficacy. A total of 1402 individuals over 18 years of age who declared not having COVID-19 at the time of the survey were interviewed. The acceptability for this hypothetical vaccine was 80.7%. In addition, the amount of WTP by Brazilian consumers for a hypothetical SARS CoV-2 vaccine was estimated at US$ 22.18(120.00 BRL). This study can contribute to decision-making to inform potential pricing for a hypothetical SARS CoV-2 vaccine.", "The goal of achieving herd immunity to the coronavirus requires high vaccination acceptance levels on the part of the population. The objectives of this study were to: 1) Measure individuals' willingness to pay (WTP) for a COVID-19 vaccine in Kenya; 2) evaluate the effect of vaccine characteristics (duration of protection and efficacy) and individuals' socioeconomic variables on WTP, and 3) estimate the aggregate demand and economic value of a COVID-19 vaccine. The contingent valuation (CV) method was used as the basis for the analyses. Data for this study were obtained from a survey of 1,050 individuals in Kenya conducted from April 7 to April 15, 2020. The survey included CV questions using a double-bounded dichotomous choice format. Results reveal that most of the individuals in Kenya (at least 96%) were willing to accept a COVID-19 vaccine. Approximately 80% of individuals were willing to pay a positive amount. Conservative estimates of individuals' mean WTP for the vaccine range from USD 49.81 to USD 68.25 (depending on vaccine characteristics). Both vaccine duration of protection and efficacy were found to influence WTP (p < .10). The perceived probability of being hospitalized, age, gender, education, location and region of residence, and household income were also found to be associated with WTP for the vaccine (p < .10). In conclusion, the COVID-19 vaccine is highly valued and accepted by the Kenyan population; however, a high percent of the population is unwilling to pay for it or is only willing to pay a low price.", "The development of a vaccine against SARS-CoV-2 infection is on the way. To prepare for public availability, the acceptability of a hypothetical COVID-19 vaccine and willingness to pay (WTP) were assessed to provide insights into future demand forecasts and pricing considerations. A cross-sectional survey was conducted from 3 to 12 April 2020. The health belief model (HBM) was used to assess predictors of the intent to receive the vaccine and the WTP. A total of 1,159 complete responses was received. The majority reported a definite intent to receive the vaccine (48.2%), followed by a probable intent (29.8%) and a possible intent (16.3%). Both items under the perceived benefits construct in the HBM, namely believe the vaccination decreases the chance of infection (OR = 2.51, 95% CI 1.19-5.26) and the vaccination makes them feel less worry (OR = 2.19, 95% CI 1.03-4.65), were found to have the highest significant odds of a definite intention to take the vaccine. The mean ± standard deviation (SD) for the amount that participants were willing to pay for a dose of COVID-19 vaccine was MYR$134.0 (SD±79.2) [US$30.66 ± 18.12]. Most of the participants were willing to pay an amount of MYR$100 [US$23] (28.9%) and MYR$50 [US$11.5] (27.2%) for the vaccine. The higher marginal WTP for the vaccine was influenced by no affordability barriers as well as by socio-economic factors, such as higher education levels, professional and managerial occupations and higher incomes. The findings demonstrate the utility of HBM constructs in understanding COVID-19 vaccination intention and WTP." ]
is mntx good for constipation?	Opioid-induced constipation (OIC) may increase the risk of fecal impaction and mortality in patients with advanced illness. Methylnaltrexone (MNTX) is efficacious for OIC.	[ "Opioid therapy is often associated with adverse effects, including opioid-induced constipation (OIC), in patients receiving opioids for cancer pain. This retrospective observational cohort study evaluated healthcare utilization and costs during the first year after initiating opioid therapy among cancer patients with (cohort 1) and without (cohort 2) constipation. This study used administrative claims data from the HealthCore Integrated Research Environment between January 1, 2006, and April 30, 2014. Eligible patients included adults ≥ 18 years with a diagnosis of cancer who initiated continuous opioid therapy (≥ 30 days). Propensity scores were used to match patients with constipation in a 1:1 ratio to those without constipation. Generalized linear models were used to evaluate healthcare utilization and costs during the 12 months after initiating opioid therapy. After matching, 1369 patients were included in each cohort. Patients with constipation were more than twice as likely as those without constipation to have an all-cause inpatient hospitalization (odds ratio [95% confidence interval (CI)], 2.47 [2.11-2.90]), or pain-related hospitalization (2.15 [1.82-2.54]) during the 12 months after initiating therapy. Mean unadjusted overall healthcare costs during the first 12 months post-index were $21,629 (95% CI, $14,850-$29,018) higher for patients with constipation than for those without constipation. For patients with constipation, total mean (SD) constipation-related costs were $9196 ($26,896). These results suggest that OIC is associated with significantly increased healthcare and economic burden in cancer pain patients and that early and ongoing recognition and management of OIC are unmet needs in this population.", "Methylnaltrexone is effective for opioid-induced constipation (OIC) in advanced illness patients. This 4-week, double-blind, randomized, placebo-controlled study investigated the effect of subcutaneous methylnaltrexone on OIC in patients receiving opioids for chronic, nonmalignant pain. Patients (N = 460) received subcutaneous methylnaltrexone 12 mg once daily (QD) or every other day (alternating with placebo) compared with placebo. Assessments included bowel movement count, time of bowel movement, straining, sense of complete evacuation, Bristol Stool Form Scales, and quality of life. Within 4 hours of first dose, 34.2% of patients in both methylnaltrexone groups had rescue-free bowel movements (RFBMs) versus 9.9% on placebo (P < .001). The estimated number needed to treat was about 4. On average, 28.9% of methylnaltrexone QD and 30.2% of methylnaltrexone alternate-day dosing resulted in RFBMs within 4 hours versus 9.4% QD and 9.3% alternate-day placebo injections (both P < .001). Both methylnaltrexone groups had significantly shorter time to first RFBM (P < .001) and greater increase in number of weekly RFBMs (P < .05) versus placebo. Adverse events included abdominal pain, diarrhea, nausea, and hyperhidrosis. Bristol Stool Form Scale scores (P = .002) and sensation of complete evacuation (P < .04) were significantly superior with methylnaltrexone QD; both methylnaltrexone groups reported no or mild straining during RFBMs in the first 2 weeks (P < .02). At 4 weeks, a significantly greater improvement in patient-reported, constipation-specific quality of life was seen in the alternate-day dosing (P < .05) and QD (P < .001) groups. We present data demonstrating that subcutaneous methylnaltrexone 12 mg given once daily (QD) or every other day provides significant relief of OIC and was generally well tolerated in patients with chronic, nonmalignant pain. These results expand on prior effectiveness observed for the treatment of OIC in advanced illness patients to a broader population.", "Little is known regarding the burden of opioid-induced constipation (OIC) among patients who suffer from cancer-related pain. A prospective longitudinal study was conducted among cancer patients in the United Kingdom (UK), Canada, and Germany, which included medical record data abstraction, Internet-based patient surveys, and physician surveys. Patients on daily opioid therapy (≥30 mg for ≥4 weeks) for treatment of cancer pain with self-reported OIC were recruited. Response to laxatives was defined by classifying participants into categories of laxative use and evaluating the prevalence of inadequate response. Descriptive statistics were used to evaluate outcomes, including the patient assessment of constipation-symptom (PAC-SYM), patient assessment of constipation-quality of life, EuroQOL-5 dimensions, and global assessment of treatment benefit, satisfaction, and willingness to continue. Recruitment was difficult for this study with only 31 participants completing the baseline survey and meeting criteria for opioid use and OIC (26 UK, 1 Canada, and 4 Germany). Fifty-two percent (n = 16) of participants were male, and all were White. Breast (23%, n = 7), pancreatic (13%, n = 4), and multiple myeloma (13%, n = 4) were the most common cancers. Mean duration of chronic pain and opioid use were 2.3 and 1.3 years, respectively. Participants reported having a mean of 4.4 bowel movements/week in the 2 weeks prior to baseline, of which a mean of 0.9 were spontaneous. Most participants (90%, n = 28) were using at least 1 lifestyle approach to manage their constipation; 65% (n = 20) were taking ≥1 over-the-counter laxative; 19% (n = 6) were taking ≥1 prescription laxative; 23% (n = 7) reported no laxative use in the prior 2 weeks. Moderate-to-severe constipation symptoms on the PAC-SYM were common, and mean scores on health-related quality of life outcomes were comparable to chronic pain populations. In this primarily UK sample, there appears to be considerable unmet OIC treatment needs among cancer patients.", "Opioids are widely accepted as treatment for moderate to severe pain, and opioid-induced constipation is one of the most common side effects of opioids. This side effect negatively affects pain management and patients' quality of life, which could result in increased healthcare utilization and costs. To assess healthcare utilization and costs (all-cause, constipation-related, and pain-related) for individuals with and without opioid-induced constipation during the 12 months after initiation of opioid therapy for noncancer pain. This retrospective cohort study was conducted using administrative claims data from HealthCore Integrated Research Environment between January 1, 2006, and June 30, 2014. The analysis was limited to patients aged ≥18 years who filled a prescription for continuous opioid treatment (≥28 days) for noncancer pain. Propensity scores were used to match opioid users with constipation (cohort 1) and opioid users without constipation (cohort 2), using a 1:1 ratio. Generalized linear models were used to estimate all-cause, constipation-related, and pain-related healthcare utilization and costs during the 12 months after the initiation of opioid therapy. After matching and balancing for all prespecified variables, 17,384 patients were retained in each cohort (mean age, 56 years; 63% female). Opioid users with constipation were twice as likely as those without constipation to have ≥1 inpatient hospitalizations (odds ratio, 2.28; 95% confidence interval [CI], 2.17-2.39) during the 12 months. The total mean adjusted overall costs per patient during the study period were $12,413 higher for patients with constipation versus those without it (95% CI, $11,726-$13,116). The total mean adjusted overall pain-related costs per patient were $6778 (95% CI, $6293-$7279) higher for the patients with constipation than those without. Among patients using opioids for noncancer pain, the annual mean constipation-related costs per patient totaled $4646 (total average plan-paid costs, $4424; total patient-paid costs, $222). Patients using opioids with newly diagnosed constipation had significantly greater healthcare utilization and costs than patients without constipation; these costs accounted for approximately 16% of the total healthcare costs per patient during the 12-month study period. Recognition and effective treatment of opioid-induced constipation may decrease healthcare utilization for patients with chronic noncancer pain and may reduce the economic burden of pain therapy.", "A case of rapid demise following an undiagnosed stercoral perforation is reported. A 57-year-old woman on chronic opioid replacement therapy presented with constipation and abdominal pain to the hospital. Following an unremarkable abdominal radiograph and admission for laxation and pain therapy, she was found dead only 18 h later. To exclude medical malpractice, a postmortem investigation was ordered. Postmortem computed tomography and autopsy revealed fatal fecal peritonitis based on a stercoral perforation of the rectosigmoid, which had been undiagnosed. This report highlights the need for early cross-sectional imaging and contributes to the data collection concerning this ever-growing, vulnerable group of patients undergoing opioid replacement.", "The main objective of this study was to compare the opioid use patterns, resource utilization, and costs of patients on opioid therapy who have constipation with those who do not. Retrospective, observational matched cohort design Patients initiating opioid therapy between Jan. 1, 1999 and Dec. 31, 2005 were identified from a longitudinal insurance claims database. Patients had > or = 30 days of opioid use and continuous plan coverage for > or = 6 months before and > or = 12 months after their index date, defined as the date of the first pharmacy claim for an opioid. Constipation was defined as having one or more ICD-9 codes of 564.0 during the follow-up period. Patterns of opioid use and resource utilization were compared between patients with constipation and a demographically matched (1:1) cohort of opioid initiators without consti- pation using t-tests and Chi-square (chi2) tests. We identified 39,485 patients, of whom 2,519 (6.4%) had constipation. Most patients with constipation were female (66%) and > or = 45 years old (68%). Compared to controls, the constipation group had significantly higher rates of concurrent use of > or = 2 opioids (p < 0.0001), discontinuation, and switching between opioids. Patients with constipation had statistically significant higher hospital admissions, emergency room visits, home health services, nursing home care, physician office visits, other outpatient/ ancillary care, and laboratory tests. Patients with constipation had significantly higher mean all-cause costs for emergency, physician visits, nursing facility, home health, and prescription drug services compared to patients without constipation. Opioid-treated patients with constipation were found to have significant differences in opioid use patterns and significantly higher health care utilization and associated costs.", "Propensity-score methods are increasingly being used to reduce the impact of treatment-selection bias in the estimation of treatment effects using observational data. Commonly used propensity-score methods include covariate adjustment using the propensity score, stratification on the propensity score, and propensity-score matching. Empirical and theoretical research has demonstrated that matching on the propensity score eliminates a greater proportion of baseline differences between treated and untreated subjects than does stratification on the propensity score. However, the analysis of propensity-score-matched samples requires statistical methods appropriate for matched-pairs data. We critically evaluated 47 articles that were published between 1996 and 2003 in the medical literature and that employed propensity-score matching. We found that only two of the articles reported the balance of baseline characteristics between treated and untreated subjects in the matched sample and used correct statistical methods to assess the degree of imbalance. Thirteen (28 per cent) of the articles explicitly used statistical methods appropriate for the analysis of matched data when estimating the treatment effect and its statistical significance. Common errors included using the log-rank test to compare Kaplan-Meier survival curves in the matched sample, using Cox regression, logistic regression, chi-squared tests, t-tests, and Wilcoxon rank sum tests in the matched sample, thereby failing to account for the matched nature of the data. We provide guidelines for the analysis and reporting of studies that employ propensity-score matching." ]
Galectin-3/Lrp1 axis regulates transcriptional programs and intracellular signaling cascades critical to osteoclast function	Bone-resorbing osteoclasts mobilize proteolytic enzymes belonging to the matrix metalloproteinase (MMP) family to directly degrade type I collagen, the dominant extracellular matrix component of skeletal tissues. While searching for additional MMP substrates critical to bone resorption, Mmp9/Mmp14 double-knockout (DKO) osteoclasts-as well as MMP-inhibited human osteoclasts-unexpectedly display major changes in transcriptional programs in tandem with compromised RhoA activation, sealing zone formation and bone resorption. Further study revealed that osteoclast function is dependent on the ability of Mmp9 and Mmp14 to cooperatively proteolyze the β-galactoside-binding lectin, galectin-3, on the cell surface. Mass spectrometry identified the galectin-3 receptor as low-density lipoprotein-related protein-1 (Lrp1), whose targeting in DKO osteoclasts fully rescues RhoA activation, sealing zone formation and bone resorption. Together, these findings identify a previously unrecognized galectin-3/Lrp1 axis whose proteolytic regulation controls both the transcriptional programs and the intracellular signaling cascades critical to mouse as well as human osteoclast function.	[ "Rac small GTPases may play an important regulatory role in osteoclastogenesis. Our in vitro and in vivo results show that both Rac1 and Rac2 are required for optimal osteoclast differentiation, but Rac1 is more critical. Rac1 is the key Rac isoform responsible for regulating ROS generation and the actin cytoskeleton during the multiple stages of osteoclast differentiation. Recent evidence suggests that the Rac small GTPases may play an important regulatory role in osteoclastogenesis. This finding is important because bisphosphonates may regulate their antiresorptive/antiosteoclast effects through the modification of Rho family of small GTPases. To elucidate the specific roles of the Rac1 and Rac2 isoforms during osteoclastogenesis, we used mice deficient in Rac1, Rac2, or both Rac1 and Rac2 in monocyte/osteoclast precursors. Macrophage-colony stimulating factor (M-CSF)- and RANKL-mediated osteoclastogenesis in vitro was studied by using bone marrow-derived mononucleated preosteoclast precursors (MOPs). The expression of osteoclast-specific markers was examined using quantitative real-time PCR and Western blot analysis. Free actin barbed ends in bone marrow MOPs after M-CSF stimulation was determined. The ability of MOPs to migrate toward M-CSF was assayed using Boyden chambers. Margin spreading on heparin sulfate-coated glass and RANKL-induced reactive oxygen species generation were also performed. Functional assays of in vitro-generated osteoclasts were ascertained using dentine sections from narwal tusks. Osteoclast levels in vivo were counted in TRACP and immunohistochemically stained distal tibial sections. In vivo microarchitexture of lumbar vertebrate was examined using microCT 3D imaging and analysis. We show here that, although both Rac isoforms are required for normal osteoclast differentiation, Rac1 deletion results in a more profound reduction in osteoclast formation in vitro because of its regulatory role in pre-osteoclast M-CSF-mediated chemotaxis and actin assembly and RANKL-mediated reactive oxygen species generation. This Rac1 cellular defect also manifests at the tissue level with increased trabecular bone volume and trabeculae number compared with wildtype and Rac2-null mice. This unique mouse model has shown for the first time that Rac1 and Rac2 play different and nonoverlapping roles during osteoclastogenesis and will be useful for identifying the key roles played by these two proteins during the multiple stages of osteoclast differentiation. Rac1 and Rac2 play different and nonoverlapping roles during osteoclastogenesis. This model showed that Rac1 is the key Rac isoform responsible for regulating ROS generation and the actin cytoskeleton during the multiple stages of osteoclast differentiation.", "In order to resorb the mineralized bone extracellular matrix, the osteoclast relies on the generation of a resorption lacuna characterized by the presence of specific proteases and a low pH. Hence, bone resorption by osteoclasts is highly dependent on lysosomes, the organelles specialized in intra- and extracellular material degradation. This is best illustrated by the fact that multiple forms of human osteopetrosis are caused by mutations in genes encoding for lysosomal proteins. Yet, until recently, the molecular mechanisms regulating lysosomal biogenesis and function in osteoclasts were poorly understood. Here we review the latest developments in the study of lysosomal biogenesis and function in osteoclasts with an emphasis on the transcriptional control of these processes.", "The osteoclast cytoskeleton is a unique structure that polarizes the cell's resorptive machinery to the bone-cell interface where it creates an isolated resorptive microenvironment consisting of an actin ring surrounding a ruffled border. This polarization process occurs under the aegis of the α(v) β(3) integrin in collaboration with the M-CSF receptor, c-Fms. When occupied, α(v) β(3) activates a canonical signaling complex consisting of c-Src, Syk, Dap12, Slp76, Vav 3, and Rac that permits the cell to spread and form actin rings. Generation of the ruffled border, the cell's resorptive organelle, is an exocytic process wherein synaptotagmin VII mediates fusion of secretory lysosomes to the bone-apposed plasma membrane. Absence of any component of this signaling pathway compromises osteoclast cytoskeletal organization and abridges bone resorption.", "Osteoporosis is a metabolic bone disorder associated with compromised bone strength and an increased risk of fracture. Inhibition of the differentiation of bone-resorbing osteoclasts is an effective strategy for the treatment of osteoporosis. Prior work by our laboratory and others has shown that MYC promotes osteoclastogenesis in vitro, but the underlying mechanisms are not well understood. In addition, the in vivo importance of osteoclast-expressed MYC in physiological and pathological bone loss is not known. Here, we have demonstrated that deletion of Myc in osteoclasts increases bone mass and protects mice from ovariectomy-induced (OVX-induced) osteoporosis. Transcriptomic analysis revealed that MYC drives metabolic reprogramming during osteoclast differentiation and functions as a metabolic switch to an oxidative state. We identified a role for MYC action in the transcriptional induction of estrogen receptor-related receptor α (ERRα), a nuclear receptor that cooperates with the transcription factor nuclear factor of activated T cells, c1 (NFATc1) to drive osteoclastogenesis. Accordingly, pharmacological inhibition of ERRα attenuated OVX-induced bone loss in mice. Our findings highlight a MYC/ERRα pathway that contributes to physiological and pathological bone loss by integrating the MYC/ERRα axis to drive metabolic reprogramming during osteoclast differentiation.", "Osteoclasts are bone-degrading cells, which play a central role in physiological bone remodeling. Unbalanced osteoclast activity is largely responsible for pathological conditions such as osteoporosis. Osteoclasts develop specialized adhesion structures, the so-called podosomes, which subsequently undergo dramatic reorganization into sealing zones. These ring-like adhesion structures, which delimit the resorption site, effectively seal the cell to the substrate forming a diffusion barrier. The structural integrity of the sealing zone is essential for the cell ability to degrade bone, yet its structural organization is poorly understood. Combining high-resolution scanning electron microscopy with fluorescence microscopy performed on the same sample, we mapped the molecular architecture of the osteoclast resorptive apparatus from individual podosomes to the sealing zone, at an unprecedented resolution. Podosomes are composed of an actin-bundle core, flanked by a ring containing adhesion proteins connected to the core via dome-like radial actin fibers. The sealing zone, hallmark of bone-resorbing osteoclasts, consists of a dense array of podosomes communicating through a network of actin filaments, parallel to the substrate and anchored to the adhesive plaque domain via radial actin fibers. The sealing zone of osteoclasts cultured on bone is made of structural units clearly related to individual podosomes. It differs from individual or clustered podosomes in the higher density and degree of inter-connectivity of its building blocks, thus forming a unique continuous functional structure connecting the cell to its extracellular milieu. Through this continuous structure, signals reporting on the substrate condition may be transmitted to the whole cell, modulating the cell response under physiological and pathological conditions.", "Bone is a dynamic tissue constantly renewed through a regulated balance between bone formation and resorption. Excessive bone degradation by osteoclasts leads to pathological decreased bone density characteristic of osteolytic diseases such as post-menopausal osteoporosis or bone metastasis. Osteoclasts are multinucleated cells derived from hematopoietic stem cells via a complex differentiation process. Their unique ability to resorb bone is dependent on the formation of the actin-rich sealing zone. Within this adhesion structure, the plasma membrane differentiates into the ruffled border where protons and proteases are secreted to demineralize and degrade bone, respectively. On the bone surface, mature osteoclasts alternate between stationary resorptive and migratory phases. These are associated with profound actin cytoskeleton reorganization, until osteoclasts die of apoptosis. In this review, we highlight the role of Rho GTPases in all the steps of osteoclasts differentiation, function, and death and conclude on their interest as targets for treatment of osteolytic pathologies.", "Cell fusion is essential for fertilization, myotube formation, and inflammation. Macrophages fuse under various circumstances, but the molecular signals involved in the distinct steps of their fusion are not fully characterized. Using null mice and derived cells, we show that the protease MT1-MMP is necessary for macrophage fusion during osteoclast and giant-cell formation in vitro and in vivo. Specifically, MT1-MMP is required for lamellipodia formation and for proper cell morphology and motility of bone marrow myeloid progenitors prior to membrane fusion. These functions of MT1-MMP do not depend on MT1-MMP catalytic activity or downstream pro-MMP-2 activation. Instead, MT1-MMP null cells show a decreased Rac1 activity and reduced membrane targeting of Rac1 and the adaptor protein p130Cas. Retroviral rescue experiments and protein binding assays delineate a signaling pathway in which MT1-MMP, via its cytosolic tail, contributes to macrophage migration and fusion by regulating Rac1 activity through an association with p130Cas." ]
24-Hour Movement: A New Paradigm for Health Behavior Research in Oncology	Increased physical activity (PA), improved sleep, and decreased sedentary behavior (SB) are essential components of supportive care for cancer survivors. However, researchers and health care professionals have achieved limited success in improving these behaviors among cancer survivors. One potential reasoning is that, over the past two decades, guidelines for promoting and measuring PA, sleep, and SB have been largely siloed. With greater understanding of these three behaviors, health behavior researchers have recently developed a new paradigm: the 24-Hour movement approach. This approach considers PA, SB, and sleep as movement behaviors along a continuum that represent low through vigorous intensity activity. Together these three behaviors form the sum of an individual's movement across a 24-hour day. While this paradigm has been studied in the general population, its usage is still limited in cancer populations. Here, we seek to highlight (a) the potential benefits of this new paradigm for clinical trial design in oncology; (b) how this approach can allow for greater integration of wearable technology as a means of assessing and monitoring patient health outside the clinical setting, improving patient autonomy through self-monitoring of movement behavior. Ultimately, implementation of the 24-Hour movement paradigm will allow health behavior research in oncology to better promote and assess critical health behaviors to support the long-term well-being for cancer patients and survivors.	[ "Kidney cancer survivors spend large quantities of time sedentary and little time physically active, which negatively impacts quality of life (QoL). This study examined (1) the association of reallocating sedentary time to sleep, light physical activity (PA), or moderate-to-vigorous PA (MVPA) on QoL in kidney cancer survivors and (2) the threshold at which results are clinically meaningful. Kidney cancer survivors (N = 463) completed a survey including the Godin Leisure-Time Exercise Questionnaire, sitting time, sleep duration, and Functional Assessment of Cancer Therapy (FACT) scales. Isotemporal substitution analyses estimated associations of reallocating sedentary time to PA and sleep on QoL. Reallocating 10 min/day of sedentary time to MVPA was significantly associated with higher scores on the Trial Outcome Index-Fatigue (B = 0.60, SE = 0.25, p = 0.02), FACT-Fatigue (B = 0.71, SE = 0.32, p = 0.03), functional well-being (B = 0.18, SE = 0.08, p = 0.02), and fatigue subscales (B = 0.35, SE = 0.15, p = 0.02). Reallocating sedentary time to sleep was significantly associated with higher FACT-General (B = 0.15, SE = 0.08, p = 0.04) and functional well-being subscale (B = 0.06, SE = 0.03, p = 0.049) scores. Reallocating sedentary time to light PA was significantly associated with higher fatigue subscale scores (B = 0.46, SE = 0.23, p = 0.045). Kidney cancer survivors would need to reallocate a minimum of about 83, 200, and 65 min/day of MVPA, sleep, and light PA, respectively, for associations to be clinically meaningful. Reallocating sedentary time to MVPA, light PA, or sleep at higher doses is associated with better fatigue and physical aspects of QoL. Interventions should consider replacing sedentary time with MVPA or light PA in a gradual manner, and improve sleep quality for kidney cancer survivors.", "The association between physical activity and colorectal adenoma is equivocal. This study was designed to assess the relationship between physical activity and colorectal adenoma recurrence. Pooled analyses from two randomized, controlled trials included 1,730 participants who completed the Arizona Activity Frequency Questionnaire at baseline, had a colorectal adenoma removed within 6 months of study registration, and had a follow-up colonoscopy during the trial. Logistic regression modeling was employed to estimate the effect of sedentary behavior, light-intensity physical activity, and moderate-vigorous physical activity on colorectal adenoma recurrence. No statistically significant trends were found for any activity type and odds of colorectal adenoma recurrence in the pooled population. However, males with the highest levels of sedentary time experienced 47% higher odds of adenoma recurrence. Compared to the lowest quartile of sedentary time, the ORs (95% CIs) for the second, third, and fourth quartiles among men were 1.23 (0.88, 1.74), 1.41 (0.99, 2.01), and 1.47 (1.03, 2.11), respectively (p(trend) = 0.03). No similar association was observed for women. This study suggests that sedentary behavior is associated with a higher risk of colorectal adenoma recurrence among men, providing evidence of detrimental effects of a sedentary lifestyle early in the carcinogenesis pathway.", "Existing data suggest that physical activity reduces colon cancer risk, but the association is not consistently observed in women. One potential explanation for this inconsistency is that hormone therapy, which is associated with lower colon cancer risk, acts as a modifier of the physical activity/colon cancer relationship. Participants in the California Teachers Study (N = 120,147), a prospective cohort of female teachers and administrators residing in California, ages 22 to 84 years at baseline and with no prior history of colon cancer were eligible for study. Between 1996 and 2002, 395 patients were diagnosed with invasive colon cancer. The relative risks (RR) associated with lifetime (high school through age 54 years or current age) and recent (past 3 years) strenuous and moderate recreational physical activity were estimated using Cox proportional hazards regression models. Combined lifetime moderate and strenuous recreational physical activity was only modestly associated with colon cancer risk in the cohort [>or=4 versus <or=0.5 h/wk/y: RR, 0.75; 95% confidence interval, 0.57-1.00; P(trend) = 0.23]. Lifetime physical activity reduced colon cancer risk among postmenopausal women who had never taken hormone therapy (>or=4 versus <or=0.5 h/wk/y: RR, 0.51; 95% confidence interval, 0.31-0.85; P(trend) = 0.02). Postmenopausal women with histories of hormone therapy use had lower colon cancer risk, but their risk was not associated with physical activity. The likelihood ratio test for interaction between hormone use and lifetime moderate plus strenuous physical activity was of borderline statistical significance (P = 0.05). We observed no effect modification by age, body mass index, smoking status, menopausal status, or folate intake. Lifetime recreational physical activity may protect against colon cancer among postmenopausal women who have never used hormone therapy. Among hormone therapy users, who have lower risk of colon cancer, recreational physical activity does not seem to provide any additional benefit. With declining rates of hormone therapy use, physical activity offers one possible means for reducing women's colon cancer risk.", "Few epidemiologic investigations of visceral adiposity and colorectal neoplasms have attempted the direct quantification of visceral fat. The authors measured visceral fat volume among middle-aged and elderly Japanese men and women who underwent colonoscopy and positron emission tomography/computed tomography for cancer screening in Tokyo, Japan, between February 2004 and February 2005, and examined the association between visceral adiposity and colorectal adenoma in 1,205 eligible subjects. Odds ratios and 95% confidence intervals for colorectal adenoma were estimated by using an unconditional logistic regression model after adjustment for potential confounders. Despite its high correlation with body mass index, visceral fat volume was associated with the prevalence of colorectal adenoma independently of body mass index in both sexes. After further adjustment for body mass index, the odds ratio of colorectal adenoma for the highest compared with the lowest quartile of visceral fat volume was 1.58 (95% confidence interval: 1.11, 2.24) for men and women combined. Conversely, body mass index was unlikely to modify the association between visceral fat volume and colorectal adenoma (P(interaction) = 0.39). These findings add to accumulating evidence that visceral adiposity exerts an important influence on the pathogenesis of colorectal neoplasms. The mechanisms of this potential association between visceral adiposity and colorectal carcinogenesis warrant further investigation.", "Obesity is associated with vitamin D insufficiency and secondary hyperparathyroidism. This study assessed whether obesity alters the cutaneous production of vitamin D(3) (cholecalciferol) or the intestinal absorption of vitamin D(2) (ergocalciferol). Healthy, white, obese [body mass index (BMI; in kg/m(2)) > or = 30] and matched lean control subjects (BMI </= 25) received either whole-body ultraviolet radiation or a pharmacologic dose of vitamin D(2) orally. Obese subjects had significantly lower basal 25-hydroxyvitamin D concentrations and higher parathyroid hormone concentrations than did age-matched control subjects. Evaluation of blood vitamin D(3) concentrations 24 h after whole-body irradiation showed that the incremental increase in vitamin D(3) was 57% lower in obese than in nonobese subjects. The content of the vitamin D(3) precursor 7-dehydrocholesterol in the skin of obese and nonobese subjects did not differ significantly between groups nor did its conversion to previtamin D(3) after irradiation in vitro. The obese and nonobese subjects received an oral dose of 50000 IU (1.25 mg) vitamin D(2). BMI was inversely correlated with serum vitamin D(3) concentrations after irradiation (r = -0.55, P: = 0.003) and with peak serum vitamin D(2) concentrations after vitamin D(2) intake (r = -0.56, P: = 0.007). Obesity-associated vitamin D insufficiency is likely due to the decreased bioavailability of vitamin D(3) from cutaneous and dietary sources because of its deposition in body fat compartments.", "High levels of sedentary behavior may negatively affect health outcomes in cancer survivors. A systematic review and meta-analysis was performed to clarify whether postdiagnosis sedentary behavior is related to survival, patient-reported outcomes, and anthropometric outcomes in cancer survivors. The Ovid MEDLINE, EMBASE, CINAHL (The Cumulative Index to Nursing and Allied Health Literature), and SPORTDiscus databases were searched from study inception to June 2019. Studies of adults who had been diagnosed with cancer that examined the association between sedentary behavior and mortality, patient-reported outcomes (eg, fatigue, depression), or anthropometric outcomes (eg, body mass index, waist circumference) were eligible for inclusion. Meta-analyses were performed to estimate hazard ratios for the highest compared with the lowest levels of sedentary behavior for all-cause and colorectal cancer-specific mortality outcomes. The ROBINS-E (Risk of Bias in Nonrandomized Studies-of Exposures tool) and the GRADE (Grading of Recommendations Assessment, Development, and Evaluation) system were used to assess the risk of bias and the strength of evidence, respectively. Thirty-three eligible publications from a total of 3569 identified articles were included in the review. A higher level of postdiagnosis sedentary behavior was associated with an increased risk of all-cause mortality (hazard ratio, 1.22; 95% CI, 1.06-1.41; heterogeneity [I2 statistic], 33.8%) as well as colorectal cancer-specific mortality (hazard ratio, 1.53; 95% CI, 1.14-2.06; I2 , 0%). No clear or consistent associations between sedentary behavior and patient-reported or anthropometric outcomes were identified. The risk of bias in individual studies ranged from moderate to serious, and the strength of evidence ranged from very low to low. Although avoiding high levels of sedentary behavior after a cancer diagnosis may improve survival, further research is required to help clarify whether the association is causal.", "Little evidence is available on the relation of physical activity with colon adenomas, a colon cancer precursor. We conducted a systematic literature review and meta-analysis of published studies (in English) through April 2010, examining physical activity or exercise and risk or prevalence of colon adenoma or polyp. Random effects models were used to estimate relative risks (RRs) and corresponding confidence intervals (CIs). A total of 20 studies were identified that examined the association and provided RRs and corresponding 95% CIs. A significant inverse association between physical activity and colon adenomas was found with an overall RR of 0.84 (CI: 0.77-0.92). The association was similar in men (RR=0.81, CI: 0.67-0.98) and women (RR=0.87, CI: 0.74-1.02). The association appeared slightly stronger in large/advanced polyps (RR=0.70, CI: 0.56-0.88). This study confirms previous reports of a significant inverse association of physical activity and colon adenoma, and suggests that physical activity can have an important role in colon cancer prevention." ]
Expression of ELFN1-AS1 and TRIM29 in gastric cancer	Long noncoding RNA (lncRNA) extracellular leucine rich repeat and fibronectin type III domain containing 1-antisense RNA 1 (ELFN1-AS1) has been found to be upregulated in various tumors. However, the biological functions of ELFN1-AS1 in gastric cancer (GC) are not entirely understood. In the present study, the expression levels of ELFN1-AS1, miR-211-3p, and TRIM29 are determined using reverse transcription-quantitative PCR. Subsequently, CCK8, EdU, and colony formation assays are performed to determine GC cell vitality. The migratory and invasive capabilities of GC cells are further evaluated using transwell invasion and cell scratch assays. Western blot analysis is performed to quantify the levels of proteins associated with GC cell apoptosis and epithelialmesenchymal transition (EMT). The competing endogenous RNA (ceRNA) activity of ELFN1-AS1 on TRIM29 through miR-211-3p is confirmed by pull-down, RIP, and luciferase reporter assays. Our study proves that ELFN1-AS1 and TRIM29 are highly expressed in GC tissues.	[ "Accumulating evidence suggests long noncoding RNAs (lncRNAs) play an important role in cancer progression. However, the function of lncRNA SNHG7 in colorectal cancer (CRC) remains unclear. In this study, SNHG7 expression was significantly upregulated in CRC tissues, especially in aggressive cases. In accordance, high level of SNHG7 was observed in CRC cell lines compared to normal colon cells. Furthermore, SNHG7 overexpression promoted the proliferation, migration, and invasion of CRC cell lines, while SNHG7 depletion inhibited invasion and cell viability in vitro. Mechanistically, knockdown of SNHG7 inhibited GALNT1 and EMT markers (E-cadherin and Vimentin). Importantly, SNHG7 directly interacted with miR-216b and downregulation of miR-216b reversed efficiently the suppression of GALNT1 induced by SNHG7 siRNA. Moreover, overexpression of SNHG7 significantly enhanced the tumorigenesis and liver metastasis of SW480 cells in vivo. SNHG7 positively regulated GALNT1 level through sponging miR-216b, and played an oncogenic role in CRC progression. Together, our study elucidated the role of SNHG7 as an miRNA sponge in CRC, and shed new light on lncRNA-directed diagnostics and therapeutics in CRC.", "Deregulation of long non-coding RNAs (lncRNAs) expression has been proven to be involved in the development and progression of cancer. However, expression pattern and prognostic value of lncRNAs in breast cancer recurrence remain unclear. Here, we analyzed lncRNA expression profiles of breast cancer patients who did or did not develop recurrence by repurposing existing microarray datasets from the Gene Expression Omnibus database, and identified 12 differentially expressed lncRNAs that were closely associated with tumor recurrence of breast cancer patients. We constructed a lncRNA-focus molecular signature by the risk scoring method based on the expression levels of 12 relapse-related lncRNAs from the discovery cohort, which classified patients into high-risk and low-risk groups with significantly different recurrence-free survival (HR = 2.72, 95% confidence interval 2.07-3.57; p = 4.8e-13). The 12-lncRNA signature also represented similar prognostic value in two out of three independent validation cohorts. Furthermore, the prognostic power of the 12-lncRNA signature was independent of known clinical prognostic factors in at least two cohorts. Functional analysis suggested that the predicted relapse-related lncRNAs may be involved in known breast cancer-related biological processes and pathways. Our results highlighted the potential of lncRNAs as novel candidate biomarkers to identify breast cancer patients at high risk of tumor recurrence." ]
Long noncoding RNAs in ventricular septal defects	Long noncoding RNAs (lncRNAs) have been shown to be involved in the regulation of numerous biological processes in embryonic development. We aimed to explore lncRNA expression profiles in ventricular septal defects (VSDs) and reveal their potential roles in heart development.	[ "In previous studies, we have demonstrated that long noncoding RNA uc.4 may influence the cell differentiation through the TGF-β signaling pathway, suppressed the heart development of zebrafish and resulting cardiac malformation. DNA methylation plays a significant role in the heart development and disordered of DNA methylation may cause disruption of control of gene promoter. In this study, methylated DNA immunoprecipitation was performed to identify the different expression levels of methylation regions. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were also performed to identify the possible biological process and pathway that uc.4 may join, associated with Rap1 signaling pathway, gonadotropin-releasing hormone signaling pathway, and Calcium signaling pathway. We found that the distribution of differentially methylated regions peaks was mainly located in intergenic and intron regions. Altogether, our result showed that differentially methylated genes are significantly expressed in uc.4-overexpression cells, providing valuable data for further exploration of the role of uc.4 in heart development.", "Although small non-coding RNAs, such as microRNAs, have well-established functions in the cell, long non-coding RNAs (lncRNAs) have only recently started to emerge as abundant regulators of cell physiology, and their functions may be diverse. A small number of studies describe interactions between small and lncRNAs, with lncRNAs acting either as inhibitory decoys or as regulatory targets of microRNAs, but such interactions are still poorly explored. To facilitate the study of microRNA-lncRNA interactions, we implemented miRcode: a comprehensive searchable map of putative microRNA target sites across the complete GENCODE annotated transcriptome, including 10 419 lncRNA genes in the current version. http://www.mircode.org [email protected] Supplementary data are available at Bioinformatics online.", "The roles of long non-coding RNA (LncRNA) have been highlighted in various development processes including congenital heart defects (CHD). Here, we characterized the molecular function of LncRNA, Moshe (1010001N08ik-203), one of the Gata6 antisense transcripts located upstream of Gata6, which is involved in both heart development and the most common type of congenital heart defect, atrial septal defect (ASD). During mouse embryonic development, Moshe was first detected during the cardiac mesoderm stage (E8.5 to E9.5) where Gata6 is expressed and continues to increase at the atrioventricular septum (E12.5), which is involved in ASD. Functionally, the knock-down of Moshe during cardiogenesis caused significant repression of Nkx2.5 in cardiac progenitor stages and resulted in the increase in major SHF lineage genes, such as cardiac transcriptional factors (Isl1, Hand2, Tbx2), endothelial-specific genes (Cd31, Flk1, Tie1, vWF), a smooth muscle actin (a-Sma) and sinoatrial node-specific genes (Shox2, Tbx18). Chromatin Isolation by RNA Purification showed Moshe activates Nkx2.5 gene expression via direct binding to its promoter region. Of note, Moshe was conserved across species, including human, pig and mouse. Altogether, this study suggests that Moshe is a heart-enriched lncRNA that controls a sophisticated network of cardiogenesis by repressing genes in SHF via Nkx2.5 during cardiac development and may play an important role in ASD.", "Pre-processing methods for two-sample long oligonucleotide arrays, specifically the Agilent technology, have not been extensively studied. The goal of this study is to quantify some of the sources of error that affect measurement of expression using Agilent arrays and to compare Agilent's Feature Extraction software with pre-processing methods that have become the standard for normalization of cDNA arrays. These include log transformation followed by loess normalization with or without background subtraction and often a between array scale normalization procedure. The larger goal is to define best study design and pre-processing practices for Agilent arrays, and we offer some suggestions. Simple loess normalization without background subtraction produced the lowest variability. However, without background subtraction, fold changes were biased towards zero, particularly at low intensities. ROC analysis of a spike-in experiment showed that differentially expressed genes are most reliably detected when background is not subtracted. Loess normalization and no background subtraction yielded an AUC of 99.7% compared with 88.8% for Agilent processed fold changes. All methods performed well when error was taken into account by t- or z-statistics, AUCs > or = 99.8%. A substantial proportion of genes showed dye effects, 43% (99% CI: 39%, 47%). However, these effects were generally small regardless of the pre-processing method. Simple loess normalization without background subtraction resulted in low variance fold changes that more reliably ranked gene expression than the other methods. While t-statistics and other measures that take variation into account, including Agilent's z-statistic, can also be used to reliably select differentially expressed genes, fold changes are a standard measure of differential expression for exploratory work, cross platform comparison, and biological interpretation and can not be entirely replaced. Although dye effects are small for most genes, many array features are affected. Therefore, an experimental design that incorporates dye swaps or a common reference could be valuable." ]
