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Isolation and in vitro propagation of human skeletal muscle progenitor cells from fetal muscle.

ABSTRACT: Skeletal muscle progenitor cells (SMPCs) are considered one of the most valuable cells for cell-based therapy targeting skeletal muscle. However, an efficient protocol for isolating and maintaining human myogenic progenitors in vitro has not been fully established. In this study, we demonstrate that human myogenic progenitors can be expanded and proliferated from human fetal muscles. Human SMPCs were prepared from fetal hind limb muscles and induced to proliferate as free-floating spheres termed myospheres in the medium containing basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF). Both myogenic progenitors and myoblast populations from human fetal muscles were effectively propagated in myospheres and passaged by a mechanical chopping. After expanding these spheres in culture, we tested whether myogenic progenitor cells can differentiate into multinucleated myotubes. The myospheres were dissociated, plated down on coverslips and cultured in the medium for terminal differentiation. We could confirm that the plated cells formed well-developed, multinucleated myotubes. This culture method using myospheres is an effective protocol to isolate and maintain SMPCs from human fetal skeletal muscles in culture.

SUBMITTER: Hosoyama T

PROVIDER: S-EPMC3620662 | biostudies-literature | 2013 Feb

REPOSITORIES: biostudies-literature

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Isolation and in vitro propagation of human skeletal muscle progenitor cells from fetal muscle.

Hosoyama Tohru T Meyer Michael G MG Krakora Dan D Suzuki Masatoshi M

Cell biology international 20130102 2

Skeletal muscle progenitor cells (SMPCs) are considered one of the most valuable cells for cell-based therapy targeting skeletal muscle. However, an efficient protocol for isolating and maintaining human myogenic progenitors in vitro has not been fully established. In this study, we demonstrate that human myogenic progenitors can be expanded and proliferated from human fetal muscles. Human SMPCs were prepared from fetal hind limb muscles and induced to proliferate as free-floating spheres termed ...[more]

PMID: 23319422

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Project description:Heterotopic ossification (HO) is defined as the formation of ectopic bone in soft tissue outside the skeletal tissue. HO is thought to result from aberrant differentiation of osteogenic progenitors within skeletal muscle. However, the precise origin of HO is still unclear. Skeletal muscle contains two kinds of progenitor cells, myogenic progenitors and mesenchymal progenitors. Myogenic and mesenchymal progenitors in human skeletal muscle can be identified as CD56(+) and PDGFR?(+) cells, respectively. The purpose of this study was to investigate the osteogenic differentiation potential of human skeletal muscle-derived progenitors. Both CD56(+) cells and PDGFR?(+) cells showed comparable osteogenic differentiation potential in vitro. However, in an in vivo ectopic bone formation model, PDGFR?(+) cells formed bone-like tissue and showed successful engraftment, while CD56(+) cells did not form bone-like tissue and did not adapt to an osteogenic environment. Immunohistological analysis of human HO sample revealed that many PDGFR?(+) cells were localized in proximity to ectopic bone formed in skeletal muscle. MicroRNAs (miRNAs) are known to regulate many biological processes including osteogenic differentiation. We investigated the participation of miRNAs in the osteogenic differentiation of PDGFR?(+) cells by using microarray. We identified miRNAs that had not been known to be involved in osteogenesis but showed dramatic changes during osteogenic differentiation of PDGFR?(+) cells. Upregulation of miR-146b-5p and -424 and downregulation of miR-7 during osteogenic differentiation of PDGFR?(+) cells were confirmed by quantitative real-time RT-PCR. Inhibition of upregulated miRNAs, miR-146b-5p and -424, resulted in the suppression of osteocyte maturation, suggesting that these two miRNAs have the positive role in the osteogenesis of PDGFR?(+) cells. Our results suggest that PDGFR?(+) cells may be the major source of HO and that the newly identified miRNAs may regulate osteogenic differentiation process of PDGFR?(+) cells.

Dataset Information

Isolation and in vitro propagation of human skeletal muscle progenitor cells from fetal muscle.

Publications

Isolation and in vitro propagation of human skeletal muscle progenitor cells from fetal muscle.

Similar Datasets

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