Clonal isolation of muscle-derived cells capable of enhancing muscle regeneration and bone healing

Several recent studies suggest the isolation of stem cells in skeletal musc le, but the functional properties of these muscle-derived stem cells is sti ll unclear. In the present study, we report the purification of muscle-deri ved stem cells from the mdx mouse, an animal model for Duchenne muscular dy strophy. We show that enrichment of desmin(+) cells using the preplate tech nique from mouse primary muscle cell culture also enriches a cell populatio n expressing CD34 and Bcl-2. The CD34(+) cells and Bcl-2(+) cells were foun d to reside within the basal lamina, where satellite cells are normally fou nd. Clonal isolation and characterization from this CD34(+)Bcl-2(+) enriche d population yielded a putative muscle-derived stem cell, mc13, that is cap able of differentiating into both myogenic and osteogenic lineage in vitro and in vivo. The mc13 cells are c-kit and CD45 negative and express: desmin , c-met and MNF, three markers expressed in early myogenic progenitors; Flk -1, a mouse homologue of KDR recently identified in humans as a key marker in hematopoietic cells with stem cell-like characteristics; and Sca-1, a ma rker for both skeletal muscle and hematopoietic stem cells. Intramuscular, and more importantly, intravenous injection of mc13 cells result in muscle regeneration and partial restoration of dystrophin in mdx mice. Transplanta tion of mc13 cells engineered to secrete osteogenic protein differentiate i n osteogenic lineage and accelerate healing of a skull defect in SCID mice. Taken together, these results suggest the isolation of a population of mus cle-derived stem cells capable of improving both muscle regeneration and bo ne healing.