RESUMEN
Regulatory T cells (Tregs) are key immune regulators that have shown promise in enhancing cardiac repair post-MI, although the mechanisms remain elusive. Here, we show that rapidly increasing Treg number in the circulation post-MI via systemic administration of exogenous Tregs improves cardiac function in male mice, by limiting cardiomyocyte death and reducing fibrosis. Mechanistically, exogenous Tregs quickly home to the infarcted heart and adopt an injury-specific transcriptome that mediates repair by modulating monocytes/macrophages. Specially, Tregs lead to a reduction in pro-inflammatory Ly6CHi CCR2+ monocytes/macrophages accompanied by a rapid shift of macrophages towards a pro-repair phenotype. Additionally, exogenous Treg-derived factors, including nidogen-1 and IL-10, along with a decrease in cardiac CD8+ T cell number, mediate the reduction of the pro-inflammatory monocyte/macrophage subset in the heart. Supporting the pivotal role of IL-10, exogenous Tregs knocked out for IL-10 lose their pro-repair capabilities. Together, this study highlights the beneficial use of a Treg-based therapeutic approach for cardiac repair with important mechanistic insights that could facilitate the development of novel immunotherapies for MI.
Asunto(s)
Interleucina-10 , Macrófagos , Ratones Endogámicos C57BL , Infarto del Miocardio , Linfocitos T Reguladores , Animales , Infarto del Miocardio/inmunología , Infarto del Miocardio/genética , Infarto del Miocardio/patología , Linfocitos T Reguladores/inmunología , Macrófagos/inmunología , Macrófagos/metabolismo , Masculino , Ratones , Interleucina-10/metabolismo , Interleucina-10/genética , Fenotipo , Miocardio/patología , Miocardio/inmunología , Miocardio/metabolismo , Monocitos/inmunología , Monocitos/metabolismo , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/inmunología , Fibrosis , Linfocitos T CD8-positivos/inmunología , Modelos Animales de Enfermedad , Ratones NoqueadosRESUMEN
Interleukin-(IL) 22 production by intestinal group 3 innate lymphoid cells (ILC3) is critical to maintain gut homeostasis. However, IL-22 needs to be tightly controlled; reduced IL-22 expression is associated with intestinal epithelial barrier defect while its overexpression promotes tumor development. Here, using a single-cell ribonucleic acid sequencing approach, we identified a core set of genes associated with increased IL-22 production by ILC3. Among these genes, programmed cell death 1 (PD-1), extensively studied in the context of cancer and chronic infection, was constitutively expressed on a subset of ILC3. These cells, found in the crypt of the small intestine and colon, displayed superior capacity to produce IL-22. PD-1 expression on ILC3 was dependent on the microbiota and was induced during inflammation in response to IL-23 but, conversely, was reduced in the presence of Notch ligand. PD-1+ ILC3 exhibited distinct metabolic activity with increased glycolytic, lipid, and polyamine synthesis associated with augmented proliferation compared with their PD-1- counterparts. Further, PD-1+ ILC3 showed increased expression of mitochondrial antioxidant proteins which enable the cells to maintain their levels of reactive oxygen species. Loss of PD-1 signaling in ILC3 led to reduced IL-22 production in a cell-intrinsic manner. During inflammation, PD-1 expression was increased on natural cytotoxicity receptor (NCR)- ILC3 while deficiency in PD-1 expression resulted in increased susceptibility to experimental colitis and failure to maintain gut barrier integrity. Collectively, our findings uncover a new function of the PD-1 and highlight the role of PD-1 signaling in the maintenance of gut homeostasis mediated by ILC3 in mice.
