Journal
STEM CELL REPORTS
Volume 3, Issue 3, Pages 516-529Publisher
CELL PRESS
DOI: 10.1016/j.stemcr.2014.07.001
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Funding
- Muscular Dystrophy Association [218371]
- Natural Sciences and Engineering Research Council [RGPIN 293170-11]
- Canadian Institutes of Health Research [MOP-89910]
- Queen Elizabeth II Graduate Scholarship in Science and Technology
- Government of Ontario Ministry of Economic Development and Innovation for the Ontario Research Fund
- Fonds de la Recherche en Santedu Quebec
- Canada Research Chair in Integrative Stem Cell Biology
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Cell therapies treating pathological muscle atrophy or damage requires an adequate quantity of muscle progenitor cells (MPCs) not currently attainable from adult donors. Here, we generate cultures of approximately 90% skeletal myogenic cells by treating human embryonic stem cells (ESCs) with the GSK3 inhibitor CHIR99021 followed by FGF2 and N2 supplements. Gene expression analysis identified progressive expression of mesoderm, somite, dermomyotome, and myotome markers, following patterns of embryonic myogenesis. CHIR99021 enhanced transcript levels of the pan-mesoderm gene T and paraxial-mesoderm genes MSGN1 and TBX6; immunofluorescence confirmed that 91% +/- 6% of cells expressed T immediately following treatment. By 7 weeks, 47% +/- 3% of cells were MYH+ve myocytes/myotubes surrounded by a 43% +/- 4% population of PAX7(+ve) MPCs, indicating 90% of cells had achieved myogenic identity without any cell sorting. Treatment of mouse ESCs with these factors resulted in similar enhancements of myogenesis. These studies establish a foundation for serum-free and chemically defined monolayer skeletal myogenesis of ESCs.
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