Human mesenchymal stem cells ectopically expressing full-length dystrophin can complement Duchenne muscular dystrophy myotubes by cell fusion

Duchenne muscular dystrophy (DMD) is the most prevalent inheritable muscle disease. It is caused by mutations in the ∼2.5-megabase dystrophin (Dys) encoding gene. Therapeutic attempts at DMD have relied on injection of allogeneic Dys-positive myoblasts. The immune rejection of these cells and their limited availability have prompted the search for alternative therapies and sources of myogenic cells. Stem cell-based gene therapy aims to restore tissue function by the transplantation of gene-corrected autologous cells. It depends on (i) the capacity of stem cells to participate in tissue regeneration and (ii) the efficient genetic correction of defective autologous stem cells. We explored the potential of bone marrow-derived human mesenchymal stem cells (hMSCs) genetically modified with the full-length Dys-coding sequence to engage in myogenesis. By tagging hMSCs with enhanced green fluorescent protein (EGFP) or the membrane dye PKH26, we demonstrated that they could participate in myotube formation when cultured together with differentiating human myoblasts. Experiments performed with EGFP-marked hMSCs and DsRed-labeled DMD myoblasts revealed that the EGFP-positive DMD myotubes were also DsRed-positive indicating that hMSCs participate in human myogenesis through cellular fusion. Finally, we showed that hMSCs transduced with a tropism-modified high-capacity hybrid viral vector encoding full-length Dys could complement the genetic defect of DMD myotubes.