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Muscle side population (SP) cells are rare myogenic progenitors distinct from satellite cells, the known tissue-specific stem cells of skeletal muscle. Studies in mice demonstrated that muscle SP cells give rise to satellite cells  in vivo . Given that muscle SP cells are heterogeneous, it has been difficult to prospectively enrich for myogenic progenitors within the SP fraction, particularly from human tissue. Further, conditions that favor the expansion of human muscle SP cells while retaining their myogenic potential have yet to be reported. In this study, human fetal muscle SP and main population (MP) cells were purified based on the expression of melanoma cell adhesion molecule (MCAM), a marker we previously reported to enrich for cells with myogenic potential. To define the relationship between MCAM expression and the degree of myogenic commitment, single cells were analyzed for the expression of myogenic-specific markers. Myogenic factors strongly associated with MCAM expression in single cells, particularly Myf5. Different MCAM+ populations, including SP cells, were expanded and assayed for fusion potential  in vitro  and engraftment potential  in vivo . All MCAM+ subpopulations fused robustly into myotubes  in vitro , whereas the MCAM− subpopulations did not. Further, MCAM+ SP cells exhibited the highest fusion potential  in vitro  and were the only fraction to engraft  in vivo , although at low levels, following propagation. Thus, MCAM can be used to prospectively enrich for myogenic muscle SP cells in human fetal muscle. Moreover, we provide evidence that human MCAM+ SP cells have intrinsic myogenic activity that is retained after propagation.

Human fetal skeletal muscle contains a myogenic side population that expresses the melanoma cell-adhesion molecule