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Homeostasis , Inmunidad Innata , Interleucina-22 , Interleucinas , Linfocitos , Ratones Noqueados , Receptor de Muerte Celular Programada 1 , Animales , Ratones , Receptor de Muerte Celular Programada 1/metabolismo , Receptor de Muerte Celular Programada 1/genética , Linfocitos/inmunología , Linfocitos/metabolismo , Interleucinas/metabolismo , Mucosa Intestinal/inmunología , Mucosa Intestinal/metabolismo , Transducción de Señal , Colitis/inmunología , Intestinos/inmunología , Ratones Endogámicos C57BL , Humanos , Modelos Animales de EnfermedadRESUMEN
Single-cell RNA sequencing (scRNA-Seq) has emerged as a powerful tool for understanding cellular heterogeneity and function. However the choice of sample multiplexing reagents can impact data quality and experimental outcomes. In this study, we compared various multiplexing reagents, including MULTI-Seq, Hashtag antibody, and CellPlex, across diverse sample types such as human peripheral blood mononuclear cells (PBMCs), mouse embryonic brain and patient-derived xenografts (PDXs). We found that all multiplexing reagents worked well in cell types robust to ex vivo manipulation but suffered from signal-to-noise issues in more delicate sample types. We compared multiple demultiplexing algorithms which differed in performance depending on data quality. We find that minor improvements to laboratory workflows such as titration and rapid processing are critical to optimal performance. We also compared the performance of fixed scRNA-Seq kits and highlight the advantages of the Parse Biosciences kit for fragile samples. Highly multiplexed scRNA-Seq experiments require more sequencing resources, therefore we evaluated CRISPR-based destruction of non-informative genes to enhance sequencing value. Our comprehensive analysis provides insights into the selection of appropriate sample multiplexing reagents and protocols for scRNA-Seq experiments, facilitating more accurate and cost-effective studies.
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Leucocitos Mononucleares , Análisis de la Célula Individual , Humanos , Animales , Ratones , RNA-Seq , Análisis de Secuencia de ARN/métodos , Análisis de la Célula Individual/métodos , Algoritmos , Perfilación de la Expresión Génica/métodosRESUMEN
Vγ9Vδ2 T cells are the largest population of γδ T cells in adults and can play important roles in providing effective immunity against cancer and infection. Many studies have suggested that peripheral Vγ9Vδ2 T cells are derived from the fetal liver and thymus and that the postnatal thymus plays little role in the development of these cells. More recent evidence suggested that these cells may also develop postnatally in the thymus. Here, we used high-dimensional flow cytometry, transcriptomic analysis, functional assays, and precursor-product experiments to define the development pathway of Vγ9Vδ2 T cells in the postnatal thymus. We identify three distinct stages of development for Vγ9Vδ2 T cells in the postnatal thymus that are defined by the progressive acquisition of functional potential and major changes in the expression of transcription factors, chemokines, and other surface markers. Furthermore, our analysis of donor-matched thymus and blood revealed that the molecular requirements for the development of functional Vγ9Vδ2 T cells are delivered predominantly by the postnatal thymus and not in the periphery. Tbet and Eomes, which are required for IFN-γ and TNFα expression, are up-regulated as Vγ9Vδ2 T cells mature in the thymus, and mature thymic Vγ9Vδ2 T cells rapidly express high levels of these cytokines after stimulation. Similarly, the postnatal thymus programs Vγ9Vδ2 T cells to express the cytolytic molecules, perforin, granzyme A, and granzyme K. This study provides a greater understanding of how Vγ9Vδ2 T cells develop in humans and may lead to opportunities to manipulate these cells to treat human diseases.
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Receptores de Antígenos de Linfocitos T gamma-delta , Subgrupos de Linfocitos T , Adulto , Humanos , Timo , Perfilación de la Expresión GénicaRESUMEN
Group heteroscedasticity is commonly observed in pseudo-bulk single-cell RNA-seq datasets and its presence can hamper the detection of differentially expressed genes. Since most bulk RNA-seq methods assume equal group variances, we introduce two new approaches that account for heteroscedastic groups, namely voomByGroup and voomWithQualityWeights using a blocked design (voomQWB). Compared to current gold-standard methods that do not account for group heteroscedasticity, we show results from simulations and various experiments that demonstrate the superior performance of voomByGroup and voomQWB in terms of error control and power when group variances in pseudo-bulk single-cell RNA-seq data are unequal.
