Genetic Complementation of Human Muscle Cells via Directed Stem Cell Fusion

Abstract

Duchenne muscular dystrophy (DMD) is caused by mutations in the X chromosome-linked DMD gene, which encodes the sarcolemma-stabilizing protein-dystrophin. Initial attempts at DMD therapy deployed muscle progenitor cells from healthy donors. The utilization of these cells is, however, hampered by their immunogenicity, while those from DMD patients are scarce and display limited ex vivo replication. Nonmuscle cells with myogenic capacity may offer valuable alternatives especially if, to allow autologous transplantation, they are amenable to genetic intervention. As a paradigm for therapeutic gene transfer by heterotypic cell fusion we are investigating whether human mesenchymal stem cells (hMSCs) can serve as donors of recombinant DMD genes for recipient human muscle cells. Here, we show that forced MyoD expression in hMSCs greatly increases their tendency to participate in human myotube formation turning them into improved DNA delivery vehicles. Efficient loading of hMSCs with recombinant DMD was achieved through a new tropism-modified high-capacity adenoviral (hcAd) vector directing striated muscle-specific synthesis of full-length dystrophin. This study introduces the principle of genetic complementation of gene-defective cells via directed cell fusion and provides an initial framework to test whether transient MyoD synthesis in autologous, gene-corrected hMSCs increases their potential for treating DMD and, possibly, other muscular dystrophies.

Histone Deacetylase Inhibition Activates Transgene Expression from Integration-Defective Lentiviral Vectors in Dividing and Non-Dividing Cells

“…The origin and the culture conditions for the human myoblasts and the human bone-marrow-derived mesenchymal stem cells (hMSCs) used in this study have been described previously (Cudré-Mauroux et al, 2003;Knaä n-Shanzer et al, 2005;Gonçalves et al, 2008;Gonçalves et al, 2011).…”

“…One day later, the cells were washed twice with PBS and maintained for 5 days in differentiation medium (DM) (Cudré-Mauroux et al, 2003;Gonçalves et al, 2008;Gonçalves et al, 2011). The resulting myotubes were then transduced with either LV.CMV.eGFP or IDLV.CMV.eGFP at 93 ng of p24 gag per well in the absence or in the presence of either 8 lM TSA or 10 mM NaBu.…”

Barriers in contribution of human mesenchymal stem cells to murine muscle regeneration

“…Evidence provided so far for the ability of MSCs to differentiate along the myogenic lineage is conflicting. Although some previous studies assigned myogenic properties to MSCs by demonstrating their in vitro and in vivo differentiation into satellite cells and myoblasts and their ability to form myotube-like structures through homotypic fusion [19,21] , others regard the myogenic reprogramming of the MSCs to be the consequence of their fusion with inherently myogenic cells [2,22,23] . Whether this contradiction can be attributed to the differences in MSC origin, the model used or the read-out methods applied, remains to be investigated.…”

Concise Review: Mesoangioblast and Mesenchymal Stem Cell Therapy for Muscular Dystrophy: Progress, Challenges, and Future Directions

“…Despite this, neither human nor canine ucMSCs expressed muscle proteins [38][39][40][41], indicating that ucMSCs lack myogenic potential in vivo in animal models for muscular dystrophy. However, forced expression of MyoD in ucMSCs enhanced skeletal muscle differentiation in vitro [42] and might also be useful to stimulate their myogenic differentiation in vivo, because this approach has been used to improve skeletal muscle differentiation of MSCs from synovium and orthopedic surgery remnants in vitro and in vivo [43,44].…”