Long-term bisphosphonate use has been linked to increased risk of pathological neck of femur fractures	Long-term bisphosphonate use has been linked to an increased risk of pathological neck of femur fractures.	[ "Several studies have identified a specific fracture in the proximal diaphysis of the femur in patients treated with bisphosphonates. The fractures typically are sustained after a low-energy mechanism with the presence of an existing characteristic stress fracture. However, it is unclear whether these patients are best treated nonoperatively or operatively. What is the likelihood of nonoperatively treated bisphosphonate-associated femoral stress fractures progressing to completion and during what time period? If prophylactic fixation is performed, do patients have a shorter hospital length-of-stay compared with patients having surgical fixation after fracture completion? We retrospectively searched for patients older than 50 years receiving bisphosphonate therapy, with either incomplete, nondisplaced stress fractures or completed, displaced fractures in the proximal diaphysis of the femur between July 2002 and April 2009. After applying exclusion criteria, we identified 34 patients with a total of 40 bisphosphonate-associated fractures. The average duration of bisphosphonate use was 77 months. Twenty-eight of 40 (70%) fractures were completed, displaced fractures. Six of the 12 nondisplaced stress fractures initially were treated nonoperatively. The remaining six stress fractures were treated with prophylactic cephalomedullary nail fixation. The minimum followup was 12 months (mean, 36.5 months; range, 12-72 months). Five of the six stress fractures treated nonoperatively progressed to fracture completion and displacement at an average of 10 months (range, 3-18 months). The average hospital stay was 3.7 days for patients treated prophylactically and 6.0 days for patients treated after fracture completion. Our data suggest nonoperative treatment of bisphosphonate-related femoral stress fractures is not a reliable way to treat these fractures as the majority progress to fracture completion. Prophylactic fixation of femoral stress fractures also reduces total hospital admission time. Level IV, therapeutic study. See Guidelines for Authors for a complete description of levels of evidence.", "We investigated the effects of inhibitors of bone resorption (estrogen, raloxifene, and alendronate) on the processes of fracture repair in ovariectomized (OVX) rats. One hundred forty female Sprague-Dawley rats at 3 months of age were either OVX or sham-operated and divided into five groups: sham control, OVX control, estrogen (17alpha-ethynyl estradiol [EE2], 0.1 mg/kg), raloxifene (Rlx, 1.0 mg/kg), and alendronate (Aln, 0.01 mg/kg) groups. Treatment began immediately after the surgery. Four weeks postovariectomy, prefracture controls were killed and bilateral osteotomies were performed on the femoral midshafts and fixed with intramedullary wires. Treatment was continued and fracture calluses were excised at 6 weeks and 16 weeks postfracture for evaluation by X-ray radiography, quantitative computed tomography (QCT,) biomechanical testing, and histomorphometry. At 6 weeks postfracture, Aln and OVX had larger calluses than other groups. Sham and OVX had higher ultimate load than EE2 and Rlx, with Aln not different from either control. Aln calluses also contained more mineral (bone mineral content [BMC]) than all other groups. By 16 weeks postfracture, OVX calluses were smaller than at 6 weeks and the dimensions for Aln had not changed. Aln had higher BMC and ultimate load than OVX, EE2, and Rlx. EE2 and Rlx had similar biomechanical properties, which were similar to sham. Interestingly, OVX and Aln animals were heavier than other groups at all time points; therefore, ultimate load was normalized by body weight to show no significant differences in strength of the whole callus between groups at either 6 weeks or 16 weeks postfracture. However, Aln strongly suppressed remodeling of the callus, resulting in the highest content of woven bone, persistent visibility of the original fracture line, and lowest content of lamellar bone, compared with other groups. Therefore, the larger Aln callus appeared to be a remarkable, morphological adaptation to secure the fracture with inferior material. In conclusion, OVX-stimulated bone turnover resulted in the fastest progression of fracture repair that was most delayed with Aln treatment, consistent with marked suppression of bone resorption and formation activity. Estrogen and Rlx had similar effects that were generally similar to sham, indicating that mild suppression of bone turnover with these agents has insignificant effects on the progression of fracture repair.", "Atypical femoral fractures differ from ordinary femoral diaphyseal or subtrochanteric fractures in several aspects. Although several authors have reported the results of surgical treatment for atypical femoral fractures, the rate of complications (e.g., delayed union, nonunion, fixation failure, and reoperation) is still high. Therefore, we reviewed principles of surgical treatment and describe useful methods for overcoming femoral bowing in these high-risk patients.", "Alendronate, an inhibitor of bone resorption, is widely used in osteoporosis treatment. However, concerns have been raised about potential oversuppression of bone turnover during long-term use. We report on nine patients who sustained spontaneous nonspinal fractures while on alendronate therapy, six of whom displayed either delayed or absent fracture healing for 3 months to 2 yr during therapy. Histomorphometric analysis of the cancellous bone showed markedly suppressed bone formation, with reduced or absent osteoblastic surface in most patients. Osteoclastic surface was low or low-normal in eight patients, and eroded surface was decreased in four. Matrix synthesis was markedly diminished, with absence of double-tetracycline label and absent or reduced single-tetracycline label in all patients. The same trend was seen in the intracortical and endocortical surfaces. Our findings raise the possibility that severe suppression of bone turnover may develop during long-term alendronate therapy, resulting in increased susceptibility to, and delayed healing of, nonspinal fractures. Although coadministration of estrogen or glucocorticoids appears to be a predisposing factor, this apparent complication can also occur with monotherapy. Our observations emphasize the need for increased awareness and monitoring for the potential development of excessive suppression of bone turnover during long-term alendronate therapy.", "Previous studies have shown that alendronate can increase bone mineral density (BMD) and prevent radiographically defined (morphometric) vertebral fractures. The Fracture Intervention Trial aimed to investigate the effect of alendronate on the risk of morphometric as well as clinically evident fractures in postmenopausal women with low bone mass. Women aged 55-81 with low femoral-neck BMD were enrolled in two study groups based on presence or absence of an existing vertebral fracture. Results for women with at least one vertebral fracture at baseline are reported here. 2027 women were randomly assigned placebo (1005) or alendronate (1022) and followed up for 36 months. The dose of alendronate (initially 5 mg daily) was increased (to 10 mg daily) at 24 months, with maintenance of the double blind. Lateral spine radiography was done at baseline and at 24 and 36 months. New vertebral fractures, the primary endpoint, were defined by morphometry as a decrease of 20% (and at least 4 mm) in at least one vertebral height between the baseline and latest follow-up radiograph. Non-spine clinical fractures were confirmed by radiographic reports. New symptomatic vertebral fractures were based on self-report and confirmed by radiography. Follow-up radiographs were obtained for 1946 women (98% of surviving participants). 78 (8.0%) of women in the alendronate group had one or more new morphometric vertebral fractures compared with 145 (15.0%) in the placebo group (relative risk 0.53 [95% Cl 0.41-0.68]). For clinically apparent vertebral fractures, the corresponding numbers were 23 (2.3%) alendronate and 50 (5.0%) placebo (relative hazard 0.45 [0.27-0.72]). The risk of any clinical fracture, the main secondary endpoint, was lower in the alendronate than in the placebo group (139 [13.6%] vs 183 [18.2%]; relative hazard 0.72 [0.58-0.90]). The relative hazards for hip fracture and wrist fracture for alendronate versus placebo were 0.49 (0.23-0.99) and 0.52 (0.31-0.87). There was no significant difference between the groups in numbers of adverse experiences, including upper-gastrointestinal disorders. We conclude that among women with low bone mass and existing vertebral fractures, alendronate is well tolerated and substantially reduces the frequency of morphometric and clinical vertebral fractures, as well as other clinical fractures.", "Bisphosphonates are effective in reducing hip and osteoporotic fractures. However, concerns about atypical femur fractures have contributed to substantially decreased bisphosphonate use, and the incidence of hip fractures may be increasing. Important uncertainties remain regarding the association between atypical femur fractures and bisphosphonates and other risk factors. We studied women 50 years of age or older who were receiving bisphosphonates and who were enrolled in the Kaiser Permanente Southern California health care system; women were followed from January 1, 2007, to November 30, 2017. The primary outcome was atypical femur fracture. Data on risk factors, including bisphosphonate use, were obtained from electronic health records. Fractures were radiographically adjudicated. Multivariable Cox models were used. The risk-benefit profile was modeled for 1 to 10 years of bisphosphonate use to compare associated atypical fractures with other fractures prevented. Among 196,129 women, 277 atypical femur fractures occurred. After multivariable adjustment, the risk of atypical fracture increased with longer duration of bisphosphonate use: the hazard ratio as compared with less than 3 months increased from 8.86 (95% confidence interval [CI], 2.79 to 28.20) for 3 years to less than 5 years to 43.51 (95% CI, 13.70 to 138.15) for 8 years or more. Other risk factors included race (hazard ratio for Asians vs. Whites, 4.84; 95% CI, 3.57 to 6.56), height, weight, and glucocorticoid use. Bisphosphonate discontinuation was associated with a rapid decrease in the risk of atypical fracture. Decreases in the risk of osteoporotic and hip fractures during 1 to 10 years of bisphosphonate use far outweighed the increased risk of atypical fracture among Whites but less so among Asians. After 3 years, 149 hip fractures were prevented and 2 bisphosphonate-associated atypical fractures occurred in Whites, as compared with 91 and 8, respectively, in Asians. The risk of atypical femur fracture increased with longer duration of bisphosphonate use and rapidly decreased after bisphosphonate discontinuation. Asians had a higher risk than Whites. The absolute risk of atypical femur fracture remained very low as compared with reductions in the risk of hip and other fractures with bisphosphonate treatment. (Funded by Kaiser Permanente and others.).", "Many risk factors for hip fractures have been suggested but have not been evaluated in a comprehensive prospective study. We assessed potential risk factors, including bone mass, in 9516 white women 65 years of age or older who had had no previous hip fracture. We then followed these women at 4-month intervals for an average of 4.1 years to determine the frequency of hip fracture. All reports of hip fractures were validated by review of x-ray films. During the follow-up period, 192 women had first hip fractures not due to motor vehicle accidents. In multivariable age-adjusted analyses, a maternal history of hip fracture doubled the risk of hip fracture (relative risk, 2.0; 95 percent confidence interval, 1.4 to 2.9), and the increase in risk remained significant after adjustment for bone density. Women who had gained weight since the age of 25 had a lower risk. The risk was higher among women who had previous fractures of any type after the age of 50, were tall at the age of 25, rated their own health as fair or poor, had previous hyperthyroidism, had been treated with long-acting benzodiazepines or anticonvulsant drugs, ingested greater amounts of caffeine, or spent four hours a day or less on their feet. Examination findings associated with an increased risk included the inability to rise from a chair without using one's arms, poor depth perception, poor contrast sensitivity, and tachycardia at rest. Low calcaneal bone density was also an independent risk factor. The incidence of hip fracture ranged from 1.1 (95 percent confidence interval, 0.5 to 1.6) per 1,000 woman-years among women with no more than two risk factors and normal calcaneal bone density for their age to 27 (95 percent confidence interval, 20 to 34) per 1,000 woman-years among those with five or more risk factors and bone density in the lowest third for their age. Women with multiple risk factors and low bone density have an especially high risk of hip fracture. Maintaining body weight, walking for exercise, avoiding long-acting benzodiazepines, minimizing caffeine intake, and treating impaired visual function are among the steps that may decrease the risk." ]
SET domain-containing 5 enzymatic activity and substrate specificity	SET domain-containing 5 (SETD5) is an uncharacterized member of the protein lysine methyltransferase family and is best known for its transcription machinery by methylating histone H3 on lysine 36 (H3K36). These well-characterized functions of SETD5 are transcription regulation, euchromatin formation, and RNA elongation and splicing. SETD5 is frequently mutated and hyperactive in both human neurodevelopmental disorders and cancer, and could be down-regulated by degradation through the ubiquitin-proteasome pathway, but the biochemical mechanisms underlying such dysregulation are rarely understood. Herein, we provide an update on the particularities of SETD5 enzymatic activity and substrate specificity concerning its biological importance, as well as its molecular and cellular impact on normal physiology and disease, with potential therapeutic options.	[ "Mouse mutants with an impaired DNA damage response frequently exhibit a set of remarkably similar defects in the HSPC compartment that are of largely unknown molecular basis. Using Mixed-Lineage-Leukemia-5 (Mll5)-deficient mice as prototypical examples, we have identified a mechanistic pathway linking DNA damage and HSPC malfunction. We show that Mll5 deficiency results in accumulation of DNA damage and reactive oxygen species (ROS) in HSPCs. Reduction of ROS efficiently reverses hematopoietic defects, establishing ROS as a major cause of impaired HSPC function. The Ink4a/Arf locus also contributes to HSPC phenotypes, at least in part via promotion of ROS. Strikingly, toxic ROS levels in Mll5-/- mice are critically dependent on type 1 interferon (IFN-1) signaling, which triggers mitochondrial accumulation of full-length Bid. Genetic inactivation of Bid diminishes ROS levels and reverses HSPC defects in Mll5-/- mice. Overall, therefore, our findings highlight an unexpected IFN-1 > Bid > ROS pathway underlying DNA damage-associated HSPC malfunction.", "Chromatin dynamics are central to the regulation of gene expression and genome stability, particularly in the presence of environmental signals or stresses that prompt rapid reprogramming of the genome to promote survival or differentiation. While numerous chromatin regulators have been implicated in modulating cellular responses to stress, gaps in our mechanistic understanding of chromatin-based changes during stress suggest that additional proteins are likely critical to these responses and the molecular details underlying their activities are unclear in many cases. We recently identified a role for the relatively uncharacterized SET domain protein Set4 in promoting cell survival during oxidative stress in Saccharomyces cerevisiae. Set4 is a member of the Set3 subfamily of SET domain proteins which are defined by the presence of a PHD finger and divergent SET domain sequences. Here, we integrate our new observations on the function of Set4 with known roles for other related family members, including yeast Set3, fly UpSET and mammalian proteins MLL5 and SETD5. We discuss outstanding questions regarding the molecular mechanisms by which these proteins control gene expression and their potential contributions to cellular responses to environmental stress.", "miR126-5p is processed from the miR126-3p/-5p duplex, which is expressed in endothelial cells and gives rise to the guide strand miR126-3p and the passenger strand miR126-5p. miR126-3p has prominent roles in vascular development and diseases, whereas the expression and physiological functions of miR126-5p are unknown. The purpose of this study was to evaluate the expression and role of miR126-5p in blood vessel endothelial cells. miR126-5p is mostly expressed in blood vessel endothelial cells in vivo and in vitro. Gain- and loss-of-function approaches revealed that miR126-5p promotes leucocyte adhesion and represses leucocyte transendothelial migration. Two distinct target genes of miR126-5p in endothelial cells were identified: the activated leucocyte cell adhesion molecule (ALCAM) gene which codes for an adhesion molecule involved in leucocyte transendothelial migration and SetD5, a gene with previously unknown functions. Using either a blocking antibody or target protectors which specifically disrupt the miRNA/mRNA target pairing, we showed that miR126-5p promotes leucocyte adhesion by controlling the expression of SetD5 and represses transendothelial migration via the regulation of ALCAM. miR126-5p controls ALCAM and SetD5 expression in vivo in separate tissues and regulates leucocyte infiltration into inflamed lungs by repressing ALCAM expression. miR126-5p is a functional, endothelial-enriched microRNA that participates in the control of leucocyte trafficking by regulating the expression of ALCAM and SetD5.", "A complex interplay between multiple chromatin modifiers is critical for cells to regulate chromatin structure and accessibility during essential DNA-templated processes such as transcription. However, the coordinated activities of these chromatin modifiers in the regulation of gene expression are not fully understood. We previously determined that the budding yeast histone H4 methyltransferase Set5 functions together with Set1, the H3K4 methyltransferase, in specific cellular contexts. Here, we sought to understand the relationship between these evolutionarily conserved enzymes in the regulation of gene expression. We generated a comprehensive genetic interaction map of the functionally uncharacterized Set5 methyltransferase and expanded the existing genetic interactome of the global chromatin modifier Set1, revealing functional overlap of the two enzymes in chromatin-related networks, such as transcription. Furthermore, gene expression profiling via RNA-Seq revealed an unexpected synergistic role of Set1 and Set5 in repressing transcription of Ty transposable elements and genes located in subtelomeric regions. This study uncovers novel pathways in which the methyltransferase Set5 participates and, more importantly, reveals a partnership between Set1 and Set5 in transcriptional repression near repetitive DNA elements in budding yeast. Together, our results define a new functional relationship between histone H3 and H4 methyltransferases, whose combined activity may be implicated in preserving genomic integrity.", "When yeast cells are challenged by a fluctuating environment, signaling networks activate differentiation programs that promote their individual or collective survival. These programs include the initiation of meiotic sporulation, the formation of filamentous growth structures, and the activation of programmed cell death pathways. The establishment and maintenance of these distinct cell fates are driven by massive gene expression programs that promote the necessary changes in morphology and physiology. While these genomic reprogramming events depend on a specialized network of transcription factors, a diverse set of chromatin regulators, including histone-modifying enzymes, chromatin remodelers, and histone variants, also play essential roles. Here, we review the broad functions of histone modifications in initiating cell fate transitions, with particular focus on their contribution to the control of expression of key genes required for the differentiation programs and chromatin reorganization that accompanies these cell fates.", "Arginine methylation is a common post-translational modification functioning as an epigenetic regulator of transcription and playing key roles in pre-mRNA splicing, DNA damage signaling, mRNA translation, cell signaling, and cell fate decision. Recently, a wealth of studies using transgenic mouse models and selective PRMT inhibitors helped define physiological roles for protein arginine methyltransferases (PRMTs) linking them to diseases such as cancer and metabolic, neurodegenerative, and muscular disorders. This review describes the recent molecular advances that have been uncovered in normal and diseased mammalian cells.", "Packaging of eukaryotic genomes into chromatin has wide-ranging effects on gene transcription. Curiously, it is commonly observed that deletion of a global chromatin regulator affects expression of only a limited subset of genes bound to or modified by the regulator in question. However, in many single-gene studies it has become clear that chromatin regulators often do not affect steady-state transcription, but instead are required for normal transcriptional reprogramming by environmental cues. We therefore have systematically investigated the effects of 83 histone mutants, and 119 gene deletion mutants, on induction/repression dynamics of 170 transcripts in response to diamide stress in yeast. Importantly, we find that chromatin regulators play far more pronounced roles during gene induction/repression than they do in steady-state expression. Furthermore, by jointly analyzing the substrates (histone mutants) and enzymes (chromatin modifier deletions) we identify specific interactions between histone modifications and their regulators. Combining these functional results with genome-wide mapping of several histone marks in the same time course, we systematically investigated the correspondence between histone modification occurrence and function. We followed up on one pathway, finding that Set1-dependent H3K4 methylation primarily acts as a gene repressor during multiple stresses, specifically at genes involved in ribosome biosynthesis. Set1-dependent repression of ribosomal genes occurs via distinct pathways for ribosomal protein genes and ribosomal biogenesis genes, which can be separated based on genetic requirements for repression and based on chromatin changes during gene repression. Together, our dynamic studies provide a rich resource for investigating chromatin regulation, and identify a significant role for the \"activating\" mark H3K4me3 in gene repression.", "Obsessive-compulsive disorder (OCD) is a chronic anxiety disorder with a substantial genetic basis and a broadly undiscovered etiology. Recent studies of de novo mutation (DNM) exome-sequencing studies for OCD have reinforced the hypothesis that rare variation contributes to the risk. We performed, to our knowledge, the first whole-genome sequencing on 53 parent-offspring families with offspring affected with OCD to investigate all rare de novo variants and insertions/deletions. We observed higher mutation rates in promoter-anchored chromatin loops (empirical P = 0.0015) and regions with high frequencies of histone marks (empirical P = 0.0001). Mutations affecting coding regions were significantly enriched within coexpression modules of genes involved in chromatin modification during human brain development. Four genes—SETD5, KDM3B, ASXL3, and FBL—had strong aggregated evidence and functionally converged on transcription’s epigenetic regulation, suggesting an important OCD risk mechanism. Our data characterized different genome-wide DNMs and highlighted the contribution of chromatin modification in the etiology of OCD.", "SET domain-containing 5 (SETD5) is an uncharacterized member of the protein lysine methyltransferase family. Although it was reported that SETD5 gene mutations are associated with the several types of human cancer, its functional role in esophageal squamous cell carcinoma (ESCC) progression has not been fully elucidated. In the present study, we used tissue samples from 147 patients with ESCC and ESCC cell lines to determine the clinicopathological significance of SETD5 in ESCC and its effects on ESCC stemness. We performed immunohistochemical staining, immunofluorescence imaging, and tumor sphere formation, colony formation, flow cytometry, wound healing, Transwell, and western blotting assays. SETD5 expression was upregulated in ESCC tissue and associated with primary tumor (pT) stage, clinical stage, lymph node metastasis, shorter overall survival rate, and disease-free survival rate. Cox regression analyses indicated that SETD5 is an independent poor prognostic factor of ESCC. In addition, SETD5 expression was correlated with cancer stemness-related protein, hypoxia-inducible factor-1α (HIF-1α), and CD68 expression. Moreover, immunofluorescence analysis revealed that SETD5 was co-localized with CD44 and SOX2 in TE10 and TE11 cells and that exposing cells to cobalt chloride increased HIF-1α, SETD5, and stemness-related protein expression in a time-dependent manner. Furthermore, SETD5 expression was significantly correlated with the expression of cell cycle-related genes and PI3K/Akt signaling pathway-related proteins. Finally, knocking down SETD5 downregulated the expression of stemness-related and PI3K/Akt signaling pathway proteins, while inhibiting tumor spheroid formation, cell proliferation, migration, and invasion in ESCC cells. These results indicate that SETD5 expression is associated with cancer stemness and that SETD5 is a potential prognostic biomarker and therapeutic target for ESCC." ]
T-cell exhaustion and stemness in cancer immunotherapy	T cells play a crucial role in the regulation of immune response and are integral to the efficacy of cancer immunotherapy. Because immunotherapy has emerged as a promising treatment for cancer, increasing attention has been focused on the differentiation and function of T cells in immune response. In this review, we describe the research progress on T-cell exhaustion and stemness in the field of cancer immunotherapy and summarize advances in potential strategies to intervene and treat chronic infection and cancer by reversing T-cell exhaustion and maintaining and increasing T-cell stemness. Moreover, we discuss therapeutic strategies to overcome T-cell immunodeficiency in the tumor microenvironment and promote continuous breakthroughs in the anticancer activity of T cells.	[ "Stem memory T cells (TSCM) have been described in mice, non-human primates and in humans, constituting approximately 2-4% of the total CD4(+) and CD8(+) T-cell population in the periphery. TSCM represent the earliest and long-lasting developmental stage of memory T cells, displaying stem cell-like properties, and exhibiting a gene profile between naïve and central memory T cells. Their self-renewal capacity and long-term survival has sparked interest in the cancer and human immunodeficiency virus (HIV) fields. How and when the formation of TSCM occurs during the immune response to pathogens and the therapeutic potential of these cells are currently being investigated. This review will explore the potential role of TSCM to be used as, or targeted by, immunotherapies and vaccines for treatment of cancer and HIV.", "Memory stem T cells (T(SCM)) constitute a long-lived, self-renewing lymphocyte population essential for the maintenance of functional immunity. The hallmarks of HIV-1 pathogenesis are CD4(+) T cell depletion and abnormal cellular activation. We investigated the impact of HIV-1 infection on the T(SCM) compartment, as well as any protective role these cells may have in disease progression, by characterizing this subset in a cohort of 113 subjects with various degrees of viral control on and off highly active antiretroviral therapy (HAART). We observed that the frequency of CD8(+) T(SCM) was decreased in all individuals with chronic, untreated HIV-1 infection and that HAART had a restorative effect on this subset. In contrast, natural controllers of HIV-1 had the highest absolute number of CD4(+) T(SCM) cells among all of the infected groups. The frequency of CD4(+) T(SCM) predicted higher CD8(+) T(SCM) frequencies, consistent with a role for the CD4(+) subset in helping to maintain CD8(+) memory T cells. In addition, T(SCM) appeared to be progenitors for effector T cells (TEM), as these two compartments were inversely correlated. Increased frequencies of CD8(+) T(SCM) predicted lower viral loads, higher CD4(+) counts, and less CD8(+) T cell activation. Finally, we found that T(SCM) express the mucosal homing integrin α4β7 and can be identified in gut-associated lymphoid tissue (GALT). The frequency of mucosal CD4(+) T(SCM) was inversely correlated with that in the blood, potentially reflecting the ability of these self-renewing cells to migrate to a crucial site of ongoing viral replication and CD4(+) T cell depletion. HIV-1 infection leads to profound impairment of the immune system. T(SCM) constitute a recently identified lymphocyte subset with stem cell-like qualities, including the ability to generate other memory T cell subtypes, and are therefore likely to play an important role in controlling viral infection. We investigated the relationship between the size of the CD8(+) T(SCM) compartment and HIV-1 disease progression in a cohort of chronically infected individuals. Our results suggest that HAART restores a normal frequency of CD8(+) T(SCM) and that the natural preservation of this subset in the setting of untreated HIV-1 infection is associated with improved viral control and immunity. Therefore, the CD8(+) T(SCM) population may represent a correlate of protection in chronic HIV-1 infection that is directly relevant to the design of T cell-based vaccines, adoptive immunotherapy approaches, or the pharmacologic induction of T(SCM).", "Chimeric antigen receptor (CAR) T cells have induced remarkable antitumor responses in B cell malignancies. Some patients do not respond because of T cell deficiencies that hamper the expansion, persistence, and effector function of these cells. We used longitudinal immune profiling to identify phenotypic and pharmacodynamic changes in CD19-directed CAR T cells in patients with chronic lymphocytic leukemia (CLL). CAR expression maintenance was also investigated because this can affect response durability. CAR T cell failure was accompanied by preexisting T cell-intrinsic defects or dysfunction acquired after infusion. In a small subset of patients, CAR silencing was observed coincident with leukemia relapse. Using a small molecule inhibitor, we demonstrated that the bromodomain and extra-terminal (BET) family of chromatin adapters plays a role in downregulating CAR expression. BET protein blockade also ameliorated CAR T cell exhaustion as manifested by inhibitory receptor reduction, enhanced metabolic fitness, increased proliferative capacity, and enriched transcriptomic signatures of T cell reinvigoration. BET inhibition decreased levels of the TET2 methylcytosine dioxygenase, and forced expression of the TET2 catalytic domain eliminated the potency-enhancing effects of BET protein targeting in CAR T cells, providing a mechanism linking BET proteins and T cell dysfunction. Thus, modulating BET epigenetic readers may improve the efficacy of cell-based immunotherapies.", "CD8 T cell responses play an important role in protection against intracellular parhogens. Memory CD8 T cells mediate rapid clearance of pathogens upon secondary infection owing to their elevated frequency, ready localization to peripheral sites of infection and their ability to rapidly expand and mount effector functions. Such potent long-lasting protective memory CD8 T cells develop in acute infections where antigen is effectively cleared. In contrast, chronic infections with persistently high viral loads are characterized by CD8 T-cell dysfunction. In this chapter we present our current understanding of signals and mechanisms that regulate the development of functional long-lived memory CD8 T cells during acute infections. This is discussed in the context of proposed models of memory differentiation and compared with CD8 T-cell exhaustion and altered T-cell homeostasis, as occurs during persistent viral infections.", "Memory stem T cells (TSCM) have been proposed as key determinants of immunologic memory. However, their exact contribution to a mounting immune response, as well as the mechanisms and timing of their in vivo generation, are poorly understood. We longitudinally tracked TSCM dynamics in patients undergoing haploidentical hematopoietic stem cell transplantation (HSCT), thereby providing novel hints on the contribution of this subset to posttransplant immune reconstitution in humans. We found that donor-derived TSCM are highly enriched early after HSCT. We showed at the antigen-specific and clonal level that TSCM lymphocytes can differentiate directly from naive precursors infused within the graft and that the extent of TSCM generation might correlate with interleukin 7 serum levels. In vivo fate mapping through T-cell receptor sequencing allowed defining the in vivo differentiation landscapes of human naive T cells, supporting the notion that progenies of single naive cells embrace disparate fates in vivo and highlighting TSCM as relevant novel players in the diversification of immunological memory after allogeneic HSCT.", "T cell differentiation requires appropriate regulation of DNA methylation. In this article, we demonstrate that the methylcytosine dioxygenase ten-eleven translocation (TET)2 regulates CD8+ T cell differentiation. In a murine model of acute viral infection, TET2 loss promotes early acquisition of a memory CD8+ T cell fate in a cell-intrinsic manner without disrupting Ag-driven cell expansion or effector function. Upon secondary recall, TET2-deficient memory CD8+ T cells demonstrate superior pathogen control. Genome-wide methylation analysis identified a number of differentially methylated regions in TET2-deficient versus wild-type CD8+ T cells. These differentially methylated regions did not occur at the loci of differentially expressed memory markers; rather, several hypermethylated regions were identified in known transcriptional regulators of CD8+ T cell memory fate. Together, these data demonstrate that TET2 is an important regulator of CD8+ T cell fate decisions.", "Suppression of T-cell responses by host-derived regulatory factors is a key event leading to viral persistence. Antibody blockade of either IL-10 or programmed death-ligand 1 (PD-L1) during viral persistence enhances T-cell function and reduces viral titers. Because blockade of these immunoregulatory networks represents a powerful approach to establish immune control during persistent infection, it is important to determine whether these immunoinhibitory factors act independently or jointly and if combined blockade of these factors further enhances T-cell immunity and viral clearance. Herein, we demonstrate that the IL-10 and PD-L1 immunosuppressive pathways are mechanistically distinct. As a result, simultaneous blockade of IL-10 and PD-L1 was significantly more effective in restoring antiviral T-cell responses than blockade of either alone, and led to substantially enhanced control of an established persistent viral infection. Thus, combinatorial blockade of multiple immune-regulatory molecules may ultimately restore the T-cell responses required to tip the balance from viral persistence to immune-mediated control or elimination of persistent infection.", "Response rates to immunotherapy in solid tumors remain low due in part to the elevated prevalence of terminally exhausted T cells, a hypofunctional differentiation state induced through persistent antigen and stress signaling. However, the mechanisms promoting progression to terminal exhaustion in the tumor remain undefined. Using the low-input chromatin immunoprecipitation sequencing method CUT&RUN, we profiled the histone modification landscape of tumor-infiltrating CD8+ T cells throughout differentiation. We found that terminally exhausted T cells had unexpected chromatin features that limit their transcriptional potential. Terminally exhausted T cells had a substantial fraction of active chromatin, including active enhancers enriched for bZIP/AP-1 transcription factor motifs that lacked correlated gene expression, which was restored by immunotherapeutic costimulatory signaling. Reduced transcriptional potential was also driven by an increase in histone bivalency, which we linked directly to hypoxia exposure. Enforced expression of the hypoxia-insensitive histone demethylase Kdm6b was sufficient to overcome hypoxia, increase function, and promote antitumor immunity. Our study reveals the specific epigenetic changes mediated by histone modifications during T cell differentiation that support exhaustion in cancer, highlighting that their altered function is driven by improper costimulatory signals and environmental factors. These data suggest that even terminally exhausted T cells may remain competent for transcription in settings of increased costimulatory signaling and reduced hypoxia.", "Cancer is associated with higher morbidity and mortality and is the second leading cause of death in the US. Further, in some nations, cancer has overtaken heart disease as the leading cause of mortality. Identification of molecular mechanisms by which cancerous cells evade T cell-mediated cytotoxic damage has led to the modern era of immunotherapy in cancer treatment. Agents that release these immune brakes have shown activity to recover dysfunctional T cells and regress various cancer. Both cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) and Programmed Death-1 (PD-1) play their role as physiologic brakes on unrestrained cytotoxic T effector function. CTLA-4 (CD 152) is a B7/CD28 family; it mediates immunosuppression by indirectly diminishing signaling through the co-stimulatory receptor CD28. Ipilimumab is the first and only FDA-approved CTLA-4 inhibitor; PD-1 is an inhibitory transmembrane protein expressed on T cells, B cells, Natural Killer cells (NKs), and Myeloid-Derived Suppressor Cells (MDSCs). Programmed Death-Ligand 1 (PD-L1) is expressed on the surface of multiple tissue types, including many tumor cells and hematopoietic cells. PD-L2 is more restricted to hematopoietic cells. Blockade of the PD-1 /PDL-1 pathway can enhance anti-tumor T cell reactivity and promotes immune control over the cancerous cells. Since the FDA approval of ipilimumab (human IgG1 k anti-CTLA-4 monoclonal antibody) in 2011, six more immune checkpoint inhibitors (ICIs) have been approved for cancer therapy. PD-1 inhibitors nivolumab, pembrolizumab, cemiplimab and PD-L1 inhibitors atezolizumab, avelumab, and durvalumab are in the current list of the approved agents in addition to ipilimumab. In this review paper, we discuss the role of each immune checkpoint inhibitor (ICI), the landmark trials which led to their FDA approval, and the strength of the evidence per National Comprehensive Cancer Network (NCCN), which is broadly utilized by medical oncologists and hematologists in their daily practice.", "Long-lived memory T cells are able to persist in the host in the absence of antigen; however, the mechanism by which they are maintained is not well understood. Recently, a subset of human T cells, stem cell memory T cells (TSCM cells), was shown to be self-renewing and multipotent, thereby providing a potential reservoir for T cell memory throughout life. However, their in vivo dynamics and homeostasis still remain to be defined due to the lack of suitable animal models. We identified T cells with a TSCM phenotype and stem cell-like properties in nonhuman primates. These cells were the least-differentiated memory subset, were functionally distinct from conventional memory cells, and served as precursors of central memory. Antigen-specific TSCM cells preferentially localized to LNs and were virtually absent from mucosal surfaces. They were generated in the acute phase of viral infection, preferentially survived in comparison with all other memory cells following elimination of antigen, and stably persisted for the long term. Thus, one mechanism for maintenance of long-term T cell memory derives from the unique homeostatic properties of TSCM cells. Vaccination strategies designed to elicit durable cellular immunity should target the generation of TSCM cells." ]
Vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and Graves disease	Vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) which were approved for emergency use have been administered on a large scale globally to contain the pandemic coronavirus disease 2019 (COVID-19) and to save lives. Vaccine safety is one of the issues under surveillance and a possible correlation between vaccines and thyroid function has been reported. However, reports of the impact of coronavirus vaccines on those with Graves' disease (GD) are rare.	[ "The autoimmune/inflammatory syndrome induced by adjuvants (ASIA) comprises four entities, including the postvaccination phenomenon, which appears after being exposed to adjuvants in vaccines that increase the immune response. There is limited information about autoimmune endocrine diseases and ASIA after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccination. Two female health care workers received a SARS-CoV-2 vaccine, and three days later developed clinical manifestations of thyroid hyperactivity, with increased thyroid hormone levels on thyroid function tests, suppressed thyroid-stimulating hormone, and elevated antithyroid antibodies. Vaccines have been shown to trigger an immune response that leads to a broad spectrum of autoimmune diseases, including autoimmune thyroid disease. Our patients met the diagnostic criteria for ASIA; they were exposed to an adjuvant (vaccine), and they developed clinical manifestations of thyroid hyperfunction within a few days, with the appearance of antithyroid antibodies, despite being healthy before vaccination. Graves' disease can occur after SARS-CoV-2 vaccination.", "West Nile virus (WNV) is a flavivirus that causes meningitis and encephalitis. There are no licensed vaccines to prevent WNV in humans. The safety and immunogenicity of a first-generation WNV DNA vaccine was demonstrated in a clinical trial and a similar DNA vaccine has been licensed for use in horses. A DNA vaccine encoding the protein premembrane and the E glycoproteins of the NY99 strain of WNV under the transcriptional control of the CMV/R promoter was evaluated in an open-label study in 30 healthy adults. Half of the subjects were age 18-50 years and half were age 51-65 years. Immune responses were assessed by enzyme-linked immunosorbent assay, neutralization assays, intracellular cytokine staining, and ELISpot. The 3-dose vaccine regimen was safe and well tolerated. Vaccine-induced T cell and neutralizing antibody responses were detected in the majority of subjects. The antibody responses seen in the older age group were of similar frequency, magnitude, and duration as those seen in the younger cohort. Neutralizing antibody responses to WNV were elicited by DNA vaccination in humans, including in older individuals, where responses to traditional vaccine approaches are often diminished. This DNA vaccine elicited T cell responses of greater magnitude when compared with an earlier-generation construct utilizing a CMV promoter. NCT00300417.", "The mode of acquisition and causes for the variable clinical spectrum of coronavirus disease 2019 (COVID-19) remain unknown. We utilized a reverse genetics system to generate a GFP reporter virus to explore severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pathogenesis and a luciferase reporter virus to demonstrate sera collected from SARS and COVID-19 patients exhibited limited cross-CoV neutralization. High-sensitivity RNA in situ mapping revealed the highest angiotensin-converting enzyme 2 (ACE2) expression in the nose with decreasing expression throughout the lower respiratory tract, paralleled by a striking gradient of SARS-CoV-2 infection in proximal (high) versus distal (low) pulmonary epithelial cultures. COVID-19 autopsied lung studies identified focal disease and, congruent with culture data, SARS-CoV-2-infected ciliated and type 2 pneumocyte cells in airway and alveolar regions, respectively. These findings highlight the nasal susceptibility to SARS-CoV-2 with likely subsequent aspiration-mediated virus seeding to the lung in SARS-CoV-2 pathogenesis. These reagents provide a foundation for investigations into virus-host interactions in protective immunity, host susceptibility, and virus pathogenesis.", "Testing of vaccine candidates to prevent infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in an older population is important, since increased incidences of illness and death from coronavirus disease 2019 (Covid-19) have been associated with an older age. We conducted a phase 1, dose-escalation, open-label trial of a messenger RNA vaccine, mRNA-1273, which encodes the stabilized prefusion SARS-CoV-2 spike protein (S-2P) in healthy adults. The trial was expanded to include 40 older adults, who were stratified according to age (56 to 70 years or ≥71 years). All the participants were assigned sequentially to receive two doses of either 25 μg or 100 μg of vaccine administered 28 days apart. Solicited adverse events were predominantly mild or moderate in severity and most frequently included fatigue, chills, headache, myalgia, and pain at the injection site. Such adverse events were dose-dependent and were more common after the second immunization. Binding-antibody responses increased rapidly after the first immunization. By day 57, among the participants who received the 25-μg dose, the anti-S-2P geometric mean titer (GMT) was 323,945 among those between the ages of 56 and 70 years and 1,128,391 among those who were 71 years of age or older; among the participants who received the 100-μg dose, the GMT in the two age subgroups was 1,183,066 and 3,638,522, respectively. After the second immunization, serum neutralizing activity was detected in all the participants by multiple methods. Binding- and neutralizing-antibody responses appeared to be similar to those previously reported among vaccine recipients between the ages of 18 and 55 years and were above the median of a panel of controls who had donated convalescent serum. The vaccine elicited a strong CD4 cytokine response involving type 1 helper T cells. In this small study involving older adults, adverse events associated with the mRNA-1273 vaccine were mainly mild or moderate. The 100-μg dose induced higher binding- and neutralizing-antibody titers than the 25-μg dose, which supports the use of the 100-μg dose in a phase 3 vaccine trial. (Funded by the National Institute of Allergy and Infectious Diseases and others; mRNA-1273 Study ClinicalTrials.gov number, NCT04283461.)." ]
9,10-epoxy-13-hydroxy-octadecenoateinduced nociceptive responses in mice and rats	Psoriasis is characterized by intense pruritus, with a subset of individuals with psoriasis experiencing thermal hypersensitivity. However, the pathophysiology of thermal hypersensitivity in psoriasis and other skin conditions remains enigmatic. Linoleic acid is an omega-6 fatty acid that is concentrated in the skin, and oxidation of linoleic acid into metabolites with multiple hydroxyl and epoxide functional groups has been shown to play a role in skin barrier function. Previously, we identified several linoleic acid‒derived mediators that were more concentrated in psoriatic lesions, but the role of these lipids in psoriasis remains unknown. In this study, we report that two such compounds-9,10-epoxy-13-hydroxy-octadecenoate and 9,10,13-trihydroxy-octadecenoate-are present as free fatty acids and induce nociceptive behavior in mice but not in rats. By chemically stabilizing 9,10-epoxy-13-hydroxy-octadecenoate and 9,10,13-trihydroxy-octadecenoate through the addition of methyl groups, we observed pain and hypersensitization in mice. The nociceptive responses suggest an involvement of the TRPA1 channel, whereas hypersensitive responses induced by these mediators may require both TRPA1 and TRPV1 channels. Furthermore, we showed that 9,10,13-trihydroxy-octadecenoate‒induced calcium transients in sensory neurons are mediated through the Gβγ subunit of an unidentified G-protein coupled receptor (GPCR). Overall, mechanistic insights from this study will guide the development of potential therapeutic targets for the treatment of pain and hypersensitivity.	[ "β-Alanine, a popular supplement for muscle building, induces itch and tingling after consumption, but the underlying molecular and neural mechanisms are obscure. Here we show that, in mice, β-alanine elicited itch-associated behavior that requires MrgprD, a G-protein-coupled receptor expressed by a subpopulation of primary sensory neurons. These neurons exclusively innervate the skin, respond to β-alanine, heat, and mechanical noxious stimuli but do not respond to histamine. In humans, intradermally injected β-alanine induced itch but neither wheal nor flare, suggesting that the itch was not mediated by histamine. Thus, the primary sensory neurons responsive to β-alanine are likely part of a histamine-independent itch neural circuit and a target for treating clinical itch that is unrelieved by anti-histamines.", "Cutaneous C-unmyelinated MRGPRD+ free nerve endings and C-LTMRs innervating hair follicles convey two opposite aspects of touch sensation: a sensation of pain and a sensation of pleasant touch. The molecular mechanisms underlying these diametrically opposite functions are unknown. Here, we used a mouse model that genetically marks C-LTMRs and MRGPRD+ neurons in combination with fluorescent cell surface labeling, flow cytometry, and RNA deep-sequencing technology (RNA-seq). Cluster analysis of RNA-seq profiles of the purified neuronal subsets revealed 486 and 549 genes differentially expressed in MRGPRD-expressing neurons and C-LTMRs, respectively. We validated 48 MRGPD- and 68 C-LTMRs-enriched genes using a triple-staining approach, and the Cav3.3 channel, found to be exclusively expressed in C-LTMRs, was validated using electrophysiology. Our study greatly expands the molecular characterization of C-LTMRs and suggests that this particular population of neurons shares some molecular features with Aβ and Aδ low-threshold mechanoreceptors.", "Unique features of sensory neuron subtypes are manifest by their distinct physiological and pathophysiological functions. Using patch-clamp electrophysiology, Ca2+ imaging, calcitonin gene-related peptide release assay from tissues, protein biochemistry approaches, and behavioral physiology on pain models, this study demonstrates the diversity of sensory neuron pathophysiology is due in part to subtype-dependent sensitization of TRPV1 and TRPA1. Differential sensitization is influenced by distinct expression of inflammatory mediators, such as prostaglandin E2 (PGE2), bradykinin (BK), and nerve growth factor (NGF) as well as multiple kinases, including protein kinase A (PKA) and C (PKC). However, the co-expression and interaction of TRPA1 with TRPV1 proved to be the most critical for differential sensitization of sensory neurons. We identified N- and C-terminal domains on TRPV1 responsible for TRPA1-TRPV1 (A1-V1) complex formation. Ablation of A1-V1 complex with dominant-negative peptides against these domains substantially reduced the sensitization of TRPA1, as well as BK- and CFA-induced hypersensitivity. These data indicate that often occurring TRP channel complexes regulate diversity in neuronal sensitization and may provide a therapeutic target for many neuroinflammatory pain conditions.", "TRPA1 and TRPV1 are crucial pain mediators, but how their interaction contributes to persistent pain is unknown. Here, we identify Tmem100 as a potentiating modulator of TRPA1-V1 complexes. Tmem100 is coexpressed and forms a complex with TRPA1 and TRPV1 in DRG neurons. Tmem100-deficient mice show a reduction in inflammatory mechanical hyperalgesia and TRPA1- but not TRPV1-mediated pain. Single-channel recording in a heterologous system reveals that Tmem100 selectively potentiates TRPA1 activity in a TRPV1-dependent manner. Mechanistically, Tmem100 weakens the association of TRPA1 and TRPV1, thereby releasing the inhibition of TRPA1 by TRPV1. A Tmem100 mutant, Tmem100-3Q, exerts the opposite effect; i.e., it enhances the association of TRPA1 and TRPV1 and strongly inhibits TRPA1. Strikingly, a cell-permeable peptide (CPP) containing the C-terminal sequence of Tmem100-3Q mimics its effect and inhibits persistent pain. Our study unveils a context-dependent modulation of the TRPA1-V1 complex, and Tmem100-3Q CPP is a promising pain therapy.", "Psoriasis is a chronic immune-mediated disease that represents a unique model for investigating inflammation at local and systemic levels. Bioactive lipid mediators (LMs) are potent compounds reported to play a role in the development and resolution of inflammation. Currently, it is not known to what extent these LMs are involved in psoriasis pathophysiology and related metabolic dysfunction. Here, we use targeted and untargeted liquid chromatography-tandem mass spectrometry approaches to quantify LMs in skin and peripheral blood from psoriasis patients and compared them with those of healthy individuals. Lesional psoriasis skin was abundant in arachidonic acid metabolites, as 8-, 12- and 15-hydroxyeicosatetraenoic acid, compared with adjacent nonlesional and skin from healthy individuals. Additionally, a linoleic acid-derived LM, 13-hydroxyoctadecadienoic acid, was significantly increased compared with healthy skin (607.9 ng/g vs. 5.4 ng/g, P = 0.001). These psoriasis skin differences were accompanied by plasma decreases in antioxidant markers, including glutathione, and impaired lipolysis characterized by lower concentrations of primary and secondary bile acids. In conclusion, our study shows that psoriasis skin and blood have disease-specific phenotype profiles of bioactive LMs represented by omega-6 fatty acid-oxidized derivatives. These findings provide insights into psoriasis pathophysiology that could potentially contribute to new biomarkers and therapeutics.", "Inflammation contributes to pain hypersensitivity through multiple mechanisms. Among the most well characterized of these is the sensitization of primary nociceptive neurons by arachidonic acid metabolites such as prostaglandins through G protein-coupled receptors. However, in light of the recent discovery that the nociceptor-specific ion channel transient receptor potential A1 (TRPA1) can be activated by exogenous electrophilic irritants through direct covalent modification, we reasoned that electrophilic carbon-containing A- and J-series prostaglandins, metabolites of prostaglandins (PG) E(2) and D(2), respectively, would excite nociceptive neurons through direct activation of TRPA1. Consistent with this prediction, the PGD(2) metabolite 15-deoxy-Delta(12,14)-prostaglandin J(2) (15dPGJ(2)) activated heterologously expressed human TRPA1 (hTRPA1-HEK), as well as a subset of chemosensitive mouse trigeminal neurons. The effects of 15dPGJ(2) on neurons were blocked by both the nonselective TRP channel blocker ruthenium red and the TRPA1 inhibitor (HC-030031), but unaffected by the TRPV1 blocker iodo-resiniferatoxin. In whole-cell patch-clamp studies on hTRPA1-HEK cells, 15dPGJ(2) evoked currents similar to equimolar allyl isothiocyanate (AITC) in the nominal absence of calcium, suggesting a direct mechanism of activation. Consistent with the hypothesis that TRPA1 activation required reactive electrophilic moieties, A- and J-series prostaglandins, and the isoprostane 8-iso-prostaglandin A(2)-evoked calcium influx in hTRPA1-HEK cells with similar potency and efficacy. It is noteworthy that this effect was not mimicked by their nonelectrophilic precursors, PGE(2) and PGD(2), or PGB(2), which differs from PGA(2) only in that its electrophilic carbon is rendered unreactive through steric hindrance. Taken together, these data suggest a novel mechanism through which reactive prostanoids may activate nociceptive neurons independent of prostaglandin receptors.", "Transient receptor potential A1 (TRPA1) ion channel senses a variety of noxious stimuli and is involved in nociception. Many TRPA1 agonists covalently modify the channel, which can lead to desensitization. The fate of modified TRPA1 and the mechanism of preserving its response to subsequent stimuli are not understood. Moreover, inflammatory signals sensitize TRPA1 by involving protein kinase A (PKA) and phospholipase C (PLC) through unknown means. We show that TRPA1-mediated nocifensive behavior can be sensitized in vivo via PKA/PLC signaling and by activating TRPA1 with the ligand mustard oil (MO). Interestingly, both stimuli increased TRPA1 membrane levels in vitro. Tetanus toxin attenuated the response to the second of two pulses of MO in neurons, suggesting that vesicle fusion increases functional surface TRPA1. Capacitance recordings suggest that MO can induce exocytosis. We propose that TRPA1 translocation to the membrane might represent one of the mechanisms controlling TRPA1 functionality upon acute activation or inflammatory signals." ]
