Development of approaches to improve the regenerative potential of muscle stem cells

Stem cell therapy is a promising treatment for diseases such as Duchenne muscular dystrophy (DMD) and ischemic heart failure. However, low survival and differentiation of transplanted cells hinders therapy. In this study, we examined ways to enhance the effectiveness of muscle cells for cardiomyoplasty by increasing antioxidant levels, explored the role of vascular endothelial growth factor (VEGF) in mechanical stimulation pre-treatment and characterized muscle derived stem cells (MDSCs) from normal and dystrophic mice. \ud First we demonstrated that increasing antioxidant levels positively correlated with the early survival of myoblasts after implantation into infarcted hearts, but did not result in long term functional benefits, indicating that early survival does not necessarily correlate with long term regeneration and repair. \ud Next we aimed to determine the effect of VEGF on mechanically stimulated MDSCs transplanted into dystrophic muscle. MDSCs were transduced with vectors carrying the LacZ reporter gene (lacZ-MDSCs), the soluble VEGF receptor Flt1 (sFlt1-MDSCs) or short hairpin RNA targeting VEGF messenger RNA (shRNA_VEGF MDSCs). They were subjected to 24 hours of cyclic strain and injected into the gastrocnemius muscles of dystrophic mdx/SCID mice. After 2 weeks, there was an increase in angiogenesis in muscles transplanted with mechanically stimulated lacZ-MDSCs compared to non-stimulated lacZ-MDSCs and sFlt1-MDSCs. Dystrophin positive myofiber regeneration and in vitro myotube differentiation were significantly lower in the shRNA_VEGF-MDSC group compared to the lacZ-MDSC and sFlt1-MDSC groups. Thus, the beneficial effects of mechanical stimulation on MDSC mediated muscle repair were lost by inhibiting VEGF. \ud Finally, we aimed to compare wild-type (wt) MDSCs with MDSCs obtained from mdx and dystrophin/utrophin double knock out (DKO) mice, which are models of muscular dystrophy. We demonstrated that wt and mdx MDSCs did not have differences in proliferation, differentiation, or VEGF secretion. We compared DKO homozygous MDSCs and DKO heterozygous MDSCs and found that DKO homo MDSCs had decreased proliferation, differentiation, and cell survival capabilities compared to DKO het MDSCs. Finally, we pre-treated DKO MDSCs with mechanical stimulation and increased their proliferation rates. In conclusion, efforts to optimize cell therapy are necessary to improve transplantation outcomes for both ischemic cardiac repair and muscular diseases.\u