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Perfilación de la Expresión Génica , Programas Informáticos , Perfilación de la Expresión Génica/métodos , Análisis de Secuencia de ARN/métodos , Análisis de Expresión Génica de una Sola Célula , Análisis de la Célula Individual/métodosRESUMEN
Female mouse embryonic stem cells (mESCs) present differently from male mESCs in several fundamental ways; however, complications with their in vitro culture have resulted in an under-representation of female mESCs in the literature. Recent studies show that the second X chromosome in female, and more specifically the transcriptional activity from both of these chromosomes due to absent X chromosome inactivation, sets female and male mESCs apart. To avoid this undesirable state, female mESCs in culture preferentially adopt an XO karyotype, with this adaption leading to loss of their unique properties in favour of a state that is near indistinguishable from male mESCs. If female pluripotency is to be studied effectively in this system, it is crucial that high-quality cultures of XX mESCs are available. Here, we report a method for better maintaining XX female mESCs in culture that also stabilises the male karyotype and makes study of female-specific pluripotency more feasible.
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Células Madre Embrionarias de Ratones , Inactivación del Cromosoma X , Masculino , Animales , Femenino , Ratones , Diferenciación Celular/fisiología , Inactivación del Cromosoma X/genética , CariotipoRESUMEN
OBJECTIVES: This study aimed to investigate the number of days following hamstring strain injury (HSI) taken to introduce high-intensity eccentric loading (HIEL) into rehabilitation based on exercise-specific progression criteria, and whether pain resolution during isometric knee flexion strength testing occurred before or after this milestone. DESIGN: Cohort study. METHODS: We included 42 men (mean⯱â¯sd; ageâ¯=â¯26⯱â¯5â¯years; heightâ¯=â¯181⯱â¯8â¯cm; massâ¯=â¯86⯱â¯12â¯kg) with HSIs, who performed fully supervised rehabilitation twice per week until they met return to play clearance criteria. Isometric knee flexion strength testing was completed before every rehabilitation session and HIEL was introduced via the Nordic hamstring exercise and unilateral slider once participants could perform a bilateral slider through full eccentric knee flexion range of motion. We reported the median (IQR) number of days following HSI taken to introduce HIEL, along with participant's pain rating during isometric knee flexion strength testing before that rehabilitation session. We also reported the median (IQR) number of days following HSI taken for participants to achieve pain resolution during isometric knee flexion. RESULTS: HIEL was introduced 5 (2-8) days following HSI, despite 35/42 participants reporting pain during isometric knee flexion strength testing immediately prior to that rehabilitation session, which was rated as 3.5 (3-5) on a 0-10 numeric rating scale. Pain resolution during isometric knee flexion strength testing was achieved 11 (9-13) days following HSI. CONCLUSION: HIEL can be safely introduced into early HSI rehabilitation based on exercise-specific progression criteria, without needing to wait for pain resolution during isometric knee flexion strength testing before doing so.
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Músculos Isquiosurales , Traumatismos de la Pierna , Traumatismos de los Tejidos Blandos , Adulto , Estudios de Cohortes , Músculos Isquiosurales/lesiones , Humanos , Masculino , Fuerza Muscular , Dolor , Adulto JovenRESUMEN
The process of epigenetic silencing, while fundamentally important, is not yet completely understood. Here we report a replenishable female mouse embryonic stem cell (mESC) system, Xmas, that allows rapid assessment of X chromosome inactivation (XCI), the epigenetic silencing mechanism of one of the two X chromosomes that enables dosage compensation in female mammals. Through a targeted genetic screen in differentiating Xmas mESCs, we reveal that the BAF complex is required to create nucleosome-depleted regions at promoters on the inactive X chromosome during the earliest stages of establishment of XCI. Without this action gene silencing fails. Xmas mESCs provide a tractable model for screen-based approaches that enable the discovery of unknown facets of the female-specific process of XCI and epigenetic silencing more broadly.