Comparison of different extraction methods of Yizhiqingxin formula and its neuroprotective effects in a mouse model of Alzheimer's disease	This study compared different extraction methods of Yizhiqingxin formula (YQF) and its neuroprotective effects based on pharmacodynamic indices such as learning and memory ability, brain tissue histopathology and morphology, and inflammatory factor expression in a mouse model of Alzheimer's disease (AD).	[ "The efficacy of ginsenoside treatment on cognitive decline in individuals with Alzheimer's disease (AD) has yet to be investigated. In this protocal, we conducted a systematic review to evaluate the effect of ginsenosides on cognitive deficits in experimental rodent AD models. We identified eligible studies by searching seven electronic databases spanning from January 1980 to October 2014. We assessed the study quality, evaluated the efficacy of ginsenoside treatment, and performed a stratified meta-analysis and meta-regression analysis to assess the influence of the study design on ginsenoside efficacy. Twelve studies fulfilled our inclusion criteria from a total of 283 publications. The overall methodological quality of these studies was poor. The meta-analysis revealed that ginsenosides have a statistically significant positive effect on cognitive performance in experimental AD models. The stratified analysis revealed that ginsenoside Rg1 had the greatest effect on acquisition and retention memory in AD models. The effect size was significantly higher for both acquisition and retention memory in studies that used female animals compared with male animals. We conclude that ginsenosides might reduce cognitive deficits in AD models. However, additional well-designed and well-reported animal studies are needed to inform further clinical investigations.", "To establish an optimized method for extracting saponins from Radix et Rhizoma Notoginseng (Sanqi). HPLC coupled with charged aerosol detector (CAD) was used for saponins determination. Peak areas of the main saponins notoginsenoside R1, ginsenosides Rg1, Re, Rb1, Rg2, Rh1 and Rd in Sanqi were monitored to evaluate the extract effectiveness. One-factor analysis was conducted to investigate the factors of particle size, solvent, extraction method and soaking time. Orthogonal experiment design L9 (3(4)) with different level of concentration of solvents, extraction time, extraction times and volume of the solvent, was implemented to determine the optimized extract condition. The optimum extract technique was that the dried and well pulverized (passed through a 80 mesh (180 +/-7.6) microm sample 1.0 g was extracted in 20 mL methanol by reflux (1.5 h) after soaked at room temperature for 0.5 h. The optimized technique is simple and high extraction efficiency. It provide basic data for further study on Sanqi.", "A solvent recycling reflux extraction process for Panax notoginseng was optimized using a design space approach to improve the batch-to-batch consistency of the extract. Saponin yields, total saponin purity, and pigment yield were defined as the process critical quality attributes (CQAs). Ethanol content, extraction time, and the ratio of the recycling ethanol flow rate and initial solvent volume in the extraction tank (RES) were identified as the critical process parameters (CPPs) via quantitative risk assessment. Box-Behnken design experiments were performed. Quadratic models between CPPs and process CQAs were developed, with determination coefficients higher than 0.88. As the ethanol concentration decreases, saponin yields first increase and then decrease. A longer extraction time leads to higher yields of the ginsenosides Rb1 and Rd. The total saponin purity increases as the ethanol concentration increases. The pigment yield increases as the ethanol concentration decreases or extraction time increases. The design space was calculated using a Monte-Carlo simulation method with an acceptable probability of 0.90. Normal operation ranges to attain process CQA criteria with a probability of more than 0.914 are recommended as follows: ethanol content of 79-82%, extraction time of 6.1-7.1 h, and RES of 0.039-0.040 min-1. Most of the results of the verification experiments agreed well with the predictions. The verification experiment results showed that the selection of proper operating ethanol content, extraction time, and RES within the design space can ensure that the CQA criteria are met." ]
Chemical evolution of thiophene-based ligands for detecting polymorphic protein aggregates	The development of ligands for detecting protein aggregates is of great interest, as these aggregated proteinaceous species are the pathological hallmarks of several devastating diseases, including Alzheimer's disease. In this regard, thiophene-based ligands have emerged as powerful tools for fluorescent assessment of these pathological entities. The intrinsic conformationally sensitive photophysical properties of poly- and oligothiophenes have allowed optical assignment of disease-associated protein aggregates in tissue sections, as well as real-time in vivo imaging of protein deposits. Herein, we recount the chemical evolution of different generations of thiophene-based ligands, and exemplify their use for the optical distinction of polymorphic protein aggregates. Furthermore, the chemical determinants for achieving a superior fluorescent thiophene-based ligand, as well as the next generation of thiophene-based ligands targeting distinct aggregated species are described. Finally, the directions for future research into the chemical design of thiophene-based ligands that can aid in resolving the scientific challenges around protein aggregation diseases are discussed.	[ "Fluorescence lifetime imaging (FLIM) uses the fact that the fluorescence lifetime of a fluorophore depends on its molecular environment but not on its concentration. Molecular effects in a sample can therefore be investigated independently of the variable, and usually unknown concentration of the fluorophore. There is a variety of technical solutions of lifetime imaging in microscopy. The technical part of this paper focuses on time-domain FLIM by multidimensional time-correlated single photon counting, time-domain FLIM by gated image intensifiers, frequency-domain FLIM by gain-modulated image intensifiers, and frequency-domain FLIM by gain-modulated photomultipliers. The application part describes the most frequent FLIM applications: Measurement of molecular environment parameters, protein-interaction measurements by Förster resonance energy transfer (FRET), and measurements of the metabolic state of cells and tissue via their autofluorescence. Measurements of local environment parameters are based on lifetime changes induced by fluorescence quenching or conformation changes of the fluorophores. The advantage over intensity-based measurements is that no special ratiometric fluorophores are needed. Therefore, a much wider selection of fluorescence markers can be used, and a wider range of cell parameters is accessible. FLIM-FRET measures the change in the decay function of the FRET donor on interaction with an acceptor. FLIM-based FRET measurement does not have to cope with problems like donor bleedthrough or directly excited acceptor fluorescence. This relaxes the requirements to the absorption and emission spectra of the donors and acceptors used. Moreover, FLIM-FRET measurements are able to distinguish interacting and noninteracting fractions of the donor, and thus obtain independent information about distances and interacting and noninteracting protein fractions. This is information not accessible by steady-state FRET techniques. Autofluorescence FLIM exploits changes in the decay parameters of endogenous fluorophores with the metabolic state of the cells or the tissue. By resolving changes in the binding, conformation, and composition of biologically relevant compounds FLIM delivers information not accessible by steady-state fluorescence techniques.", "Protein deposits composed of specific proteins or peptides are associated with several neurodegenerative diseases and fluorescent ligands able to detect these pathological hallmarks are vital. Here, we report the synthesis of a class of thiophene-based ligands, denoted proteophenes, with different amino acid side-chain functionalities along the conjugated backbone, which display selectivity towards specific disease-associated protein aggregates in tissue sections with Alzheimer's disease (AD) pathology. The selectivity of the ligands towards AD associated pathological hallmarks, such as aggregates of the amyloid-β (Aβ) peptide or tau filamentous inclusions, was highly dependent on the chemical nature of the amino acid functionality, as well as on the location of the functionality along the pentameric thiophene backbone. Finally, the concept of synthesizing donor-acceptor-donor proteophenes with distinct photophysical properties was shown. Our findings provide the structural and functional basis for the development of new thiophene-based ligands that can be utilized for optical assignment of different aggregated proteinaceous species in tissue sections.", "Our understanding of the molecular structures of amyloid fibrils that are associated with neurodegenerative diseases, of mechanisms by which disease-associated peptides and proteins aggregate into fibrils, and of structural properties of aggregation intermediates has advanced considerably in recent years. Detailed molecular structural models for certain fibrils and aggregation intermediates are now available. It is now well established that amyloid fibrils are generally polymorphic at the molecular level, with a given peptide or protein being capable of forming a variety of distinct, self-propagating fibril structures. Recent results from structural studies and from studies involving cell cultures, transgenic animals, and human tissue provide initial evidence that molecular structural variations in amyloid fibrils and related aggregates may correlate with or even produce variations in disease development. This article reviews our current knowledge of the structural and mechanistic aspects of amyloid formation, as well as current evidence for the biological relevance of structural variations.", "A sensor is provided that detects single-stranded deoxyribonucleic acid (ssDNA) with a specific base sequence. The ssDNA sequence sensor comprises an aqueous solution containing a cationic water-soluble conjugated polymer [in this case, poly(9,9-bis(6'-N,N,N-trimethylammonium)-hexyl)-fluorene phenylene), 1] with a ssDNA labeled with a dye (in this case, fluorescein). The emission of light from the sensor solution with the wavelength characteristic of the probe oligonucleotide indicates the presence of ssDNA with a specific base sequence complementary to that of the probe ssDNA-fluorescein. Maximum energy transfer from 1 to the signaling chromophore occurs when the ratio of polymer chains to DNA strands is approximately 1:1. Energy transfer from 1 results in a fluorescein emission that is more intense than that observed by direct excitation of the chromophore. Furthermore, the decrease in energy transfer upon addition of electrolyte indicates that electrostatic forces dominate the interactions between 1 and DNA.", "Prions cause transmissible spongiform encephalopathies for which no treatment exists. Prions consist of PrP(Sc), a misfolded and aggregated form of the cellular prion protein (PrP(C)). We explore the antiprion properties of luminescent conjugated polythiophenes (LCPs) that bind and stabilize ordered protein aggregates. By administering a library of structurally diverse LCPs to the brains of prion-infected mice via osmotic minipumps, we found that antiprion activity required a minimum of five thiophene rings bearing regularly spaced carboxyl side groups. Solid-state nuclear magnetic resonance analyses and molecular dynamics simulations revealed that anionic side chains interacted with complementary, regularly spaced cationic amyloid residues of model prions. These findings allowed us to extract structural rules governing the interaction between LCPs and protein aggregates, which we then used to design a new set of LCPs with optimized binding. The new set of LCPs showed robust prophylactic and therapeutic potency in prion-infected mice, with the lead compound extending survival by >80% and showing activity against both mouse and hamster prions as well as efficacy upon intraperitoneal administration into mice. These results demonstrate the feasibility of targeted chemical design of compounds that may be useful for treating diseases of aberrant protein aggregation such as prion disease.", "The molecular architecture of amyloids formed in vivo can be interrogated using luminescent conjugated oligothiophenes (LCOs), a unique class of amyloid dyes. When bound to amyloid, LCOs yield fluorescence emission spectra that reflect the 3D structure of the protein aggregates. Given that synthetic amyloid-β peptide (Aβ) has been shown to adopt distinct structural conformations with different biological activities, we asked whether Aβ can assume structurally and functionally distinct conformations within the brain. To this end, we analyzed the LCO-stained cores of β-amyloid plaques in postmortem tissue sections from frontal, temporal, and occipital neocortices in 40 cases of familial Alzheimer's disease (AD) or sporadic (idiopathic) AD (sAD). The spectral attributes of LCO-bound plaques varied markedly in the brain, but the mean spectral properties of the amyloid cores were generally similar in all three cortical regions of individual patients. Remarkably, the LCO amyloid spectra differed significantly among some of the familial and sAD subtypes, and between typical patients with sAD and those with posterior cortical atrophy AD. Neither the amount of Aβ nor its protease resistance correlated with LCO spectral properties. LCO spectral amyloid phenotypes could be partially conveyed to Aβ plaques induced by experimental transmission in a mouse model. These findings indicate that polymorphic Aβ-amyloid deposits within the brain cluster as clouds of conformational variants in different AD cases. Heterogeneity in the molecular architecture of pathogenic Aβ among individuals and in etiologically distinct subtypes of AD justifies further studies to assess putative links between Aβ conformation and clinical phenotype.", "Metastable oligomeric and protofibrillar forms of amyloidogenic proteins have been implicated as on-pathway assembly intermediates in amyloid formation and as the major toxic species in a number of amyloid diseases including Alzheimer's disease. We describe here a chemical biology approach to structural analysis of Abeta protofibrils. Library screening yielded several molecules that stimulate Abeta aggregation. One of these compounds, calmidazolium chloride (CLC), rapidly and efficiently converts Abeta(1-40) monomers into clusters of protofibrils. As monitored by electron microscopy, these protofibrils persist for days when incubated in PBS at 37 degrees C, with a slow transition to fibrillar structures apparent only after several weeks. Like normal protofibrils, the CLC-Abeta aggregates exhibit a low thioflavin T response. Like Abeta fibrils, the clustered protofibrils bind the anti-amyloid Ab WO1. The CLC-Abeta aggregates exhibit the same protection from hydrogen-deuterium exchange as do protofibrils isolated from a spontaneous Abeta fibril formation reaction: approximately 12 of the 39 Abeta(1-40) backbone amide protons are protected from exchange in the protofibril, compared with approximately twice that number in amyloid fibrils. Scanning proline mutagenesis analysis shows that the Abeta molecule in these protofibrillar assemblies exhibits the same flexible N and C termini as do mature amyloid fibrils. The major difference in Abeta conformation between fibrils and protofibrils is added structural definition in the 22-29 segment in the fibril. Besides aiding structural analysis, compounds capable of facilitating oligomer and protofibril formation might have therapeutic potential, if they act to sequester Abeta in a form and/or location that cannot engage the toxic pathway.", "Molecular probes for selective identification of protein aggregates are important to advance our understanding of the molecular pathogenesis underlying cerebral amyloidoses. Here we report the chemical design of pentameric thiophene derivatives, denoted luminescent conjugated oligothiophenes (LCOs), which could be used for real-time visualization of cerebral protein aggregates in transgenic mouse models of neurodegenerative diseases by multiphoton microscopy. One of the LCOs, p-FTAA, could be utilized for ex vivo spectral assignment of distinct prion deposits from two mouse-adapted prion strains. p-FTAA also revealed staining of transient soluble pre-fibrillar non-thioflavinophilic Abeta-assemblies during in vitro fibrillation of Abeta peptides. In brain tissue samples, Abeta deposits and neurofibrillary tangles (NFTs) were readily identified by a strong fluorescence from p-FTAA and the LCO staining showed complete co-localization with conventional antibodies (6E10 and AT8). In addition, a patchy islet-like staining of individual Abeta plaque was unveiled by the anti-oligomer A11 antibody during co-staining with p-FTAA. The major hallmarks of Alzheimer's disease, namely, Abeta aggregates versus NFTs, could also be distinguished because of distinct emission spectra from p-FTAA. Overall, we demonstrate that LCOs can be utilized as powerful practical research tools for studying protein aggregation diseases and facilitate the study of amyloid origin, evolution and maturation, Abeta-tau interactions, and pathogenesis both ex vivo and in vivo.", "Conjugated polymers (CPs) have been used for creating bioimaging tools or biosensors that provide a direct link between spectral signal and different biological processes. The detection schemes of these sensors are mainly employing the efficient light harvesting properties or the conformation sensitive optical properties of the CPs. Hence, the presence of biomolecules or biological events can be detected through fluorescence resonance energy transfer (FRET) between the CP and an acceptor molecule, or through their impact on the conformation of the conjugated backbone, which is seen as an alteration of the optical properties of the CP. In this review, the utilization of CPs for sensitive detection of DNA and protein conformational changes will be presented. The main part will be focused on the specific binding of CPs to protein deposits associated with protein misfolding diseases, such as Alzheimer's disease (AD), and the discovery that tailor-made CPs can be used for in vivo optical imaging of protein aggregates will be discussed. The unique optical properties of CPs can be used as molecular tools for sensitive detection of genetic material and for characterization of the pathological hallmarks associated with protein misfolding disorders, such as AD. CPs are novel molecular tools that can be used for sensitive bioimaging of biological processes and these tools offer the possibility to study biological events in a complementary fashion to conventional techniques. This article is part of a Special Issue entitled Nanotechnologies - Emerging Applications in Biomedicine.", "Amyloid beta(40) (Abeta(40)) is the most abundant Abeta peptide in the brain. The cerebrospinal fluid (CSF) level of Abeta(40) might therefore be considered to most closely reflect the total Abeta load in the brain. Both in Alzheimer's disease (AD) and in normal aging the Abeta load in the brain has a large inter-individual variability. Relating Abeta(42) to Abeta(40) levels might consequently provide a more valid measure for reflecting the change in Abeta metabolism in dementia patients than the CSF Abeta(42) concentrations alone. This measure may also improve differential diagnosis between AD and other dementia syndromes, such as vascular dementia (VaD), dementia with Lewy bodies (DLB), and frontotemporal dementia (FTD). To investigate the diagnostic value of the CSF Abeta(42)/Abeta(40) ratio in differentiating AD from controls, VaD, DLB and FTD. We analysed the CSF Abeta(42)/Abeta(40) ratio, phosphorylated tau(181) and total tau in 69 patients with AD, 26 patients with VaD, 16 patients with DLB, 27 patients with FTD, and 47 controls. Mean Abeta(40) levels were 2850 pg/ml in VaD and 2830 pg/ml in DLB patients, both significantly lower than in AD patients (3698 pg/ml; p<0.01). Abeta(40) levels in AD patients were not significantly different from those in controls (4035 pg/ml; p=0.384). The Abeta(42)/Abeta(40) ratio was significantly lower in AD patients than in all other groups (p <0.001, ANCOVA). Differentiating AD from VaD, DLB and non-AD dementia improved when the Abeta(42)/Abeta(40) ratio was used instead of Abeta(42) concentrations alone (p<0.01) The Abeta(42)/Abeta(40) ratio performed equally well as the combination of Abeta(42), phosphorylated tau(181) and total tau in differentiating AD from FTD and non-AD dementia. The diagnostic performance of the latter combination was not improved when the Abeta(42)/Abeta(40) ratio was used instead of Abeta(42) alone. The CSF Abeta42/Abeta40 ratio improves differentiation of AD patients from VaD, DLB and non-AD dementia patients, when compared to Abeta42 alone, and is a more easily interpretable alternative to the combination of Abeta42, p-tau and t-tau when differentiating AD from either FTD or non-AD dementia." ]
Microbial safety has become a research hotspot with the development of manned space technology	Microbial safety has become a research hotspot with the development of manned space technology.	[ "For unsuspecting bacteria, the difference between life and death depends upon efficient and specific responses to various stressors. Facing a much larger world, microbes are invariably challenged with ever-changing environments where temperature, pH, chemicals, and nutrients are in a constant state of flux. Only those that are able to rapidly reprogram themselves and express subsets of genes needed to overcome the stress will survive and outcompete neighboring microbes. Recently, low shear stress, emulating microgravity (MG) experienced in space, has been characterized in a number of microorganisms including fungi and prokaryotes ranging from harmless surrogate organisms to bona fide pathogens. Interestingly, MG appears to induce a plethora of effects ranging from enhanced pathogenicity in several Gram-negative enterics to enhanced biofilm formation. Furthermore, MG-exposed bacteria appeared better able to handle subsequent stressors including: osmolarity, pH, temperature, and antimicrobial challenge while yeast exhibited aberrant budding post-MG-exposure. This review will focus on MG-induced alterations of virulence in various microbes with the emphasis placed on bacteria.", "Candida albicans is an opportunistic human pathogen that is capable of causing superficial and systemic infections in immunocompromised patients. Extracts of Sapindus saponaria have been used as antimicrobial agents against various organisms. In the present study, we used a combination of two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) to identify the changes in protein abundance of C. albicans after exposure to the minimal inhibitory concentration (MIC) and sub-minimal inhibitory concentration (sub-MIC) of the butanolic extract (BUTE) of S. saponaria and also to fluconazole. A total of six different proteins with greater than 1.5 fold induction or repression relative to the untreated control cells were identified among the three treatments. In general, proteins/enzymes involved with the glycolysis (GPM1, ENO1, FBA1), amino acid metabolism (ILV5, PDC11) and protein synthesis (ASC1) pathways were detected. In conclusion, our findings reveal antifungal-induced changes in protein abundance of C. albicans. By using the previously identified components of the BUTE of S. saponaria(e.g., saponins and sesquiterpene oligoglycosides), it will be possible to compare the behavior of compounds with unknown mechanisms of action, and this knowledge will help to focus the subsequent biochemical work aimed at defining the effects of these compounds.", "The polymorphic fungus Candida albicans is a member of the normal human microbiome. In most individuals, C. albicans resides as a lifelong, harmless commensal. Under certain circumstances, however, C. albicans can cause infections that range from superficial infections of the skin to life-threatening systemic infections. Several factors and activities have been identified which contribute to the pathogenic potential of this fungus. Among them are molecules which mediate adhesion to and invasion into host cells, the secretion of hydrolases, the yeast-to-hypha transition, contact sensing and thigmotropism, biofilm formation, phenotypic switching and a range of fitness attributes. Our understanding of when and how these mechanisms and factors contribute to infection has significantly increased during the last years. In addition, novel virulence mechanisms have recently been discovered. In this review we present an update on our current understanding of the pathogenicity mechanisms of this important human pathogen.", "The space environment could have impacts on a variety of characteristics of microorganism such as cell metabolism, drug resistance, and virulence. However, relevant mechanisms need to be clarified. In the present study, the effect of a space environment on Escherichia coli was investigated. E. coli strains were sent to space for 398 h on the Shenzhou VIII and ground simulation was conducted as control. After the flight, a mutant strain LCT-EC67 was selected for further analysis. Although no changes in hemolysis, morphology or antibiotic sensitivity were observed, the mutant strain showed elevated carbon source utilization compared with the control group. Genomic and proteomic analyses showed that 801 genes were upregulated and 825 genes were downregulated. In addition, 167 proteins were overexpressed and 92 proteins were downregulated using a cut-off fold-change value of 1.4 and a p < 0.05. The changed proteins were associated with metabolic functions such as alanine and glutamate metabolism, arginine and proline metabolism, and fatty acid and propanoate metabolism. E. coli showed alterations at gene and protein levels mainly regarding biochemical metabolism after spaceflight.", "Candida albicans is an opportunistic pathogenic yeast, which could become pathogenic in various stressful environmental factors including the spaceflight environment. In this study, we aim to explore the phenotypic changes and possible mechanisms of C. albicans after exposure to spaceflight conditions. The effect of C. albicans after carried on the \"SJ-10\" satellite for 12 days was evaluated by proliferation, morphology, environmental resistance and virulence experiment. The result showed that the proliferation rate, biofilm formation, antioxidant capacity, cytotoxicity and filamentous morphology of C. albicans were increased in the spaceflight group compared to the control group. Proteomics and metabolomics technologies were used to analyze the profiles of proteins and metabolites in C. albicans under spaceflight conditions. Proteomic analysis identified 548 up-regulated proteins involved in the ribosome, DNA replication, base excision repair and sulfur metabolism in the spaceflight group. Moreover, 332 down-regulated proteins related to metabolic processes were observed. The metabolomic analysis found five differentially expressed metabolites. The combined analysis of proteomic and metabolomic revealed the accumulation of cysteine and methionine in C. albicans after spaceflight. Mechanisms that could explain the results in the phenotypic experiment of C. albicans were found through proteomic and metabolomic analysis. And our data provide an important basis for the assessment of the risk that C. albicans could cause under spaceflight environment.", "Changes in bacterial physiology caused by the combined action of the magnetic force and microgravity were studied in Escherichia coli grown using a specially developed device aboard the International Space Station. The morphology and metabolism of E. coli grown under spaceflight (SF) or combined spaceflight and magnetic force (SF + MF) conditions were compared with ground cultivated bacteria grown under standard (control) or magnetic force (MF) conditions. SF, SF + MF, and MF conditions provided the up-regulation of Ag43 auto-transporter and cell auto-aggregation. The magnetic force caused visible clustering of non-sedimenting bacteria that formed matrix-containing aggregates under SF + MF and MF conditions. Cell auto-aggregation was accompanied by up-regulation of glyoxylate shunt enzymes and Vitamin B12 transporter BtuB. Under SF and SF + MF but not MF conditions nutrition and oxygen limitations were manifested by the down-regulation of glycolysis and TCA enzymes and the up-regulation of methylglyoxal bypass. Bacteria grown under combined SF + MF conditions demonstrated superior up-regulation of enzymes of the methylglyoxal bypass and down-regulation of glycolysis and TCA enzymes compared to SF conditions, suggesting that the magnetic force strengthened the effects of microgravity on the bacterial metabolism. This strengthening appeared to be due to magnetic force-dependent bacterial clustering within a small volume that reinforced the effects of the microgravity-driven absence of convectional flows.", "The methionine salvage pathway is universally used to regenerate methionine from 5'-methylthioadenosine, a byproduct of certain reactions involving S-adenosylmethionine. We identified and verified the genes encoding the enzymes of all steps in this cycle in a commonly used eukaryotic model system: the yeast Saccharomyces cerevisiae. The genes encoding 5'-methylthioribose-1-phosphate isomerase and 5'-methylthioribulose-1-phosphate dehydratase are herein named MRI1 and MDE1, respectively. The 5'-methylthioadenosine phosphorylase was verified as Meu1p, the 2,3-dioxomethiopentane-1-phosphate enolase/phosphatase as Utr4p and the aci-reductone dioxygenase as Adi1p. The homologue of the enolase/phosphatase gene, YNL010w, was excluded from its candidate role in the cycle. The methodology used involved auxotrophic growth tests and analysis of intracellular 5'-methylthioadenosine in deletion mutants. The last step, a transamination of 4-methylthio-2-oxobutyrate to yield methionine, was found to be a highly redundant step. It was catalysed by amino acid transaminases, mainly coupled with aromatic and branched chain amino acids as amino donors, but also with proline, lysine and glutamate/glutamine. The aromatic amino acid transaminases, Aro8p and Aro9p, and the branched chain amino acid transaminases, Bat1p and Bat2p, seemed to be the main enzymes exhibiting 4-methylthio-2-oxobutyrate transaminase activity. Bat2p was found to be less specific and used proline, lysine, tyrosine and glutamate as amino donors in addition to the branched chain amino acids. Thus, for the first time, all enzymes of the methionine salvage pathway were identified in a eukaryote." ]
Peptigoglycan sensing and fitness/virulence regulation in Gram-negatives	Given the growing clinical-epidemiological threat posed by the phenomenon of antibiotic resistance, new therapeutic options are urgently needed, especially against top nosocomial pathogens such as those within the ESKAPE group. In this scenario, research is pushed to explore therapeutic alternatives and, among these, those oriented toward reducing bacterial pathogenic power could pose encouraging options. However, the first step in developing these antivirulence weapons is to find weak points in the bacterial biology to be attacked with the goal of dampening pathogenesis. In this regard, during the last decades some studies have directly/indirectly suggested that certain soluble peptidoglycan-derived fragments display virulence-regulatory capacities, likely through similar mechanisms to those followed to regulate the production of several β-lactamases: binding to specific transcriptional regulators and/or sensing/activation of two-component systems. These data suggest the existence of intra- and also intercellular peptidoglycan-derived signaling capable of impacting bacterial behavior, and hence likely exploitable from the therapeutic perspective. Using the well-known phenomenon of peptidoglycan metabolism-linked β-lactamase regulation as a starting point, we gather and integrate the studies connecting soluble peptidoglycan sensing with fitness/virulence regulation in Gram-negatives, dissecting the gaps in current knowledge that need filling to enable potential therapeutic strategy development, a topic which is also finally discussed.	[ "β-Lactam antibiotics were the earliest discovered and are the most widely used group of antibiotics that work by inactivating penicillin-binding proteins to inhibit peptidoglycan biosynthesis. As one of the most efficient defense strategies, many bacteria produce β-lactam-degrading enzymes, β-lactamases, whose biochemical functions and regulation have been extensively studied. A signal transduction pathway for β-lactamase induction by β-lactam antibiotics, consisting of the major peptidoglycan recycling enzymes and the LysR-type transcriptional regulator, AmpR, has been recently unveiled in some bacteria. Because inactivation of some of these proteins, especially the permease AmpG and the β-hexosaminidase NagZ, results in substantially elevated susceptibility to the antibiotics, these have been recognized as potential therapeutic targets. Here, we show a contrasting scenario in Shewanella oneidensis, in which the homologue of AmpR is absent. Loss of AmpG or NagZ enhances β-lactam resistance drastically, whereas other identified major peptidoglycan recycling enzymes are dispensable. Moreover, our data indicate that there exists a parallel signal transduction pathway for β-lactamase induction, which is independent of either AmpG or NagZ.", "The envelope of Gram-negative bacteria is an essential compartment that constitutes a protective and permeability barrier between the cell and its environment. The envelope also hosts the cell wall, a mesh-like structure made of peptidoglycan (PG) that determines cell shape and provides osmotic protection. Since the PG must grow and divide in a cell-cycle-synchronized manner, its synthesis and remodeling are tightly regulated. Here, we discovered that PG homeostasis is intimately linked to the levels of activation of the Cpx system, an envelope stress response system traditionally viewed as being involved in protein quality control in the envelope. We first show that Cpx is activated when PG integrity is challenged and that this activation provides protection to cells exposed to antibiotics inhibiting PG synthesis. By rerouting the outer membrane lipoprotein NlpE, a known Cpx activator, to a different envelope subcompartment, we managed to manipulate Cpx activation levels. We found that Cpx overactivation leads to aberrant cellular morphologies, to an increased sensitivity to β-lactams, and to dramatic division and growth defects, consistent with a loss of PG homeostasis. Remarkably, these phenotypes were largely abrogated by the deletion of ldtD, a Cpx-induced gene involved in noncanonical PG cross-linkage, suggesting that this transpeptidase is an important link between PG homeostasis and the Cpx system. Altogether our data show that fine-tuning of an envelope quality control system constitutes an important layer of regulation of the highly organized cell wall structure. The envelope of Gram-negative bacteria is essential for viability. First, it includes the cell wall, a continuous polymer of peptidoglycan (PG) that determines cell morphology and protects against osmotic stress. Moreover, the envelope constitutes a protective barrier between the cell interior and the environment. Therefore, mechanisms called envelope stress response systems (ESRS) exist to monitor and defend envelope integrity against harmful conditions. Cpx is a major ESRS that detects and manages the accumulation of misfolded proteins in the envelope of Escherichia coli. We found that this protein quality control system also plays a fundamental role in the regulation of PG assembly. Strikingly, the level of Cpx response is critical, as an excessive activation leads to phenotypes associated with a loss of cell wall integrity. Thus, by contributing to PG homeostasis, the Cpx system lies at the crossroads between key processes of bacterial life, including cell shape, growth, division, and antibiotic resistance.", "A mechanism for bacteria to monitor the status of their vital cell wall peptidoglycan is suggested by the convergence of two phenomena: peptidoglycan recycling and beta-lactamase induction. ampG and ampD, genes essential for beta-lactamase regulation, are here shown to be required for recycling as well. Cells lacking either AmpG or AmpD lose up to 40% of their peptidoglycan per generation, whereas Escherichia coli normally suffers minimal losses and instead recycles 40 or 50% of the tripeptide, L-alanyl-D-glutamyl-meso-diaminopimelic acid, from its peptidoglycan each generation. The ampG mutant releases peptidoglycan-derived material into the medium. In contrast, the ampD mutant accumulates a novel cell wall muropeptide, 1,6-anhydro N-acetylmuramyl-L-alanyl-D-glutamyl-meso-diaminopimelic acid (anhMurNAc-tripeptide), in its cytoplasm. This work suggests that AmpG is the permease for a large muropeptide and AmpD is a novel cytosolic N-acetylmuramyl-L-alanine amidase that cleaves anhMurNAc-tripeptide to release tripeptide, which is then recycled. These results also suggest that the phenomenon of beta-lactamase induction is regulated by the level of muropeptide(s) in the cytoplasm, since an ampD mutation that results in beta-lactamase expression even in the absence of a beta-lactamase inducer coincides with accumulation of anhMurNAc-tripeptide. The transcriptional regulator AmpR is presumably converted into an activator for beta-lactamase production by sensing the higher level of muropeptide(s). This may be an example of a general mechanism for signaling the progress of external events such as cell wall maturation, cell division or cell wall damage.", "Benzyl[14C]penicillin binds to six proteins with molecular weights between 40000 and 91000 in the inner membrane of Escherichia coli. Two additional binding proteins with molecular weights of 29000 and 32000 were sometimes detected. All proteins were accessible to benzyl[14C]penicillin in whole cells. Proteins 5 and 6 released bound benzyl[14C]penicillin with half times of 5 and 19 min at 30 degrees C but the other binding proteins showed less than 50% release during a 60-min period at 30 degrees C. The rate of release of bound penicillin from some of the proteins was greatly stimulated by 2-mercaptoethanol and neutral hydroxylamine. Release of benzyl[14C]penicillin did not occur if the binding proteins were denatured in anionic detergent and so was probably enzymic. No additional binding proteins were detected with two [14C]cephalosporins. These beta-lactams bound to either all or some of those proteins to which benzyl[14C]penicillin bound. No binding proteins have been detected in the outer membrane of E coli with any beta-[14C]lactam. The binding of a range of unlabelled penicillins and cephalosporins were studied by measuring their competition for the binding of benzyl[14C]penicillin to the six penicillin-binding proteins. These results, together with those obtained by direct binding experiments with beta-[14C]lactams, showed that penicillins bind to all six proteins but that at least some cephalosporins fail to bind, or bind very slowly, to proteins 2, 5 and 6, although they bind to the other proteins. Since these cephalosporins inhibited cell division and caused cell lysis at concentrations where we could detect no binding to proteins 2, 5 and 6, we believe that these latter proteins are not the target at which beta-lactams bind to elicit the above physiological responses. The binding properties of proteins 1, 3, and 4 correlate reasonably well with those expected for the above killing targets.", "The phenomenon of peptidoglycan recycling is reviewed. Gram-negative bacteria such as Escherichia coli break down and reuse over 60% of the peptidoglycan of their side wall each generation. Recycling of newly made peptidoglycan during septum synthesis occurs at an even faster rate. Nine enzymes, one permease, and one periplasmic binding protein in E. coli that appear to have as their sole function the recovery of degradation products from peptidoglycan, thereby making them available for the cell to resynthesize more peptidoglycan or to use as an energy source, have been identified. It is shown that all of the amino acids and amino sugars of peptidoglycan are recycled. The discovery and properties of the individual proteins and the pathways involved are presented. In addition, the possible role of various peptidoglycan degradation products in the induction of beta-lactamase is discussed.", "The clinical and epidemiological threat of the growing antimicrobial resistance in Gram-negative pathogens, particularly for β-lactams, the most frequently used and relevant antibiotics, urges research to find new therapeutic weapons to combat the infections caused by these microorganisms. An essential previous step in the development of these therapeutic solutions is to identify their potential targets in the biology of the pathogen. This is precisely what we sought to do in this review specifically regarding the barely exploited field analyzing the interplay among the biology of the peptidoglycan and related processes, such as β-lactamase regulation and virulence. Hence, here we gather, analyze, and integrate the knowledge derived from published works that provide information on the topic, starting with those dealing with the historically neglected essential role of the Gram-negative peptidoglycan in virulence, including structural, biogenesis, remodeling, and recycling aspects, in addition to proinflammatory and other interactions with the host. We also review the complex link between intrinsic β-lactamase production and peptidoglycan metabolism, as well as the biological costs potentially associated with the expression of horizontally acquired β-lactamases. Finally, we analyze the existing evidence from multiple perspectives to provide useful clues for identifying targets enabling the future development of therapeutic options attacking the peptidoglycan-virulence interconnection as a key weak point of the Gram-negative pathogens to be used, if not to kill the bacteria, to mitigate their capacity to produce severe infections.", "The history of modern medicine cannot be written apart from the history of the antibiotics. Antibiotics are cytotoxic secondary metabolites that are isolated from Nature. The antibacterial antibiotics disproportionately target bacterial protein structure that is distinct from eukaryotic protein structure, notably within the ribosome and within the pathways for bacterial cell-wall biosynthesis (for which there is not a eukaryotic counterpart). This review focuses on a pre-eminent class of antibiotics-the β-lactams, exemplified by the penicillins and cephalosporins-from the perspective of the evolving mechanisms for bacterial resistance. The mechanism of action of the β-lactams is bacterial cell-wall destruction. In the monoderm (single membrane, Gram-positive staining) pathogen Staphylococcus aureus the dominant resistance mechanism is expression of a β-lactam-unreactive transpeptidase enzyme that functions in cell-wall construction. In the diderm (dual membrane, Gram-negative staining) pathogen Pseudomonas aeruginosa a dominant resistance mechanism (among several) is expression of a hydrolytic enzyme that destroys the critical β-lactam ring of the antibiotic. The key sensing mechanism used by P. aeruginosa is monitoring the molecular difference between cell-wall construction and cell-wall deconstruction. In both bacteria, the resistance pathways are manifested only when the bacteria detect the presence of β-lactams. This review summarizes how the β-lactams are sensed and how the resistance mechanisms are manifested, with the expectation that preventing these processes will be critical to future chemotherapeutic control of multidrug resistant bacteria.", "In Citrobacter freundii and Enterobacter cloacae, synthesis of AmpC beta-lactamase is inducible by the addition of beta-lactams to the growth medium. Spontaneous mutants that constitutively overproduce the enzyme occur at a high frequency. When the C. freundii ampC beta-lactamase gene is cloned into Escherichia coli together with the regulatory gene ampR, beta-lactamase expression from the clone is inducible. Spontaneous cefotaxime-resistant mutants were selected from an E. coli strain carrying the cloned C. freundii ampC and ampR genes on a plasmid. Virtually all isolates had chromosomal mutations leading to semiconstitutive overproduction of beta-lactamase. The mutation ampD2 in one such mutant was caused by an IS1 insertion into the hitherto unknown ampD gene, located between nadC and aroP at minute 2.4 on the E. coli chromosome. The wild-type ampD allele cloned on a plasmid could fully trans-complement beta-lactamase-overproducing mutants of both E. coli and C. freundii, restoring the wild-type phenotype of highly inducible enzyme synthesis. This indicates that these E. coli and C. freundii mutants have their lesions in ampD. We hypothesize that induction of beta-lactamase synthesis is caused by blocking of the AmpD function by the beta-lactam inducer and that this leads directly or indirectly to an AmpR-mediated stimulation of ampC expression.", "Pseudomonas aeruginosa is one of the first causes of acute nosocomial and chronic infections in patients with underlying respiratory pathologies such as cystic fibrosis (CF). It has been proposed that P. aeruginosa accumulates mutations driving to peptidoglycan modifications throughout the development of the CF-associated infection, as a strategy to lower the immune detection hence ameliorating the chronic persistence. As well, some studies dealing with peptidoglycan modifications driving to a better survival within the host have been published in other gram-negatives. According to these facts, the gram-negative peptidoglycan could be considered as a pathogen-associated molecular pattern with very important implications regarding the host's detection-response, worthy to dissect in detail. For this reason, in this work we characterized for the first time the peptidoglycans of three large collections [early CF, late CF and acute infection (bloodstream) P. aeruginosa strains] from qualitative (HPLC), quantitative and inflammatory capacity-related perspectives. The final goal was to identify composition trends potentially supporting the cited strategy of evasion/resistance to the immune system and providing information regarding the differential intrinsic adaptation depending on the type of infection. Although we found several punctual strain-specific particularities, our results indicated a high degree of inter-collection uniformity in the peptidoglycan-related features and the absence of trends amongst the strains studied here. These results suggest that the peptidoglycan of P. aeruginosa is a notably conserved structure in natural isolates regardless of transitory changes that some external conditions could force. Finally, the inverse correlation between the relative amount of stem pentapeptides within the murein sacculus and the resistance to immune lytic attacks against the peptidoglycan was also suggested by our results. Altogether, this work is a major step ahead to understand the biology of peptidoglycan from P. aeruginosa natural strains, hopefully useful in future for therapeutic alternatives design.", "The three components of the pyruvate dehydrogenase complex of Escherichia coli are encoded by three linked genes, ace E (pyruvate dehydrogenase, E1), aceF (dihydrolipoamide acetyltransferase, E2) and lpd (lipoamide dehydrogenase, E3, situated close to the nadC (quinolinate phosphoribosyltransferase) and aroP (general aromatic amino acid permease) genes with the gene order: nadC-aroP-aceE-aceF-lpd. Several types of transducing phages, lambda nadC and lambda lpd, carrying the nadC and lpd genes were isolated from populations of artificially constructed transducing phages containing R.HindIII or R.EcoRI fragments of bacterial DNA, by selecting for their ability to complement the metabolic lesions of the corresponding mutants. The cloned fragments were extended to include a functional ace operon by in vivo methods involving prophage insertion into the nadC-lpd region and aberrant excision to yield lambda nadC-lpd and lambda lpd-ace phages. These contained overlapping segments of bacterial DNA capable of expressing the aceE, aceF and lpd genes. A physical map of a 20 kilobase pairs (kb) segment of bacterial DNA encoding the entire nadC-lpd region, bounded by R.HindIII and R.EcoRI targets and possessing several internal restriction targets, R.HindIII (3) and R.EcoRI (2), was constructed. Using a combination of nutritional and enzymological studies with dilysogens and genetic analysis with ace mutants the approximate positions of the genes specifying the pyruvate dehydrogenase complex were traced to a 9.5 kb segment of the restriction map. The cloned lpd gene was expressed in the complete absence of a functional ace operon and when the major lambda promoters were repressed. This confirms that the lpd gene can be independently transcribed from its own promoter." ]
