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Single-nucleus RNA-seq on undifferentiated human KD3 myoblasts and differentiated myotubes and mononucleated cells.

ABSTRACT: We report the application of single-nucleus-based sequencing technology for high-throughput profiling of transcriptome in immortazalized human myoblast KD3. By obtaining over sixty billion bases of sequence from mRNA, we generated comprehensive transcriptome profiles from KD3 undifferentiated myoblast and differentiated multi-nucleated myotube and mono-nucleated cells. We find that the data from single-nucleus RNA-seq is consistent with the transcriptome from single-cell RNA-seq. The pri-mRNA expression characterized by single-nucleus RNA-seq can reflect the actual miRNA level in the whole cell.

ORGANISM(S): Homo sapiens

PROVIDER: GSE79920 | GEO | 2016/06/30

SECONDARY ACCESSION(S): PRJNA317461

REPOSITORIES: GEO

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Single-nucleus RNA-seq on undifferentiated human KD3 myoblasts and differentiated myotubes and mononucleated cells.

Project description:We report the application of single-nucleus-based sequencing technology for high-throughput profiling of transcriptome in immortazalized human myoblast KD3. By obtaining over sixty billion bases of sequence from mRNA, we generated comprehensive transcriptome profiles from KD3 undifferentiated myoblast and differentiated multi-nucleated myotube and mono-nucleated cells. We find that the data from single-nucleus RNA-seq is consistent with the transcriptome from single-cell RNA-seq. The pri-mRNA expression characterized by single-nucleus RNA-seq can reflect the actual miRNA level in the whole cell. Examination of transcriptome in 1 cell type in 3 differential stages.

2016-06-30 | E-GEOD-79920 | biostudies-arrayexpress

Expression profiling FSHD vs. control myoblasts and myotubes

Project description:The gene expression pathways leading to muscle pathology in facioscapulohumeral dystrophy (FSHD) remain to be elucidated. This muscular dystrophy is caused by a contraction of an array of tandem 3.3-kb repeats (D4Z4) at 4q35.2. We compared expression of control and FSHD myoblasts and myotubes (three preparations each) on exon microarrays (Affymetrix Human Exon 1.0 ST) and validated FSHD-specific differences for representative genes by qRT-PCR on additional myoblast cell strains. The FSHD and control myoblasts used for these experiments were shown to grow and differentiate into myotubes equally efficiently as control myoblasts. There were no significant FSHD-control differences in RNA levels for MYOD1 and MYOG at the myoblast and myotube stages and for MYF5 and MYF6 at the myoblast stage. In contrast, 295 other genes were dysregulated at least 2-fold in FSHD vs. control myoblasts (p <0.01, adjusted for multiple comparisons). Remarkably, only 10% of the FSHD-associated gene dysregulation at the myoblast stage was downregulation. At the myotube stage, about ten times as many genes exhibited FSHD-associated downregulated as at the myoblast stage and twice as many genes displayed FSHD-associated upregulation. The FSHD-related changes in RNA levels appear to be due to posttranscriptional as well as transcriptional alterations. Among the prominently dysregulated pathways were signaling and oxidative stress pathways. By comparing expression profiles of control myoblasts and myotubes to each other and to 19 non-muscle cell types profiled identically, our study also revealed many new myogenesis associations for genes not previously annotated as muscle-specific. Keywords: Disease state analysis and time course for differentiation

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Transcriptomic profile of hiPSC-derived myoblasts and myotubes coated on distinct substrates

Project description:Driving efficient and pure skeletal muscle cell differentiation from pluripotent stem cells has been challenging. Here, we report an optimized protocol that generates skeletal muscle progenitor cells with high efficiency and purity, in a short period of time. Human induced pluripotent stem cells (hiPSC) and mouse embryonic stem cells (mESC) were specified into the mesodermal myogenic fate using distinct and species-specific protocols. We used a specific Maturation Medium to promote terminal differentiation of both human and mouse myoblast populations, and generated myotubes associated with a large pool of cell cycle-arrested PAX7+ cells. We further show that myotube maturation is modulated by the dish coating properties, cell density and percentage of myogenic progenitor cells. Given the high efficiency in the generation of myogenic progenitors and differentiated myofibers, this protocol provides an attractive strategy for tissue engineering, modeling of muscle dystrophies and evaluation of new therapeutic approaches in vitro.

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Expression data from untreated vs. BMP4-treated cultured wild-type and MuSK null muscle cells (myoblasts and myotubes)

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Project description:We sought to determine the effects of over-expression of Gli1 on gene expression in C2C12 myotube cultures. C2C12 myoblasts were induced to differentiate for 4 days. At that time, when >80% of nuclei were incorporated into multi-nucleated syncitial myotubes, we infected the cultures with recombinant adenovirus expressing GFP alone or GFP and a full length human Gli1. Media was changed 12 hours later. Cultures were lysed 60 hours after the initial infection. Gli1 over-expression induces de-differentiation of myotubes and proliferation of myoblasts.

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Expression data from C2C12 myotubes infected with adenovirus expressing Gli1

2013-12-01 | E-GEOD-37702 | biostudies-arrayexpress

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2017-04-07 | GSE84158 | GEO

IGR_MYOGENIC_STUDY_PD

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