ESTABLISHMENT OF LONG-TERM MYOGENIC CULTURES FROM PATIENTS WITH DUCHENNE MUSCULAR-DYSTROPHY BY RETROVIRAL TRANSDUCTION OF A TEMPERATURE-SENSITIVE SV40 LARGE T-ANTIGEN

We have established long-term human myogenic cultures from adult human skeletal muscle biopsies by infecting primary explant cultures with a n amphotropic retroviral construct encoding a temperature-sensitive SV 40 large T antigen, tsA58-U19. Infected myoblasts expressed the large T antigen and showed greatly enhanced proliferative capacity when cult ured at 33 degrees C, compared with noninfected cells. When the infect ed cultures were incubated at 39 degrees C, the cells withdrew from cy cle, aligned, and fused to form multinucleated myotubes which expresse d certain antigens that are similarly expressed in nontransduced diffe rentiating muscle cells. Myogenic clones with greatly increased prolif erative capacity were generated, for the first time, from biopsies obt ained from Duchenne muscular dystrophy patients as well as from normal , dystrophin-positive individuals. Cell lines produced by this approac h may prove valuable for in vitro studies of myogenesis and for invest igating the cellular and molecular consequences of inherited muscle di seases. (C) 1996 Academic Press, Inc.