Duodenal mucinous adenocarcinoma in a patient with immunodeficiency virus	Duodenal mucinous adenocarcinoma is a rare type of small bowel carcinoma. It is not commonly encountered; hence little knowledge exists about its presentation, diagnosis, and management. The diagnosis is mostly made by either esophagogastroduodenoscopy (EGD) or intra-operatively. Some of the main symptoms are abdominal pain, nausea, vomiting, weight loss, or signs and symptoms of upper gastrointestinal bleeding. Therefore, this is a serious condition that healthcare providers and patients should be aware of to reduce its severity and improve prognosis. We present a case of duodenal mucinous adenocarcinoma in a patient with immunodeficiency virus.	[ "A 65-year-old woman presented with abdominal pain, weight loss, fatigue, and microcytic anemia. Esophagogastroduodenoscopy, until the second part of duodenum, was normal. Ultrasound and computed tomography demonstrated a solid mass in the distal duodenum. A repeat endoscopy confirmed an ulcerative, intraluminar mass in the third and fourth part of the duodenum. Segmental resection of the third and fourth portion of the duodenum was performed. Histology revealed an adenocarcinoma. On the 4th postoperative day, the patient developed severe acute pancreatitis leading to multiple organ failure and died on the 30th postoperative day.", "Duodenal adenocarcinoma is a rare but aggressive malignancy. Given its rarity, previous studies have traditionally combined duodenal adenocarcinoma (DA) with either other periampullary cancers or small bowel adenocarcinomas, limiting the available data to guide treatment decisions. Nevertheless, management primarily involves complete surgical resection when technically feasible. Surgery may require pancreaticoduodenectomy or segmental duodenal resection; either are acceptable options as long as negative margins are achievable and an adequate lymphadenectomy can be performed. Adjuvant chemotherapy and radiation are important components of multi-modality treatment for patients at high risk of recurrence. Further research would benefit from multi-institutional trials that do not combine DA with other periampullary or small bowel malignancies. The purpose of this article is to perform a comprehensive review of DA with special focus on the surgical management and principles.", "Patients with duodenal polyps associated with familial adenomatous polyposis (FAP) have a considerable risk of developing duodenal carcinoma. Prophylactic resection of the duodenum for Spigelman stage III disease is the treatment of choice to prevent progression to cancer. Pancreaticoduodenectomy and pancreas-preserving total duodenectomy (PPTD) are the techniques that have been described for the surgical treatment of duodenal polyposis. We report the first case of laparoscopic PPTD in a patient with previous total colectomy for FAP and Spigelman stage III duodenal polyposis. A laparoscopic total dissection of the duodenum was carried out and the restoration was achieved performing pancreatico-biliary-jejunostomy and gastrojejunostomy. The postoperative period was uneventful. Laparoscopic PPTD can be performed safely in selected cases for the management of FAP.", "Other than that in the duodenum, adenocarcinoma in the small bowel is rare. The present study describes a case of adenocarcinoma with adenoma in the jejunum. A 70-year-old male was admitted to hospital due to dehydration induced by abdominal discomfort and difficulty with oral intake. Computed tomography revealed a tumor in the upper side of the jejunum, which was subsequently resected. The tumor contained adenocarcinoma and adenoma. The protein expression of p53 and Ki-67 was analyzed in the normal mucosa, adenoma and adenocarcinoma. The number of epithelial cells expressing p53 and Ki-67 was found to increase in the adenoma tissue compared with that in the normal mucosa. In the adenocarcinoma tissue, the number of cells expressing p53 and Ki-67 further increased, suggesting that an adenoma-adenocarcinoma sequence may occur in the small bowel, similar to that observed in the large bowel." ]
Lipopolysaccharide-accelerated autoimmune glomerulonephritis in NZBWF1 mice	Lipopolysaccharide (LPS)-accelerated autoimmune glomerulonephritis (GN) in NZBWF1 mice is a preclinical model potentially applicable for investigating lipidome-modulating interventions against lupus. LPS can be expressed as one of two chemotypes: smooth LPS (S-LPS) or rough LPS (R-LPS) which is devoid of O-antigen polysaccharide sidechain. Since these chemotypes differentially affect toll-like receptor 4 (TLR4)-mediated immune cell responses, these differences may influence GN induction.	[ "Tris (dibenzylideneacetone) dipalladium (Tris DBA), a small-molecule palladium complex, has been shown to inhibit cell growth and proliferation in pancreatic cancer, lymphocytic leukemia, and multiple myeloma. In the current study, we examined the therapeutic effects of Tris DBA on glomerular cell proliferation, renal inflammation, and immune cells. Treatment of accelerated and severe lupus nephritis (ASLN) mice with Tris DBA resulted in improved renal function, albuminuria, and pathology, including measurements of glomerular cell proliferation, cellular crescents, neutrophils, fibrinoid necrosis, and tubulointerstitial inflammation in the kidneys as well as scoring for glomerulonephritis activity. The treated ASLN mice also showed significantly decreased glomerular IgG, IgM, and C3 deposits. Furthermore, the compound was able to 1) inhibit bone marrow-derived dendritic cell-mediated T cell functions and reduce serum anti-dsDNA autoantibody levels; 2) differentially regulate autophagy and both the priming and activation signals of the NLRP3 inflammasome; and 3) suppress the phosphorylation of JNK, ERK, and p38 MAPK signaling pathways. Tris DBA improved ASLN in mice through immunoregulation by blunting the MAPK (ERK, JNK)-mediated priming signal of the NLRP3 inflammasome and by regulating the autophagy/NLRP3 inflammasome axis. These results suggest that the pure compound may be a drug candidate for treating the accelerated and deteriorated type of lupus nephritis.", "A growing number of reports indicate that anti-inflammatory actions of fish oil (FO) are beneficial against systemic lupus erythematosus (SLE). However, the majority of pre-clinical studies were performed using 5-20% FO, which is higher than the clinically relevant dose for lupus patients. The present study was performed in order to determine the effective low dose of FDA-approved concentrated FO (Lovaza®) compared to the commonly used FO-18/12 (18-Eicosapentaenoic acid [EPA]/12-Docosahexaenoic acid [DHA]). We examined the dose-dependent response of Lovaza® (1% and 4%) on an SLE mouse strain (NZBxNZW)F1 and compared the same with 1% and 4% placebo, as well as 4% FO-18/12, maintaining standard chow as the control. Results show for the first time that 1% Lovaza® extends maximal lifespan (517 d) and 4% Lovaza® significantly extends both the median (502 d) and maximal (600 d) life span of (NZBxNZW)F1 mice. In contrast, FO-18/12 extends only median lifespan (410 d) compared to standard chow diet (301 d). Additionally, 4% Lovaza® significantly decreased anti-dsDNA antibodies, reduced glomerulonephritis and attenuated lipopolysaccharide-induced pro-inflammatory cytokines (IL-1β, IL-6, TNF-α) in splenocytes compared to placebo. 4% Lovaza® was also shown to reduce the expression of inflammatory cytokines, including IL-1β, IL-6 and TNF-α, while increasing renal anti-oxidant enzymes in comparison to placebo. Notably, NFκB activation and p65 nuclear translocation were lowered by 4% Lovaza® compared to placebo. These data indicate that 1% Lovaza® is beneficial, but 4% Lovaza® is more effective in suppressing glomerulonephritis and extending life span of SLE-prone short-lived mice, possibly via reducing inflammation signaling and modulating oxidative stress.", "Polyunsaturated fatty acids (PUFAs) are essential FAs for human health. Cytochrome P450 oxygenates PUFAs to produce anti-inflammatory and pain-resolving epoxy fatty acids (EpFAs) and other oxylipins whose epoxide ring is opened by the soluble epoxide hydrolase (sEH/Ephx2), resulting in the formation of toxic and pro-inflammatory vicinal diols (dihydroxy-FAs). Pharmacological inhibition of sEH is a promising strategy for the treatment of pain, inflammation, cardiovascular diseases, and other conditions. We tested the efficacy of a potent, selective sEH inhibitor, 1-trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl) urea (TPPU), in an animal model of multiple sclerosis (MS), experimental autoimmune encephalomyelitis (EAE). Prophylactic TPPU treatment significantly ameliorated EAE without affecting circulating white blood cell counts. TPPU accumulated in the spinal cords (SCs), which was correlated with plasma TPPU concentration. Targeted lipidomics in EAE SCs and plasma identified that TPPU blocked production of dihydroxy-FAs efficiently and increased some EpFA species including 12(13)-epoxy-octadecenoic acid (12(13)-EpOME) and 17(18)-epoxy-eicosatrienoic acid (17(18)-EpETE). TPPU did not alter levels of cyclooxygenase (COX-1/2) metabolites, while it increased 12-hydroxyeicosatetraenoic acid (12-HETE) and other 12/15-lipoxygenase metabolites. These analytical results are consistent with sEH inhibitors that reduce neuroinflammation and accelerate anti-inflammatory responses, providing the possibility that sEH inhibitors could be used as a disease modifying therapy, as well as for MS-associated pain relief.", "To examine associations between dietary intake of omega-3 (n-3; generally antiinflammatory) and omega-6 (n-6; generally proinflammatory) fatty acids and patient-reported outcomes in systemic lupus erythematosus (SLE). This study was based on the population-based Michigan Lupus Epidemiology and Surveillance cohort. Estimates of n-3 and n-6 intake were derived from Diet History Questionnaire II items (past year with portion size version). Patient-reported outcomes included self-reported lupus activity (Systemic Lupus Activity Questionnaire [SLAQ]). Multivariable regression, adjusted for age, sex, race, and body mass index, was used to assess associations between absolute intake of n-3 and n-6, as well as the n-6:n-3 ratio, and patient-reported outcomes. Among 456 SLE cases, 425 (93.2%) were female, 207 (45.4%) were African American, and the mean ± SD age was 52.9 ± 12.3 years. Controlling for potential confounders, the average SLAQ score was significantly higher by 0.3 points (95% confidence interval [95% CI] 0.1, 0.6; P = 0.013) with each unit increase of the n-6:n-3 ratio. Both lupus activity and Patient-Reported Outcomes Measurement Information System (PROMIS) sleep disturbance scores were lower with each 1-gram/1,000 kcal increase of n-3 fatty acids (SLAQ regression coefficient β = -0.8 [95% CI -1.6, 0.0]; P = 0.055; PROMIS sleep β = -1.1 [95% CI -2.0, -0.2]; P = 0.017). Higher n-3 intakes were nonsignificantly associated with lower levels of depressive symptoms and comorbid fibromyalgia, and with higher quality of life, whereas results for the n6:n3 ratio trended in the opposite direction. This population-based study suggests that higher dietary intake of n-3 fatty acids and lower n-6:n-3 ratios are favorably associated with patient-reported outcomes in SLE, particularly self-reported lupus activity and sleep quality.", "Epidemiological and preclinical evidence supports that omega-3 dietary fatty acids (fish oil) reduce the risks of macular degeneration and cancers, but the mechanisms by which these omega-3 lipids inhibit angiogenesis and tumorigenesis are poorly understood. Here we show that epoxydocosapentaenoic acids (EDPs), which are lipid mediators produced by cytochrome P450 epoxygenases from omega-3 fatty acid docosahexaenoic acid, inhibit VEGF- and fibroblast growth factor 2-induced angiogenesis in vivo, and suppress endothelial cell migration and protease production in vitro via a VEGF receptor 2-dependent mechanism. When EDPs (0.05 mg · kg(-1) · d(-1)) are coadministered with a low-dose soluble epoxide hydrolase inhibitor, EDPs are stabilized in circulation, causing ~70% inhibition of primary tumor growth and metastasis. Contrary to the effects of EDPs, the corresponding metabolites derived from omega-6 arachidonic acid, epoxyeicosatrienoic acids, increase angiogenesis and tumor progression. These results designate epoxyeicosatrienoic acids and EDPs as unique endogenous mediators of an angiogenic switch to regulate tumorigenesis and implicate a unique mechanistic linkage between omega-3 and omega-6 fatty acids and cancers.", "To determine the prevalence trends of osteoarthritis (OA), rheumatoid arthritis (RA), and other types of arthritis in the United States from 1999 to 2014. We analyzed data on 43 706 community-dwelling adults aged 20 years and older who participated in the 1999-2014 National Health and Nutrition Examination Surveys. We accounted for survey design and sampling weights so that estimates were nationally representative. We assessed temporal trends in age-standardized arthritis prevalence by using joinpoint regression. Age-adjusted prevalence of arthritis was 24.7% (OA = 9.7%; RA = 4.2%; other arthritis = 2.8%; \"don't know\" type = 8.0%). Prevalence of OA increased from 6.6% to 14.3%, whereas RA prevalence decreased from 5.9% to 3.8%. Increase in OA prevalence was significant in both genders; in non-Hispanic Whites, non-Hispanic Blacks, and Hispanics; and in people with high socioeconomic status. Decrease in RA prevalence was more pronounced in men, non-Hispanic Blacks, and participants with low income or obesity. Between 1999 and 2014, nearly one quarter of American adults reported arthritis. The prevalence of OA has more than doubled over time, whereas RA prevalence has declined.", "Costimulatory and adhesion molecules are known to play a major role in the pathogenesis of systemic lupus erythematosus. Since fish oil and calorie restriction have been reported to attenuate the development of disease in lupus prone (NZBxNZW)F1 mice, the objective of this study was to assess the expression of these key inflammatory molecules in these mice fed diets differing in n-6 and n-3 fatty acid content and fed either food restricted or ad libitum. Age-associated increases in the expression of CD28, ICAM-1, and PGP-1 molecules that are involved in the recruitment of inflamed lymphocytes into the kidney were attenuated in mice restricted in food intake. The increase in costimulatory (CD80 and CD86) and adhesion (ICAM-1, PGP-1, LFA-1, and Mac-1) in peripheral blood mononuclear cells was also attenuated by food restriction and to a lesser extent by fish oil alone. Interestingly, amelioration of lupus (laminin expression and proteinuria) correlated with the above beneficial effects and could be seen even in 24-month-old mice. In summary, food restriction and fish oil delay the onset of lupus disease and increase life span in B/W mice by prolonging the maintenance of a youthful immune phenotype.", "Epoxyeicosatrienoic acids (EET) and related epoxy fatty acids (EpFA) are endogenous anti-inflammatory compounds, which are converted by the soluble epoxide hydrolase (sEH) to dihydroxylethersatrienoic acids (DHETs) with lessened biological effects. Inhibition of sEH is used as a strategy to increase EET levels leading to lower inflammation. Rheumatoid arthritis is a chronic autoimmune disease that leads to destruction of joint tissues. This pathogenesis involves a complex interplay between the immune system, and environmental factors. Here, we investigate the effects of inhibiting sEH with 1-trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl) urea (TPPU) on a collagen-induced arthritis model. The treatment with TPPU ameliorates hyperalgesia, edema, and decreases the expression of important pro-inflammatory cytokines of Th1 and Th17 profiles, while increasing Treg cells. Considering the challenges to control RA, this study provides robust data supporting that inhibition of the sEH is a promising target to treat arthritis.", "Occupational exposure to respirable crystalline silica (cSiO2, quartz) is etiologically linked to systemic lupus erythematosus (lupus) and other human autoimmune diseases (ADs). In the female NZBWF1 mouse, a widely used animal model that is genetically prone to lupus, short-term repeated intranasal exposure to cSiO2 triggers premature initiation of autoimmune responses in the lungs and kidneys. In contrast to cSiO2's triggering action, consumption of the ω-3 polyunsaturated fatty acid docosahexaenoic acid (DHA) prevents spontaneous onset of autoimmunity in this mouse strain. The aim of this study was to test the hypothesis that consumption of DHA will prevent cSiO2-triggered autoimmunity in the female NZBWF1 mouse. Mice (6 wk old) were fed isocaloric AIN-93G diets containing 0.0, 0.4, 1.2 or 2.4% DHA. Two wk after initiating feeding, mice were intranasally instilled with 1 mg cSiO2 once per wk for 4 wk and maintained on experimental diets for an additional 12 wk. Mice were then sacrificed and the lung, blood and kidney assessed for markers of inflammation and autoimmunity. DHA was incorporated into lung, red blood cells and kidney from diet in a concentration-dependent fashion. Dietary DHA dose-dependently suppressed cSiO2-triggered perivascular leukocyte infiltration and ectopic lymphoid tissue neogenesis in the lung. DHA consumption concurrently inhibited cSiO2-driven elevation of proinflammatory cytokines, B-cell proliferation factors, IgG and anti-dsDNA Ig in both bronchoalveolar lavage fluid and plasma. DHA's prophylactic effects were further mirrored in reduced proteinuria and glomerulonephritis in cSiO2-treated mice. Taken together, these results reveal that DHA consumption suppresses cSiO2 triggering of autoimmunity in female NZBWF1 mice as manifested in the lung, blood and kidney. Our findings provide novel insight into how dietary modulation of the lipidome might be used to prevent or delay triggering of AD by cSiO2. Such knowledge opens the possibility of developing practical, low-cost preventative strategies to reduce the risk of initiating AD and subsequent flaring in cSiO2-exposed individuals. Additional research in this model is required to establish the mechanisms by which DHA suppresses cSiO2-induced autoimmunity and to ascertain unique lipidome signatures predictive of susceptibility to cSiO2-triggered AD.", "Staphylococcal septic arthritis remains a serious medical concern due to rapid and sustained production of inflammatory cytokines that leads to progressive and irreversible joint destruction with high mortality rate in patients despite adequate antibiotics treatment. TNF-α signalling via TNFR-1 contributes to arthritic destruction by aggravating inflammation. Impact of TNFR-2 signalling is not well established in this aspect. Hence the objective of our study was to evaluate the role of dual neutralization TNFR-1 and TNFR-2 in the pathogenesis of S. aureus infection induced septic arthritis. Mice were infected with live S. aureus (5 × 106 cells/ml) followed by administration of TNFR-1and TNFR-2 neutralizing antibody. To measure arthritis index and osteoclastogenesis, histology result in joint tissue and TRAP staining images of arthritis joints have been performed respectively. Maximum reduction in the joint and paw swelling was observed in infected mice treated with both TNFR-1 and TNFR-2 antibody. NF-κB signalling was found to be mainly regulated by TNFR-1 whereas TNFR-2 significantly modulated JNK pathway. Lowest levels of inflammatory cytokines like TNF-α, IL-1β, IL-6, and IFN-γ were observed in both serum and synovial tissues signifying maximum protection in S. aureus arthritis during combination treatment. However IFN-γ and IL-10 levels were significantly altered by TNFR-2 neutralization that indicates both pro and anti inflammatory role of TNFR-2 respectively. Highest decrement in ROS concentration, iNOS expression with least MPO and lysozyme activity was detected in case of combined neutralization. During the early phase of infection all the aforesaid inflammatory parameters remained elevated due to lack of IL-10 as a result of TNFR-2 neutralization as IL-10 negatively modulates pro inflammatory cytokines. Increase in inflammatory cytokines during early phase might also be responsible for decreased bacterial count in TNFR-2 neutralized groups. Thus it can be suggested that combined administration of TNFR-1 and TNFR-2 antibody has a beneficial effect against the severity of S. aureus induced arthritis." ]
Development of multiple control strategies to mitigate the potential risk of host cell proteins in a concentrated fed-batch mode manufactured product	Multiple control strategies, including a downstream purification process with well-controlled parameters and a comprehensive release or characterization for intermediates or drug substances, were implemented to mitigate the potential risk of host cell proteins (HCPs) in one concentrated fed-batch (CFB) mode manufactured product. A host cell process specific enzyme-linked immunosorbent assay (ELISA) method was developed for the quantitation of HCPs. The method was fully validated and showed good performance including high antibody coverage. This was confirmed by 2D Gel-Western Blot analysis. Furthermore, a LC-MS/MS method with non-denaturing digestion and a long gradient chromatographic separation coupled with data dependent acquisition (DDA) on a Thermo/QE-HF-X mass spectrometer was developed as an orthogonal method to help identify the specific types of HCPs in this CFB product. Because of the high sensitivity, selectivity and adaptability of the new developed LC-MS/MS method, significantly more species of HCP contaminants were able to be identified. Even though high levels of HCPs were observed in the harvest bulk of this CFB product, the development of multiple processes and analytical control strategies may greatly mitigate potential risks and reduce HCPs contaminants to a very low level. No high-risk HCP was identified and the total amount of HCPs was very low in the CFB final product.	[ "Host cell proteins (HCPs) are the predominant class of impurities during manufacturing of therapeutic proteins. Previous reports have successfully shown that HCP characterization by LC-MS/MS ultimately leads to drug products of superior safety and quality. Here, we present two sample preparation strategies to approach the wide dynamic range required and compared them systematically to a standard protocol. First, we describe PreOmics fractionation as an effective 2D offline strategy. Second, we evaluate an alternative digestion approach specifically designed for purified antibodies - native (nondenaturing) digestion. Both protocols increased detection sensitivity as shown by two low level HCP models. Out of a 5 ppm spike of eight common HCPs into antibody product, all spiked proteins were positively identified. Additionally, by Universal Proteomics Standard 1 (UPS-1) spiking we obtained a comprehensive coverage of 77% below 10 ppm for the native digestion. Furthermore, we were able to detect 27% to 173% more HCPs in protein A elution pools of five different antibodies and to reject new concerns of HCP coprecipitation by pellet digestion. Although it encounters new challenges, the native digestion is very attractive due its simplicity and comparability to 2D workflows. However, for complex samples such as mock transfected cell culture fermentation, best results were obtained with peptide fractionation. This study highlights the advantages of both methods and their value to facilitate LC-MS/MS approaches to become an even more powerful tool for HCP profiling.", "Sales of monoclonal antibody (mAbs) therapies exceeded $ 40 billion in 2010 and are expected to reach $ 70 billion by 2015. The majority of the approved antibodies are targeting cancer and autoimmune diseases with the top 5 grossing antibodies populating these two areas. In addition over 100 monoclonal antibodies are in Phase II and III of clinical development and numerous others are in various pre-clinical and safety studies. Commercial production of monoclonal antibodies is one of the few biotechnology manufacturing areas that has undergone significant improvements and standardization over the last ten years. Platform technologies have been established based on the structural similarities of these molecules and the regulatory requirements. These improvements include better cell lines, advent of high-performing media free of animal-derived components, and advances in bioreactor and purification processes. In this chapter we will examine the progress made in antibody production as well as discuss the future of manufacturing for these molecules, including the emergence of single use technologies.", "During the production of recombinant protein products, such as monoclonal antibodies, manufacturers must demonstrate clearance of host cell impurities and contaminants to appropriate levels prior to use in the clinic. These include host cell DNA and RNA, product related contaminants such as aggregates, and importantly host cell proteins (HCPs). Despite the importance of HCP removal, the identity and dynamics of these proteins during cell culture and downstream processing (DSP) are largely unknown. Improvements in technologies such as SELDI-TOF mass spectrometry alongside the gold standard technique of ELISA has allowed semi-quantification of the total HCPs present. However, only recently have techniques been utilized in order to identify those HCPs present and align this with the development of approaches to monitor the dynamics of HCPs during both fermentation and downstream processing. In order to enable knowledge based decisions with regards to improving HCP clearance it is vital to identify potential problematic HCPs on a cell line and product specific basis. Understanding the HCP dynamics will in the future help provide a platform to rationally manipulate and engineer and/or select suitable recombinant CHO cell lines and downstream processing steps to limit problematic HCPs.", "Host cell proteins (HCPs) are process-related impurities that may copurify with biopharmaceutical drug products. Within this class of impurities there are some that are more problematic. These problematic HCPs can be considered high-risk and can include those that are immunogenic, biologically active, or enzymatically active with the potential to degrade either product molecules or excipients used in formulation. Some have been shown to be difficult to remove by purification. Why should the biopharmaceutical industry worry about these high-risk HCPs? What approach could be taken to understand the origin of its copurification and address these high-risk HCPs? To answer these questions, the BioPhorum Development Group HCP Workstream initiated a collaboration among its 26-company team with the goal of industry alignment around high-risk HCPs. The information gathered through literature searches, company experiences, and surveys were used to compile a list of frequently seen problematic/high-risk HCPs. These high-risk HCPs were further classified based on their potential impact into different risk categories. A step-by-step recommendation is provided for establishing a comprehensive control strategy based on risk assessments for monitoring and/or eliminating the known impurity from the process that would be beneficial to the biopharmaceutical industry.", "Host cell proteins (HCPs) constitute a major group of impurities for biologic drugs produced using cell culture technology. HCPs are required to be closely monitored and adequately removed in the downstream process. However, HCPs are a complex mixture of proteins with significantly diverse molecular and immunological properties. An overall understanding of the composition of HCPs and changes in their molecular properties upon changes in upstream and harvest process conditions can greatly facilitate downstream process design. This article describes the use of a comparative proteomic profiling method viz. two-dimensional difference gel electrophoresis (2D-DIGE) to examine HCP composition in the harvest stream of CHO cell culture. The effect of upstream process parameters such as cell culture media, bioreactor control strategy, feeding strategy, and cell culture duration/cell viability on HCP profile was examined using this technique. Among all the parameters studied, cell viability generated the most significant changes on the HCP profile. 2D-DIGE was also used to compare the HCP differences between monoclonal antibody producing and null cell cultures. The HCP species in production cell culture was found to be well represented in null cell culture, which confirms the suitability of using the null cell culture for immunoassay reagent generation. 2D-DIGE is complimentary to the commonly used HCP immunoassay. It provides a direct comparison of the changes in HCP composition under different conditions and can reveal properties (pI, MW) of individual species, whereas the immunoassay sensitively quantifies total HCP amount in a given sample.", "This review focusses on affinity purification of immunoglobulins, a methodology which is a powerful tool to obtain pure and intact antibodies. Affinity techniques allow antibody purification both in a single step chromatographic procedure as well as in complex purification protocols depending on the intention to use the target antibody. The purification strategies for antibodies by interaction with affinity ligands such as antibodies and Fe receptors or low molecular weight compounds are described." ]
lncRNA-mediated pathogenesis of diesel exhaust particles	Exposure to diesel exhaust particles (DEP) has been linked to a variety of adverse health effects, including increased morbidity and mortality from cardiovascular diseases, chronic obstructive pulmonary disease (COPD), metabolic syndrome, and lung cancer. The epigenetic changes caused by air pollution have been associated with increased health risks. However, the exact molecular mechanisms underlying the lncRNA-mediated pathogenesis induced by DEP exposure have not been revealed.	[ "We recently showed that the mammalian genome encodes >1,000 large intergenic noncoding (linc)RNAs that are clearly conserved across mammals and, thus, functional. Gene expression patterns have implicated these lincRNAs in diverse biological processes, including cell-cycle regulation, immune surveillance, and embryonic stem cell pluripotency. However, the mechanism by which these lincRNAs function is unknown. Here, we expand the catalog of human lincRNAs to approximately 3,300 by analyzing chromatin-state maps of various human cell types. Inspired by the observation that the well-characterized lincRNA HOTAIR binds the polycomb repressive complex (PRC)2, we tested whether many lincRNAs are physically associated with PRC2. Remarkably, we observe that approximately 20% of lincRNAs expressed in various cell types are bound by PRC2, and that additional lincRNAs are bound by other chromatin-modifying complexes. Also, we show that siRNA-mediated depletion of certain lincRNAs associated with PRC2 leads to changes in gene expression, and that the up-regulated genes are enriched for those normally silenced by PRC2. We propose a model in which some lincRNAs guide chromatin-modifying complexes to specific genomic loci to regulate gene expression.", "Histone lysine demethylation modification is a critical epigenetic modification. Lysine demethylase 2A (KDM2A), a Jumonji C domain-containing demethylase, demethylates the dimethylated H3 lysine 36 (H3K36) residue and exerts little or no activity on monomethylated and trimethylated H3K36 residues. KDM2A expression is regulated by several factors, such as microRNAs, and the phosphorylation of KDM2A also plays a vital role in its function. KDM2A mainly recognizes the unmethylated region of CpG islands and subsequently demethylates histone H3K36 residues. In addition, KDM2A recognizes and binds to phosphorylated proteins, and promotes their ubiquitination and degradation. KDM2A plays an important role in chromosome remodeling and gene transcription, and is involved in cell proliferation and differentiation, cell metabolism, heterochromosomal homeostasis and gene stability. Notably, KDM2A is crucial for tumorigenesis and progression. In the present review, the documented biological functions of KDM2A in physiological and pathological processes are comprehensively summarized.", "The incidence of diabetes and its associated micro- and macrovascular complications is greatly increasing worldwide. The most prevalent vascular complications of both type 1 and type 2 diabetes include nephropathy, retinopathy, neuropathy and cardiovascular diseases. Evidence suggests that both genetic and environmental factors are involved in these pathologies. Clinical trials have underscored the beneficial effects of intensive glycaemic control for preventing the progression of complications. Accumulating evidence suggests a key role for epigenetic mechanisms such as DNA methylation, histone post-translational modifications in chromatin, and non-coding RNAs in the complex interplay between genes and the environment. Factors associated with the pathology of diabetic complications, including hyperglycaemia, growth factors, oxidant stress and inflammatory factors can lead to dysregulation of these epigenetic mechanisms to alter the expression of pathological genes in target cells such as endothelial, vascular smooth muscle, retinal and cardiac cells, without changes in the underlying DNA sequence. Furthermore, long-term persistence of these alterations to the epigenome may be a key mechanism underlying the phenomenon of 'metabolic memory' and sustained vascular dysfunction despite attainment of glycaemic control. Current therapies for most diabetic complications have not been fully efficacious, and hence a study of epigenetic mechanisms that may be involved is clearly warranted as they can not only shed novel new insights into the pathology of diabetic complications, but also lead to the identification of much needed new drug targets. In this review, we highlight the emerging role of epigenetics and epigenomics in the vascular complications of diabetes and metabolic memory.", "The human genome contains many thousands of long noncoding RNAs (lncRNAs). While several studies have demonstrated compelling biological and disease roles for individual examples, analytical and experimental approaches to investigate these genes have been hampered by the lack of comprehensive lncRNA annotation. Here, we present and analyze the most complete human lncRNA annotation to date, produced by the GENCODE consortium within the framework of the ENCODE project and comprising 9277 manually annotated genes producing 14,880 transcripts. Our analyses indicate that lncRNAs are generated through pathways similar to that of protein-coding genes, with similar histone-modification profiles, splicing signals, and exon/intron lengths. In contrast to protein-coding genes, however, lncRNAs display a striking bias toward two-exon transcripts, they are predominantly localized in the chromatin and nucleus, and a fraction appear to be preferentially processed into small RNAs. They are under stronger selective pressure than neutrally evolving sequences-particularly in their promoter regions, which display levels of selection comparable to protein-coding genes. Importantly, about one-third seem to have arisen within the primate lineage. Comprehensive analysis of their expression in multiple human organs and brain regions shows that lncRNAs are generally lower expressed than protein-coding genes, and display more tissue-specific expression patterns, with a large fraction of tissue-specific lncRNAs expressed in the brain. Expression correlation analysis indicates that lncRNAs show particularly striking positive correlation with the expression of antisense coding genes. This GENCODE annotation represents a valuable resource for future studies of lncRNAs.", "Compared with normal cells, tumor cells display distinct metabolic characteristics. Long non-coding RNAs (lncRNAs), a large class of regulatory RNA molecules with limited or no protein-coding capacity, play key roles in tumorigenesis and progression. Recent advances have revealed that lncRNAs play a vital role in cell metabolism by regulating the reprogramming of the metabolic pathways in cancer cells. LncRNAs could regulate various metabolic enzymes that integrate cell malignant transformation and metabolic reprogramming. In addition to the known functions of lncRNAs in regulating glycolysis and glucose homeostasis, recent studies also implicate lncRNAs in amino acid and lipid metabolism. These observations reveal the high complexity of the malignant metabolism. Elucidating the metabolic-related functions of lncRNAs will provide a better understanding of the regulatory mechanisms of metabolism and thus may provide insights for the clinical development of cancer diagnostics, prognostics and therapeutics.", "Metabolic diseases result from multiple genetic and environmental factors. We report here that one manner in which environmental factors can contribute to metabolic disease progression is through modification to chromatin. We demonstrate that high fat diet leads to chromatin remodeling in the livers of C57BL/6J mice, as compared with mice fed a control diet, and that these chromatin changes are associated with changes in gene expression. We further show that the regions of greatest variation in chromatin accessibility are targeted by liver transcription factors, including HNF4α, CCAAT/enhancer-binding protein α (CEBP/α), and FOXA1. Repeating the chromatin and gene expression profiling in another mouse strain, DBA/2J, revealed that the regions of greatest chromatin change are largely strain-specific and that integration of chromatin, gene expression, and genetic data can be used to characterize regulatory regions. Our data indicate dramatic changes in the epigenome due to diet and demonstrate strain-specific dynamics in chromatin remodeling.", "Misregulation of angiotensin II (Ang II) actions can lead to atherosclerosis and hypertension. Evaluating transcriptomic responses to Ang II in vascular smooth muscle cells (VSMCs) is important to understand the gene networks regulated by Ang II, which might uncover previously unidentified mechanisms and new therapeutic targets. To identify all transcripts, including novel protein-coding and long noncoding RNAs, differentially expressed in response to Ang II in rat VSMCs using transcriptome and epigenome profiling. De novo assembly of transcripts from RNA-sequencing revealed novel protein-coding and long noncoding RNAs (lncRNAs). The majority of the genomic loci of these novel transcripts are enriched for histone H3 lysine-4-trimethylation and histone H3 lysine-36-trimethylation, 2 chromatin modifications found at actively transcribed regions, providing further evidence that these are bonafide transcripts. Analysis of transcript abundance identified all protein-coding and lncRNAs regulated by Ang II. We further discovered that an Ang II-regulated lncRNA functions as the host transcript for miR-221 and miR-222, 2 microRNAs implicated in cell proliferation. Additionally, small interfering RNA-mediated knockdown of Lnc-Ang362 reduced proliferation of VSMCs. These data provide novel insights into the epigenomic and transcriptomic effects of Ang II in VSMCs. They provide the first identification of Ang II-regulated lncRNAs, which suggests functional roles for these lncRNAs in mediating cellular responses to Ang II. Furthermore, we identify an Ang II-regulated lncRNA that is responsible for the production of 2 microRNAs implicated in VSMC proliferation. These newly identified noncoding transcripts could be exploited as novel therapeutic targets for Ang II-associated cardiovascular diseases." ]
Association between non-alcoholic fatty liver disease and juvenile obesity	The prevalence of juvenile obesity is increasing, reaching epidemic proportions, presenting a link not only to NAFLD (non-alcoholic fatty liver disease) but to abnormal lipid profiles and liver enzyme abnormalities. Liver ultrasonography is a sensitive and specific tool for the recognition of NAFLD. This study aims to assess the association between NAFLD and juvenile obesity and to determine the other related changes in a set of indicators, including lipid profile abnormalities and serum transaminases. The sample included 470 obese and 210 non-obese individuals aged 6-16. Anthropometric measures were assessed, with the serum lipid profile and liver transaminases, and abdominal ultrasonography was used to detect NAFLD. Fatty liver was found in 38% of the obese subjects and none of the non-obese subjects. Within obese subjects, mean body mass index (BMI) and waist circumference increased significantly in patients with NAFLD compared to those without fatty liver. Moreover, LDL (low-density lipoprotein), CHOL (cholesterol), and serum liver enzymes were significantly higher in the presence of NAFLD. In conclusion, NAFLD commonly associates with juvenile obesity, relating to obesity and the abnormal lipid profile (including elevated CHOL and LDL) among obese people, reflecting elevated liver transaminases, which increase the risk of cirrhosis.	[ "Nonalcoholic fatty liver disease (NAFLD) is a growing epidemic that encompasses three distinct clinical phenotypes: uncomplicated fatty liver, nonalcoholic steatohepatitis (NASH) and NASH-related cirrhosis with its complications, including hepatocellular carcinoma. To date, no pharmacological treatments have been approved and lifestyle modifications including reduced caloric intake targeting a 7%-10% weight loss from baseline assessment represent the standard approach. Mediterranean diet has been recommended as the best dietary pattern since it is easy to follow and, independently of caloric intake its nutritional components have beneficial metabolic effects that not only improve steatosis but also risk factors for cardiovascular events, the leading cause of morbidity/mortality in individuals with NAFLD. Other dietary patterns such as ketogenic diet and Dietary Approach to Stop Hypertension (DASH) diet can be used in patients with NAFLD. Recently, intermittent fasting diets have gained popularity among healthy individuals and have been proposed as a safe and effective treatment for the metabolic syndrome in experimental and in a few human studies. In this narrative review, we aim to summarize the evidence for the available dietary approaches for patients with NAFLD.", "Obesity and overweight are accompanied with several different chronic diseases. Overweight and obesity can be measured by using body mass index (BMI) and is also used widely as an index of relative adiposity among any population. The aim of the study is to evaluate the prevalence of overweight and obesity among general population in Al-Kharj, Saudi Arabia. Cross-sectional analysis was undertaken from a representative sample (N = 1019) of the Al Kharj population. Anthropometric measurements including the waist circumference (in centimeters), height (in meters), and weight (in kilograms) of the subjects were undertaken by means of standard apparatus. SPSS 24.0 was utilized for statistical analysis of the data. Majority of respondents in this study were overweight and obese (54.3%) compared with 45.7% being non-obese. A linear positive association of increasing BMI with older age groups was present in males and females. Men had larger waist circumference, weight and height measures as compared with their female counterparts. Regression analysis showed increasing age, being married and high serum cholesterol to be the significant predictors of overweight and obesity while gender, education level, job status, and having diabetes were not. The obesity-overweight prevalence in the Saudi population is high mainly across both genders. However, the associated factors are potentially preventable and modifiable. The regional barriers to lifestyle modifications and interventions to encourage active lifestyles, especially among adolescents to limit the occurrence of obesity and ultimately promote health and wellbeing, are warranted. Furthermore, prospective studies are needed in future to confirm the aetiological nature of such associations.", "Physical inactivity and obesity are modifiable risk factors for many chronic diseases, including cardiovascular disease, diabetes mellitus, osteoporosis, osteoarthritis, and depression. Both physical inactivity and obesity are associated with low-grade systemic inflammation that may contribute to the inflammatory processes present in many chronic diseases. In asthma, almost no studies are available in which physical inactivity has been studied using performance-based instruments. In contrast, the association between obesity and a higher prevalence of asthma has often been suggested in a large number of studies. In chronic obstructive pulmonary disease (COPD) physical inactivity has been demonstrated in a few studies that used performance-based instruments; this was associated with the higher COPD Global Initiative on Obstructive Lung Disease (GOLD) stages and a higher degree of systemic inflammation, independent of body mass index. In contrast to physical inactivity, obesity in COPD is associated with the lower GOLD stages. Additionally, obesity is associated with the chronic obstructive phenotype and features of the metabolic syndrome. To elucidate the independent relation of physical inactivity and obesity with systemic inflammation, performance-based studies of physical inactivity in asthma and COPD are highly needed.", "Nonalcoholic fatty liver disease (NAFLD) is a multifactorial metabolic disorder that was first described in 1980. It has been prevalent and on the rise for many years and is associated with other metabolic disorders such as obesity and type 2 diabetes mellitus (T2DM). NAFLD can be best described as a metabolic dysfunction that stems from insulin resistance-induced hepatic lipogenesis. This lipogenesis increases oxidative stress and hepatic inflammation and is often potentiated by genetic and gut microbiome dysfunction. As NAFLD progresses from simple steatosis to non-alcoholic steatohepatitis (NASH) and to cirrhosis and hepatocellular carcinoma (HCC), the odds of complications including cardiovascular disease (CVD), chronic kidney disease (CKD), and overall mortality increase. The aim of this review is to describe the metabolic causes and consequences of NAFLD while examining the risks that each stage of NAFLD poses. In this review, the etiology of \"lean\" NAFLD, the impact of obesity, T2DM, genetics, and microbiome dysbiosis on NAFLD progression are all explored. This review will also discuss the core issue behind the progression of NAFLD: insulin resistance (IR). Upon describing the causes and consequences of NAFLD, the effectiveness of diet modification, lifestyle changes, and glucagon-like peptide 1 receptor (GLP-1) agonists to retard NAFLD progression and stem the rate of complications is examined.", "The effectiveness of pulmonary rehabilitation (PR) has not yet been established in patients with asthma - chronic obstructive pulmonary disease overlap (ACO) depending on their nutritional status. We aimed to evaluate the effectiveness of a short-term PR program in patients with comorbid asthma, chronic obstructive pulmonary disease (COPD), and obesity. We included 40 ACO patients and divided them into 3 groups according to body mass index (BMI) and then subdivided them into PR (n=21) and control (n=19) groups. The COPD Assessment Test (CAT), the Asthma Control Test (ACT), and the modified Medical Research Council dyspnea scale (mMRS) were used to evaluate symptoms levels. BODE index (body mass index, forced expiratory volume in one second, dyspnoea, and 6-min walk distance) was used to evaluate the effectiveness of pulmonary rehabilitation. In addition, spirometry and bioimpedansometry were performed. All measurements were done before and after a 6-month PR program. A significantly lower decline in the BODE index was observed in overweight patients (decreased by 43.6% compared to baseline and lower by 40.7% compared to the control group). The six-minute walking test (6MWT) significantly increased in all groups (p<0.001). There was a decrease in total CAT score by 25.4% and by 31.2% in the overweight group (p<0.001). The BMI decreased more in the obese group (by 9.4% compared to baseline). Our study result showed that early use of PR program significantly improves functional capacity and BODE index, leads to dyspnea and CAT scores reduction and improvement in pulmonary function, cause a decrease in BMI, body fat percentage, and visceral fat level, and an increase in muscle mass in overweight and obese patients with ACO." ]
