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Assessment of the myogenic stem cell compartment following transplantation of Pax3/Pax7-induced embryonic stem cell-derived progenitors.

ABSTRACT: An effective long-term cell therapy for skeletal muscle regeneration requires donor contribution to both muscle fibers and the muscle stem cell pool. Although satellite cells have these abilities, their therapeutic potential so far has been limited due to their scarcity in adult muscle. Myogenic progenitors obtained from Pax3-engineered mouse embryonic stem (ES) cells have the ability to generate myofibers and to improve the contractility of transplanted muscles in vivo, however, whether these cells contribute to the muscle stem cell pool and are able to self-renew in vivo are still unknown. Here, we addressed this question by investigating the ability of Pax3, which plays a critical role in embryonic muscle formation, and Pax7, which is important for maintenance of the muscle satellite cell pool, to promote the derivation of self-renewing functional myogenic progenitors from ES cells. We show that Pax7, like Pax3, can drive the expansion of an ES-derived myogenic progenitor with significant muscle regenerative potential. We further demonstrate that a fraction of transplanted cells remains mononuclear, and displays key features of skeletal muscle stem cells, including satellite cell localization, response to reinjury, and contribution to muscle regeneration in secondary transplantation assays. The ability to engraft, self-renew, and respond to injury provide foundation for the future therapeutic application of ES-derived myogenic progenitors in muscle disorders.

SUBMITTER: Darabi R

PROVIDER: S-EPMC3325545 | biostudies-literature | 2011 May

REPOSITORIES: biostudies-literature

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Assessment of the myogenic stem cell compartment following transplantation of Pax3/Pax7-induced embryonic stem cell-derived progenitors.

Darabi Radbod R Santos Filipe N C FN Filareto Antonio A Pan Weihong W Koene Ryan R Rudnicki Michael A MA Kyba Michael M Perlingeiro Rita C R RC

Stem cells (Dayton, Ohio) 20110501 5

An effective long-term cell therapy for skeletal muscle regeneration requires donor contribution to both muscle fibers and the muscle stem cell pool. Although satellite cells have these abilities, their therapeutic potential so far has been limited due to their scarcity in adult muscle. Myogenic progenitors obtained from Pax3-engineered mouse embryonic stem (ES) cells have the ability to generate myofibers and to improve the contractility of transplanted muscles in vivo, however, whether these c ...[more]

PMID: 21374762

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Project description:BackgroundFacioscapulohumeral muscular dystrophy (FSHD) is most commonly inherited in an autosomal dominant pattern and caused by the abnormal expression of DUX4 in skeletal muscle. The DUX4 transcription factor has DNA binding domains similar to several paired class homeotic transcription factors, but only myogenic factors PAX3 and PAX7 rescue cell viability when co-expressed with DUX4 in mouse myoblasts. This observation suggests competition for DNA binding sites in satellite cells might limit muscle repair and may be one aspect of DUX4-associated myotoxicity. The competition hypothesis requires that DUX4 and PAX3/7 be expressed in the same cells at some point during development or in adult tissues. We modeled myogenesis using human isogenic iPS and ES cells and examined expression patterns of DUX4, PAX3, and PAX7 to determine if conditions that promote PAX3 and PAX7 expression in cell culture also promote DUX4 expression in the same cells.MethodsIsogenic iPSCs were generated from human fibroblasts of two FSHD-affected individuals with somatic mosaicism. Clones containing the shortened FSHD-causing D4Z4 array or the long non-pathogenic array were isolated from the same individuals. We also examined myogenesis in commercially available hES cell lines derived from FSHD-affected and non-affected embryos. DUX4, PAX3, and PAX7 messenger RNAs (mRNAs) were quantified during a 40-day differentiation protocol, and antibodies were used to identify cell types in different stages of differentiation to determine if DUX4 and PAX3 or PAX7 are present in the same cells.ResultsHuman iPS and ES cells differentiated into skeletal myocytes as evidenced by Titin positive multinucleated fibers appearing toward the end of a 40-day differentiation protocol. PAX3 and PAX7 were expressed at similar times during differentiation, and DUX4 positive nuclei were seen at terminal stages of differentiation in cells containing the short D4Z4 arrays. Nuclei that expressed both DUX4 and PAX3, or DUX4 and PAX7 were not observed after examining immunostained nuclei at five different time points during myogenic differentiation of pluripotent cells.ConclusionsWe conclude that DUX4, PAX3, and PAX7 have distinct expression patterns during myogenic differentiation of stem cells. Our findings are consistent with the hypothesis that muscle damage in FSHD is due to DUX4-mediated toxicity causing destruction of terminally differentiated myofibers. While these studies examine DUX4, PAX3, and PAX7 expression patterns during stem cell myogenesis, they should not be generalized to tissue repair in adult muscle tissue.

Dataset Information

Assessment of the myogenic stem cell compartment following transplantation of Pax3/Pax7-induced embryonic stem cell-derived progenitors.

Publications

Assessment of the myogenic stem cell compartment following transplantation of Pax3/Pax7-induced embryonic stem cell-derived progenitors.

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