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ARN Largo no Codificante , Inactivación del Cromosoma X , Animales , Cromatina/genética , Compensación de Dosificación (Genética) , Epigénesis Genética , Femenino , Ratones , ARN Largo no Codificante/genética , Cromosoma X/genética , Inactivación del Cromosoma X/genéticaRESUMEN
BACKGROUND: Single-cell RNA-sequencing (scRNA-seq) technologies and associated analysis methods have rapidly developed in recent years. This includes preprocessing methods, which assign sequencing reads to genes to create count matrices for downstream analysis. While several packaged preprocessing workflows have been developed to provide users with convenient tools for handling this process, how they compare to one another and how they influence downstream analysis have not been well studied. RESULTS: Here, we systematically benchmark the performance of 10 end-to-end preprocessing workflows (Cell Ranger, Optimus, salmon alevin, alevin-fry, kallisto bustools, dropSeqPipe, scPipe, zUMIs, celseq2, and scruff) using datasets yielding different biological complexity levels generated by CEL-Seq2 and 10x Chromium platforms. We compare these workflows in terms of their quantification properties directly and their impact on normalization and clustering by evaluating the performance of different method combinations. While the scRNA-seq preprocessing workflows compared vary in their detection and quantification of genes across datasets, after downstream analysis with performant normalization and clustering methods, almost all combinations produce clustering results that agree well with the known cell type labels that provided the ground truth in our analysis. CONCLUSIONS: In summary, the choice of preprocessing method was found to be less important than other steps in the scRNA-seq analysis process. Our study comprehensively compares common scRNA-seq preprocessing workflows and summarizes their characteristics to guide workflow users.
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Benchmarking/métodos , Análisis de Secuencia de ARN/métodos , Análisis de la Célula Individual/métodos , Flujo de Trabajo , Análisis por Conglomerados , Perfilación de la Expresión Génica/métodos , RNA-Seq , Programas Informáticos , TranscriptomaRESUMEN
A key benefit of long-read nanopore sequencing technology is the ability to detect modified DNA bases, such as 5-methylcytosine. The lack of R/Bioconductor tools for the effective visualization of nanopore methylation profiles between samples from different experimental groups led us to develop the NanoMethViz R package. Our software can handle methylation output generated from a range of different methylation callers and manages large datasets using a compressed data format. To fully explore the methylation patterns in a dataset, NanoMethViz allows plotting of data at various resolutions. At the sample-level, we use dimensionality reduction to look at the relationships between methylation profiles in an unsupervised way. We visualize methylation profiles of classes of features such as genes or CpG islands by scaling them to relative positions and aggregating their profiles. At the finest resolution, we visualize methylation patterns across individual reads along the genome using the spaghetti plot and heatmaps, allowing users to explore particular genes or genomic regions of interest. In summary, our software makes the handling of methylation signal more convenient, expands upon the visualization options for nanopore data and works seamlessly with existing methylation analysis tools available in the Bioconductor project. Our software is available at https://bioconductor.org/packages/NanoMethViz.
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Metilación de ADN/genética , Genómica/métodos , Secuenciación de Nanoporos/métodos , Análisis de Secuencia de ADN/métodos , Programas Informáticos , Animales , Humanos , RatonesRESUMEN
Blood vessel growth and remodelling are essential during embryonic development and disease pathogenesis. The diversity of endothelial cells (ECs) is transcriptionally evident and ECs undergo dynamic changes in gene expression during vessel growth and remodelling. Here, we investigated the role of the histone acetyltransferase HBO1 (KAT7), which is important for activating genes during development and for histone H3 lysine 14 acetylation (H3K14ac). Loss of HBO1 and H3K14ac impaired developmental sprouting angiogenesis and reduced pathological EC overgrowth in the retinal endothelium. Single-cell RNA sequencing of retinal ECs revealed an increased abundance of tip cells in Hbo1-deficient retinas, which led to EC overcrowding in the retinal sprouting front and prevented efficient tip cell migration. We found that H3K14ac was highly abundant in the endothelial genome in both intra- and intergenic regions, suggesting that HBO1 acts as a genome organiser that promotes efficient tip cell behaviour necessary for sprouting angiogenesis. This article has an associated 'The people behind the papers' interview.