Megalin imports testosterone bound to sex hormone binding globulin into prostate cells	Vitamin D deficiency is associated with an increased risk of prostate cancer mortality and is hypothesized to contribute to prostate cancer aggressiveness and disparities in African American populations. The prostate epithelium was recently shown to express megalin, an endocytic receptor that internalizes circulating globulin-bound hormones, which suggests regulation of intracellular prostate hormone levels. This contrasts with passive diffusion of hormones that is posited by the free hormone hypothesis. Here, we demonstrate that megalin imports testosterone bound to sex hormone-binding globulin into prostate cells. Prostatic loss of	[ "We sought to determine whether there are racial differences in androgenic stimulation within the prostate tissue microenvironment. Steroid hormones were extracted from snap frozen tissue obtained intraoperatively from radical prostatectomy specimens of 36 black and 59 white Americans. Testosterone, dihydrotestosterone (DHT), androstenedione (ASD), dehydroepiandrosterone, dehydroepiandrosterone sulfate, sex hormone-binding globulin (SHBG) and prostate specific antigen (PSA) were measured using radioimmunoassay. The Wilcoxon 2 group test was performed to compare clinical characteristics and tissue steroid levels between white and black Americans. Nonparametric rank ANOVA was used to consider race and other clinical factors in a multivariable way. Black and white American men were similar with respect to serum PSA, and pathological grade and stage. However, black men were younger (p = 0.01) and had a significantly higher body mass index (p = 0.02). Black and white men had similar testosterone and DHT. However, black men had higher ASD (p = 0.006) and SHBG (p = 0.009). Racial differences in ASD (p = 0.015) and SHBG (p = 0.008) persisted after controlling for age, body mass index, PSA, and pathological Gleason sum and stage. Tissue levels of testosterone and DHT did not differ by race. However, black men had higher tissue ASD and SHBG than white men. Higher tissue ASD did not result in a greater conversion of ASD to testosterone in the prostate of black men. Higher tissue SHBG may activate the androgen receptor through cyclic adenosine monophosphate dependent pathways.", "Although estrogen receptor beta (ERβ) has been implicated in prostate cancer (PCa) progression, its potential role in health disparity of PCa remains elusive. The objective of this study was to examine serum estrogens and prostate tumor ERβ expression and examine their correlation with clinical and pathological parameters in African American (AA) versus Caucasian American (CA) men. The circulating 17β-estradiol (E2) was measured by enzyme immunoassay in blood procured from racially stratified normal subjects and PCa patients. Differential expression profile analysis of ERβ was analyzed by quantitative immunohistochemistry using ethnicity-based tissue microarray encompassing 300 PCa tissue cores. In situ ERβ expression was validated by quantitative reverse transcription-PCR in matched microdissected normal prostate epithelium and tumor cells and datasets extracted from independent cohorts. In comparison with normal age-matched subjects, circulating E2 levels were significantly elevated in all PCa patients. Further analysis demonstrates an increase in blood E2 levels in AA men in both normal and PCa in comparison with age- and stage-matched counterparts of CA decent. Histochemical score analysis reveals intense nuclear immunoreactivity for ERβ in tumor cores of AA men than in CA men. Gene expression analysis in microdissected tumors corroborated the biracial differences in ERβ expression. Gene expression analysis from independent cohort datasets revealed correlation between ERβ expression and PCa progression. However, unlike in CA men, adjusted multivariate analysis showed that ERβ expression correlates with age at diagnosis and low prostate-specific antigen recurrence-free survival in AA men. Taken together, our results suggest that E2-ERβ axis may have potential clinical utility in PCa diagnosis and clinical outcome among AA men.", "Sex steroid hormones are associated with chronic diseases and mortality with risk associations that differ between racial and ethnic groups. However, it is currently unclear whether sex steroid hormone levels differ between black and white men. The aim of this study was to assess racial variation in circulating testosterone, free testosterone, sex hormone-binding globulin (SHBG) and estradiol levels in men. We searched PubMed for articles comparing circulating hormones in black and white men. A meta-analysis was performed using weighted mean differences (WMD) to compare hormones levels between black and white men. Fifteen eligible studies were identified; three did not report adjusted means. After age adjustment, free testosterone levels were significantly higher in black than in white men (WMD = 4.07 pg/mL, 95% CI 1.26, 6.88). Depending on the free testosterone concentration in white men, this WMD translates into a racial difference ranging from 2.5 to 4.9%. Total testosterone (WMD = 0.10 ng/mL, 95% CI -0.02, 0.22), estradiol (WMD = 0.67 pg/mL, 95% CI -0.04, 1.38) and SHBG (WMD = -0.45 nmol/L, 95% CI -1.75, 0.85) concentrations did not differ comparing blacks with whites. After adjustment for age, black men have a modestly but significantly 2.5 to 4.9% higher free testosterone level than white men. Based on previous studies on effects of sex steroid hormones on risk of chronic diseases or mortality, this modest difference is unlikely to explain racial differences in disease risk.", "Prostate cancer is the most common malignancy in males and is the second most common cause of cancer mortality in American men. Polymorphisms have been identified in two genes, the 17-hydroxylase cytochrome P450 gene (CYP17) and the steroid 5-reductase type II gene (SRD5A2) that are involved with androgen biosynthesis and metabolism. The CYP17 A2 allele contains a T-->C transition in the 5' promoter region that creates an additional Sp1-type (CCACC box) promoter site. The SRD5A2 valine to leucine (V89L) polymorphism has been correlated with lower dihydroxytestosterone levels. We tested genotypes in 108 prostate cases and 167 controls along with samples (n = 340) from several different ethnic groups. The CYP17 A2 allele (combined A1/A2 and A2/A2 genotypes) occurred at a higher frequency in Caucasian patients with prostate cancer (70%) than in Caucasian clinical control urology patients (57%), suggesting that the A2 allele may convey increased risk for prostate cancer [odds ratio (OR) = 1.7, 95% confidence interval (CI) = 1.0-3.0]. Blacks and Caucasians had a similar frequency of the A2 genotype (16 and 17%, respectively) while Taiwanese had the highest frequency (27%). The SRD5A2 leucine genotype was most frequent in Taiwanese (28%), intermediate in Caucasians (8.5%) and lowest in Blacks (2.5%). Genotypes having a SRD5A2 leucine allele were somewhat more common in prostate cancer cases (56%) than in controls (49%) (OR = 1.4, 95% CI = 0.8-2.2) but this difference was not significant. These results support the hypothesis that some allelic variants of genes involved in androgen biosynthesis and metabolism may be associated with prostate cancer risk.", "Each year, the American Cancer Society estimates the numbers of new cancer cases and deaths that will occur in the United States and compiles the most recent data on cancer incidence, mortality, and survival. Incidence data, available through 2014, were collected by the Surveillance, Epidemiology, and End Results Program; the National Program of Cancer Registries; and the North American Association of Central Cancer Registries. Mortality data, available through 2015, were collected by the National Center for Health Statistics. In 2018, 1,735,350 new cancer cases and 609,640 cancer deaths are projected to occur in the United States. Over the past decade of data, the cancer incidence rate (2005-2014) was stable in women and declined by approximately 2% annually in men, while the cancer death rate (2006-2015) declined by about 1.5% annually in both men and women. The combined cancer death rate dropped continuously from 1991 to 2015 by a total of 26%, translating to approximately 2,378,600 fewer cancer deaths than would have been expected if death rates had remained at their peak. Of the 10 leading causes of death, only cancer declined from 2014 to 2015. In 2015, the cancer death rate was 14% higher in non-Hispanic blacks (NHBs) than non-Hispanic whites (NHWs) overall (death rate ratio [DRR], 1.14; 95% confidence interval [95% CI], 1.13-1.15), but the racial disparity was much larger for individuals aged <65 years (DRR, 1.31; 95% CI, 1.29-1.32) compared with those aged ≥65 years (DRR, 1.07; 95% CI, 1.06-1.09) and varied substantially by state. For example, the cancer death rate was lower in NHBs than NHWs in Massachusetts for all ages and in New York for individuals aged ≥65 years, whereas for those aged <65 years, it was 3 times higher in NHBs in the District of Columbia (DRR, 2.89; 95% CI, 2.16-3.91) and about 50% higher in Wisconsin (DRR, 1.78; 95% CI, 1.56-2.02), Kansas (DRR, 1.51; 95% CI, 1.25-1.81), Louisiana (DRR, 1.49; 95% CI, 1.38-1.60), Illinois (DRR, 1.48; 95% CI, 1.39-1.57), and California (DRR, 1.45; 95% CI, 1.38-1.54). Larger racial inequalities in young and middle-aged adults probably partly reflect less access to high-quality health care. CA Cancer J Clin 2018;68:7-30. © 2018 American Cancer Society.", "Twin studies suggest a heritable component to circulating sex steroid hormones and sex hormone-binding globulin (SHBG). In the NCI-Breast and Prostate Cancer Cohort Consortium, 874 SNPs in 37 candidate genes in the sex steroid hormone pathway were examined in relation to circulating levels of SHBG (N = 4720), testosterone (N = 4678), 3 alpha-androstanediol-glucuronide (N = 4767) and 17beta-estradiol (N = 2014) in Caucasian men. rs1799941 in SHBG is highly significantly associated with circulating levels of SHBG (P = 4.52 x 10(-21)), consistent with previous studies, and testosterone (P = 7.54 x 10(-15)), with mean difference of 26.9 and 14.3%, respectively, comparing wild-type to homozygous variant carriers. Further noteworthy novel findings were observed between SNPs in ESR1 with testosterone levels (rs722208, mean difference = 8.8%, P = 7.37 x 10(-6)) and SRD5A2 with 3 alpha-androstanediol-glucuronide (rs2208532, mean difference = 11.8%, P = 1.82 x 10(-6)). Genetic variation in genes in the sex steroid hormone pathway is associated with differences in circulating SHBG and sex steroid hormones.", "Each year, the American Cancer Society estimates the numbers of new cancer cases and deaths expected in the United States in the current year and compiles the most recent data on cancer incidence, mortality, and survival based on incidence data from the National Cancer Institute, the Centers for Disease Control and Prevention, and the North American Association of Central Cancer Registries and mortality data from the National Center for Health Statistics. A total of 1,660,290 new cancer cases and 580,350 cancer deaths are projected to occur in the United States in 2013. During the most recent 5 years for which there are data (2005-2009), delay-adjusted cancer incidence rates declined slightly in men (by 0.6% per year) and were stable in women, while cancer death rates decreased by 1.8% per year in men and by 1.5% per year in women. Overall, cancer death rates have declined 20% from their peak in 1991 (215.1 per 100,000 population) to 2009 (173.1 per 100,000 population). Death rates continue to decline for all 4 major cancer sites (lung, colorectum, breast, and prostate). Over the past 10 years of data (2000-2009), the largest annual declines in death rates were for chronic myeloid leukemia (8.4%), cancers of the stomach (3.1%) and colorectum (3.0%), and non-Hodgkin lymphoma (3.0%). The reduction in overall cancer death rates since 1990 in men and 1991 in women translates to the avoidance of approximately 1.18 million deaths from cancer, with 152,900 of these deaths averted in 2009 alone. Further progress can be accelerated by applying existing cancer control knowledge across all segments of the population, with an emphasis on those groups in the lowest socioeconomic bracket and other underserved populations.", "Despite considerable racial and geographical differences in human phenotypes and in the incidence of diseases that may be associated with sex steroid action, there are few data concerning variation in sex steroid levels among populations. We designed an international study to determine the degree to which geography and race influence sex steroid levels in older men. Using mass spectrometry, concentrations of serum androgens, estrogens, and sex steroid precursors/metabolites were measured in 5003 older men from five countries. SHBG levels were assessed using radioimmunoassay. There was substantial geographical variation in the levels of sex steroids, precursors, and metabolites, as well as SHBG. For instance, Asian men in Hong Kong and Japan, but not in the United States, had levels of total testosterone approximately 20% higher than in other groups. Even greater variation was present in levels of estradiol, SHBG, and dihydrotestosterone. Group differences in body mass index did not explain most geographical differences. In addition, body mass index-independent racial differences were present; Black men had higher levels of estrogens (estradiol, estrone), and Asian men had lower levels of glucuronidated androgen metabolites. On a global scale, there are important geographical and racial differences in the concentrations of serum sex steroids and SHBG in older men.", "The primary objective of the REDUCE (REduction by DUtasteride of prostate Cancer Events) Follow-Up Study was to collect data on the occurrence of newly diagnosed prostate cancers for 2 years beyond the 4-year REDUCE study. The 4-year REDUCE study evaluated prostate cancer risk reduction in men taking dutasteride. This 2-year observational study followed men from REDUCE with a clinic visit shortly after study conclusion and with up to 2 annual telephone calls during which patient reported data were collected regarding prostate cancer events, chronic medication use, prostate specific antigen levels and serious adverse events. No study drug was provided and all biopsies during the 2-year followup were performed for cause. The primary objective was to collect data on the occurrence of new biopsy detectable prostate cancers. Secondary end points included assessment of Gleason score and serious adverse events. A total of 2,751 men enrolled in the followup study with numbers similar to those of the REDUCE former treatment groups (placebo and dutasteride). Few new prostate cancers were detected during the 2-year followup period in either former treatment group. A greater number of cancers were detected in the former dutasteride group than in the former placebo group (14 vs 7 cases). No Gleason score 8-10 prostate cancers were detected in either former treatment group based on central pathology review. No new safety issues were identified during the study. Two years of followup of the REDUCE study cohort demonstrated a low rate of new prostate cancer diagnoses in the former placebo and dutasteride treated groups. No new Gleason 8-10 cancers were detected.", "Age trends in estradiol and estrone levels in men and how lifestyle factors, comorbid conditions, testosterone, and sex hormone-binding globulin affect these age trends remain poorly understood, and were examined in men of the Framingham Heart Study. Estrone and estradiol concentrations were measured in morning fasting samples using liquid chromatography tandem mass spectrometry in men of Framingham Offspring Generation. Free estradiol was calculated using a law of mass action equation. There were 1,461 eligible men (mean age [±SD] 61.1±9.5 years and body mass index [BMI] 28.8±4.5kg/m(2)). Total estradiol and estrone were positively associated with age, but free estradiol was negatively associated with age. Age-related increase in total estrone was greater than that in total estradiol. Estrone was positively associated with smoking, BMI, and testosterone, and total and free estradiol with diabetes, BMI, testosterone, and comorbid conditions; additionally, free estradiol was associated negatively with smoking. Collectively, age, BMI, testosterone, and other health and behavioral factors explained only 18% of variance in estradiol, and 9% of variance in estrone levels. Men in the highest quintile of estrone levels had significantly higher age and BMI, and a higher prevalence of smoking, diabetes, and cardiovascular disease than others, whereas those in the highest quintile of estradiol had higher BMI than others. Total estrone and estradiol levels in men, measured using liquid chromatography tandem mass spectrometry, revealed significant age-related increases that were only partially accounted for by cross-sectional differences in BMI, diabetes status, and other comorbidities and health behaviors. Longitudinal studies are needed to confirm these findings." ]
The rs13702 variant in the lipoprotein lipase gene is linked to non-alcoholic fatty liver disease	Progression of alcohol-associated liver disease (ALD) is driven by genetic predisposition. The rs13702 variant in the lipoprotein lipase (LPL) gene is linked to non-alcoholic fatty liver disease. We aimed at clarifying its role in ALD.	[ "It is unclear if hepatocellular carcinoma (HCC) risk declines over time after hepatitis C virus (HCV) eradication. We analyzed changes in HCC annual incidence over time following HCV eradication and identified dynamic markers of HCC risk. We identified 48,135 patients who initiated HCV antiviral treatment from 2000 through 2015 and achieved a sustained virologic response (SVR) in the Veterans Health Administration (29,033 treated with direct-acting antiviral [DAA] agents and 19,102 treated with interferon-based regimens). Patients were followed after treatment until February 14, 2019 (average 5.4 years), during which 1509 incident HCCs were identified. Among patients with cirrhosis before treatment with DAAs (n = 9784), those with pre-SVR fibrosis-4 (FIB-4) scores ≥3.25 had a higher annual incidence of HCC (3.66%/year) than those with FIB-4 scores <3.25 (1.16%/year) (adjusted hazard ratio 2.14; 95% confidence interval 1.66-2.75). In DAA-treated patients with cirrhosis and FIB-4 scores ≥3.25, annual HCC risk decreased from 3.8%/year in the first year after SVR to 2.4%/year by the fourth year (P=.01). In interferon-treated patients with FIB-4 scores ≥3.25, annual HCC risk remained above 2%/year, even 10 years after SVR. A decrease in FIB-4 scores from ≥3.25 pre-SVR to <3.25 post-SVR was associated with an approximately 50% lower risk of HCC, but the absolute annual risk remained above 2%/year. Patients without cirrhosis before treatment (n = 38,351) had a low risk of HCC, except for those with pre-SVR FIB-4 scores ≥3.25 (HCC risk 1.22%/year) and post-SVR FIB-4 scores ≥3.25 (HCC risk 2.39%/year); risk remained high for many years after SVR. Patients with cirrhosis before an SVR to treatment for HCV infection continue to have a high risk for HCC (>2%/year) for many years, even if their FIB-4 score decreases, and should continue surveillance. Patients without cirrhosis but with FIB-4 scores ≥3.25 have a high enough risk to merit HCC surveillance, especially if FIB-4 remains ≥3.25 post-SVR.", "Aside from the multikinase inhibitor sorafenib, there are no effective systemic therapies for the treatment of advanced hepatocellular carcinoma. To assess the efficacy of everolimus in patients with advanced hepatocellular carcinoma for whom sorafenib treatment failed. EVOLVE-1 was a randomized, double-blind, phase 3 study conducted among 546 adults with Barcelona Clinic Liver Cancer stage B or C hepatocellular carcinoma and Child-Pugh A liver function whose disease progressed during or after sorafenib or who were intolerant of sorafenib. Patients were enrolled from 17 countries between May 2010 and March 2012. Randomization was stratified by region (Asia vs rest of world) and macrovascular invasion (present vs absent). Everolimus, 7.5 mg/d, or matching placebo, both given in combination with best supportive care and continued until disease progression or intolerable toxicity. Per the 2:1 randomization scheme, 362 patients were randomized to the everolimus group and 184 patients to the placebo group. The primary end point was overall survival. Secondary end points included time to progression and the disease control rate (the percentage of patients with a best overall response of complete or partial response or stable disease). No significant difference in overall survival was seen between treatment groups, with 303 deaths (83.7%) in the everolimus group and 151 deaths (82.1%) in the placebo group (hazard ratio [HR], 1.05; 95% CI, 0.86-1.27; P = .68; median overall survival, 7.6 months with everolimus, 7.3 months with placebo). Median time to progression with everolimus and placebo was 3.0 months and 2.6 months, respectively (HR, 0.93; 95% CI, 0.75-1.15), and disease control rate was 56.1% and 45.1%, respectively (P = .01). The most common grade 3/4 adverse events for everolimus vs placebo were anemia (7.8% vs 3.3%, respectively), asthenia (7.8% vs 5.5%, respectively), and decreased appetite (6.1% vs 0.5%, respectively). No patients experienced hepatitis C viral flare. Based on central laboratory results, hepatitis B viral reactivation was experienced by 39 patients (29 everolimus, 10 placebo); all cases were asymptomatic, but 3 everolimus recipients discontinued therapy. Everolimus did not improve overall survival in patients with advanced hepatocellular carcinoma whose disease progressed during or after receiving sorafenib or who were intolerant of sorafenib. clinicaltrials.gov Identifier: NCT01035229.", "The precise staging of hepatocellular carcinoma (HCC) based on the size and number of lesions that predict recurrence after orthotopic liver transplantation (OLT) has not been clearly established. We therefore analyzed the outcome of 70 consecutive patients with cirrhosis and HCC who underwent OLT over a 12-year period at our institution. Pathologic tumor staging of the explanted liver was based on the American Tumor Study Group modified Tumor-Node-Metastases (TNM) Staging Classification. Tumor recurrence occurred in 11.4% of patients after OLT. The Kaplan-Meier survival rates at 1 and 5 years were 91.3% and 72.4%, respectively, for patients with pT1 or pT2 HCC; and 82.4% and 74.1%, respectively, for pT3 tumors (P =.87). Patients with pT4 tumors, however, had a significantly worse 1-year survival of 33.3% (P =.0001). An alpha-fetoprotein (AFP) level > 1,000 ng/mL, total tumor diameter > 8 cm, age > or = 55 years and poorly differentiated histologic grade were also significant predictors for reduced survival in univariate analysis. Only pT4 stage and total tumor diameter remained statistically significant in multivariate analysis. Patients with HCC meeting the following criteria: solitary tumor < or = 6.5 cm, or < or = 3 nodules with the largest lesion < or = 4.5 cm and total tumor diameter < or = 8 cm, had survival rates of 90% and 75.2%, at 1 and 5 years, respectively, after OLT versus a 50% 1-year survival for patients with tumors exceeding these limits (P =.0005). We conclude that the current criteria for OLT based on tumor size may be modestly expanded while still preserving excellent survival after OLT.", "Liver cancer remains a global health challenge, with an estimated incidence of >1 million cases by 2025. Hepatocellular carcinoma (HCC) is the most common form of liver cancer and accounts for ~90% of cases. Infection by hepatitis B virus and hepatitis C virus are the main risk factors for HCC development, although non-alcoholic steatohepatitis associated with metabolic syndrome or diabetes mellitus is becoming a more frequent risk factor in the West. Moreover, non-alcoholic steatohepatitis-associated HCC has a unique molecular pathogenesis. Approximately 25% of all HCCs present with potentially actionable mutations, which are yet to be translated into the clinical practice. Diagnosis based upon non-invasive criteria is currently challenged by the need for molecular information that requires tissue or liquid biopsies. The current major advancements have impacted the management of patients with advanced HCC. Six systemic therapies have been approved based on phase III trials (atezolizumab plus bevacizumab, sorafenib, lenvatinib, regorafenib, cabozantinib and ramucirumab) and three additional therapies have obtained accelerated FDA approval owing to evidence of efficacy. New trials are exploring combination therapies, including checkpoint inhibitors and tyrosine kinase inhibitors or anti-VEGF therapies, or even combinations of two immunotherapy regimens. The outcomes of these trials are expected to change the landscape of HCC management at all evolutionary stages.", "Lipids are a complex and diverse group of molecules with crucial roles in many physiological processes, as well as in the onset, progression, and maintenance of cancers. Fatty acids and cholesterol are the building blocks of lipids, orchestrating these crucial metabolic processes. In the liver, lipid alterations are prevalent as a cause and consequence of chronic hepatitis B and C virus infections, alcoholic hepatitis, and non-alcoholic fatty liver disease and steatohepatitis. Recent developments in lipidomics have also revealed that dynamic changes in triacylglycerols, phospholipids, sphingolipids, ceramides, fatty acids, and cholesterol are involved in the development and progression of primary liver cancer. Accordingly, the transcriptional landscape of lipid metabolism suggests a carcinogenic role of increasing fatty acids and sterol synthesis. However, limited mechanistic insights into the complex nature of the hepatic lipidome have so far hindered the development of effective therapies.", "Hepatocellular carcinoma (HCC) is a highly heterogeneous disease, and prior attempts to develop genomic-based classification for HCC have yielded highly divergent results, indicating difficulty in identifying unified molecular anatomy. We performed a meta-analysis of gene expression profiles in data sets from eight independent patient cohorts across the world. In addition, aiming to establish the real world applicability of a classification system, we profiled 118 formalin-fixed, paraffin-embedded tissues from an additional patient cohort. A total of 603 patients were analyzed, representing the major etiologies of HCC (hepatitis B and C) collected from Western and Eastern countries. We observed three robust HCC subclasses (termed S1, S2, and S3), each correlated with clinical parameters such as tumor size, extent of cellular differentiation, and serum alpha-fetoprotein levels. An analysis of the components of the signatures indicated that S1 reflected aberrant activation of the WNT signaling pathway, S2 was characterized by proliferation as well as MYC and AKT activation, and S3 was associated with hepatocyte differentiation. Functional studies indicated that the WNT pathway activation signature characteristic of S1 tumors was not simply the result of beta-catenin mutation but rather was the result of transforming growth factor-beta activation, thus representing a new mechanism of WNT pathway activation in HCC. These experiments establish the first consensus classification framework for HCC based on gene expression profiles and highlight the power of integrating multiple data sets to define a robust molecular taxonomy of the disease.", "Hepatitis C virus (HCV) is the main cause of chronic hepatitis worldwide. Chronic hepatitis ultimately results in the development of hepatocellular carcinoma (HCC). However, the mechanism of hepatocarcinogenesis in chronic HCV infection is still unclear. The ability of the core protein of HCV to modulate gene transcription, cell proliferation and cell death may be involved in the pathogenesis of HCC. Here, we report the development of HCC in two independent lines of mice transgenic for the HCV core gene, which develop hepatic steatosis early in life as a histological feature characteristic of chronic hepatitis C. After the age of 16 months, mice of both lines developed hepatic tumors that first appeared as adenomas containing fat droplets in the cytoplasm. Then HCC, a more poorly-differentiated neoplasia, developed from within the adenomas, presenting in a 'nodule-in-nodule' manner without cytoplasmic fat droplets; this closely resembled the histopathological characteristics of the early stage of HCC in patients with chronic hepatitis C. These results indicate that the HCV core protein has a chief role in the development of HCC, and that these transgenic mice provide good animal models for determining the molecular events in hepatocarcinogenesis with HCV infection.", "Hepatocellular carcinoma (HCC) is the most common primary liver malignancy. Here, we performed high-resolution copy-number analysis on 125 HCC tumors and whole-exome sequencing on 24 of these tumors. We identified 135 homozygous deletions and 994 somatic mutations of genes with predicted functional consequences. We found new recurrent alterations in four genes (ARID1A, RPS6KA3, NFE2L2 and IRF2) not previously described in HCC. Functional analyses showed tumor suppressor properties for IRF2, whose inactivation, exclusively found in hepatitis B virus (HBV)-related tumors, led to impaired TP53 function. In contrast, inactivation of chromatin remodelers was frequent and predominant in alcohol-related tumors. Moreover, association of mutations in specific genes (RPS6KA3-AXIN1 and NFE2L2-CTNNB1) suggested that Wnt/β-catenin signaling might cooperate in liver carcinogenesis with both oxidative stress metabolism and Ras/mitogen-activated protein kinase (MAPK) pathways. This study provides insight into the somatic mutational landscape in HCC and identifies interactions between mutations in oncogene and tumor suppressor gene mutations related to specific risk factors.", "Non-alcoholic fatty liver disease (NAFLD) is frequent in patients with features of the metabolic syndrome (MetS), obesity, or type 2 diabetes. Lipoprotein lipase (LPL) is the main driver of triglyceride (TG) hydrolysis in chylomicrons and very-low density lipoproteins (VLDL). In some patients with MetS, dysfunction of this pathway can lead to plasma TG values > 10 mmol/L (multifactorial chylomicronemia or MCS). Chylomicronemia also characterizes LPL deficiency (LPLD), a rare autosomal recessive disease called familial chylomicronemia syndrome (FCS), which is associated with an increased risk of recurrent pancreatitis. This study aims to investigate the expression of NAFLD, as assessed by transient elastography, in MCS and FCS subjects. Data were obtained from 38 subjects with chylomicronemia; 19 genetically confirmed FCS and 19 sex- and age-matched MCS. All participants underwent liver ultrasonography and stiffness measurement after a 4-h fast using transient elastography (FibroScan®, Echosens, Waltham, MA, USA). NAFLD (controlled attenuation parameter (CAP) > 280 dB/m) was observed in 42.1% of FCS and 73.7% of MCS subjects (p = 0.05). FCS subjects had lower body mass index (BMI) than MCS. Only 25% of FCS subjects with NAFLD had a BMI ≥ 30 compared to 64.3% in MCS (p = 0.004). In FCS, NAFLD occurred even in the presence of very low (≤18 kg/m2) BMI. In both FCS and MCS, CAP was negatively associated with acute pancreatitis risk. In this study, NAFLD was commonly observed in both FCS and MCS subjects and occurred independently of the BMI and fasting glucose values in FCS; NAFLD was associated with a lower occurrence of acute pancreatitis episodes.", "Endoplasmic reticulum (ER) stress has been implicated in the pathogenesis of viral hepatitis, insulin resistance, hepatosteatosis, and nonalcoholic steatohepatitis (NASH), disorders that increase risk of hepatocellular carcinoma (HCC). To determine whether and how ER stress contributes to obesity-driven hepatic tumorigenesis we fed wild-type (WT) and MUP-uPA mice, in which hepatocyte ER stress is induced by plasminogen activator expression, with high-fat diet. Although both strains were equally insulin resistant, the MUP-uPA mice exhibited more liver damage, more immune infiltration, and increased lipogenesis and, as a result, displayed classical NASH signs and developed typical steatohepatitic HCC. Both NASH and HCC development were dependent on TNF produced by inflammatory macrophages that accumulate in the MUP-uPA liver in response to hepatocyte ER stress." ]
Population dynamics of H9N2 avian influenza viruses in live-poultry markets in China	H9N2 avian influenza viruses are endemic and persistent in China, but those that are prevalent in different provinces are also causes of wide epidemics, related to the spread of wild birds and the cross-regional trade in live poultry. For the past 4 years, beginning in 2018, we have sampled a live-poultry market in Foshan, Guangdong, in this ongoing study. In addition to the prevalence of H9N2 avian influenza viruses in China during this period, we identified isolates from the same market belonging to clade A and clade B, which diverged in 2012-2013, and clade C, which diverged in 2014-2016, respectively. An analysis of population dynamics revealed that, after a critical divergence period from 2014 to 2016, the genetic diversity of H9N2 viruses peaked in 2017. Our spatiotemporal dynamics analysis found that clade A, B, and C, which maintain high rates of evolution, have different prevalence ranges and transmission paths. Clades A and B were mainly prevalent in East China in the early stage, and then spread to Southern China, becoming epidemic with clade C. Strains from different regions converge at the same live-poultry market to communicate, which may be one reasons the H9N2 viruses are difficult to eradicate and increasingly dominant throughout China. Selection pressure and molecular analysis have demonstrated that single amino acid polymorphisms at key receptor binding sites 156, 160, and 190 under positive selection pressure, suggesting that H9N2 viruses are undergoing mutations to adapt to new hosts. Live-poultry markets are important because people who visit them have frequent contact with poultry, H9N2 viruses from different regions converge at these markets and spread through contact between live birds and humans, generating increased risks of human exposure to these viruses and threatening public health safety. Thus, it is important to reducing the cross-regional trade of live poultry and strengthening the monitoring of avian influenza viruses in live-poultry markets to reduce the spread of avian influenza viruses.	[ "Several zoonotic influenza A viruses detected in humans contain genes derived from avian H9N2 subtypes. We uncovered a Eurasian avian-like H1N1 swine influenza virus with polymerase basic 1 and matrix gene segments derived from the H9N2 subtype, suggesting that H9N2 viruses are infecting pigs and reassorting with swine influenza viruses in China.", "Influenza H9N2 subtype viruses and their reassortants (such as H7N9) are posing increasing threats to birds and humans in China. During 2009-2013, multiple novel subtype viruses with H9N2 original genes emerged in China. Yet, the genetic evolution of H9N2 viruses in various host organisms in China has not been systematically investigated since 2009. In the present study, we performed large-scale sequence analysis of H9N2 viral genomes from public databases, representing the spectrum of viruses isolated from birds, mammals and humans in China from 1994 to 2013, and updated the clade classification for each segment. We identified 117 distinct genotypes in 730 H9N2 viruses. We analyzed the sequences of all eight segments in each virus and found three important time points: the years 2000, 2006 and 2010. In the periods divided by these years, genotypic diversity, geographic distribution and host range changed considerably. Genotypic diversity fluctuated greatly in 2000 and 2006. Since 2010, a single genotype became predominant in poultry throughout China, and the eastern coastal region became the newly identified epidemic center. Throughout their 20-year prevalence in China, H9N2 influenza viruses have emerged and adapted from aquatic birds to chickens. The minor avian species and wild birds exacerbated H9N2 genotypes by providing diversified genes, and chickens were the most prevalent vector in which the viruses evolved and expanded their prevalence. It is the necessity for surveillance and disease control on live-bird markets, poultry farms and wild-bird habitats in China.", "Variations in the potential glycosylation sites were observed in hemagglutinin (HA) sequences of H9N2 avian influenza virus isolated in China, deposited in the Influenza Virus Resource of NCBI before 2017, which showed a deleted glycosylation site at amino acid residue 218, and an introduced glycosylation site at amino acid residue 313. Based on the variations in the glycosylation sites at these amino acids, H9N2 avian influenza viruses could be divided into three categories. Firstly, most of the H9N2 influenza viruses were 218G+ viruses; less 313G+ viruses were isolated between 1997 and 2004. Secondly, the occurrence of the 218G+/313G+ viruses increased, while the 218G+/313G- viruses decreased from 2005 to 2012. Thirdly, from 2013 to 2016, the 218G-/313G+ viruses were predominant compared to the 218G+/313G+ viruses. Here, based on an F/98 virus backbone, a 218G+/313G- virus, two reassortment viruses were generated, and named rF/HA218G+/313G+ and rF/HA 218G+/313G-, respectively. HA protein migration demonstrated that the glycosylation sites at amino acid residues 313 and 218 were both functional. The absence of the glycosylation site at amino acid residue 218 and the presence of the glycosylation site at amino acid residue 313 increased antibody binding and moderately prevented the virus from escaping neutralization with homologous antisera. Additionally, compared to the F/98 virus (218G+/313G-), the viruses rF/HA218G+/313G+ or rF/HA218G-/313G+ showed significantly increased infectivity of MDCK cells, chicken embryo eggs, and trachea and lung tissue of SPF chickens, but did not display differences in airborne spread in chickens or infectivity of mice compared with its parental virus F/98.", "Highly pathogenic avian influenza (HPAI) virus (H5N1) has appeared in >60 countries and continues to evolve and diversify at a concerning rate. Because different names have been used to describe emerging lineages of the virus, this study describes a unified nomenclature system to facilitate discussion and comparison of subtype H5N1 lineages.", "The nonsynonymous (amino acid-altering) to synonymous (silent) substitution rate ratio (omega = d(N)/d(S)) provides a measure of natural selection at the protein level, with omega = 1, >1, and <1, indicating neutral evolution, purifying selection, and positive selection, respectively. Previous studies that used this measure to detect positive selection have often taken an approach of pairwise comparison, estimating substitution rates by averaging over all sites in the protein. As most amino acids in a functional protein are under structural and functional constraints and adaptive evolution probably affects only a few sites at a few time points, this approach of averaging rates over sites and over time has little power. Previously, we developed codon-based substitution models that allow the omega ratio to vary either among lineages or among sites. In this paper we extend previous models to allow the omega ratio to vary both among sites and among lineages and implement the new models in the likelihood framework. These models may be useful for identifying positive selection along prespecified lineages that affects only a few sites in the protein. We apply those branch-site models as well as previous branch- and site-specific models to three data sets: the lysozyme genes from primates, the tumor suppressor BRCA1 genes from primates, and the phytochrome (PHY) gene family in angiosperms. Positive selection is detected in the lysozyme and BRCA genes by both the new and the old models. However, only the new models detected positive selection acting on lineages after gene duplication in the PHY gene family. Additional tests on several data sets suggest that the new models may be useful in detecting positive selection after gene duplication in gene family evolution.", "The role of Africa in the dynamics of the global spread of a zoonotic and economically-important virus, such as the highly pathogenic avian influenza (HPAI) H5Nx of the Gs/GD lineage, remains unexplored. Here we characterise the spatiotemporal patterns of virus diffusion during three HPAI H5Nx intercontinental epidemic waves and demonstrate that Africa mainly acted as an ecological sink of the HPAI H5Nx viruses. A joint analysis of host dynamics and continuous spatial diffusion indicates that poultry trade as well as wild bird migrations have contributed to the virus spreading into Africa, with West Africa acting as a crucial hotspot for virus introduction and dissemination into the continent. We demonstrate varying paths of avian influenza incursions into Africa as well as virus spread within Africa over time, which reveal that virus expansion is a complex phenomenon, shaped by an intricate interplay between avian host ecology, virus characteristics and environmental variables.", "DendroPy is a cross-platform library for the Python programming language that provides for object-oriented reading, writing, simulation and manipulation of phylogenetic data, with an emphasis on phylogenetic tree operations. DendroPy uses a splits-hash mapping to perform rapid calculations of tree distances, similarities and shape under various metrics. It contains rich simulation routines to generate trees under a number of different phylogenetic and coalescent models. DendroPy's data simulation and manipulation facilities, in conjunction with its support of a broad range of phylogenetic data formats (NEXUS, Newick, PHYLIP, FASTA, NeXML, etc.), allow it to serve a useful role in various phyloinformatics and phylogeographic pipelines. The stable release of the library is available for download and automated installation through the Python Package Index site (http://pypi.python.org/pypi/DendroPy), while the active development source code repository is available to the public from GitHub (http://github.com/jeetsukumaran/DendroPy)." ]
Effect of dietary methionine restriction on antioxidant function and inflammatory responses in lipopolysaccharide-challenged broilers	In this study, we investigated the effect of dietary methionine restriction (MR) on the antioxidant function and inflammatory responses in lipopolysaccharide (LPS)-challenged broilers reared at high stocking density. A total of 504 one-day-old male Arbor Acre broiler chickens were randomly divided into four treatments: 1) CON group, broilers fed a basal diet; 2) LPS group, LPS-challenged broilers fed a basal diet; 3) MR1 group, LPS-challenged broilers fed a methionine-restricted diet (0.3% methionine); and 4) MR2 group, LPS-challenged broilers fed a methionine-restricted diet (0.4% methionine). LPS-challenged broilers were intraperitoneally injected with 1 mg/kg body weight (BW) of LPS at 17, 19, and 21 days of age, whereas the CON group was injected with sterile saline. The results showed that: LPS significantly increased the liver histopathological score (	[ "Long-Evans female rats three weeks, three months and 13-14 months of age were placed on tryptophan-deficient diets for periods ranging from a few months to nearly two years. Growth was interupted during the period of tryptophan-deficiency, but when the animals were returned to a complete diet, they gained weight and grew to normal size. Ability to reproduce, as indicated by litter production, was present at 17-28 months of age in rats which had been deprived of tryptophan, whereas no controls over 17 months of age produced any offspring. Other signs of delayed aging in the experimental group included, at advanced ages, greater longevity, as well as later onset in the appearance of obvious tumors, and better coat condition and hair regrowth. Many of these effects were also seen in pair-fed controls (fed a diet equal in amount to that eaten by the tryptophan-deprived rats, but with 1-tryptophan added). It is hypothesized that tryptophan deficiency delays growth, development and maturation of the central nervous system (CNS), in particular, by decreasing the levels of the neurotransmitter serotonin, for which tryptophan is the necessary precursor. In a parallel experiment, chronic treatment with d, 1-parachlorophenylalanine, an inhibitor of brain serotonin synthesis, from weaning until adulthood, also inhibited growth (body weight) and delayed sexual maturation (age of vaginal opening). These observations suggest that diets deficient in tryptophan or restricted in calories can affect maturation and aging by interfering with CNS protein synthesis, or neurotransmitter metabolism, or both.", "This study was conducted to investigate the effects of oridonin (ORI) on growth performance and antioxidant capacity in broiler chickens that were repeatedly challenged with lipopolysaccharide (LPS). A total of 384 one-day-old male Arbor Acre broiler chickens were randomly assigned to 8 treatments with 6 replicate cages per treatment and 8 birds per replicate. There were 4 dietary treatments: the control group (birds fed the basal diet), the ORI 50 group, the ORI 80 group, and the ORI 100 group (the basal diet supplemented with 50, 80, and 100 mg/kg oridonin, respectively). Broilers were intraperitoneally injected with either 250 μg/kg BW LPS or an equivalent amount of sterile saline at 16, 18, and 20 d of age. LPS decreased the average daily weight gain (ADG), the average daily feed intake (ADFI), and the feed conversion ratio (FCR) of broiler chickens (P < 0.05); oridonin supplementation had no effects on performance whether before or after LPS injection (P > 0.05). LPS stimulation increased the relative weight of the spleen and bursa (P < 0.05); oridonin inclusion markedly attenuated the increased spleen index (P < 0.05). Additionally, the LPS-induced increases in the concentrations of malondialdehyde (MDA) and decreases in activities of total superoxide dismutase (T-SOD), total antioxidant capacity (T-AOC) and catalase (CAT) were dramatically attenuated by oridonin in both the serum and liver (P < 0.05). Furthermore, LPS down-regulated the mRNA expressions of nuclear factor erythroid 2-related factor 2 (Nrf2), copper and zinc superoxide dismutase (Cu/Zn-SOD), manganese superoxide dismutase (Mn-SOD), glutathione peroxidase (GPx1), and CAT in the liver (P < 0.05), However, oridonin inclusion increased the liver mRNA expression levels of Nrf2, Cu/Zn-SOD, Mn-SOD, CAT, and GPx1 (P < 0.05). It was concluded that the dietary oridonin supplementation at an optimum dose of 100 mg/kg improves the antioxidant capacity in broilers, as evidenced by the decrease in MDA and the increase in total SOD activities and mRNA expression levels of the liver antioxidant genes, although the effects on growth performance was negligible.", "In the present study, two experiments were conducted to investigate the effect of dietary L-arginine (Arg) supplementation on the inflammatory response and innate immunity of broiler chickens. Expt 1 was designed as a 2 × 3 factorial arrangement (n 8 cages/treatment; n 6 birds/cage) with three dietary Arg concentrations (1.05, 1.42 and 1.90%) and two immune treatments (injection of lipopolysaccharide (LPS) or saline) given at an interval of 48 h between 14 and 21 d of age. In Expt 2, correlation between dietary Arg concentration (0.99, 1.39, 1.76, 2.13 or 2.53%) and percentage of circulating B cells (percentage of circulating lymphocytes) was determined. In Expt 1, LPS injection decreased body-weight gain and feed intake and increased feed conversion ratio of the challenged broilers (14-21 d; P< 0.05). LPS injection suppressed (P< 0.05) the percentages of splenic CD11+ and B cells (percentages of splenic lymphocytes) and phagocytic activity of splenic heterophils and macrophages; Arg supplementation linearly decreased the percentages of CD11+, CD14+ and B cells in the spleen (P< 0.10). LPS injection increased (P< 0.05) the expression of IL-1β and IL-6 mRNA in the spleen and caecal tonsils. Arginine supplementation decreased (P< 0.05) the expression of IL-1β, Toll-like receptor 4 (TLR4) and PPAR-γ mRNA in the spleen and IL-1β, IL-10, TLR4 and NF-κB mRNA in the caecal tonsils. In Expt 2, increasing dietary Arg concentrations linearly and quadratically reduced the percentage of circulating B cells (P< 0.01). Collectively, Arg supplementation attenuated the overexpression of pro-inflammatory cytokines probably through the suppression of the TLR4 pathway and CD14+ cell percentage. Furthermore, excessive Arg supplementation (1.76%) suppressed the percentages of circulating and splenic B cells.", "With few exceptions, nutritional and dietary interventions generally impact upon both old-age quality of life and longevity. The life prolonging effects, commonly observed with dietary restriction reportedly are linked to alterations in protein intake and specifically limiting the dietary intake of certain essential amino acids. There is however a paucity of data methodically evaluating the various essential amino acids on health- and lifespan and the mechanisms involved. Rodent diets containing either lower methionine content, or tryptophan, than that found in commercially available chow, appear to elicit beneficial effects. It is unclear whether all of these favorable effects associated with restricted intake of methionine and tryptophan are due to their specific unique properties or if restriction of other essential amino acids, or proteins in general, may produce similar results. Considerably more work remains to be done to elucidate the mechanisms by which limiting these vital molecules may delay the onset of age-associated diseases and improve quality of life at older ages.", "Inflammation is a crucial component of various stress-induced responses that contributes to the pathogenesis of major depressive disorder (MDD). Depressive-like behavior (DLB) is characterized by decreased mobility and depressive behavior that occurs in systemic infection induced by Lipopolysaccharide (LPS) in experimental animals and is considered as a model of exacerbation of MDD. We assessed the effects of melatonin on behavioral changes and inflammatory cytokine expression in hippocampus of mice in LPS-induced DLB, as well as its effects on NLR Family Pyrin Domain Containing 3 (NLRP3) inflammasome activation, oxidative stress and pyroptotic cell death in murine microglia in vitro. Intraperitoneal 5 mg/kg dose of LPS was used to mimic depressive-like behaviors and melatonin was given at a dose of 500 mg/kg for 4 times with 6 h intervals, starting at 2 h before LPS administration. Behavioral assessment was carried out at 24 h post-LPS injection by tail suspension and forced swimming tests. Additionally, hippocampal cytokine and NLRP3 protein levels were estimated. Melatonin increased mobility time of LPS-induced DLB mice and suppressed NLRP3 expression and interleukin-1β (IL-1β) cleavage in the hippocampus. Immunofluorescence staining of hippocampal tissue showed that NLRP3 is mainly expressed in ionized calcium-binding adapter molecule 1 (Iba1) -positive microglia. Our results show that melatonin prevents LPS and Adenosine triphosphate (ATP) induced NLRP3 inflammasome activation in murine microglia in vitro, evidenced by inhibition of NLRP3 expression, Apoptosis-associated speck-like protein containing a CARD (ASC) speck formation, caspase-1 cleavage and interleukin-1β (IL-1β) maturation and secretion. Additionally, melatonin inhibits pyroptosis, production of mitochondrial and cytosolic reactive oxygen species (ROS) and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling. The beneficial effects of melatonin on NLRP3 inflammasome activation were associated with nuclear factor erythroid 2-related factor 2 (Nrf2) and Silent information regulator 2 homolog 1 (SIRT1) activation, which were reversed by Nrf2 siRNA and SIRT1 inhibitor treatment.", "Newly weaned female rats fed diets severely deficient in the essential amino acid tryptophan show marked delays in reproductive aging, with conception and delivery occurring as late as 36 months. The rate of aging in these rats seems inversely related to both their early growth rates and the accessibility of brain tryptophan. The subsequent age retardation may depend on a reduction in both early cell loss and rate of brain maturation.", "In a previous phase I clinical trial of dietary methionine (MET) restriction with cystemustine treatment for melanoma or glioma, we determined the optimal MET-free diet duration to be 1 day. On this basis, a phase II clinical trial was initiated to evaluate safety and efficacy of this combination. Twenty-two patients (20 with metastatic melanoma and 2 with recurrent gloma) received a median of 4 cycles of the association of a 1-day MET-free diet with cystemustine (60 mg/m(2)) every two weeks. This association was well tolerated (toxicity and nutritional status). Toxicity remained mainly hematological and consisted of WHO grade 3-4 thrombocytopenia, leucopenia and neutropenia in 36, 27 and 27% of patients respectively. The median disease-free survival was 1.8 months and the median survival was 4.6 months, with 2 long-duration stabilizations. The plasmatic MET depletion obtained was of 40 + or - 31%." ]
