EFFECT OF PLATELET-DERIVED GROWTH-FACTOR ISOFORMS ON THE MIGRATION OFMOUSE EMBRYO LIMB MYOGENIC CELLS

The effect of platelet-derived growth factor (PDGF) isoforms on limb m yoblast migration was examined in vitro. Using Blindwell Chemotaxis ch ambers, the ability of PDGF-AA, -AB and -BB to stimulate the migration of myoblasts, obtained from the proximal region of 11.5 day mouse for elimb buds, was examined. Immunocytochemistry, with the anti-sarcomeri c myosin antibody MF-20, was used to identify the myogenic cells in th e heterogeneous cell population. Myoblasts, suspended in PDGF-free med ium in the upper chamber, migrated across the polycarbonate filter of the Blindwell chamber to 1-10 ng/ml PDGF-AB and 1-100 ng/ml PDGF-BB si tuated in the lower well. At 1-10 ng/ml of either PDGF-AB or -BB migra tion increased in a dose-dependent manner. PDGF-AA, however, was unabl e to elicit a significant locomotory response in forelimb myoblasts. A Checkerboard assay, with various concentrations of PDGF-AA, -AB or -B B in the upper and lower wells of the chamber, indicated that -AB and -BB but not -AA stimulated the random migration of limb myoblasts. The differential effect of PDGF isoforms on myoblast migration was compar ed with other aspects of skeletal muscle development. At 0.1-10 ng/ml all three isoforms were able to stimulate an increase in the number of differentiated myoblasts, indicated by the expression of sarcomeric m yosin, on examination after 48 h when cultured at low density. In high density cultures, however, these isoforms inhibited myoblast fusion w hen compared to the spontaneous fusion observed in untreated cultures. Immunohistochemical studies of both cultured limb cells and cryosecti ons of 11.5 day whole limbs revealed that myoblasts expressed both PDG F alpha- and beta-receptors which suggests that the action of PDGF iso forms on limb myoblasts is receptor-mediated. Finally, having demonstr ated that the PDGF-B monomer stimulates migration in limb myoblasts, b y immunohistochemistry, the presence of PDGF-B was confirmed and its d istribution examined in the 11.5 day forelimb.