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Histona Acetiltransferasas/metabolismo , Neovascularización Patológica/metabolismo , Acetilación , Animales , Movimiento Celular/fisiología , Células Cultivadas , Desarrollo Embrionario/fisiología , Células Endoteliales/metabolismo , Femenino , Histonas/metabolismo , Células Endoteliales de la Vena Umbilical Humana , Humanos , Lisina/metabolismo , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BLRESUMEN
BACKGROUND: Pooling cells from multiple biological samples prior to library preparation within the same single-cell RNA sequencing experiment provides several advantages, including lower library preparation costs and reduced unwanted technological variation, such as batch effects. Computational demultiplexing tools based on natural genetic variation between individuals provide a simple approach to demultiplex samples, which does not require complex additional experimental procedures. However, to our knowledge these tools have not been evaluated in cancer, where somatic variants, which could differ between cells from the same sample, may obscure the signal in natural genetic variation. RESULTS: Here, we performed in silico benchmark evaluations by combining raw sequencing reads from multiple single-cell samples in high-grade serous ovarian cancer, which has a high copy number burden, and lung adenocarcinoma, which has a high tumor mutational burden. Our results confirm that genetic demultiplexing tools can be effectively deployed on cancer tissue using a pooled experimental design, although high proportions of ambient RNA from cell debris reduce performance. CONCLUSIONS: This strategy provides significant cost savings through pooled library preparation. To facilitate similar analyses at the experimental design phase, we provide freely accessible code and a reproducible Snakemake workflow built around the best-performing tools found in our in silico benchmark evaluations, available at https://github.com/lmweber/snp-dmx-cancer.
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Neoplasias , Proyectos de Investigación , Biblioteca de Genes , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Humanos , Neoplasias/genética , ARN , Programas InformáticosRESUMEN
BACKGROUND: DNA methylation dynamics in the brain are associated with normal development and neuropsychiatric disease and differ across functionally distinct brain regions. Previous studies of genome-wide methylation differences among human brain regions focus on limited numbers of individuals and one to two brain regions. RESULTS: Using GTEx samples, we generate a resource of DNA methylation in purified neuronal nuclei from 8 brain regions as well as lung and thyroid tissues from 12 to 23 donors. We identify differentially methylated regions between brain regions among neuronal nuclei in both CpG (181,146) and non-CpG (264,868) contexts, few of which were unique to a single pairwise comparison. This significantly expands the knowledge of differential methylation across the brain by 10-fold. In addition, we present the first differential methylation analysis among neuronal nuclei from basal ganglia tissues and identify unique CpG differentially methylated regions, many associated with ion transport. We also identify 81,130 regions of variably CpG methylated regions, i.e., variable methylation among individuals in the same brain region, which are enriched in regulatory regions and in CpG differentially methylated regions. Many variably methylated regions are unique to a specific brain region, with only 202 common across all brain regions, as well as lung and thyroid. Variably methylated regions identified in the amygdala, anterior cingulate cortex, and hippocampus are enriched for heritability of schizophrenia. CONCLUSIONS: These data suggest that epigenetic variation in these particular human brain regions could be associated with the risk for this neuropsychiatric disorder.