How do I get a list of all the elements in a list?	Calcium (Ca	[ "Our knowledge about genes involved in the control of basal motor activity that may contribute to the pathology of the hyperactivity disorders, e.g., attention deficit hyperactivity disorder (ADHD), is limited. Disruption of monoamine neurotransmitter signaling through G protein-coupled receptors (GPCR) is considered to be a major contributing factor to the etiology of the ADHD. Genetic association evidence and functional data suggest that regulators of G protein signaling proteins of the R7 family (R7 RGS) that form obligatory complexes with type 5 G protein beta subunit (Gβ5) and negatively regulate signaling downstream from monoamine GPCRs may play a role in controlling hyperactivity. To test this hypothesis, we conducted behavioral, pharmacological, and neurochemical studies using a genetic mouse model that lacked Gβ5, a subunit essential for the expression of the entire R7 RGS family. Elimination of Gβ5-RGS complexes led to a striking level of hyperactivity that far exceeds activity levels previously observed in animal models. This hyperactivity was accompanied by motor learning deficits and paradoxical behavioral sensitization to a novel environment. Neurochemical studies indicated that Gβ5-RGS-deficient mice had higher sensitivity of inhibitory GPCR signaling and deficits in basal levels, release, and reuptake of dopamine. Surprisingly, pharmacological treatment with monoamine reuptake inhibitors failed to alter hyperactivity. In contrast, blockade of NMDA receptors reversed the expression of hyperactivity in Gβ5-RGS-deficient mice. These findings establish that Gβ5-RGS complexes are critical regulators of monoamine-NMDA receptor signaling cross-talk and link these complexes to disorders that manifest as hyperactivity, impaired learning, and motor dysfunctions.", "Recent systematic reviews have encouraged the psychiatric research community to reevaluate the contours of schizophrenia epidemiology. This paper provides a concise overview of three related systematic reviews on the incidence, prevalence, and mortality associated with schizophrenia. The reviews shared key methodological features regarding search strategies, analysis of the distribution of the frequency estimates, and exploration of the influence of key variables (sex, migrant status, urbanicity, secular trend, economic status, and latitude). Contrary to previous interpretations, the incidence of schizophrenia shows prominent variation between sites. The median incidence of schizophrenia was 15.2/100,000 persons, and the central 80% of estimates varied over a fivefold range (7.7-43.0/100,000). The rate ratio for males:females was 1.4:1. Prevalence estimates also show prominent variation. The median lifetime morbid risk for schizophrenia was 7.2/1,000 persons. On the basis of the standardized mortality ratio, people with schizophrenia have a two- to threefold increased risk of dying (median standardized mortality ratio = 2.6 for all-cause mortality), and this differential gap in mortality has increased over recent decades. Compared with native-born individuals, migrants have an increased incidence and prevalence of schizophrenia. Exposures related to urbanicity, economic status, and latitude are also associated with various frequency measures. In conclusion, the epidemiology of schizophrenia is characterized by prominent variability and gradients that can help guide future research.", "Autism spectrum disorders (ASDs) are complex, lifelong, neurodevelopmental conditions of largely unknown cause. They are much more common than previously believed, second in frequency only to mental retardation among the serious developmental disorders. Although a heritable component has been demonstrated in ASD etiology, putative risk genes have yet to be identified. Environmental risk factors may also play a role, perhaps via complex gene-environment interactions, but no specific exposures with significant population effects are known. A number of endogenous biomarkers associated with autism risk have been investigated, and these may help identify significant biologic pathways that, in turn, will aid in the discovery of specific genes and exposures. Future epidemiologic research should focus on expanding population-based descriptive data on ASDs, exploring candidate risk factors in large well-designed studies incorporating both genetic and environmental exposure data and addressing possible etiologic heterogeneity in studies that can stratify case groups and consider alternate endophenotypes.", "Autism spectrum disorder (ASD) is a heterogeneous group of neurodevelopmental disorders without any defined uniting pathophysiology. Ca2+ signaling is emerging as a potential node in the genetic architecture of the disorder. We previously reported decreased inositol trisphosphate (IP3)-mediated Ca2+ release from the endoplasmic reticulum in several rare monogenic syndromes highly comorbid with autism - fragile X and tuberous sclerosis types 1 and 2 syndromes. We now extend those findings to a cohort of subjects with sporadic ASD without any known mutations. We developed and applied a high throughput Fluorometric Imaging Plate Reader (FLIPR) assay to monitor agonist-evoked Ca2+ signals in human primary skin fibroblasts. Our results indicate that IP3 -mediated Ca2+ release from the endoplasmic reticulum in response to activation of purinergic receptors is significantly depressed in subjects with sporadic as well as rare syndromic forms of ASD. We propose that deficits in IP3-mediated Ca2+ signaling represent a convergent hub function shared across the spectrum of autistic disorders - whether caused by rare highly penetrant mutations or sporadic forms - and holds promise as a biomarker for diagnosis and novel drug discovery.", "Disrupted-in-schizophrenia-1 (DISC1) has emerged as a convincing susceptibility gene for multiple mental disorders, but its mechanistic link to the pathogenesis of schizophrenia related psychiatric conditions is yet to be further understood. Here, we showed that DISC1 localizes to the outer surface of the endoplasmic reticulum (ER). EXOC1, a subunit of the exocyst complex, interacted with DISC1 and affected its recruitment to inositol-1,4,5-trisphosphate receptor 1 (IP3R1). Notably, knockdown of DISC1 and EXOC1 elicited an exaggerated ER calcium response upon stimulation of IP3R agonists. Similar abnormal ER calcium responses were observed in hippocampal neurons from DISC1-deficient mutant mice. Moreover, perturbation of ER calcium dynamics upon DISC1 knockdown was effectively reversed by treatment with antipsychotic drugs, such as clozapine and haloperidol. These results collectively indicate that DISC1 is a regulatory factor in ER calcium dynamics, linking a perturbed intracellular calcium signaling and schizophrenia pathogenesis.", "Autism spectrum disorders (ASDs) are childhood neurodevelopmental disorders with complex genetic origins. Previous studies focusing on candidate genes or genomic regions have identified several copy number variations (CNVs) that are associated with an increased risk of ASDs. Here we present the results from a whole-genome CNV study on a cohort of 859 ASD cases and 1,409 healthy children of European ancestry who were genotyped with approximately 550,000 single nucleotide polymorphism markers, in an attempt to comprehensively identify CNVs conferring susceptibility to ASDs. Positive findings were evaluated in an independent cohort of 1,336 ASD cases and 1,110 controls of European ancestry. Besides previously reported ASD candidate genes, such as NRXN1 (ref. 10) and CNTN4 (refs 11, 12), several new susceptibility genes encoding neuronal cell-adhesion molecules, including NLGN1 and ASTN2, were enriched with CNVs in ASD cases compared to controls (P = 9.5 x 10(-3)). Furthermore, CNVs within or surrounding genes involved in the ubiquitin pathways, including UBE3A, PARK2, RFWD2 and FBXO40, were affected by CNVs not observed in controls (P = 3.3 x 10(-3)). We also identified duplications 55 kilobases upstream of complementary DNA AK123120 (P = 3.6 x 10(-6)). Although these variants may be individually rare, they target genes involved in neuronal cell-adhesion or ubiquitin degradation, indicating that these two important gene networks expressed within the central nervous system may contribute to the genetic susceptibility of ASD.", "Preconception and prenatal nutrition is critical for fetal brain development. However, its associations with offspring neurodevelopmental disorders are not well understood. This study aims to systematically review the associations of preconception and prenatal nutrition with offspring risk of neurodevelopmental disorders. We searched the PubMed and Embase for articles published through March 2019. Nutritional exposures included nutrient intake or status, food intake, or dietary patterns. Neurodevelopmental outcomes included autism spectrum disorders (ASD), attention deficit disorder-hyperactivity (ADHD) and intellectual disabilities. A total of 2169 articles were screened, and 20 articles on ASD and 17 on ADHD were eventually reviewed. We found an overall inverse association between maternal folic acid or multivitamin supplementation and children's risk of ASD; a meta-analysis including six prospective cohort studies estimated an RR of ASD of 0.64 (95% CI: 0.46, 0.90). Data on associations of other dietary factors and ASD, ADHD and related outcomes were inconclusive and warrant future investigation. Future studies should integrate comprehensive and more objective methods to quantify the nutritional exposures and explore alternative study design such as Mendelian randomization to evaluate potential causal effects." ]
Multi-fiber metrics analysis of the extracellular matrix in hypertrophic scars	The hypertrophic scar is an aberrant form of wound healing process, whose clinical efficacy is limited by a lack of understanding of its pathophysiology. Remodeling of collagen and elastin fibers in the extracellular matrix (ECM) is closely associated with scar progression. Herein, we perform label-free multiphoton microscopy (MPM) of both fiber components from human skin specimens and propose a multi-fiber metrics (MFM) analysis model for mapping the structural remodeling of the ECM in hypertrophic scars in a highly-sensitive, three-dimensional (3D) manner. We find that both fiber components become wavier and more disorganized in scar tissues, while content accumulation is observed from elastin fibers only. The 3D MFM analysis can effectively distinguish normal and scar tissues with better than 95% in accuracy and 0.999 in the area under the curve value of the receiver operating characteristic curve. Further, unique organizational features with orderly alignment of both fibers are observed in scar-normal adjacent regions, and an optimized combination of features from 3D MFM analysis enables successful identification of all the boundaries. This imaging and analysis system uncovers the 3D architecture of the ECM in hypertrophic scars and exhibits great translational potential for evaluating scars	[ "New high-dimensional, single-cell technologies offer unprecedented resolution in the analysis of heterogeneous tissues. However, because these technologies can measure dozens of parameters simultaneously in individual cells, data interpretation can be challenging. Here we present viSNE, a tool that allows one to map high-dimensional cytometry data onto two dimensions, yet conserve the high-dimensional structure of the data. viSNE plots individual cells in a visual similar to a scatter plot, while using all pairwise distances in high dimension to determine each cell's location in the plot. We integrated mass cytometry with viSNE to map healthy and cancerous bone marrow samples. Healthy bone marrow automatically maps into a consistent shape, whereas leukemia samples map into malformed shapes that are distinct from healthy bone marrow and from each other. We also use viSNE and mass cytometry to compare leukemia diagnosis and relapse samples, and to identify a rare leukemia population reminiscent of minimal residual disease. viSNE can be applied to any multi-dimensional single-cell technology.", "Management of hypertrophic scars and keloids has advanced from crude, invasive methods such as gross excision and radiation to intralesional or topical agents that act on a cellular level. There is no universally accepted treatment regimen and no evidence-based literature to guide management. Our objectives are to present a list of available treatment regimens, their proposed mechanisms of action, and supporting evidence and to perform a meta-analysis of clinical trials to identify treatments with a better-than-even likelihood of improvement. We conducted a PubMed search through October 2005, identifying clinical studies of various treatments for hypertrophic scars and keloids. We graded the quality of each study, delineated the results into favorable vs nonfavorable, and calculated the statistical significance of the findings. The meta-analysis of 70 treatment series for various clinical measures showed a 70% chance of improvement with treatment; however, the mean amount of improvement to be expected was around 60%. There was no statistically significant difference between treatments. Most treatments for keloidal and hypertrophic scarring offer minimal likelihood of improvement. The magnitude of likely permanent improvement in any sign or symptom may be clinically meaningful but far short of cure. Novel therapies deserve further investigation but remain without proven benefit to date.", "Scars are a natural part of dermal healing processes and consist of networks of fibrous collagen tissue, laid down in response to injury to the dermis. In some genetically susceptible individuals, the process of scar tissue formation is excessive, and there is an over-secretion of collagen. This causes the formation of benign tumours (raised) scars known as keloid and hypertrophic scars. Scars can have many significant functional, cosmetic and psychological sequelae. Steroids are used therapeutically in the management of abnormal scars; however, this is associated with a variety of adverse effects. Their intralesional administration is the most widely used and most effective treatment modality for raised skin scars today. Despite this, our understanding of the way in which they work is poor and literature on the topic is, to date, lacking. Further work is needed to clarify the exact mechanisms that bring about abnormal scarring, to aid our understanding of the disease and facilitate the development of more evidence-based treatment strategies.", "With recent advances in cancer therapeutics, there is a great need for improved imaging methods for characterizing cancer onset and progression in a quantitative and actionable way. Collagen, the most abundant extracellular matrix protein in the tumor microenvironment (and the body in general), plays a multifaceted role, both hindering and promoting cancer invasion and progression. Collagen deposition can defend the tumor with immunosuppressive effects, while aligned collagen fiber structures can enable tumor cell migration, aiding invasion and metastasis. Given the complex role of collagen fiber organization and topology, imaging has been a tool of choice to characterize these changes on multiple spatial scales, from the organ and tumor scale to cellular and subcellular level. Macroscale density already aids in the detection and diagnosis of solid cancers, but progress is being made to integrate finer microscale features into the process. Here we review imaging modalities ranging from optical methods of second harmonic generation (SHG), polarized light microscopy (PLM), and optical coherence tomography (OCT) to the medical imaging approaches of ultrasound and magnetic resonance imaging (MRI). These methods have enabled scientists and clinicians to better understand the impact collagen structure has on the tumor environment, at both the bulk scale (density) and microscale (fibrillar structure) levels. We focus on imaging methods with the potential to both examine the collagen structure in as natural a state as possible and still be clinically amenable, with an emphasis on label-free strategies, exploiting intrinsic optical properties of collagen fibers.", "Keloid and hypertrophic scars are common and are caused by a proliferation of dermal tissue following skin injury. They cause functional and psychological problems for patients, and their management can be difficult. The use of silicon gel sheeting to prevent and treat hypertrophic scarring is still relatively new, and started in 1981 with treatment of burn scars. To determine the effectiveness of silicon gel sheeting for: (1) prevention of hypertrophic or keloid scarring in people with newly healed wounds (e.g. post surgery); (2) treatment of established scarring in people with existing keloid or hypertrophic scars. Trials were identified from searches of the Cochrane Wounds Group Specialised Register (searched September 2005), the Cochrane Central Register of Controlled Trials (The Cochrane Library Issue 3, 2005); MEDLINE (1989 to June 2002); EMBASE (1988 to May 2002); CINAHL (1982 to May 2002) and reference lists of articles and relevant reviews. The major supplier of silicon gel sheeting (Smith and Nephew) was approached for details of unpublished, ongoing and recently published trials. Any randomised or quasi-randomised controlled trials, or controlled clinical trials comparing silicon gel sheeting for prevention or treatment of hypertrophic or keloid scars against no treatment, placebo, or any other treatment type except surgery. All relevant trials were assessed for methodological quality. Data were extracted independently by both reviewers using a standardized form, and the results cross-checked. All trials, meeting the selection criteria were assessed for methodological quality. Thirteen trials, involving 559 people, ranging in age from 2 to 81 years, were included in the review. The trials compared adhesive silicon gel sheeting with control; non-silicon gel sheeting; silicon gel plates with added Vitamin E; laser therapy; triamcinolone acetonide injection, and non-adhesive silicon gel sheeting. In the prevention studies, when compared with a no treatment option; whilst silicon gel sheeting reduced the incidence of hypertrophic scarring in people prone to scarring, (RR 0.46, 95% CI 0.21 to 0.98) these studies were highly susceptible to bias. Silicon gel sheeting produced a statistically significant improvement in scar elasticity, (RR 8.60, 95% CI 2.55 to 29.02), but again these studies were highly susceptible to bias. Trials evaluating silicon gel sheeting as a treatment for hypertrophic and keloid scarring are of poor quality and highly susceptible to bias. There is weak evidence of a benefit of silicon gel sheeting as a prevention for abnormal scarring in high risk individuals but the poor quality of research means a great deal of uncertainty prevails.", "The depth of a burn wound and/or its healing potential are the most important determinants of the therapeutic management and of the residual morbidity or scarring. Traditionally, burn surgeons divide burns into superficial which heal by rapid re-epithelialization with minimal scarring and deep burns requiring surgical therapy. Clinical assessment remains the most frequent technique to measure the depth of a burn wound although this has been shown to be accurate in only 60-75% of the cases, even when carried out by an experienced burn surgeon. In this article we review all current modalities useful to provide an objective assessment of the burn wound depth, from simple clinical evaluation to biopsy and histology and to various perfusion measurement techniques such as thermography, vital dyes, video angiography, video microscopy, and laser Doppler techniques. The different needs according to the different diagnostic situations are considered. It is concluded that for the initial emergency assessment, the use of telemetry and simple burn photographs are the best option, that for research purposes a wide range of different techniques can be used but that, most importantly, for the actual treatment decisions, laser Doppler imaging is the only technique that has been shown to accurately predict wound outcome with a large weight of evidence. Moreover this technique has been approved for burn depth assessment by regulatory bodies including the FDA.", "Prestress in tissue is currently detected through destructive methods which obviate both in vivo and longitudinal assessment. We hypothesized that prestress could be detected and quantified by analyzing the microstructure of the extracellular matrix at different spatial scales using non-invasive and non-destructive optical imaging. A simple model of tissue prestress was created using fibroblast-mediated contraction of collagen gels around a central mandrel. Using a quantitative, multiscale, image processing technique, termed generalized image correlation spectroscopy (GICS) of second harmonic images, collagen fiber number and alignment at three different length scales characteristic of the collagen fibril, collagen fiber, and cell were analyzed. GICS fiber alignment (σ(maj/min)) was significantly different across load state, level of prestress, and length scale. The largest fiber ratio, and thus highest alignment, was seen in prestressed, externally loaded gels at a length scale equivalent to the size of the fibroblast cells. Alignment at both fiber and cell scale correlated with prestress in this model. We conclude that GICS of second harmonic images of collagen can predict prestress, and that microstructural organization at the collagen fiber and cell scale are the primary determinants of prestress in cellularized collagen gels.", "Developments in miniaturized microscopes have enabled visualization of brain activities and structural dynamics in animals engaging in self-determined behaviors. However, it remains a challenge to resolve activity at single dendritic spines in freely behaving animals. Here, we report the design and application of a fast high-resolution, miniaturized two-photon microscope (FHIRM-TPM) that accomplishes this goal. With a headpiece weighing 2.15 g and a hollow-core photonic crystal fiber delivering 920-nm femtosecond laser pulses, the FHIRM-TPM is capable of imaging commonly used biosensors (GFP and GCaMP6) at high spatiotemporal resolution (0.64 μm laterally and 3.35 μm axially, 40 Hz at 256 × 256 pixels for raster scanning and 10,000 Hz for free-line scanning). We demonstrate the microscope's robustness with hour-long recordings of neuronal activities at the level of spines in mice experiencing vigorous body movements.", "With rare exceptions, spontaneous tumors originate from a single cell. Yet, at the time of clinical diagnosis, the majority of human tumors display startling heterogeneity in many morphological and physiological features, such as expression of cell surface receptors, proliferative and angiogenic potential. To a substantial extent, this heterogeneity might be attributed to morphological and epigenetic plasticity, but there is also strong evidence for the co-existence of genetically divergent tumor cell clones within tumors. In this perspective, we summarize the sources of intra-tumor phenotypic heterogeneity with emphasis on genetic heterogeneity. We review experimental evidence for the existence of both intra-tumor clonal heterogeneity as well as frequent evolutionary divergence between primary tumors and metastatic outgrowths. Furthermore, we discuss potential biological and clinical implications of intra-tumor clonal heterogeneity.", "The flood of high-dimensional data resulting from mass cytometry experiments that measure more than 40 features of individual cells has stimulated creation of new single cell computational biology tools. These tools draw on advances in the field of machine learning to capture multi-parametric relationships and reveal cells that are easily overlooked in traditional analysis. Here, we introduce a workflow for high dimensional mass cytometry data that emphasizes unsupervised approaches and visualizes data in both single cell and population level views. This workflow includes three central components that are common across mass cytometry analysis approaches: (1) distinguishing initial populations, (2) revealing cell subsets, and (3) characterizing subset features. In the implementation described here, viSNE, SPADE, and heatmaps were used sequentially to comprehensively characterize and compare healthy and malignant human tissue samples. The use of multiple methods helps provide a comprehensive view of results, and the largely unsupervised workflow facilitates automation and helps researchers avoid missing cell populations with unusual or unexpected phenotypes. Together, these methods develop a framework for future machine learning of cell identity." ]
Time-dependent alcohol and mitomycin C in cultured rat limbal stem cells	Ethanol and mitomycin C (MMC) are clinically used to treat corneal diseases such as LASEK and LASIK surgery. In this study, we investigated the effects of time-dependent alcohol and MMC in cultured rat limbal stem cells (LSCs) to determine the appropriate time for the use of this compound in the clinical setting.	[ "To assess relationships between energy, nutrient and food intakes, alcohol consumption, smoking status and body mass index (BMI), and serum concentrations of beta-carotene, alpha-tocopherol, vitamin C, selenium and zinc. Data on health status, alcohol consumption, smoking habits, anthropometric data and biochemical measurements were obtained in 1821 women aged 35-60 y and 1307 men aged 45-60 y, participant to the SU.VI.MAX Study. Data on dietary intake were available on a subsample who reported six 24-h dietary records during the first 18 months of the study. Women had higher baseline serum beta-carotene and vitamin C concentrations and lower concentration for serum vitamin E, zinc and selenium than men. In women, younger age was associated with lowered mean concentration of serum beta-carotene, vitamin E and selenium. In men, only differences were observed for serum zinc, which was lower in older men. Current smokers of both sexes had significantly lower concentrations of serum beta-carotene, vitamin C and selenium, and, only in women, of vitamin E, than nonsmokers. Alcohol consumers had lower concentrations of serum beta-carotene and higher selenium concentrations. Serum beta-carotene and vitamin C concentrations were lower in obese subjects. There were positive associations of dietary beta-carotene, vitamin C and E with their serum concentrations. Age, nutrient and alcohol intakes, serum cholesterol, BMI and smoking status explained 15.2% of the variance of serum beta-carotene in men and 13.9% in women, and 10.8 and 10.0% for serum vitamin C, and 26.3 and 28.6% for serum vitamin E, respectively. Serum antioxidant nutrient concentrations are primarily influenced by sex, age, obesity, tobacco smoking, alcohol consumption and especially dietary intake of those antioxidant nutrients.", "Toxic optic neuropathy (TON) is a disease entity which is not only underdiagnosed, but also often diagnosed at a stage when recovery of vision is not possible. This article gives an overview of common causes, clinical features, and management of TON.", "In this article, we present a review of ocular conditions related to alcohol consumption. A search of the literature published from 1952 to March 2020 was performed. The titles and abstracts were screened and the eligible studies were selected. PubMed, ISI Web of Knowledge database, Scopus, Embase, and the Cochrane Library were searched. We categorized the relationship between alcohol intake and ocular conditions by the type of ocular exposure to alcohol. Accordingly, ocular findings following acute alcohol intoxication, optic neuropathy following methanol toxicity, congenital conditions related to maternal alcohol consumption, and ocular disease related to chronic alcoholism are discussed. The main feature of alcohol intoxication in the eye is abnormal eye movement. Acute optic neuropathy secondary to methyl alcohol consumption is a serious ocular disease with permanent vision loss or scotoma. Prenatal exposure to ethanol may end in fetal alcohol spectrum disease, where ocular findings are a constant component. The association between chronic alcohol consumption and increased risks of cataract, age-related macular degeneration, diabetic retinopathy, different types of optic neuropathy, impairment of visual quality, retinal vascular disease, and ocular surface disease has also been reported. Along with detrimental medical and social effects, the role of alcohol consumption in different ocular conditions should be considered, as alcohol-induced visual disturbances may contribute to the heavy burden of alcohol abuse on the healthcare system and overall quality of life.", "Maternal ethanol exposure during pregnancy may cause fetal alcohol spectrum disorders (FASD). FASD is the leading cause of mental retardation. The most deleterious effect of fetal alcohol exposure is inducing neuroapoptosis in the developing brain. Ethanol-induced loss of neurons in the central nervous system underlies many of the behavioral deficits observed in FASD. The cerebellum is one of the brain areas that are most susceptible to ethanol during development. Ethanol exposure causes a loss of both cerebellar Purkinje cells and granule cells. This review focuses on the toxic effect of ethanol on cerebellar granule cells (CGC) and the underlying mechanisms. Both in vitro and in vivo studies indicate that ethanol induces apoptotic death of CGC. The vulnerability of CGC to ethanol-induced death diminishes over time as neurons mature. Several mechanisms for ethanol-induced apoptosis of CGC have been suggested. These include inhibition of N-methyl-D-aspartate receptors, interference with signaling by neurotrophic factors, induction of oxidative stress, modulation of retinoid acid signaling, disturbance of potassium channel currents, thiamine deficiency, and disruption of translational regulation. Cultures of CGC provide an excellent system to investigate cellular/molecular mechanisms of ethanol-induced neurodegeneration and to evaluate interventional strategies. This review will also discuss the approaches leading to neuroprotection against ethanol-induced neuroapoptosis." ]
Motivational interviewing among Black women at high risk for adverse birth outcomes	Motivational interviewing (MI) is an evidence-based strategy to modify health behaviors, including some risk factors for adverse birth outcomes. Black women, who have disproportionately high rates of adverse birth outcomes, have reported mixed preferences on MI. This study explored the acceptability of MI among Black women who are at high risk for adverse birth outcomes.	[ "Health disparities necessitate exploration of how race moderates response to smoking cessation treatment. Data from a randomized clinical trial of Motivational Interviewing (MI) for smoking cessation induction were used to explore differential treatment response between African American (AA) vs Non-Black (NB) smokers. Adult tobacco smokers (138 AA vs 66 NB) with low desire to quit were randomly assigned to four sessions of MI or health education (HE). Outcomes (e.g., quit attempts) were assessed 3- and 6-months. There was evidence of a Race by Treatment interaction such that MI was less effective than HE in AA smokers. Mean Cohen's d for the interaction effect was -0.32 (95% CI [-0.44, -0.20]). However, the race interaction could be accounted for by controlling for baseline relationship status and communication preference (wants directive approach). MI may be less effective for smoking cessation induction in AA vs NB smokers when compared to another active and more directive therapy. The differential response between races may be explained by psychosocial variables. MI may not be an ideal choice for all African American smokers. Patients' relationship status and preference for a directive counseling approach might explain disparities in response to MI treatment.", "Maternal morbidity and mortality remains a significant health care concern in the United States, as the rates continue to rise despite efforts to improve maternal health. In 2013, the United States ranked 60th in maternal mortality worldwide. We review the definitions, rates, trends, and top causes of severe maternal morbidity and mortality, as well as risk factors for adverse maternal outcomes. We describe current local and national initiatives in place to reduce maternal morbidity and mortality and offer suggestions for future research.", "Motivational interviewing (MI), a patient-centered behavioral counseling style, is a common behavioral intervention strategy. Because intervention outcomes are highly dependent on patient responsiveness to intervention strategy, we evaluated MI perceptions among rural African American women with type 2 diabetes before a physical activity intervention. Four moderator-led focus groups were conducted with patients aged 21-50 years who had never participated in a MI intervention and who receive diabetes care in a rural community health center. Patients were asked to share their perceptions of an MI consultation after viewing a DVD-based example. They were also asked to discuss their physical activity perceptions and readiness. A comprehensive content analysis based on grounded theory was performed by two raters in order to identify main themes. Although patients (n = 31) had an appreciation for physical activity benefits and high levels of physical activity readiness, themes related to physical activity barriers and lack of motivation were pervasive. Patients regarded the MI consultation as an effective health communication but the patient-centeredness of the approach was negatively perceived. Compared with MI, patients agreed that more traditional paternalistic approaches (i.e., physician-led interactions) were more representative of \"good counseling\" and more familiar to them. Patients shared deeply about personal experiences and provided words of encouragement to one another. Physical activity interventions including rural African-American women should include activities that focus on barrier management and increasing motivation. MI might be an appropriate behavioral counseling model when added to a more traditional cognitive-behavioral physical activity intervention that is group-based and tailored to patients' communication preferences and the clinical setting.", "This second paper in the Born Too Soon supplement presents a review of the epidemiology of preterm birth, and its burden globally, including priorities for action to improve the data. Worldwide an estimated 11.1% of all livebirths in 2010 were born preterm (14.9 million babies born before 37 weeks of gestation), with preterm birth rates increasing in most countries with reliable trend data. Direct complications of preterm birth account for one million deaths each year, and preterm birth is a risk factor in over 50% of all neonatal deaths. In addition, preterm birth can result in a range of long-term complications in survivors, with the frequency and severity of adverse outcomes rising with decreasing gestational age and decreasing quality of care. The economic costs of preterm birth are large in terms of immediate neonatal intensive care, ongoing long-term complex health needs, as well as lost economic productivity. Preterm birth is a syndrome with a variety of causes and underlying factors usually divided into spontaneous and provider-initiated preterm births. Consistent recording of all pregnancy outcomes, including stillbirths, and standard application of preterm definitions is important in all settings to advance both the understanding and the monitoring of trends. Context specific innovative solutions to prevent preterm birth and hence reduce preterm birth rates all around the world are urgently needed. Strengthened data systems are required to adequately track trends in preterm birth rates and program effectiveness. These efforts must be coupled with action now to implement improved antenatal, obstetric and newborn care to increase survival and reduce disability amongst those born too soon.", "To critically review, analyze, and synthesize the literature on parenting stress among caregivers of children with asthma, cancer, cystic fibrosis, diabetes, epilepsy, juvenile rheumatoid arthritis, and/or sickle cell disease. Method PsychInfo, MEDLINE, and Cumulative Index to Nursing and Allied Health Literature were searched according to inclusion criteria. Meta-analysis of 13 studies and qualitative analysis of 96 studies was conducted. Results Caregivers of children with chronic illness reported significantly greater general parenting stress than caregivers of healthy children (d = .40; p = ≤.0001). Qualitative analysis revealed that greater general parenting stress was associated with greater parental responsibility for treatment management and was unrelated to illness duration and severity across illness populations. Greater parenting stress was associated with poorer psychological adjustment in caregivers and children with chronic illness. Conclusion Parenting stress is an important target for future intervention. General and illness-specific measures of parenting stress should be used in future studies.", "Shared decision-making (SDM), a collaborative approach that includes and respects patients' preferences for involvement in decision-making about their treatment, is increasingly advocated. However, in the practice of clinical psychiatry, implementing SDM seems difficult to accomplish. Although the number of studies related to psychiatric patients' preferences for involvement is increasing, studies have largely focused on understanding patients in public mental healthcare settings. Thus, investigating patient preferences for involvement in both public and private settings is of particular importance in psychiatric research. The objectives of this study were to identify different latent class typologies of patient preferences for involvement in the decision-making process, and to investigate how patient characteristics predict these typologies in mental healthcare settings. We conducted latent class analysis (LCA) to identify groups of psychiatric outpatients with similar preferences for involvement in decision-making to estimate the probability that each patient belonged to a certain class based on sociodemographic, clinical and health belief variables. The LCA included 224 consecutive psychiatric outpatients' preferences for involvement in treatment decisions in public and private psychiatric settings. The LCA identified three distinct preference typologies, two collaborative and one passive, accounting for 78% of the variance. Class 1 (26%) included collaborative men aged 34-44 years with an average level of education who were treated by public services for a depressive disorder, had high psychological reactance, believed they controlled their disease and had a pharmacophobic attitude. Class 2 (29%) included collaborative women younger than 33 years with an average level of education, who were treated by public services for an anxiety disorder, had low psychological reactance or health control belief and had an unconcerned attitude toward medication. Class 3 (45%) included passive women older than 55 years with lower education levels who had a depressive disorder, had low psychological reactance, attributed the control of their disease to their psychiatrists and had a pharmacophilic attitude. Our findings highlight how psychiatric patients vary in pattern of preferences for treatment involvement regarding demographic variables and health status, providing insight into understanding the pattern of preferences and comprising a significant advance in mental healthcare research.", "Little is known about the correlates of physical activity among African-American women living in the southeastern United States. The purpose of this study was to assess the relationship of personal, social, cultural, environmental, and policy variables with physical activity among women in ethnic minority groups. The Women and Physical Activity Survey was used in a telephone interview of 917 African-American women living in two counties in South Carolina. The sample of women was selected by random-digit dialing. Approximately one third (34.1%) of the women met current recommendations for moderate or vigorous physical activity, 49.4% were insufficiently active, and 16.5% were inactive. Meeting the recommendations or engaging in insufficient activity (versus inactive) was related to attaining higher educational levels, being married or with a partner; being in excellent or very good health, having greater self-efficacy, seeing people exercise in the neighborhood, having more favorable ratings of women who exercise (social issues score), having lower social role strain, and reporting the presence of sidewalks or lighter traffic in the neighborhood. Multiple factors influence physical activity. Interventions to increase physical activity should use multilevel approaches that incorporate the personal, social environmental, and physical environmental factors related to participation in physical activity." ]
Femtosecond laser direct writing mask method for acoustofluidic device preparation	Surface acoustic wave (SAW)-based acoustofluidic devices have shown broad applications in microfluidic actuation and particle/cell manipulation. Conventional SAW acoustofluidic device fabrication generally includes photolithography and lift-off processes and thus requires accessing cleanroom facilities and expensive lithography equipment. In this paper, we report a femtosecond laser direct writing mask method for acoustofluidic device preparation. By micromachining of steel foil to form the mask and direct evaporation of metal on the piezoelectric substrate using the mask, the interdigital transducer (IDT) electrodes of the SAW device are generated. The minimum spatial periodicity of the IDT finger is about 200 μm, and the preparation for LiNbO	[ "Precise and selective manipulation of colloids and biological cells has long been motivated by applications in materials science, physics and the life sciences. Here we introduce our harmonic acoustics for a non-contact, dynamic, selective (HANDS) particle manipulation platform, which enables the reversible assembly of colloidal crystals or cells via the modulation of acoustic trapping positions with subwavelength resolution. We compose Fourier-synthesized harmonic waves to create soft acoustic lattices and colloidal crystals without using surface treatment or modifying their material properties. We have achieved active control of the lattice constant to dynamically modulate the interparticle distance in a high-throughput (>100 pairs), precise, selective and reversible manner. Furthermore, we apply this HANDS platform to quantify the intercellular adhesion forces among various cancer cell lines. Our biocompatible HANDS platform provides a highly versatile particle manipulation method that can handle soft matter and measure the interaction forces between living cells with high sensitivity.", "Regenerative medicine and drug development require large numbers of high-quality cells, usually delivered from in vitro culturing. During culturing, the appearance of unwanted cells and an inability to remove them without damaging or losing most if not all the surrounding cells in the culture reduce the overall quality of the cultured cells. This is a key problem in cell culturing, as is the inability to sample cells from a culture as desired to verify the quality of the culture. Here, we report a method to locally remove cells from an adherent cell culture using a 100.4 MHz focused surface acoustic wave (SAW) device. After exposing a plated C2C12 mouse myoblast cell culture to phosphate buffered solution (PBS), ultrasound from the SAW device transmitted into the cell culture via a coupling water droplet serves to detach a small grouping of cells. The cells are removed from an area 6 × 10-3 mm2, equivalent to about 12 cells, using a SAW device-Petri dish water gap of 1.5 mm, a PBS immersion time of 300 s, and an input voltage of 75 V to the SAW device. Cells were released as desired 90% of the time, releasing the cells from the target area nine times out of ten runs. In the one trial in ten that fails, the cells partially release and remain attached due to inter-cellular binding. By making it possible to target and remove small groups of cells as desired, the quality of cell culturing may be significantly improved. The small group of cells may be considered a colony of iPS cells. This targeted cell removal method may facilitate sustainable, contamination-free, and automated refinement of cultured cells.", "We present a piezoelectric transducer for standing wave surface acoustic wave nebulization (SW-SAWN). The transducer nebulizes nonvolatile analytes present in bulk fluid into ambient air after which the aerosolized drops are sampled by mass spectrometry (MS) for detection. Furthermore, we report for the first time integration of anisotropic ratchet conveyors (ARCs) on the SAWN transducer surfaces to automate the sample preparation and droplet delivery process. The ARCs employ micro-sized hydrophilic patterns on hydrophobic Cytop coatings. Moving, positioning, merging, and mixing of droplets at a designated nebulization location are demonstrated. To create the ARCs, we adopt parylene C as a stencil mask so that the hydrophobicity of the Cytop does not degrade during the microfabrication process. MS measurements with the SAWN chip are performed under different input frequencies. The SAWN transducer can provide a controllable nebulization rate by varying the input nebulization frequency while maintaining a reasonable signal to noise ratio for MS detection.", "An anisotropic ratchet conveyor is an asymmetric, periodic, micropatterned surface that propels droplets when vibrated with a sinusoidal signal at certain frequencies and amplitudes. For each input frequency, there is a threshold amplitude beyond which the droplet starts to move. In this paper, we study the parameters that initiate droplet motion and the relationship between the input frequency and threshold amplitude among droplets with different volume, density, viscosity, and surface tension. Through this investigation we demonstrate how nondimensionalization reveals consistent behavior for droplets of different volumes. Finally, we propose a compact model that captures the essential features of the system to describe how a pure vertical vibration results in horizontal droplet motion. This model provides an intuitive understanding of the underlying physics and explains how the surface asymmetry is the key for lateral droplet motion.", "When considering incident investigations and security checks focused on energetic materials, there is an ongoing need for rapid, on-scene chemical identification. Currently applied methods are not capable of meeting all requirements, and hence, portable mass spectrometry is an interesting alternative although many instrumental challenges still exist. To be able to analyze explosives with mass spectrometry outside the traditional laboratory, suitable ambient ionization methods need to be developed. Ideally such methods are also easily implemented in the field requiring limited to no power sources, gas supplies, flow controllers, and heating devices. For this reason, the potential of SAWN (surface acoustic wave nebulization) for the ambient ionization and subsequent mass spectrometric (MS) analysis of organic explosives was investigated in this study. Excellent sensitivity was observed for nitrate-based organic explosives when operating the MS in negative mode. No dominant adduct peaks were observed for the peroxides TATP and HMTD with SAWN-MS in positive mode. The MS spectra indicate extensive fragmentation of the peroxide explosives even under the mild ionization conditions provided by SAWN. The potential of SAWN-MS was demonstrated with the correct identification of nitrate-based organic explosives in pre- and post-explosion case samples in only a fraction of the time and effort required for the regular laboratory analysis. Results show that SAWN-MS can convincingly identify intact organic energetic compounds and mixtures but that sensitivity is not always sufficient to detect traces of explosives in post-explosion residues." ]
