Overexpression of connexin 43 in skeletal myoblasts: Relevance to cell transplantation to the heart

Objective: Skeletal myoblast transplantation is a promising strategy for tr eating end-stage heart failure. One potential problem in the development of functional, synchronously contracting grafts is the degree of intercellula r communication between grafted myoblasts and host cardiomyocytes. Thus it is expected that enhancement of intercellular gap junction formation would result in improved efficiency of skeletal myoblast transplantation. In this study we investigated whether myoblasts overexpressing connexin 43, a majo r cardiac gap junction protein, would enhance this intercellular communicat ion. Methods and Results: L6 rat skeletal myoblast cell lines overexpressing con nexin 43 were generated by means of gene transfection and clonal selection. Connexin 43 overexpression of these myoblasts, which continued both in und ifferentiated and differentiated states (Lip to 17-fold greater protein lev el in comparison with control-transfected myoblasts, as measured with Weste rn blotting), was observed on cell surfaces where gap junctions should exis t. Both dye microinjection and scrape loading with fluorescent dyes showed enhancement in intercellular dye transfer between connexin 43-transfected m yoblasts compared with that found in control-transfected cells. Morphologic ally, these myoblasts fused and differentiated into multinucleated myotubes more rapidly, demonstrating a higher level of cellular creatine kinase act ivity as a marker of myogenic differentiation throughout the culture period compared with that of control-transfected myoblasts. Conclusions: We have generated connexin 43-overexpressing skeletal myoblast cell lines that resulted in improved formation of functional intercellular gap junctions, which could be relevant to synchronous contraction of graft ed myoblasts in the heart. In addition, these cells demonstrated more rapid differentiation, which would also be advantageous in a graft for transplan tation to the heart.