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Encéfalo/metabolismo , Metilación de ADN , Variación Genética , Patrón de Herencia , Carácter Cuantitativo Heredable , Islas de CpG , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Hipocampo/metabolismo , Humanos , Trastornos Mentales/diagnóstico , Trastornos Mentales/etiología , Neuronas , Especificidad de Órganos/genéticaRESUMEN
OBJECTIVE: The primary aim was to compare time from acute hamstring strain injury (HSI) to return-to-play (RTP) clearance following a standardized rehabilitation protocol performed within either pain-free or pain-threshold limits. Secondary aims were to compare isometric knee flexor strength, biceps femoris long head (BFLH) fascicle length, fear of movement, and reinjury occurrence at the 6-month follow-up between pain-free and pain-threshold groups. DESIGN: Randomized controlled trial. METHODS: Forty-three men with acute HSIs were randomly allocated to a pain-free (n = 22) or pain-threshold (n = 21) rehabilitation group. Days from HSI to RTP clearance, isometric knee flexor strength, BFLH fascicle length, fear of movement, and reinjury occurrence at the 6-month follow-up were reported. RESULTS: Median time from HSI to RTP clearance was 15 days (95% confidence interval [CI]: 13, 17) in the pain-free group and 17 days (95% CI: 11, 24) in the pain-threshold group, which was not significantly different (P = .37). Isometric knee flexor strength recovery at 90° of hip and 90° of knee flexion was greater in the pain-threshold group at RTP clearance by 15% (95% CI: 1%, 28%) and by 15% (95% CI: 1%, 29%) at 2-month follow-up, respectively. Improvement in BFLH fascicle length from baseline was 0.91 cm (95% CI: 0.34, 1.48) greater at 2-month follow-up in the pain-threshold group. Two reinjuries occurred in both the pain-free and pain-threshold groups between RTP clearance and the 6-month follow-up. CONCLUSION: Pain-threshold rehabilitation did not accelerate RTP clearance, but resulted in greater recovery of isometric knee flexor strength and better maintenance of BFLH fascicle length, compared to pain-free rehabilitation. J Orthop Sports Phys Ther 2020;50(2):91-103. Epub 28 Jun 2019. doi:10.2519/jospt.2020.8895.
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Traumatismos en Atletas/rehabilitación , Músculos Isquiosurales/lesiones , Umbral del Dolor , Esguinces y Distensiones/fisiopatología , Esguinces y Distensiones/rehabilitación , Adulto , Traumatismos en Atletas/fisiopatología , Traumatismos en Atletas/psicología , Método Doble Ciego , Miedo , Estudios de Seguimiento , Músculos Isquiosurales/anatomía & histología , Músculos Isquiosurales/fisiología , Humanos , Contracción Isométrica/fisiología , Rodilla/fisiología , Masculino , Movimiento/fisiología , Fuerza Muscular/fisiología , Recurrencia , Volver al Deporte , Factores de Riesgo , Esguinces y Distensiones/psicología , Adulto JovenRESUMEN
Correct annotation metadata is critical for reproducible and accurate RNA-seq analysis. When files are shared publicly or among collaborators with incorrect or missing annotation metadata, it becomes difficult or impossible to reproduce bioinformatic analyses from raw data. It also makes it more difficult to locate the transcriptomic features, such as transcripts or genes, in their proper genomic context, which is necessary for overlapping expression data with other datasets. We provide a solution in the form of an R/Bioconductor package tximeta that performs numerous annotation and metadata gathering tasks automatically on behalf of users during the import of transcript quantification files. The correct reference transcriptome is identified via a hashed checksum stored in the quantification output, and key transcript databases are downloaded and cached locally. The computational paradigm of automatically adding annotation metadata based on reference sequence checksums can greatly facilitate genomic workflows, by helping to reduce overhead during bioinformatic analyses, preventing costly bioinformatic mistakes, and promoting computational reproducibility. The tximeta package is available at https://bioconductor.org/packages/tximeta.
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Biología Computacional/métodos , Perfilación de la Expresión Génica , RNA-Seq , Algoritmos , Animales , Drosophila melanogaster , Genómica , Humanos , Ratones , Modelos Estadísticos , Reconocimiento de Normas Patrones Automatizadas , Lenguajes de Programación , Reproducibilidad de los Resultados , Programas Informáticos , TranscriptomaRESUMEN
MOTIVATION: Bioinformatic analysis of single-cell gene expression data is a rapidly evolving field. Hundreds of bespoke methods have been developed in the past few years to deal with various aspects of single-cell analysis and consensus on the most appropriate methods to use under different settings is still emerging. Benchmarking the many methods is therefore of critical importance and since analysis of single-cell data usually involves multi-step pipelines, effective evaluation of pipelines involving different combinations of methods is required. Current benchmarks of single-cell methods are mostly implemented with ad-hoc code that is often difficult to reproduce or extend, and exhaustive manual coding of many combinations is infeasible in most instances. Therefore, new software is needed to manage pipeline benchmarking. RESULTS: The CellBench R software facilitates method comparisons in either a task-centric or combinatorial way to allow pipelines of methods to be evaluated in an effective manner. CellBench automatically runs combinations of methods, provides facilities for measuring running time and delivers output in tabular form which is highly compatible with tidyverse R packages for summary and visualization. Our software has enabled comprehensive benchmarking of single-cell RNA-seq normalization, imputation, clustering, trajectory analysis and data integration methods using various performance metrics obtained from data with available ground truth. CellBench is also amenable to benchmarking other bioinformatics analysis tasks. AVAILABILITY AND IMPLEMENTATION: Available from https://bioconductor.org/packages/CellBench.
