Project description:Skeletal muscle accounts for the largest proportion of human body mass, on average, and is a key tissue in complex diseases and mobility. It is composed of several different cell and muscle fiber types. Here, we optimize single-nucleus ATAC-seq (snATAC-seq) to map skeletal muscle cell-specific chromatin accessibility landscapes in frozen human and rat samples, and single-nucleus RNA-seq (snRNA-seq) to map cell-specific transcriptomes in human. We additionally perform multi-omics profiling (gene expression and chromatin accessibility) on human and rat muscle samples.
Project description:Skeletal muscle is a complex heterogeneous tissue comprised of diverse muscle fiber and non-fiber cell types that, in addition to movement, influences other systems such as immunity, metabolism and cognition. We investigated gene expression patterns of resident human skeletal muscle cells using both single-cell RNA-seq and RNA-seq of single muscle fiber dissections from vastus lateralis. We generated transcriptome profiles of the major multinucleated human skeletal muscle fiber-types as well as 11 human skeletal muscle mononuclear cell types, including immune, endothelial, pericyte and satellite cells. We delineated two fibro-adipogenic progenitor cell subtypes that may contribute to heterotopic ossification and muscular dystrophy fibrosis under pathological conditions. An important application of cell type signatures is for computational deconvolution of cell type specific gene expression changes using data from bulk transcriptome experiments. Analysis of transcriptome data from a 12 week resistance exercise training study using these human skeletal muscle cell-type signatures revealed significant changes in specific mononuclear cell-type proportions related to age, sex, acute exercise and training. This characterization of human skeletal muscle cell types will resolve cell-type specific changes in large-scale physical activity muscle transcriptome studies and can further the understanding of the diverse effects of exercise and the pathophysiology of muscle disease.
Project description:Skeletal muscle repair and maintenance are directly and indirectly supported by stromal populations such as vascular cells and fibro-adiopgenic progenitors (FAPs), a subset of which express Twist2 and possess direct myogenic potential in rodents. However, less is understood of the complexity and heterogeneity of human skeletal muscle stromal cells. To this end, we performed single-cell RNA-sequencing on ~2000 cells isolated from the human semitendinosus muscle of young individuals. This demonstrated the presence of two poorly described cell populations. First, we detected a vascular-related cell type that expressed pericyte and pan-endothelial genes that we localized to large blood vessels within skeletal muscle cross-sections and termed endothelial-like pericytes (ELPCs). RNA velocity analysis demonstrated that ELPCs may represent a "transition state" between endothelial cells and pericytes. Analysis of published single-cell RNA-seq datasets revealed evidence for ELPCs in trunk and heart musculature, which showed transcriptional similarity. Additionally, we identified a subset of FAPs expressing TWIST2 mRNA and protein. Human TWIST2-expressing cells were anatomically and transcriptionally comparable to mouse Twist2 cells as they were restricted to the myofiber interstitium, expressed fibrogenic genes but lacked satellite cell markers and colocalized with the FAPs marker PDGFRα in human muscle cross-sections. Taken together, these results highlight the complexity of stromal cells residing in human skeletal muscle and support the utility of single-cell RNA-sequencing for discovery and characterization of poorly described cell populations.
2022-11-01 | GSE216544 | GEO
Project description:Single cell RNA-seq analysis of skeletal muscle cells from Prrx1-ERT tdTomato mice
Project description:Skeletal muscle accounts for the largest proportion of human body mass, on average, and is a key tissue in complex diseases and mobility. It is composed of several different cell and muscle fiber types. Here, we optimize single-nucleus ATAC-seq (snATAC-seq) to map skeletal muscle cell-specific chromatin accessibility landscapes in frozen human and rat samples, and single-nucleus RNA-seq (snRNA-seq) to map cell-specific transcriptomes in human. We additionally perform multi-omics profiling (gene expression and chromatin accessibility) on human and rat muscle samples. We capture type I and type II muscle fiber signatures, which are generally missed by existing single-cell RNA-seq methods. We perform cross-modality and cross-species integrative analyses on 33,862 nuclei and identify seven cell types ranging in abundance from 59.6% to 1.0% of all nuclei. We introduce a regression-based approach to infer cell types by comparing transcription start site-distal ATAC-seq peaks to reference enhancer maps and show consistency with RNA-based marker gene cell type assignments. We find heterogeneity in enrichment of genetic variants linked to complex phenotypes from the UK Biobank and diabetes genome wide association studies in cell-specific ATAC-seq peaks, with the most striking enrichment patterns in muscle mesenchymal stem cells (~3.5% of nuclei). Finally, we overlay these chromatin accessibility maps on GWAS data to nominate causal cell types, SNPs, transcription factor motifs, and target genes for type 2 diabetes signals. These chromatin accessibility profiles for human and rat skeletal muscle cell types are a useful resource for nominating causal GWAS SNPs and cell types.
Project description:Identfification of MEF2A target genes using ChIP-exo in skeletla muscle and primary cardiomyocytes. Identfification of MEF2A target genes using ChIP-exo and RNA-seq in skeletal muscle and primary cardiomyocytes. MEF2 plays a profound role in the regulation of transcription in cardiac and skeletal muscle lineages. To define the overlapping and unique MEF2A genomic targets, we utilized ChIP-exo analysis of cardiomyocytes and skeletal myoblasts. Of the 2783 and 1648 MEF2A binding peaks in skeletal myoblasts and cardiomyocytes, respectively, 294 common binding sites were identified. Genomic targets were compared to differentially expressed genes in RNA-seq analysis of MEF2A depleted myogenic cells. MEF2A target genes were identified in 48 hr DM C2C12 myoblasts cells and primary cardiomyocytes using ChIP-exo. Binding profiles on MEF2A in each cell type were compared. Cross sectional-analysis between ChIP-exo identified targets and RNA-seq analysis of MEF2A deplted myoblasts was also done.