Pathogen-Induced Changes in Plant Development	Plants, as sessile organisms, are constantly exposed to pathogens in nature. Plants rely on physical barriers, constitutive chemical defenses, and sophisticated inducible immunity to fight against pathogens. The output of these defense strategies is highly associated with host development and morphology. Successful pathogens utilize various virulence strategies to colonize, retrieve nutrients, and cause disease. In addition to the overall defense-growth balance, the host-pathogen interactions often lead to changes in the development of specific tissues/organs. In this review, we focus on recent advances in understanding the molecular mechanisms of pathogen-induced changes in plants' development. We discuss that changes in host development could be a target of pathogen virulence strategies or an active defense strategy of plants. Current and ongoing research about how pathogens shape plant development to increase their virulence and causes diseases could give us novel views on plant disease control.	[ "Plant-pathogen interactions feature complex signaling exchanges between host and microbes that ultimately determine association outcomes. Plants deploy pattern recognition receptors to perceive pathogen-associated molecular patterns, mount pattern-triggered immunity (PTI), and fend off potential pathogens. In recent years an increasing number of defense-signaling components have been identified along with a mechanistic understanding of their regulation during immune responses. Post-translational modifications (PTMs) are now thought to play a crucial role in regulating defense signaling. In a bid to suppress PTI and infect their host, pathogens have evolved large repertoires of effectors that trigger susceptibility and allow colonization of host tissues. While great progress has been made in elucidating defense-signaling networks in plants and the activities of effectors in immune suppression, a critical gap exists in our understanding of effector mechanism-of-action. Given the importance of PTMs in the regulation of defense signaling, we will explore the question: how do effectors modify the post-translational status of host proteins and thus interfere with host processes required for immunity? We will consider how emerging proteomics-based experimental strategies may help us answer this important question and ultimately open the pathogens' effector black box.", "Multicellular eukaryotes coevolve with microbial pathogens, which exert strong selective pressure on the immune systems of their hosts. Plants and animals use intracellular proteins of the nucleotide-binding domain, leucine-rich repeat (NLR) superfamily to detect many types of microbial pathogens. The NLR domain architecture likely evolved independently and convergently in each kingdom, and the molecular mechanisms of pathogen detection by plant and animal NLRs have long been considered to be distinct. However, microbial recognition mechanisms overlap, and it is now possible to discern important key trans-kingdom principles of NLR-dependent immune function. Here, we attempt to articulate these principles. We propose that the NLR architecture has evolved for pathogen-sensing in diverse organisms because of its utility as a tightly folded \"hair trigger\" device into which a virtually limitless number of microbial detection platforms can be integrated. Recent findings suggest means to rationally design novel recognition capabilities to counter disease.", "With the centenary of the first descriptions of 'hypersensitiveness' following pathogenic challenge upon us, it is appropriate to assess our current understanding of the hypersensitive response (HR) form of cell death. In recent decades our understanding of the initiation, associated signalling, and some important proteolytic events linked to the HR has dramatically increased. Genetic approaches are increasingly elucidating the function of the HR initiating resistance genes and there have been extensive analyses of death-associated signals, calcium, reactive oxygen species (ROS), nitric oxide, salicylic acid, and now sphingolipids. At the same time, attempts to draw parallels between mammalian apoptosis and the HR have been largely unsuccessful and it may be better to consider the HR to be a distinctive form of plant cell death. We will consider if the HR form of cell death may occur through metabolic dysfunction in which malfunctioning organelles may play a major role. This review will highlight that although our knowledge of parts of the HR is excellent, a comprehensive molecular model is still to be attained.", "Plants rely on PAMP-triggered immunity (PTI) and effector-triggered immunity (ETI) to detect invading pathogens and subsequently activate defense mechanisms. Recently, four Nature papers (Yuan et al., Ngou et al., Pruitt et al., and Tian et al.)demonstrated that important components in PTI and ETI are required for both PTI and ETI, and PTI and ETI potentiate each other to achieve stronger plant defenses.", "At the colonization site of a foreign entity, plant cells alter their trajectory of growth and development. The resulting structure - a plant gall - accommodates various needs of the foreigner, which are phylogenetically diverse: viruses, bacteria, protozoa, oomycetes, true fungi, parasitic plants, and many types of animals, including rotifers, nematodes, insects, and mites. The plant species that make galls also are diverse. We assume gall production costs the plant. All is well if the foreigner provides a gift that makes up for the cost. Nitrogen-fixing nodule-inducing bacteria provide nutritional services. Gall wasps pollinate fig trees. Unfortunately for plants, most galls are made for foes, some of which are deeply studied pathogens and pests: Agrobacterium tumefaciens, Rhodococcus fascians, Xanthomonas citri, Pseudomonas savastanoi, Pantoea agglomerans, 'Candidatus' phytoplasma, rust fungi, Ustilago smuts, root knot and cyst nematodes, and gall midges. Galls are an understudied phenomenon in plant developmental biology. We propose gall inception for discovering unifying features of the galls that plants make for friends and foes, talk about molecules that plants and gall-inducers use to get what they want from each other, raise the question of whether plants colonized by arbuscular mycorrhizal fungi respond in a gall-like manner, and present a research agenda.", "Age-related resistance (ARR) is a plant defense response characterized by enhanced resistance to certain pathogens in mature plants relative to young plants. In Arabidopsis thaliana the transition to flowering is associated with ARR competence, suggesting that this developmental event is the switch that initiates ARR competence in mature plants (Rusterucci et al. in Physiol Mol Plant Pathol 66:222-231, 2005). The association of ARR and the floral transition was examined using flowering-time mutants and photoperiod-induced flowering to separate flowering from other developmental events that occur as plants age. Under short-day conditions, late-flowering plant lines ld-1 (luminidependens-1), soc1-2 (suppressor of overexpression of co 1-2), and FRI (+) (FRIGIDA) displayed ARR before the transition to flowering occurred. Early-flowering svp-31, svp-32 (short vegetative phase), and Ws-2 were ARR-defective, whereas early-flowering tfl1-14 (terminal flower 1-14) displayed ARR at the same time as Col-0. While svp-31, svp-32 and Ws-2 produced few rosette leaves, tfl1-14 produced a rosette leaf number similar to Col-0, suggesting that the development of a minimum number of rosette leaves is necessary to initiate ARR competence under short-day conditions. Photoperiod-induced transient expression of FT (FLOWERING LOCUS T) caused precocious flowering in short-day-grown Col-0 but this was not associated with ARR competence. Under long-day conditions co-9 (constans-9) mutants did not flower but displayed an ARR response at the same time as Col-0. This study suggests that SVP is required for the ARR response and that the floral transition is not the developmental event that regulates ARR competence.", "Cuticle thickness of leaves varies > 100 times across species, yet its dry mass cost and ecological benefits are poorly understood. It has been repeatedly demonstrated that thicker cuticle is not superior as a water barrier, implying that other functions must be important. Here, we measured the mechanical properties, dry mass and density of isolated cuticle from 13 evergreen woody species of Australian forests. Summed adaxial and abaxial cuticle membrane mass per unit leaf area (CMA) varied from 2.95 to 27.4 g m(-2) across species, and accounted for 6.7-24% of lamina dry mass. Density of cuticle varied only from 1.04 to 1.24 g cm(-3) ; thus variation in CMA was mostly due to variation in cuticle thickness. Thicker cuticle was more resistant to tearing. Tensile strength and modulus of elasticity of cuticle were much higher than those of leaf laminas, with significant differences between adaxial and abaxial cuticles. While cuticle membranes were thin, they could account for a significant fraction of leaf dry mass due to their high density. The substantial cost of thicker cuticle is probably offset by increased mechanical resistance which might confer longer leaf lifespans among evergreen species." ]
Calcifying epithelial odontogenic tumor involving the right mandibular molar-premolar	Calcifying epithelial odontogenic tumor (CEOT), also known as Pindborg tumor, is a rare benign but locally aggressive odontogenic neoplasm, accounts for <1% of all odontogenic tumors. CEOT is usually seen in the posterior area of the mandible in-between 30 and 50 years of age without definite sex predilection. A painless, slow-growing swelling with bone expansion is the most common clinical feature of CEOT. Radiographically, it presents as a mixed radiographic lesion may or may not be associated with any impacted tooth. Confirmation of the diagnosis is made by histopathological examination. The tumor has a recurrence rate of 10%-20% and so periodic follow-up is necessary. A unique case of CEOT involving the right mandibular molar-premolar in a 25-year-old female patient with clinical behavior, radiological, histopathological features and surgical managements is discussed herewith.	[ "The calcifying epithelial odontogenic tumor (CEOT) is a rare entity and represents less than 1% of all odontogenic tumors. Dr. J J Pindborg (1958) first described four cases of this unusual lesion; subsequently Shafer et al coined the term Pindborg tumor. This lesion is a locally aggressive benign odontogenic neoplasm arising from epithelial tissue. It occurs most commonly in 4(th)-5(th)-6(th) decade of life and bears no gender predilection. A case of CEOT in a 50-year-old male arising in the left body region is described.", "Calcifying epithelial odontogenic tumor (CEOT) is a rare, benign, odontogenic tumor arising from the odontogenic epithelium and accounts for approximately 1% of all odontogenic tumors. Clear cell variant of CEOT is a distinct entity and has more aggressive biological behavior and higher chances of recurrence. Here, we present a unique case of clear cell variant of CEOT involving the left side of the maxillary alveolus in a 73-year-old female patient with thorough clinical, radiological, and histological details.", "To analyse systematically the radiographic features of the calcifying epithelial odontogenic tumour (CEOT). Clinical and radiological features of 67 cases of CEOT (four new and 63 from the literature) were analysed. There were 27 (41%) males and 39 (59%) females with a male-to-female ratio of 1 : 1.5. Age ranged from 13 - 77 years (mean 43.5 years), with a peak in the fourth and fifth decades. The mandible was involved in 74% of the cases and 69% of all lesions were in the posterior area. The mixed radiolucent-radiopaque pattern was the most frequent (65%), with 32% radiolucent and 3% radiopaque. Coronal clustering of the radiopaque material was found in 12% and in one case, a 'driven snow' pattern of the radiopaque material was clearly recognizable. Lesions were unilocular in 58%, multilocular in 27% and not loculated in 15%. The unilocular type was more frequent in the maxilla than in the mandible. Borders were well defined and corticated in 20%, defined but not corticated in 59% and diffuse in 21%. Sixty per cent were associated with impacted teeth. Lesions larger than 3 cm tended to be more frequent in the mandible, mixed radiolucent-radiopaque and to have a higher proportion of diffuse borders than the smaller lesions. Radiographic features which have been considered characteristic of CEOT, coronal clustering and 'driven snow' patterns, are seen in only a small percentage of cases.", "A review of the world literature has revealed 113 cases of the calcifying epithelial odontogenic tumor. The clinical, radiographic, and histopathologic features are analyzed. Various treatments that have been carried out are related to the frequency of recurrence. The theories of histogenesis are discussed and an attempt is made to explain the nature and origin of the amyloid-like substance." ]
Immunothrombosis as a biomarker for early sepsis diagnosis	Sepsis, a medical emergency, is the overwhelming host response to infection leading to organ failure. The pathophysiology of this heterogeneous disease includes an inflammatory response that stimulates a complex interaction between endothelial and complements with associated coagulation abnormalities. Despite a more comprehensive understanding of sepsis pathophysiology, there exists a translational gap to improve sepsis diagnosis clinically. Many of the proposed biomarkers to diagnose sepsis lack sufficient specificity and sensitivity to be used in routine clinical practice. There has also been a lack of progress in diagnostic tools due to the focus on the inflammatory pathway. Inflammation and coagulation are known to be linked to the innate immune response. Early immunothrombotic changes could result in the early switch from infection to sepsis and aid in sepsis diagnosis. This review integrates both preclinical and clinical studies that highlight sepsis pathophysiology providing a framework for how the development of immunothrombosis could be used as a starting point to investigate biomarkers for early sepsis diagnosis.	[ "Complement-activated human plasma causes generation of tissue factor in human leukocytes. This phenomenon appears to be related to the fifth component of complement (C5) as demonstrated by the use of C5 deficient-plasma and suppression of activity with antibody to C5. Isolation of the chemotactic factor from activated serum or trypsinization of purified C5 reproduces the phenomenon. These data provide evidence for a direct link between complement products and activation of the coagulation system. Because chemotactic peptides from C5 can be generated by a variety of enzymes, our findings suggest a relationship between complement, coagulation, and inflammation.", "Impairment of the protein C anticoagulation pathway is critical to the thrombosis associated with sepsis and to the development of purpura fulminans in meningococcemia. We studied the expression of thrombomodulin and the endothelial protein C receptor in the dermal microvasculature of children with severe meningococcemia and purpuric or petechial lesions. We assessed the integrity of the endothelium and the expression of thrombomodulin and the endothelial protein C receptor in biopsy specimens of purpuric lesions from 21 children with meningococcal sepsis (median age, 41 months), as compared with control skin-biopsy specimens. The expression of endothelial thrombomodulin and of the endothelial protein C receptor was lower in the patients with meningococcal sepsis than in the controls, both in vessels with thrombosis and in vessels without thrombosis. On electron microscopical examination, the endothelial cells were generally intact in both thrombosed and nonthrombosed vessels. Plasma thrombomodulin levels in the children with meningococcal sepsis (median, 6.4 ng per liter) were higher than those in the controls (median, 3.6 ng per liter; P=0.002). Plasma levels, protein C antigen, protein S antigen, and antithrombin antigen were lower than those in the controls. In two patients treated with unactivated protein C concentrate, activated protein C was undetectable at the time of admission, and plasma levels remained low. In severe meningococcal sepsis, protein C activation is impaired, a finding consistent with down-regulation of the endothelial thrombomodulin-endothelial protein C receptor pathway.", "The endothelium provides an essential and selective membrane barrier that regulates the movement of water, solutes, gases, macromolecules and the cellular elements of the blood from the tissue compartment in health and disease. Its structure and continuous function is essential for life for all vertebrate organisms. Recent evidence indicates that the endothelial surface does not have a passive role in systemic inflammatory states such as septic shock. In fact, endothelial cells are in dynamic equilibrium with a myriad of inflammatory mediators and elements of the innate immune and coagulation systems to orchestrate the host response in sepsis. The barrier function of the endothelial surface is almost uniformly impaired in septic shock, and it is likely that this contributes to adverse outcomes. In this review, we will highlight recent advances in the understanding of the signalling events that regulate endothelial function and molecular events that induce endothelial dysfunction in sepsis. Endothelial barrier repair strategies as a treatment for sepsis include modulation of C5a, high-mobility group box 1 and VEGF receptor 2; stimulation of angiopoietin-1, sphingosine 1 phosphate receptor 1 and Slit; and a number of other innovative approaches.", "Evidence for the involvement of the sixth component of complement (C6) in normal blood coagulation is provided by the description of a coagulation abnormality in rabbits with a genetic C6 deficiency and by its correction with highly purified preparations of C6. Whole blood clotting time in glass or plastic was prolonged and prothrombin consumption was decreased in blood from the deficient animals. Other parameters of blood coagulation were normal, including prothrombin time, partial thromboplastin time, specific clotting factor activities, platelet factor III function, platelet count, and bleeding time. Clotting time and prothrombin consumption became normal when physiologic amounts of highly purified C6 were added to the deficient blood. Partial consumption of C6 hemolytic activity, with a time course similar to the consumption of prothrombin, was demonstrated during the clotting of normal human blood.", "Background Biomarkers can facilitate the diagnosis of sepsis, enabling early management and improving outcomes. Lipopolysaccharide-binding protein (LBP) has been reported as a biomarker for the detection of infection, but its diagnostic value is controversial. In this study, we assessed the diagnostic accuracy of LBP for sepsis in the emergency department (ED) patients, comparing it with more established biomarkers of sepsis, including procalcitonin (PCT) and C-reactive protein (CRP). Methods LBP and other sepsis biomarkers, including PCT and CRP, were measured on admission in 102 adult patients presenting with suspected infection . Classification of patients was performed using the recently updated definition for sepsis (Sepsis-3). The diagnostic accuracy of LBP, CRP and PCT for sepsis was evaluated by using receiver operating characteristic curve (ROC) analysis. Results A total of 49 patients were classified as having sepsis. In these patients, median (interquartile range) LBP (41.8 [41.1] µg/dL vs. 26.2 [25] µg/dL), CRP (240 [205] mg/L vs. 160 [148] mg/dL) and PCT (5.19 [13.68] µg/L vs. 0.39 [1.09] µg/L) were significantly higher than in patients classified as not having sepsis ( P < 0.001 for all three biomarkers). ROC curve analysis and area under curve (AUC) revealed a value of 0.701 for LBP, similar to CRP (0.707) and lower than that for PCT (0.844) ( P = 0.012). Conclusion In adult ED patients with suspected infection, the diagnostic accuracy for sepsis of LBP is similar to that of CRP but lower than that of PCT.", "Previous studies indicated that factor VIIa, in complex with tissue factor, readily activates either factor X or factor IX in the presence of calcium ions. In order to assess the relative physiological importance of the activation of factor IX versus the activation of factor X by recombinant factor VIIa, we have obtained steady-state kinetic parameters for the factor VIIa catalyzed activation of factor IX and factor X under a variety of cofactor conditions that include calcium alone, calcium and phospholipids, calcium, phospholipids, and tissue factor apoprotein, and calcium and cell-surface tissue factor. Calcium alone stimulated the activation of factors IX and X by factor VIIa maximally at 1 and 2.5 mM, respectively. In the presence of 25 microM phospholipids, maximal rates of factor IX and factor X activation were achieved at 2.5-5 mM calcium. With calcium alone, or with phospholipid and calcium, the initial rates of factor IX activation by factor VIIa were significantly higher than that observed for factor X. Kinetic studies revealed that the Km for the factor VIIa catalyzed activation of factor IX was essentially constant in the presence of 5 mM calcium and 1-500 microM phospholipid, whereas the Km for factor X activation varied with phospholipid concentration, reaching a minimum at 7-20 microM phospholipid. At all concentrations of added phospholipid, the kcat/Km ratio for the activation of factor IX by factor VIIa appeared to be considerably greater than that observed for the activation of factor X.(ABSTRACT TRUNCATED AT 250 WORDS)", "Among immune cells in responding to sepsis, macrophages and neutrophils have been extensively studied, while the contribution of T lymphocytes and natural killer T (NKT) cells is less well characterized. Here we monitored tissue specific changes of T cell subsets in male C57BL/6 mice subjected to sham operation or cecal ligation and puncture (CLP) to induce polymicrobial sepsis. Thymus, spleen, liver, lungs and blood were processed and analyzed 20h later. Total lymphocyte count showed a significant reduction in septic thymus, spleen and blood but not in lungs and liver. The septic thymi were hypocellular with severe reduction in cell numbers of immature CD4(+)CD8(+) subset. CD4(+) T and CD8(+) T lymphocyte numbers in septic spleens were also significantly reduced, but the frequency of CD4(+)CD25(+) Tregs was significantly increased. In addition, naïve and Tcm CD4(+) T cell numbers were significantly reduced in the septic spleens. By contrast, in septic liver the CD8(+) T cell numbers were significantly increased, whereas NKT cell numbers were reduced, but more activated with increased CD69 and CD25 expression. In the septic lungs, the CD4(+) T and CD8(+) T cell numbers showed no significant change, whereas they were severely reduced in the septic blood. Overall, this study provides important information on the alterations of different T-cell subsets in various tissues after sepsis." ]
Associations between a laboratory-based frailty index and adverse health outcomes	Early frailty may be captured by a frailty index (FI) based entirely on vital signs and laboratory tests. Our aim was to examine associations between a laboratory-based FI (FI-Lab) and adverse health outcomes, and investigate how this changed with age.	[ "Frailty is a significant healthcare challenge in China. However, the relationship between frailty and the prognosis of older people in China remains unclear. The present study aimed to evaluate the prevalence of frailty and determine if the frailty index, a comprehensive geriatric assessment, was associated with the prognosis of older people in a Chinese population. Data were drawn from the Beijing Longitudinal Study of Aging, a representative cohort study with an 8-year follow-up. Evaluations based on the use of the frailty index were performed in a cohort of 1808 people aged 60 years and over residing in Beijing urban and rural areas. The initial survey was conducted in 2004, with follow-up surveys at 3, 5, and 8 years. Mortality data for all individuals were collected and analyzed. The frailty index and the age of individuals showed the same trend, with a higher frailty index expected as age increased. Respondents with the same frailty index level differed across factors such as sex and location. Male individuals, rural dwellers, and older individuals showed higher frailty rates than female individuals, urban dwellers, and younger individuals. Frailty is a condition associated with problems across multiple physiological systems. The frailty index increases with age, and may be a significant tool for evaluation of the prognosis of older people in China.", "Previous meta-analyses of cohort studies indicate a J-shaped relationship between alcohol consumption and allcause mortality, with reduced risk for low-volume drinkers. However, low-volume drinkers may appear healthy only because the \"abstainers\" with whom they are compared are biased toward ill health. The purpose of this study was to determine whether misclassifying former and occasional drinkers as abstainers and other potentially confounding study characteristics underlie observed positive health outcomes for lowvolume drinkers in prospective studies of all-cause mortality. A systematic review and meta-regression analysis of studies investigating alcohol use and mortality risk after controlling for quality-related study characteristics was conducted in a population of 3,998,626 individuals, among whom 367,103 deaths were recorded. Without adjustment, meta-analysis of all 87 included studies replicated the classic J-shaped curve, with low-volume drinkers (1.3-24.9 g ethanol per day) having reduced mortality risk (RR = 0.86, 95% CI [0.83, 0.90]). Occasional drinkers (<1.3 g per day) had similar mortality risk (RR = 0.84, 95% CI [0.79, 0.89]), and former drinkers had elevated risk (RR = 1.22, 95% CI [1.14, 1.31]). After adjustment for abstainer biases and quality-related study characteristics, no significant reduction in mortality risk was observed for low-volume drinkers (RR = 0.97, 95% CI [0.88, 1.07]). Analyses of higher-quality bias-free studies also failed to find reduced mortality risk for low-volume alcohol drinkers. Risk estimates for occasional drinkers were similar to those for low- and medium-volume drinkers. Estimates of mortality risk from alcohol are significantly altered by study design and characteristics. Meta-analyses adjusting for these factors find that low-volume alcohol consumption has no net mortality benefit compared with lifetime abstention or occasional drinking. These findings have implications for public policy, the formulation of low-risk drinking guidelines, and future research on alcohol and health.", "To determine whether commonly used frailty scales exhibit shared characteristics when applied to a representative sample of middle-aged and older Europeans. Secondary analysis of the Survey of Health, Ageing, and Retirement in Europe (SHARE). Eleven European countries. Community-dwelling adults (N = 27,527; mean age 65.3 ± 10.5, 55% female). Frailty was assessed using SHARE-operationalized versions of seven frailty scales: Edmonton Frail Scale, FRAIL scale, Groningen Frailty Indicator, frailty phenotype, Tilburg Frailty Indicator, a 70-item frailty index (FI), and a 44-item frailty index based on Comprehensive Geriatric Assessment. All frailty scales demonstrated right-skewed density distributions. On all scales, frailty scores increased nonlinearly with age, between 1% (FRAIL) and 3.6% (FI) per year on a log scale. Frailty scores on all scales exhibited dose-response relationships with 5-year mortality. On all scales, women had higher frailty scores than men of the same age but demonstrated better survival than did men with the same frailty score. On all scales except the frailty phenotype, 99% of participants had scores below the scale's theoretical maximum. On each frailty scale, frailty score increased nonlinearly with age, mortality risk increased with frailty score, and women had higher scores than men but demonstrated better survival. Each scale except the frailty phenotype demonstrated an upper limit to frailty below the scale's theoretical maximum. Across commonly used frailty scales, these characteristics are common in nature but differ in magnitude.", "Sedentary behaviours are associated with adverse health outcomes in middle-aged and older adults, even among those who exercise. We examined whether the degree of frailty affects the association between sedentary behaviours and higher risk of mortality. In this prospective cohort study, we used data from 3141 community-dwelling adults 50 years of age or older from the 2003/04 and 2005/06 cohorts of the US National Health and Nutrition Examination Survey. Time engaged in sedentary behaviours was measured using uniaxial accelerometers, and frailty was based on a 46-item frailty index. Mortality data were linked up to 2011. We used Cox proportional hazard models to estimate the hazard ratio (HR) of sedentary behaviour. We found that for people with low levels of frailty (frailty index score ≤ 0.1), sedentary time was not predictive of mortality, regardless of physical activity level (adjusted HR 0.90, 95% confidence interval [CI] 0.70-1.15). Among people who were vulnerable (0.1 < frailty index score ≤ 0.2) or frail (frailty index score > 0.2), sedentary time was associated with higher mortality only among those who were physically inactive (not meeting the criterion for moderate physical activity) (HR 1.16, 95% CI 1.02-1.33 for the group defined by 0.1 < frailty index score ≤ 0.2; HR 1.27, 95% CI 1.11-1.46 for the group defined by 0.2 < frailty index score ≤ 0.3; HR 1.34, 95% CI 1.19-1.50 for frailty index score > 0.3). The effect of sedentary behaviours on mortality varied by level of frailty. Adults with the highest frailty level experienced the greatest adverse impact. Low frailty levels (frailty index score ≤ 0.1) seemed to eliminate the increased risk of mortality associated with prolonged sitting, even among people who did not meet recommended physical activity guidelines.", "Chronic pain has a strong association with major depressive disorder (MDD), but there is a relative paucity of studies on the association between chronic multisite pain and bipolar disorder (BD). Such studies are required to help elucidate the complex biological and psychological overlap between pain and mood disorders. The aim of this study is to investigate the relationship between chronic multisite pain and mood disorder across the unipolar-bipolar spectrum. We conducted a cross-sectional study of 149,611 UK Biobank participants. Self-reported depressive and bipolar features were used to categorise participants into MDD and BD groups and a non-mood disordered comparison group. Multinomial logistic regression was used to establish whether there was an association between extent of chronic pain (independent variable) and mood disorder category (dependent variable), using no pain as the referent category, and adjusting for a wide range of potential sociodemographic, lifestyle and comorbidity confounders. Multisite pain was significantly more prevalent in participants with BD and MDD, for example, 4-7 pain sites: BD 5.8%, MDD 4.5%, and comparison group 1.8% (p < 0.001). A relationship was observed between extent of chronic pain and risk of BD and persisted after adjusting for confounders (relative to individuals with no chronic pain): 2-3 sites RRR of BD 1.84 (95% CI 1.61, 2.11); 4-7 sites RRR of BD 2.39 (95% CI 1.88, 3.03) and widespread pain RRR of BD 2.37 (95% CI 1.73, 3.23). A similar relationship was observed between chronic pain and MDD: 2-3 sites RRR of MDD 1.59 (95% CI 1.54, 1.65); 4-7 sites RRR of MDD 2.13 (95% CI 1.98, 2.30); widespread pain RRR of MDD 1.86 (95% CI 1.66, 2.08). Individuals who report chronic pain and multiple sites of pain are more likely to have MDD and are at higher risk of BD. These findings highlight an important aspect of comorbidity in MDD and BD and may have implications for understanding the shared neurobiology of chronic pain and mood disorders.", "The aging population in Latin America is characterized by not optimal conditions for good health, experiencing high burden of comorbidity, which contribute to increase the frequency of frailty; thus, identification should be a priority, to classify patients at high risk to develop its negative consequences. The objective of this analysis was to validate the FRAIL instrument to measure frailty in Mexican elderly population, from the database of the Mexican Health and Aging Study (MHAS). Prospective, population study in Mexico, that included subjects of 60 years and older who were evaluated for the variables of frailty during the year 2001 (first wave of the study). Frailty was measured with the five-item FRAIL scale (fatigue, resistance, ambulation, illnesses, and weight loss). The robust, pre-frail or intermediate, and the frail group were considered when they had zero, one, and at least two components, respectively. Mortality, hospitalizations, falls, and functional dependency were evaluated during 2003 (second wave of the study). Relative risk was calculated for each complications, as well as hazard ratio (for mortality) through Cox regression model and odds ratio with logistic regression (for the rest of the outcomes), adjusted for covariates. The state of frailty was independently associated with mortality, hospitalizations, functional dependency, and falls. The pre-frailty state was only independently associated with hospitalizations, functional dependency, and falls. Frailty measured through the FRAIL scale, is associated with an increase in the rate of mortality, hospitalizations, dependency in activities of daily life, and falls.", "Frailty is associated with older age and multimorbidity (two or more long-term conditions); however, little is known about its prevalence or effects on mortality in younger populations. This paper aims to examine the association between frailty, multimorbidity, specific long-term conditions, and mortality in a middle-aged and older aged population. Data were sourced from the UK Biobank. Frailty phenotype was based on five criteria (weight loss, exhaustion, grip strength, low physical activity, slow walking pace). Participants were deemed frail if they met at least three criteria, pre-frail if they fulfilled one or two criteria, and not frail if no criteria were met. Sociodemographic characteristics and long-term conditions were examined. The outcome was all-cause mortality, which was measured at a median of 7 years follow-up. Multinomial logistic regression compared sociodemographic characteristics and long-term conditions of frail or pre-frail participants with non-frail participants. Cox proportional hazards models examined associations between frailty or pre-frailty and mortality. Results were stratified by age group (37-45, 45-55, 55-65, 65-73 years) and sex, and were adjusted for multimorbidity count, socioeconomic status, body-mass index, smoking status, and alcohol use. 493 737 participants aged 37-73 years were included in the study, of whom 16 538 (3%) were considered frail, 185 360 (38%) pre-frail, and 291 839 (59%) not frail. Frailty was significantly associated with multimorbidity (prevalence 18% [4435/25 338] in those with four or more long-term conditions; odds ratio [OR] 27·1, 95% CI 25·3-29·1) socioeconomic deprivation, smoking, obesity, and infrequent alcohol consumption. The top five long-term conditions associated with frailty were multiple sclerosis (OR 15·3; 99·75% CI 12·8-18·2); chronic fatigue syndrome (12·9; 11·1-15·0); chronic obstructive pulmonary disease (5·6; 5·2-6·1); connective tissue disease (5·4; 5·0-5·8); and diabetes (5·0; 4·7-5·2). Pre-frailty and frailty were significantly associated with mortality for all age strata in men and women (except in women aged 37-45 years) after adjustment for confounders. Efforts to identify, manage, and prevent frailty should include middle-aged individuals with multimorbidity, in whom frailty is significantly associated with mortality, even after adjustment for number of long-term conditions, sociodemographics, and lifestyle. Research, clinical guidelines, and health-care services must shift focus from single conditions to the requirements of increasingly complex patient populations. CSO Catalyst Grant and National Health Service Research for Scotland Career Research Fellowship.", "The concept of a frailty index, developed in Canadian elderly populations as an indicator of biological age as opposed to chronological age, was tested in an elderly Chinese population to determine whether it is applicable in a different ethnic and cultural setting. A data set including 62 physical, psychological, and socioeconomic variables from a cohort of 2,032 persons 70 years and older (999 men, 1,033 women) was used. The distribution of the index was evaluated using the Cramer-von Mises goodness-of-fit test, and multiple linear regression was used to assess its relationship with age and sex. A biological age for each participant was calculated based on an inverse regression of age on mean frailty index and sex. The Cox proportional hazards regression model was used to assess the ability of biological age to predict death. The distribution of the frailty index most closely resembled a Weibull distribution. The frailty index increased with age until the mid-80s, when it leveled off, and was higher in women than men for each age group. The distribution of biological age is wider than that for chronological age, and it strongly predicted death. Women had an estimated 20% lesser chance of dying at a given time than did men of the same chronological age and degree of frailty. The study confirms the robustness of the concept and method of calculating the frailty index developed in elderly Canadian populations. It also suggests that the sex difference in life expectancy may have an underlying genetic basis independent of frailty.", "Recent research has shown that markers of biological age, such as leukocyte telomere length (LTL), epigenetic clocks and the frailty index (FI) are predictive of mortality and age-related diseases. However, whether these markers associate with the need for care in old age, thereby having utility in reflecting dependency, is unclear. This study was undertaken to analyze whether LTL, two epigenetic clocks-the DNA methylation age (DNAmAge) and DNAm PhenoAge-and the FI are associated with the need for regular care in up to 604 individuals (aged 48-94 years) participating in the Swedish Adoption/Twin Study of Aging. Need for regular care was defined as receiving formal or informal help in daily routines at least once per week. Logistic regression adjusted for age, sex and education was used in the analysis. The predictive accuracies, assessed as the area under the curve (AUC) for the significant biological age measures were further compared to the accuracies of the limitations in activities of daily living (ADL) and instrumental ADL (IADL). Neither LTL nor the epigenetic clocks were associated with the need for care, whereas the FI was; odds ratio for 10% increase in FI 3.54 (95% confidence interval 2.32-5.41). The FI also demonstrated higher predictive accuracy than the ADL score (FI AUC 0.80 vs. ADL score AUC 0.62; p < 0.001 for equality of the AUCs), whereas the difference between FI AUC (0.80) and IADL score AUC (0.75) was not significant (p = 0.238). The FI might thus be a useful marker for the need for care." ]
Genetic factors and clinical phenotypes in Vietnamese Parkinson's disease	Genetic factors play a crucial role in the pathogenesis of Parkinson's disease (PD). However, no comprehensive study has described genetic alterations in Vietnamese patients diagnosed with PD. This study aimed to identify genetic causes and their association with clinical phenotypes in a Vietnamese PD cohort.	[ "Parkinson's disease is a complex neurodegenerative disorder for which both rare and common genetic variants contribute to disease risk, onset, and progression. Mutations in more than 20 genes have been associated with the disease, most of which are highly penetrant and often cause early onset or atypical symptoms. Although our understanding of the genetic basis of Parkinson's disease has advanced considerably, much remains to be done. Further disease-related common genetic variability remains to be identified and the work in identifying rare risk alleles has only just begun. To date, genome-wide association studies have identified 90 independent risk-associated variants. However, most of them have been identified in patients of European ancestry and we know relatively little of the genetics of Parkinson's disease in other populations. We have a limited understanding of the biological functions of the risk alleles that have been identified, although Parkinson's disease risk variants appear to be in close proximity to known Parkinson's disease genes and lysosomal-related genes. In the past decade, multiple efforts have been made to investigate the genetic architecture of Parkinson's disease, and emerging technologies, such as machine learning, single-cell RNA sequencing, and high-throughput screens, will improve our understanding of genetic risk.", "Mutations in GBA1, the gene encoding the lysosomal enzyme glucocerebrosidase, are among the most common known genetic risk factors for the development of Parkinson disease and related synucleinopathies. A great deal is known about GBA1, as mutations in GBA1 are causal for the rare autosomal storage disorder Gaucher disease. Over the past decades, significant progress has been made in understanding the genetics and cell biology of glucocerebrosidase. A least 495 different mutations, found throughout the 11 exons of the gene are reported, including both common and rare variants. Mutations in GBA1 may lead to degradation of the protein, disruptions in lysosomal targeting and diminished performance of the enzyme in the lysosome.Gaucher disease is phenotypically diverse and has both neuronopathic and non-neuronopathic forms. Both patients with Gaucher disease and heterozygous carriers are at increased risk of developing Parkinson disease and Dementia with Lewy Bodies, although our understanding of the mechanism for this association remains incomplete. There appears to be an inverse relationship between glucocerebrosidase and α-synuclein levels, and even patients with sporadic Parkinson disease have decreased glucocerebrosidase. Glucocerebrosidase may interact with α-synuclein to maintain basic cellular functions, or impaired glucocerebrosidase could contribute to Parkinson pathogenesis by disrupting lysosomal homeostasis, enhancing endoplasmic reticulum stress or contributing to mitochondrial impairment. However, the majority of patients with GBA1 mutations never develop parkinsonism, so clearly other risk factors play a role. Treatments for Gaucher disease have been developed that increase visceral glucocerebrosidase levels and decrease lipid storage, although they have yet to properly address the neurological defects associated with impaired glucocerebrosidase. Mouse and induced pluripotent stem cell derived models have improved our understanding of glucocerebrosidase function and the consequences of its deficiency. These models have been used to test novel therapies including chaperone proteins, histone deacetylase inhibitors, and gene therapy approaches that enhance glucocerebrosidase levels and could prove efficacious in the treatment of forms of parkinsonism. Consequently, this rare monogenic disorder, Gaucher disease, provides unique insights directly applicable to our understanding and treatment of Parkinson disease, a common and complex neurodegenerative disorder.", "1·8 billion people of diverse ethnicities and cultures live in the Western Pacific Region. The increasing longevity of populations in this region is a major contributor to the exponential increase in Parkinson's disease prevalence worldwide. Differences exist between Parkinson's disease in the Western Pacific Region and in Europe and North America that might provide important insights into our understanding of the disease and approaches to management. For example, some genetic factors (such as LRRK2 mutations or variants) differ, environmental exposures might play differential roles in modulating the risk of Parkinson's disease, and fewer dyskinesias are reported, with some differences in the profile of non-motor symptoms and comorbidities. Gaps in awareness of the disease and inequitable access to treatments pose challenges. Further improvements in infrastructure, clinical governance, and services, and concerted collaborative efforts in training and research, including greater representation of the Western Pacific Region in clinical trials, will improve care of patients with Parkinson's disease in this region and beyond.", "Parkinson's disease can be caused by rare familial genetic mutations, but in most cases it is likely to result from an interaction between multiple genetic and environmental risk factors. Over recent years, many variants in a growing number of genes involved in the pathogenesis of Parkinson's disease have been identified. Mutations in several genes have been shown to cause familial parkinsonism. In this review, we discuss 12 of them (SNCA, LRRK2, Parkin, PINK1, DJ1, ATP13A2, PLA2G6, FBXO7, UCHL1, GIGYF2, HTRA2, and EIF4G1). Additionally, six genes have been shown conclusively to be risk factors for sporadic Parkinson's disease, and are also discussed (GBA, MAPT, BST1, PARK16, GAK, and HLA). Many more genes and genetic loci have been suggested, but need confirmation. There is evidence that pathways involved in the rare familial forms also play a role in the sporadic form, and that the respective genes might also be risk factors for sporadic Parkinson's disease. The identification of genes involved in the development of Parkinson's disease will improve our understanding of the underlying molecular mechanisms, and will hopefully lead to new drug targets and treatment strategies.", "To examine the influence of the glucocerebrosidase (GBA) mutation carrier state on age at onset of Parkinson's disease (PD), the motor phenotype and cognitive function at baseline assessment in a large cohort of UK patients. We also analysed the prevalence of mood and behavioural problems that may confound the assessment of cognitive function. We prospectively recruited patients with PD in the Tracking Parkinson's study. We fully sequenced the GBA gene in all recently diagnosed patients (≤3.5 years). We examined cognitive (Montreal Cognitive Assessment) and motor (Movement Disorder Society Unified Parkinson's Disease Rating Scale part 3) function at a baseline assessment, at an average of 1.3 years after diagnosis. We used logistic regression to determine predictors of PD with mild cognitive impairment and PD with dementia. We studied 1893 patients with PD: 48 (2.5%) were heterozygous carriers for known Gaucher's disease (GD) causing pathogenic mutations; 117 (6.2%) had non-synonymous variants, previously associated with PD, and 28 (1.5%) patients carried variants of unknown significance in the GBA gene. L444P was the most common pathogenic GBA mutation. Patients with pathogenic GBA mutations were on average 5 years younger at disease onset compared with non-carriers (P=0.02). PD patients with GD-causing mutations did not have an increased family risk of PD. Patients with GBA mutations were more likely to present with the postural instability gait difficulty phenotype compared with non-carriers (P=0.02). Patients carrying pathogenic mutations in GBA had more advanced Hoehn and Yahr stage after adjustment for age and disease duration compared with non-carriers (P=0.005). There were no differences in cognitive function between GBA mutation carriers and non-carriers at this early disease stage. Our study confirms the influence of GBA mutations on the age of onset, disease severity and motor phenotype in patients with PD. Cognition did not differ between GBA mutation carriers and non-carriers at baseline, implying that cognitive impairment/dementia, reported in other studies at a later disease stage, is not present in recently diagnosed cases. This offers an important window of opportunity for potential disease-modifying therapy that may protect against the development of dementia in GBA-PD. NCT02881099; Results.", "The objective of this work was to investigate survival, dementia, and genotype-phenotype correlations in patients with Parkinson's disease (PD) with and without mutations on the glucocerebrosidase gene (GBA). We included 2,764 unrelated consecutive PD patients: 123 GBA carriers (67 mild-p.N370S and 56 severe mainly p.L444P) and 2,641 noncarriers. Brain perfusion and dopamine transporter imaging was analyzed, including dementia with Lewy Bodies (DLB) as an additional control group. Multivariable analysis adjusted by sex, age at onset, and disease duration attributed to GBA carriers a greater risk for dementia (hazard ratio [HR] = 3.16; p < 0.001) and death (HR = 1.85; p = 0.002) than noncarriers. When dementia was introduced in the model as a time-dependent covariate, the mortality risk remained greater in carriers (HR = 1.65; p = 0.016), suggesting that other clinical features are likely to contribute to reduced survival. At last examination, GBA carriers had worse motor symptoms, particularly nondopaminergic features. Carriers of severe mutations had greater risk for dementia compared to mild mutations (p < 0.001), but similar mortality risk. Consistent with clinical data, GBA carriers showed reduced posterior parietal and occipital cortical synaptic activity and nigrostriatal function than PD noncarriers. Neuroimaging features of carriers of mild mutations overlapped with PD noncarriers, whereas carriers of severe mutations were closer to DLB. Survival is reduced in GBA carriers compared to noncarriers; this seems to be partially independent from the increased risk for early dementia. The risk for dementia is strongly modulated by type of mutation. In the clinical continuum between PD and DLB, patients with GBA mutations seem to localize midway, with carriers of severe mutations closer to DLB than to idiopathic PD. Ann Neurol 2016;80:662-673.", "VarSome.com is a search engine, aggregator and impact analysis tool for human genetic variation and a community-driven project aiming at sharing global expertise on human variants. VarSome is freely available at http://varsome.com. Supplementary data are available at Bioinformatics online." ]