Asunto(s)
RNA-Seq , Análisis de la Célula Individual , Biología Computacional , Análisis de Secuencia de ARN , Programas Informáticos , Secuenciación del ExomaRESUMEN
STUDY DESIGN: Randomized controlled trial. BACKGROUND: Conventional guidelines recommend hamstring strain injury (HSI) rehabilitation should only be performed and progressed in complete absence of pain, despite lack of comparison to alternative approaches. OBJECTIVES: The primary aim of this study was to compare the number of days from acute HSI to return to play (RTP) clearance following a standardised rehabilitation protocol performed within either pain-free or pain-threshold limits. The secondary aims were to compare isometric knee flexor strength, biceps femoris long head (BFlh) fascicle length, fear of movement and re-injury during a six-month follow-up between pain-free and pain-threshold groups. METHODS: Forty-three men with acute HSIs were randomly allocated to either a pain-free (n=22) or pain-threshold (n=21) rehabilitation group. Days from HSI to RTP clearance, isometric knee flexor strength, BFlh fascicle length, fear of movement and re-injuries within six-month follow-up were reported. RESULTS: The median time from HSI to RTP clearance was 15 days (95% CI = 13 to 17) in the pain-free group and 17 days (95% CI = 11 to 24) in the pain-threshold group, which was not significantly different (p = 0.37). Recovery of isometric knee flexor strength at 90/90 degrees of hip/knee flexion was greater in the pain-threshold group at RTP clearance by 15% (95% CI = 1 to 28) and by 15% (95% CI = 1 to 29) at two-month follow-up. BFlh fascicles were 0.91cm (95% CI = 0.34 to 1.48) longer at two-month follow-up in the pain-threshold group. Two re-injuries occurred in both the pain-free and pain-threshold group during six-month follow-up. CONCLUSION: Pain-threshold rehabilitation did not accelerate RTP clearance but did result in greater recovery of isometric knee flexor strength and better maintenance of BFlh fascicle length improvements compared to pain-free rehabilitation. J Orthop Sports Phys Ther, Epub 28 Jun 2019. doi:10.2519/jospt.2019.8895.
RESUMEN
Coding variants in epigenetic regulators are emerging as causes of neurological dysfunction and cancer. However, a comprehensive effort to identify disease candidates within the human epigenetic machinery (EM) has not been performed; it is unclear whether features exist that distinguish between variation-intolerant and variation-tolerant EM genes, and between EM genes associated with neurological dysfunction versus cancer. Here, we rigorously define 295 genes with a direct role in epigenetic regulation (writers, erasers, remodelers, readers). Systematic exploration of these genes reveals that although individual enzymatic functions are always mutually exclusive, readers often also exhibit enzymatic activity (dual-function EM genes). We find that the majority of EM genes are very intolerant to loss-of-function variation, even when compared to the dosage sensitive transcription factors, and we identify 102 novel EM disease candidates. We show that this variation intolerance is driven by the protein domains encoding the epigenetic function, suggesting that disease is caused by a perturbed chromatin state. We then describe a large subset of EM genes that are coexpressed within multiple tissues. This subset is almost exclusively populated by extremely variation-intolerant genes and shows enrichment for dual-function EM genes. It is also highly enriched for genes associated with neurological dysfunction, even when accounting for dosage sensitivity, but not for cancer-associated EM genes. Finally, we show that regulatory regions near epigenetic regulators are genetically important for common neurological traits. These findings prioritize novel disease candidate EM genes and suggest that this coexpression plays a functional role in normal neurological homeostasis.