Receptor tyrosine kinase-like orphan receptor 1 as a tumor therapeutic target	The receptor tyrosine kinase-like orphan receptor 1 (ROR1) is a membrane receptor that plays a key role in development. It is highly expressed during the embryonic stage and relatively low in some normal adult tissues. Malignancies such as leukemia, lymphoma, and some solid tumors overexpress ROR1, making it a promising target for cancer treatment. Moreover, immunotherapy with autologous T-cells engineered to express a ROR1-specific chimeric antigen receptor (ROR1 CAR-T cells) has emerged as a personalized therapeutic option for patients with tumor recurrence after conventional treatments. However, tumor cell heterogeneity and tumor microenvironment (TME) hinder successful clinical outcomes. This review briefly describes the biological functions of ROR1 and its relevance as a tumor therapeutic target, as well as the architecture, activity, evaluation, and safety of some ROR1 CAR-T cells used in basic research and clinical trials. Finally, the feasibility of applying the ROR1 CAR-T cell strategy in combination with therapies targeting other tumor antigens or with inhibitors that prevent tumor antigenic escape is also discussed.	[ "Receptor-tyrosine-kinase-like orphan receptor 1 (ROR1) is expressed during embryogenesis and by certain leukemias, but not by normal adult tissues. Here we show that the neoplastic cells of many human breast cancers express the ROR1 protein and high-level expression of ROR1 in breast adenocarcinoma was associated with aggressive disease. Silencing expression of ROR1 in human breast cancer cell lines found to express this protein impaired their growth in vitro and also in immune-deficient mice. We found that ROR1 could interact with casein kinase 1 epsilon (CK1ε) to activate phosphoinositide 3-kinase-mediated AKT phosphorylation and cAMP-response-element-binding protein (CREB), which was associated with enhanced tumor-cell growth. Wnt5a, a ligand of ROR1, could induce ROR1-dependent signaling and enhance cell growth. This study demonstrates that ROR1 is expressed in human breast cancers and has biological and clinical significance, indicating that it may be a potential target for breast cancer therapy.", "Receptor tyrosine kinases (RTKs) participate in numerous developmental decisions. Ror RTKs are a family of orphan receptors that are related to muscle specific kinase (MuSK) and Trk neurotrophin receptors. MuSK assembles acetylcholine receptors at the neuromuscular junction, and Trk receptors function in the developing nervous system (reviewed in [3-5]). Rors have been identified in nematodes, insects and mammals. Recent studies have begun to shed light on Ror function during development. In most species, Rors are expressed in many tissue types during development. Analyses of mutants that are defective in the single nematode Ror demonstrate a role in cell migration and in orienting cell polarity. Mice lacking one of the two Ror gene products display defects in bone and heart formation. Similarly, two different human bone development disorders, dominant brachydactyly B and recessive Robinow syndrome, result from mutations in one of the human Ror genes.", "Increased levels of total tumor-infiltrating lymphocytes (TILs) are generally associated with good prognosis in several breast cancer subtypes. Subtypes of TILs impact both tumor cells and immune cells in a variety of different ways, leading to either a pro-tumor or antitumor effect. Tumor-infiltrating CD8+ T cells and natural killer (NK) cells perform as effector cells against tumor cells and are associated with better clinical outcome. Immunotherapy approaches that improve the antitumor activity and proliferation of CD8+ T and NK cells include PD-1/PD-L1 blockade, CAR T cell therapy, or ex vivo-stimulated NK cells. A subset of CD8+ T cells, tissue-resident memory T cells, has also recently been associated with good prognosis in breast cancer patients, and has potential to serve as a predictive biomarker and therapeutic target. Tumor-infiltrating B cells also secrete apoptosis-inducing IgG antibodies and can act as antigen-presenting cells to prime CD4+ and CD8+ T cells. On the other hand, regulatory T and regulatory B cells modulate the immune response from CD8+ T cells and NK cells by secreting immunosuppressive cytokines and inhibiting maturation of antigen-presenting cells (APCs). These regulatory cells are typically associated with poor prognosis, therefore rendering suppression of their regulatory function a key immunotherapeutic strategy.", "Despite its revolutionary success in hematological malignancies, chimeric antigen receptor T (CAR-T) cell therapy faces disappointing clinical results in solid tumors. The poor efficacy has been partially attributed to the lack of understanding in how CAR-T cells function in a solid tumor microenvironment. Hypoxia plays a critical role in cancer progression and immune editing, which potentially results in solid tumors escaping immunosurveillance and CAR-T cell-mediated cytotoxicity. Mechanistic studies of CAR-T cell biology in a physiological environment has been limited by the complexity of tumor-immune interactions in clinical and animal models, as well as by a lack of reliable in vitro models. A microdevice platform that recapitulates a 3D tumor section with a gradient of oxygen and integrates fluidic channels surrounding the tumor for CAR-T cell delivery is engineered. The design allows for the evaluation of CAR-T cell cytotoxicity and infiltration in the heterogeneous oxygen landscape of in vivo solid tumors at a previously unachievable scale in vitro.", "Neuroblastoma is the most common inheritable, solid neoplasm in children found under the age of 7 and accounts for approximately 7% of childhood cancers. A common treatment that has been prescribed for over a decade is retinoid therapy [using all‑trans retinoic acid (RA)]. Treatment with this differentiating agent has been revealed to progress the cells from their stem‑cell state to a mature neuronal state gaining classical neuronal characteristics, including the suppression of proliferation. However, the molecular mechanism underlying the action of RA treatment remains to be elucidated. In the present study, a novel mechanism of RA‑induced differentiation via regulation of receptor tyrosine kinase‑like orphan receptor 1 (ROR1) is reported. ROR1 is overexpressed in neuroblastoma but significantly downregulated in mature differentiated neurons. Hence, it was hypothesized that RA may modulate ROR1 leading to differentiation and termination of cancerous properties. Immunoblotting revealed that following RA treatment, ROR1 levels initially increased then sharply decreased by 96 h. This was paired with synaptophysin, a mature neuron marker, sharply increasing concurrently, providing evidence of differentiation by 96 h. Investigation of the ROR1 pathway confirmed ROR1‑dependent downstream activation of the PI3K/AKT signaling axis, a growth pathway previously demonstrated to promote differentiation. Chromatin immunoprecipitation revealed an increase in RAR binding to the promoters of ROR1 and its endogenous ligand, Wnt5a. This research provided compelling evidence that RA is able to modulate the expression of ROR1 and Wnt5a to promote differentiation through the expression of synaptophysin. This data combined with the overarching data from the scientific community regarding proliferation and other proliferative factors in early‑stage neurons provides a more in‑depth model of the process of differentiation in neurons.", "Neurite elongation and branching are key cellular events during brain development as they underlie the formation of a properly wired neuronal network. Here we report that the receptor tyrosine kinases Ror1 and Ror2 modulate the growth of neurites as well as their branching pattern in hippocampal neurons. Upon Ror1 or Ror2 suppression using antisense oligonucleotides or RNA interference (RNAi), neurons extended shorter and less branched minor processes when compared to those in control cells. In addition, Ror-depleted cells elongated longer, albeit less branched, axons than seen in control cells. Conversely, Ror overexpression both in non-neuronal cells and in hippocampal neurons resulted in the enhanced extension of short and highly branched processes. These phenotypes were accompanied by changes in the microtubule-associated proteins MAP1B and MAP2. Taken together, these results support a novel role for Ror receptors as modulators of neurite extension in central neurons.", "Data regarding the efficacy of treatment with ibrutinib-rituximab, as compared with standard chemoimmunotherapy with fludarabine, cyclophosphamide, and rituximab, in patients with previously untreated chronic lymphocytic leukemia (CLL) have been limited. In a phase 3 trial, we randomly assigned (in a 2:1 ratio) patients 70 years of age or younger with previously untreated CLL to receive either ibrutinib and rituximab for six cycles (after a single cycle of ibrutinib alone), followed by ibrutinib until disease progression, or six cycles of chemoimmunotherapy with fludarabine, cyclophosphamide, and rituximab. The primary end point was progression-free survival, and overall survival was a secondary end point. We report the results of a planned interim analysis. A total of 529 patients underwent randomization (354 patients to the ibrutinib-rituximab group, and 175 to the chemoimmunotherapy group). At a median follow-up of 33.6 months, the results of the analysis of progression-free survival favored ibrutinib-rituximab over chemoimmunotherapy (89.4% vs. 72.9% at 3 years; hazard ratio for progression or death, 0.35; 95% confidence interval [CI], 0.22 to 0.56; P<0.001), and the results met the protocol-defined efficacy threshold for the interim analysis. The results of the analysis of overall survival also favored ibrutinib-rituximab over chemoimmunotherapy (98.8% vs. 91.5% at 3 years; hazard ratio for death, 0.17; 95% CI, 0.05 to 0.54; P<0.001). In a subgroup analysis involving patients without immunoglobulin heavy-chain variable region (IGHV) mutation, ibrutinib-rituximab resulted in better progression-free survival than chemoimmunotherapy (90.7% vs. 62.5% at 3 years; hazard ratio for progression or death, 0.26; 95% CI, 0.14 to 0.50). The 3-year progression-free survival among patients with IGHV mutation was 87.7% in the ibrutinib-rituximab group and 88.0% in the chemoimmunotherapy group (hazard ratio for progression or death, 0.44; 95% CI, 0.14 to 1.36). The incidence of adverse events of grade 3 or higher (regardless of attribution) was similar in the two groups (in 282 of 352 patients [80.1%] who received ibrutinib-rituximab and in 126 of 158 [79.7%] who received chemoimmunotherapy), whereas infectious complications of grade 3 or higher were less common with ibrutinib-rituximab than with chemoimmunotherapy (in 37 patients [10.5%] vs. 32 [20.3%], P<0.001). The ibrutinib-rituximab regimen resulted in progression-free survival and overall survival that were superior to those with a standard chemoimmunotherapy regimen among patients 70 years of age or younger with previously untreated CLL. (Funded by the National Cancer Institute and Pharmacyclics; E1912 ClinicalTrials.gov number, NCT02048813.)." ]
Inflammation, gut microbiome disruption, and the severity of SARS-CoV-2 disease	The SARS-CoV-2 virus (severe acute respiratory syndrome coronavirus 2), and the disease it causes (COVID-19), have had a profound impact on global human society and threaten to continue to have such an impact with newly emerging variants. Because of the widespread effects of SARS-CoV-2, understanding how lifestyle choices impact the severity of disease is imperative. This review summarizes evidence for an involvement of chronic, non-resolving inflammation, gut microbiome disruption (dysbiosis with loss of beneficial microorganisms), and impaired viral defenses, all of which are associated with an imbalanced lifestyle, in severe disease manifestations and post-acute sequelae of SARS-CoV-2 (PASC). Humans' physiological propensity for uncontrolled inflammation and severe COVID-19 are briefly contrasted with bats' low propensity for inflammation and their resistance to viral disease. This insight is used to identify positive lifestyle factors with the potential to act in synergy for restoring balance to the immune response and gut microbiome, and thereby protect individuals against severe COVID-19 and PASC. It is proposed that clinicians should consider recommending lifestyle factors, such as stress management, balanced nutrition and physical activity, as preventative measures against severe viral disease and PASC.	[ "Antimicrobial resistance is a hidden threat lurking behind the COVID-19 pandemic which has claimed thousands of lives prior to the emergence of the global outbreak. With a pandemic on the scale of COVID-19, antimicrobial resistance has the potential to become a double-edged sword with the overuse of antibiotics having the potential of taking us back to the pre-antibiotic era. Antimicrobial resistance is majorly attributed to widespread and unnecessary use of antibiotics, among other causes, which has facilitated the emergence and spread of resistant pathogens. Our study aimed to conduct a rapid review of national treatment guidelines for COVID-19 in 10 African countries (Ghana, Kenya, Uganda, Nigeria, South Africa, Zimbabwe, Botswana, Liberia, Ethiopia, and Rwanda) and examined its implication for antimicrobial resistance response on the continent. Our findings revealed that various antibiotics, such as azithromycin, doxycycline, clarithromycin, ceftriaxone, erythromycin, amoxicillin, amoxicillin-clavulanic acid, ampicillin, gentamicin, benzylpenicillin, piperacillin/tazobactam, ciprofloxacin, ceftazidime, cefepime, vancomycin, meropenem, and cefuroxime among others, were recommended for use in the management of COVID-19. This is worrisome in that COVID-19 is a viral disease and only a few COVID-19 patients would have bacterial co-infection. Our study highlighted the need to emphasize prudent and judicious use of antibiotics in the management of COVID-19 in Africa.", "Although COVID-19 is primarily a respiratory illness, there is mounting evidence suggesting that the GI tract is involved in this disease. We investigated whether the gut microbiome is linked to disease severity in patients with COVID-19, and whether perturbations in microbiome composition, if any, resolve with clearance of the SARS-CoV-2 virus. In this two-hospital cohort study, we obtained blood, stool and patient records from 100 patients with laboratory-confirmed SARS-CoV-2 infection. Serial stool samples were collected from 27 of the 100 patients up to 30 days after clearance of SARS-CoV-2. Gut microbiome compositions were characterised by shotgun sequencing total DNA extracted from stools. Concentrations of inflammatory cytokines and blood markers were measured from plasma. Gut microbiome composition was significantly altered in patients with COVID-19 compared with non-COVID-19 individuals irrespective of whether patients had received medication (p<0.01). Several gut commensals with known immunomodulatory potential such as Faecalibacterium prausnitzii, Eubacterium rectale and bifidobacteria were underrepresented in patients and remained low in samples collected up to 30 days after disease resolution. Moreover, this perturbed composition exhibited stratification with disease severity concordant with elevated concentrations of inflammatory cytokines and blood markers such as C reactive protein, lactate dehydrogenase, aspartate aminotransferase and gamma-glutamyl transferase. Associations between gut microbiota composition, levels of cytokines and inflammatory markers in patients with COVID-19 suggest that the gut microbiome is involved in the magnitude of COVID-19 severity possibly via modulating host immune responses. Furthermore, the gut microbiota dysbiosis after disease resolution could contribute to persistent symptoms, highlighting a need to understand how gut microorganisms are involved in inflammation and COVID-19.", "Wild animals play an integral and complex role in the economies and ecologies of many countries across the globe, including those of West and Central Africa, the focus of this policy perspective. The trade in wild meat, and its role in diets, have been brought into focus as a consequence of discussions over the origins of COVID-19. As a result, there have been calls for the closure of China's \"wet markets\"; greater scrutiny of the wildlife trade in general; and a spotlight has been placed on the potential risks posed by growing human populations and shrinking natural habitats for animal to human transmission of zoonotic diseases. However, to date there has been little attention given to what the consequences of the COVID-19 economic shock may be for the wildlife trade; the people who rely on it for their livelihoods; and the wildlife that is exploited. In this policy perspective, we argue that the links between the COVID-19 pandemic, rural livelihoods and wildlife are likely to be more complex, more nuanced, and more far-reaching, than is represented in the literature to date. We develop a causal model that tracks the likely implications for the wild meat trade of the systemic crisis triggered by COVID-19. We focus on the resulting economic shockwave, as manifested in the collapse in global demand for commodities such as oil, and international tourism services, and what this may mean for local African economies and livelihoods. We trace the shockwave through to the consequences for the use of, and demand for, wild meats as households respond to these changes. We suggest that understanding and predicting the complex dynamics of wild meat use requires increased collaboration between environmental and resource economics and the ecological and conservation sciences.", "Recent studies suggest an association between particulate matter (PM) air pollution and gastrointestinal (GI) disease. In addition to direct deposition, PM can be indirectly deposited in oropharynx via mucociliary clearance and upon swallowing of saliva and mucus. Within the GI tract, PM may alter the GI epithelium and gut microbiome. Our goal was to determine the effect of PM on gut microbiota in a murine model of PM exposure via inhalation. C57BL/6 mice were exposed via inhalation to either concentrated ambient particles or filtered air for 8-h per day, 5-days a week, for a total of 3-weeks. At exposure's end, GI tract tissues and feces were harvested, and gut microbiota was analyzed. Alpha-diversity was modestly altered with increased richness in PM-exposed mice compared to air-exposed mice in some parts of the GI tract. Most importantly, PM-induced alterations in the microbiota were very apparent in beta-diversity comparisons throughout the GI tract and appeared to increase from the proximal to distal parts. Changes in some genera suggest that distinct bacteria may have the capacity to bloom with PM exposure. Exposure to PM alters the microbiota throughout the GI tract which maybe a potential mechanism that explains PM induced inflammation in the GI tract.", "To understand the clinical characteristics of COVID-19 patients with clinically diagnosed bacterial co-infection (CDBC), and therefore contributing to their early identification and prognosis estimation. 905 COVID-19 patients from 7 different centers were enrolled. The demography data, clinical manifestations, laboratory results, and treatments were collected accordingly for further analyses. Around 9.5% of the enrolled COVID-19 patients were diagnosed with CDBC. Older patients or patients with cardiovascular comorbidities have increased CDBC probability. Increased body temperature, longer fever duration, anhelation, gastrointestinal symptoms, illness severity, intensive care unit attending, ventilation treatment, glucocorticoid therapy, longer hospitalization time are correlated to CDBC. Among laboratory results, increased white blood cell counting (mainly neutrophil), lymphocytopenia, increased procalcitonin, erythrocyte sedimentation rate, C-reaction protein, D-dimer, blood urea nitrogen, lactate dehydrogenase, brain natriuretic peptide, myoglobin, blood sugar and decreased albumin are also observed, indicating multiple system functional damage. Radiology results suggested ground glass opacity mixed with high density effusion opacities and even pleural effusion. The aged COVID-19 patients with increased inflammatory indicators, worse lymphopenia and cardiovascular comorbidities are more likely to have clinically diagnosed bacterial co-infection. Moreover, they tend to have severer clinical manifestations and increased probability of multiple system functional damage.", "Coronavirus disease 2019 (COVID-19) is the third deadly coronavirus infection of the 21st century that has proven to be significantly more lethal than its predecessors, with the number of infected patients and deaths still increasing daily. From December 2019 to July 2021, this virus has infected nearly 200 million people and led to more than 4 million deaths. Our understanding of COVID-19 is constantly progressing, giving better insight into the heterogeneous nature of its acute and long-term effects. Recent literature on the long-term health consequences of COVID-19 discusses the need for a comprehensive understanding of the multisystemic pathophysiology, clinical predictors, and epidemiology to develop and inform an evidence-based, multidisciplinary management approach. A PubMed search was completed using variations on the term post-acute COVID-19. Only peer-reviewed studies in English published by July 17, 2021 were considered for inclusion. All studies discussed in this text are from adult populations unless specified (as with multisystem inflammatory syndrome in children). The preliminary evidence on the pulmonary, cardiovascular, neurological, hematological, multisystem inflammatory, renal, endocrine, gastrointestinal, and integumentary sequelae show that COVID-19 continues after acute infection. Interdisciplinary monitoring with holistic management that considers nutrition, physical therapy, psychological management, meditation, and mindfulness in addition to medication will allow for the early detection of post-acute COVID-19 sequelae symptoms and prevent long-term systemic damage. This review serves as a guideline for effective management based on current evidence, but clinicians should modify recommendations to reflect each patient's unique needs and the most up-to-date evidence. The presence of long-term effects presents another reason for vaccination against COVID-19.", "The number of microbes on Earth may be 1030, exceeding all other diversity. A small number of these can infect people and cause disease. The diversity of parasitic organisms likely correlates with the hosts they live in and the number mammal hosts for zoonotic infections increases with species richness among mammalian orders. Thus, while habitat loss and fragmentation may reduce species diversity, the habitat encroachment by people into species-rich areas may increase the exposure of people to novel infectious agents from wildlife. Here, we present a theoretical framework that exploits the species-area relationship to link the exposure of people to novel infections with habitat biodiversity. We model changes in human exposure to microbes through defined classes of habitat fragmentation and predict that increased habitat division intrinsically increases the hazard from microbes for all modelled biological systems. We apply our model to African tropical forests as an example. Our results suggest that it is possible to identify high-risk areas for the mitigation and surveillance of novel disease emergence and that mitigation measures may reduce this risk while conserving biodiversity.", "The recent novel coronavirus disease (COVID-19) outbreak, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is seeing a rapid increase in infected patients worldwide. The host immune response to SARS-CoV-2 appears to play a critical role in disease pathogenesis and clinical manifestations. SARS-CoV-2 not only activates antiviral immune responses, but can also cause uncontrolled inflammatory responses characterized by marked pro-inflammatory cytokine release in patients with severe COVID-19, leading to lymphopenia, lymphocyte dysfunction, and granulocyte and monocyte abnormalities. These SARS-CoV-2-induced immune abnormalities may lead to infections by microorganisms, septic shock, and severe multiple organ dysfunction. Therefore, mechanisms underlying immune abnormalities in patients with COVID-19 must be elucidated to guide clinical management of the disease. Moreover, rational management of the immune responses to SARS-CoV-2, which includes enhancing anti-viral immunity while inhibiting systemic inflammation, may be key to successful treatment. In this review, we discuss the immunopathology of COVID-19, its potential mechanisms, and clinical implications to aid the development of new therapeutic strategies against COVID-19.", "Coronavirus disease 2019 (COVID-19), which has been declared a pandemic, has exhibited a wide range of severity worldwide. Although this global variation is largely affected by socio-medical situations in each country, there is also high individual-level variation attributable to elderliness and certain underlying medical conditions, including high blood pressure, diabetes, and obesity. As both elderliness and the aforementioned chronic conditions are often associated with an altered gut microbiota, resulting in disrupted gut barrier integrity, and gut symptoms have consistently been associated with more severe illness in COVID-19 patients, it is possible that dysfunction of the gut as a whole influences COVID-19 severity. This article summarizes the accumulating evidence that supports the hypothesis that an altered gut microbiota and its associated leaky gut may contribute to the onset of gastrointestinal symptoms and occasionally to additional multiorgan complications that may lead to severe illness by allowing leakage of the causative coronavirus into the circulatory system.", "Type I interferon (IFNα/β) pathways are fine-tuned to elicit antiviral protection while minimizing immunopathology; however, the initiating stimuli, target tissues, and underlying mechanisms are unclear. Using models of physiological and dysregulated IFNα/β receptor (IFNAR1) surface expression, we show here that IFNAR1-dependent signals set the steady-state IFN signature in both hematopoietic and stromal cells. Increased IFNAR1 levels promote a lung environment refractory to early influenza virus replication by elevating the baseline interferon signature. Commensal microbiota drive the IFN signature specifically in lung stroma, as shown by antibiotic treatment and fecal transplantation. Bone marrow chimera experiments identify lung stromal cells as crucially important for early antiviral immunity and stroma-immune cell interaction for late antiviral resistance. We propose that the microbiota-driven interferon signature in lung epithelia impedes early virus replication and that IFNAR1 surface levels fine-tune this signature. Our findings highlight the interplay between bacterial and viral exposure, with important implications for antibiotic use." ]
[Clinical consensus on chronic constipation in the elderly].	It is of significant importance to scientifically assess and efficiently address chronic constipation in the elderly population. Therefore, organizing some domestic experts of geriatrics and gastroenterology, we have formulated this consensus to reference for clinical staff.	[ "The purposes of this study were: (1) to examine the efficacy of anorectal biofeedback (AB) for constipation compared to a biofeedback control (BC) treatment and (2) to examine the extent to which self-reported childhood sexual/physical abuse predicted biofeedback outcome. Twenty-one patients with pelvic floor dyssynergia were randomized to either (1) an AB arm, where patients learned to isolate the anal sphincter using an electromyography probe, or (2) a BC arm that controlled for the nonspecific effects of biofeedback, where patients learned to relax trapezius or temporalis muscles with EMG feedback. Both treatments were delivered by registered nurses for six sessions. Prior to randomization and post-treatment, patients completed the validated Constipation Severity Instrument and two measures of quality of life (QOL), the Irritable Bowel Syndrome-QOL, and the SF-36. Generalized estimating equations examined the within-group and between-group differences over time. Pre- and post-treatment data were obtained for six AB and nine BC patients. AB patients' overall constipation severity scores decreased by 35.5% (vs. 15.3%), and their obstructive defecation symptom scores decreased by 37.9% (vs. 19.7%) compared to BC. A similar pattern was shown on the IBS-QOL. On the SF-36 Mental Health Composite (MCS), AB scores improved 28.0% compared to BC scores, which worsened 12.7%. Those without (vs. with) a childhood sexual/physical abuse history showed improvement on the MCS post-biofeedback. While our sample was statistically underpowered, AB produced clinical improvements in constipation severity and QOL.", "Although a range of guidelines for the diagnosis and treatment of chronic constipation has been carried out, there was very little information about the understanding on constipation. The aim of the present study was to estimate the understanding of constipation symptoms and the diagnosis and management of constipation by clinical physicians in China. Participants were physicians and researchers in the field of gastroenterology in China who were scheduled to attend the National Conference on gastrointestinal motility (Constipation). Based on the recommendation of the Rome Foundation Board, the self-reported questionnaire was constructed. Although most of the opinions on symptoms of constipation were consistent, there were still some differences. Opinions on the Bristol stool form during constipation were discordant, 34% of the doctors thought that it was type 1 and type 2, while 46%of the doctors suggested that type 3 should also be considered constipation. There was no significant difference between them (P = 0.05); We investigated the interpretation on the duration of defecation prolonged, 27% of the doctors suggested it should be longer than 10 minutes, 22% of the doctors suggested it should be longer than 20 minutes, and other 22% of the doctors suggested it should be time of defecation became longer compared to previously bowel habits, there was no significant difference among them (P = 0.38).Only 36% of the doctors thought that psychotherapy was most important in the treatment of severe constipation, while 37% of the doctors thought that medication treatment was most important in the treatment of severe constipation, there was no significant difference between them (P = 0.895). We were able to obtain valuable information about current views on symptoms of constipation and the diagnosis and treatment of constipation among Chinese doctors. Although most of the opinions were consistent there were still some differences. This study indicated that in practice in China there was a need for further study on the role of constipation symptoms and there may also be a need for better establishment of consensus guidelines for constipation.", "To survey the emotional and sleeping status of patients with chronic constipation (CC) and analyze the relationship between psychological status and constipated symptoms. From January 2009 to April 2010, 5 centers and 25 stratified hospitals were selected as the representatives of different regions of mainland China. The CC questionnaires including constipated symptoms, emotional and sleeping status, previous treatments and self-reported impact of constipation on health status, etc. Questionnaires were completed by well-trained physicians or investigators during face-to-face interviews. CC was diagnosed in accordance with the Rome III criteria. A total of 909 valid questionnaires analyzed. There were 258 males and 651 females with a mean age of (49 ± 19) years. 41.5% (377 cases) reported \"tense feelings\" and 38.3% (348 cases) \"felt downcast\" over the past 3 months. The patients feeling tense and(or) downcast \"frequently\" and \"most of time\" were around 11.3% (103 cases) and 9.4% (85 cases). And 43.8% (398 cases) patients reported sleeping disorders over the past 3 months. Regional differences existed in the comorbidities of psychological and sleeping disorders in CC patients, especially in those from tertiary hospitals. And it was the highest in Beijing area for tense feelings and downcast. The sleeping disorders were the most common in the patients from secondary hospitals, of which 66.1% (37/56) and 65.0% (39/60) were from Wuhan and Xi'an respectively. They were higher than Beijing and Guangzhou (39.7% (23/58), 29.0% (9/31), all P = 0.001). The patients from rural primary clinics suffered more sleeping disorders than those from urban primary cares (P = 0.026). About 35.0% (318 cases) and 28.4% (258 cases) patients reported their constipation was related with emotional and sleeping disorders. The comorbidities of psychological and sleeping disorders were more common in severe constipated patients than mild and moderate counterparts and resulted in more hospital visits (both P = 0.000). The CC patients often have the comorbidities of psychological and sleeping disorders with regional differences. The patients from the hospitals at various levels may present different spectrums of comorbidities of psychological and sleeping disorders. And the severity of CC influences the moods, sleeps and hospital visits.", "The aim was to research the changes in pelvic floor morphology and corresponding visceras in patients with outlet obstructive constipation (OOC). Thirty-eight patients with OOC and 12 healthy volunteers were enrolled in this study. With simultaneous pelvicography and colpocystodefecography (PCCD), including pelvicography, vaginal opacification, voiding cystography and defecography, pelvic floor morphology was observed and the anorectal angle, the level of the perineum, peritoneum and bladder were measured. Thirty-seven cases of internal rectal prolapse (IRP), 5 cases of rectocele (RC) and 5 cases of spastic pelvic floor syndrome SPFS were diagnosed by PCCD. 12 IRP, 4 RC and 1 SPFS were detected by common physical examination. All of these were confirmed by PCCD. Moreover, PCCD found 9 pelvic floor hernia or peritoneoceles, 6 cystoceles, 3 descending perineum syndromes and 10 uterine prolapses. Compared with controls, OOC patients had a significantly large anorectal angle during defecation, abnormal descending of the perineum at rest and during defecation, and a deep pouch of Douglas during defecation. Some patients with urinary system symptoms may have had an abnormal descent of the bladder during rest and defecation. Simultaneous PCCD has a higher positive ratio than the common physical examination in diagnosing IRP and RC, and provides information for the diagnosis of pelvic floor hernia or peritoneocele, cystocele or uterine prolapse. PCCD is helpful in the selection of a proper surgical procedure." ]
Frailty Index and Postoperative Complications of Aged Patients with Nodular Goiter	To explore the correlation between frailty index (FI) and postoperative complications of aged patients with nodular goiter (NG).	[ "We investigated whether preoperative frailty among older noncardiac surgical patients provides information about the development of postoperative delirium that is in addition to traditional geriatric risk factors. One-third of patients had a frailty score ≥3, which is considered \"frail\" in others' research. Twenty-five percent of patients developed postoperative delirium, which was measured using the confusion assessment method. Multivariable logistic regression showed that age, activities of daily living dependence, instrumental activities of daily living dependence, and cognitive functioning did not contribute significantly to the prediction of postoperative delirium. Only preoperative symptoms of depression (odds ratio=1.42; 95% confidence interval=1.06-1.91; P=0.018) and the frailty score (odds ratio=1.84; 95% confidence interval=1.07-3.1; P=0.028) were independently associated with the development of postoperative delirium.", "Frailty is considered highly prevalent in old age and to confer high risk for falls, disability, hospitalization, and mortality. Frailty has been considered synonymous with disability, comorbidity, and other characteristics, but it is recognized that it may have a biologic basis and be a distinct clinical syndrome. A standardized definition has not yet been established. To develop and operationalize a phenotype of frailty in older adults and assess concurrent and predictive validity, the study used data from the Cardiovascular Health Study. Participants were 5,317 men and women 65 years and older (4,735 from an original cohort recruited in 1989-90 and 582 from an African American cohort recruited in 1992-93). Both cohorts received almost identical baseline evaluations and 7 and 4 years of follow-up, respectively, with annual examinations and surveillance for outcomes including incident disease, hospitalization, falls, disability, and mortality. Frailty was defined as a clinical syndrome in which three or more of the following criteria were present: unintentional weight loss (10 lbs in past year), self-reported exhaustion, weakness (grip strength), slow walking speed, and low physical activity. The overall prevalence of frailty in this community-dwelling population was 6.9%; it increased with age and was greater in women than men. Four-year incidence was 7.2%. Frailty was associated with being African American, having lower education and income, poorer health, and having higher rates of comorbid chronic diseases and disability. There was overlap, but not concordance, in the cooccurrence of frailty, comorbidity, and disability. This frailty phenotype was independently predictive (over 3 years) of incident falls, worsening mobility or ADL disability, hospitalization, and death, with hazard ratios ranging from 1.82 to 4.46, unadjusted, and 1.29-2.24, adjusted for a number of health, disease, and social characteristics predictive of 5-year mortality. Intermediate frailty status, as indicated by the presence of one or two criteria, showed intermediate risk of these outcomes as well as increased risk of becoming frail over 3-4 years of follow-up (odds ratios for incident frailty = 4.51 unadjusted and 2.63 adjusted for covariates, compared to those with no frailty criteria at baseline). This study provides a potential standardized definition for frailty in community-dwelling older adults and offers concurrent and predictive validity for the definition. It also finds that there is an intermediate stage identifying those at high risk of frailty. Finally, it provides evidence that frailty is not synonymous with either comorbidity or disability, but comorbidity is an etiologic risk factor for, and disability is an outcome of, frailty. This provides a potential basis for clinical assessment for those who are frail or at risk, and for future research to develop interventions for frailty based on a standardized ascertainment of frailty.", "Whether patients who subsequently develop early postoperative delirium have a genetic predisposition that renders them at risk for postoperative delirium has not been determined. The authors conducted a nested cohort study to include patients aged > or = 65 yr who were scheduled to undergo major noncardiac surgery requiring anesthesia. A structured interview was conducted preoperatively and for the first 2 days postoperatively to determine the presence of delirium, defined using the Confusion Assessment Method. Blood was drawn for measurement of the apolipoprotein genotypes. Bivariate tests of association were conducted between delirium and apolipoprotein genotypes and other potentially important risk factors. Variables that had significant bivariate association with postoperative delirium were entered in a forward multivariable logistic regression model. Of the 190 patients studied, 15.3% developed delirium on both days 1 and 2 after surgery. Forty-six patients (24.2%) had at least one copy of the apolipoprotein e4 allele. The presence of one copy of the e4 allele was associated with an increased risk of early postoperative delirium (28.3% vs. 11.1%; P = 0.005). Even after adjusting for covariates, patients with one copy of the e4 allele were still more likely to have an increased risk of early postoperative delirium (odds ratio, 3.64; 95% confidence interval, 1.51-8.77) compared with those without the e4 allele. Apolipoprotein e4 carrier status was associated with an increased risk for early postoperative delirium after controlling for known demographic and clinical risk factors. These results suggest that genetic predisposition plays a role and may interact with anesthetic/surgical factors contributing to the development of early postoperative delirium.", "To assess the association between frailty and incident Alzheimer's disease (AD) and cognitive decline. Frailty is common in older persons and associated with adverse health outcomes. Study subjects included 823 older persons without dementia who participated in the Rush Memory and Aging Project, a longitudinal study of aging, and underwent annual assessments of frailty, cognition, and diagnostic evaluation for AD. During a 3-year follow-up, 89 of 823 participants developed AD. In a proportional hazards model, both baseline level of frailty and annual rate of change in frailty were associated with an increased risk of incident AD. Each additional one tenth of a unit increase on the frailty scale at baseline was associated with >9% increased risk of AD (hazard ratio: 2.44; 95% confidence interval (CI): 1.49, 3.37); each one tenth of a unit increase in annual rate of change in frailty was associated with a 12% increased risk of AD (hazard ratio: 3.30; 95% CI: 1.52, 7.13). These results were unchanged in analyses controlling for vascular risk factors and vascular diseases. Results were similar with a categorical measure of frailty instead of a continuous measure. Further, linear mixed-effects models showed that the level of and rate of change in frailty were also associated with the rate of cognitive decline. Increasing frailty is associated with incident AD and the rate of cognitive decline in older persons. These findings suggest that frailty and AD may share similar etiologies." ]
Natural products for the modulation of intervertebral disc degeneration	Intervertebral discs (IVDs) play a crucial role in maintaining normal vertebral anatomy as well as mobile function. Intervertebral disc degeneration (IDD) is a common clinical symptom and is an important cause of low back pain (LBP). IDD is initially considered to be associated with aging and abnormal mechanical loads. However, over recent years, researchers have discovered that IDD is caused by a variety of mechanisms, including persistent inflammation, functional cell loss, accelerated extracellular matrix decomposition, the imbalance of functional components, and genetic metabolic disorders. Of these, inflammation is thought to interact with other mechanisms and is closely associated with the production of pain. Considering the key role of inflammation in IDD, the modulation of inflammation provides us with new options for mitigating the progression of degeneration and may even cause reversal. Many natural substances possess anti-inflammatory functions. Due to the wide availability of such substances, it is important that we screen and identify natural agents that are capable of regulating IVD inflammation. In fact, many studies have demonstrated the potential clinical application of natural substances for the regulation of inflammation in IDD; some of these have been proven to have excellent biosafety. In this review, we summarize the mechanisms and interactions that are responsible for inflammation in IDD and review the application of natural products for the modulation of degenerative disc inflammation.	[ "Intervertebral disc degeneration (IDD) is a globally occurring disease that represents a significant cause of socioeconomic problems. Currently, the main method for treating IDD is surgery, including discectomy and vertebral fusion. Several in vitro experiments demonstrated that platelet-rich plasma (PRP) could stimulate cell proliferation and extracellular matrix regeneration. Additionally, in vivo experiments have proven that PRP injection could restore intervertebral disc height. Clinical studies demonstrated that PRP injection could significantly relieve patient pain. However, further studies are still required to clarify the roles of PRP in IDD prevention and treatment. This review is aimed at summarizing and critically analyzing the current evidence regarding IDD treatment with PRP.", "Degeneration of the intervertebral discs (IVDs) is a major contributor to back, neck and radicular pain. IVD degeneration is characterized by increases in levels of the proinflammatory cytokines TNF, IL-1α, IL-1β, IL-6 and IL-17 secreted by the IVD cells; these cytokines promote extracellular matrix degradation, chemokine production and changes in IVD cell phenotype. The resulting imbalance in catabolic and anabolic responses leads to the degeneration of IVD tissues, as well as disc herniation and radicular pain. The release of chemokines from degenerating discs promotes the infiltration and activation of immune cells, further amplifying the inflammatory cascade. Leukocyte migration into the IVD is accompanied by the appearance of microvasculature tissue and nerve fibres. Furthermore, neurogenic factors, generated by both disc and immune cells, induce expression of pain-associated cation channels in the dorsal root ganglion. Depolarization of these ion channels is likely to promote discogenic and radicular pain, and reinforce the cytokine-mediated degenerative cascade. Taken together, an enhanced understanding of the contribution of cytokines and immune cells to these catabolic, angiogenic and nociceptive processes could provide new targets for the treatment of symptomatic disc disease. In this Review, the role of key inflammatory cytokines during each of the individual phases of degenerative disc disease, as well as the outcomes of major clinical studies aimed at blocking cytokine function, are discussed.", "Inflammation has been demonstrated to be the key factor for intervertebral disc degeneration (IVD), which remains a major public health problem. Isofraxidin is a coumarin compound that possesses strong anti-inflammatory activity. However, the role of isofraxidin in IVD remains unclear. The aim of this study was to evaluate the effects of isofraxidin on inflammatory response in human nucleus pulposus cells (NPCs) exposed to interleukin-1β (IL-1β). The results proved that isofraxidin attenuated the IL-1β-induced significant increases in inflammatory mediators and cytokines including nitric oxide (NO), inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), prostaglandin E2 (PGE2), tumor necrosis factor alpha (TNF-α), and IL-6. Besides, isofraxidin also inhibited the induction effect of IL-1β on matrix metalloproteinases (MMP)-3 and MMP-13. Moreover, the NF-κB activation caused by IL-1β was significantly inhibited by isofraxidin treatment. These findings suggested that isofraxidin alleviates IL-1β-induced inflammation in NPCs. Our work provided an idea that isofraxidin might act as a novel preventive role in IVD.", "Intervertebral disc (IVD) degeneration is characterised by catabolic and inflammatory processes that contribute largely to tissue degradation and chronic back pain. The disc cells are responsible for the pathological production of pro-inflammatory cytokines and catabolic enzymes leading to degeneration. However, this phenotypical change is poorly understood. Growing evidence in animal and human studies implicates Toll-like receptors (TLR) and their activation through danger-associated alarmins, found increasingly in degenerating IVDs. TLR signalling results in the release of pro-inflammatory cytokines and proteolytic enzymes that can directly cause IVD degeneration and back pain. This review aims to summarise the current literature on TLR activation in IVD degeneration and discuss potential treatment modalities to alleviate the inflammatory phenotype of disc cells in order to arrest IVD degeneration and back pain.", "Platelet-rich plasma (PRP) is becoming a promising strategy to treat early intervertebral disc degeneration (IDD) in clinics. Pure PRP without leukocytes (P-PRP) may decrease the catabolic and inflammatory changes in the early degenerated intervertebral discs. The aim of this study was to investigate the effects of P-PRP on nucleus pulposus-derived stem cells (NPSCs) isolated from early degenerated intervertebral discs in vitro. NPSCs isolated from early degenerated discs of rabbits were treated with P-PRP or leukocyte-platelet-rich PRP (L-PRP) in vitro, followed by measuring cell proliferation, stem cell marker expression, inflammatory gene expression, and anabolic and catabolic protein expression by immunostaining, quantitative real-time polymerase chain reaction, Western blot, and enzyme-linked immunosorbent assay. Cell proliferation was induced by P-PRP in a dose-dependent manner with maximum proliferation at 10% P-PRP dose. P-PRP induced differentiation of NPSCs into active nucleus pulposus cells. P-PRP mainly increased the expression of anabolic genes and relative proteins, aggrecan (AGC), collagen types II (Col II), while L-PRP predominantly increased the expression of catabolic and inflammatory genes, matrix metalloproteinase-1 (MMP-1), MMP-13, interleukin-1 beta (IL-1β), IL-6, tumor necrosis factor alpha (TNF-α), and protein production of IL-1β and TNF-α. Leukocytes in PRP activate inflammatory and catabolic effects on NPSCs from early degenerated intervertebral discs. Hence, P-PRP may be a more suitable therapeutic strategy for early IDD.", "Intervertebral disc degeneration (IDD) is widely considered one of the main causes of low back pain, which is a chronic progressive disease closely related to inflammation and degeneration of nucleus pulposus (NP) cells. Baicalein is a natural bioactive compound with anti-inflammatory effects in different diseases, including inhibition of the inflammatory response in chondrocytes, whose morphology and avascular supply are similar to those of NP cells. Therefore, we hypothesized that baicalein may have a therapeutic effect on IDD by suppressing the inflammatory response. In vitro, NP cells were pretreated with baicalein for 2 h and then incubated with IL-1β for 24 h. We found that baicalein not only inhibited the overexpression of inflammatory cytokine production, including NO, PGE2, TNF-α, and IL-6, but also suppressed the expression of COX-2 and iNOS. The IL-1β-induced overexpression of MMP13 and ADAMTS5 and degradation of aggrecan and type II collagen were reversed by baicalein in a dose-dependent manner. Mechanistically, we found that baicalein suppressed the IL-1β-induced activation of the NF-κB and MAPK pathways. Moreover, an in vivo study demonstrated that baicalein treatment could ameliorate IDD in a puncture-induced rat model. Thus, baicalein has great value as a potential therapeutic agent for IDD.", "Intervertebral disc degeneration is the main cause of low back pain. However, its pathomechanism has not been fully clarified yet. Previous studies have indicated that inflammation may lead to apoptosis of nucleus pulposus cells and break the balance between anabolism and catabolism of the nucleus pulposus extracellular matrix. The purpose of this study is to explore the mitigative effect of oxymatrine on extracellular matrix degradation and apoptosis of nucleus pulposus cells after interleukin-1 beta-induced inflammation, and its possible signaling pathway. We examined the gene and protein levels of collagen II, aggrecan, and MMPs (MMP2/3/9/13) and interleukin 6 in nucleus pulposus cells. The results demonstrated that oxymatrine could reduce extracellular matrix degradation and apoptosis of nucleus pulposus cells; interleukin-1 beta prompted the expression of MMPs and interleukin 6 through TLR4/NF-κB axis, while oxymatrine reduced the expression of MMPs and TNF-α induced by interleukin-1 beta. Moreover, TAK 242, as a small molecule inhibitor of TLR4 signaling, was used to detect the effect of oxymatrine on the TLR4/NF-κB signaling. The final experimental results show that oxymatrine could reduce the inflammatory response of nucleus pulposus cells and degradation of nucleus pulposus tissue. Oxymatrine may be a potential medicine to reduce disc inflammation and relieve intervertebral disc degeneration by inhibiting the TLR4/NF-κB signal pathway. Currently, drug therapy is a potential treatment for patients with intervertebral disc degeneration. In the present research, oxymatrine intervenes in intervertebral disc degeneration effectively via regulating inflammation in intervertebral disc degeneration rats. Our research highlights the therapeutic potential of oxymatrine in the treatment of intervertebral disc degeneration." ]

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