Basic fibroblast growth factor mediates neointimal formation in porcine aortic explants

grantor: University of TorontoNeointimal formation was studied in a porcine aortic explant model that exhibits intimal smooth muscle cell (ISMC) accumulation beneath an intact endothelium (nondenuded) after seven days in culture. Removal of the endothelium prior to culture (denuded explant) results in limited neointimal formation, unless denuded explants are incubated with clay 5 nondenuded conditioned media. The concentration of a number of known SMC mitogens was similar in conditioned media from nondenuded and denuded explants, indicating that these mitogens were not the soluble factor in nondenuded conditioned media which induced neointimal formation in denuded explants. Since basic fibroblast growth factor (bFGF) was found in greatest abundance, I hypothesized that this mitogen mediated early neointimal formation. Basic FGF immunoreactivity in tissue sections and mitogenic activity of bFGF in conditioned media were similar in denuded and nondenuded explants. However, the addition of exogenous bFGF (1 ng/mL) enhanced neointimal formation in nondenuded explants but not in denuded explants, indicating an FGF receptor dependent mechanism. Western blots of intimal lysates showed a time dependent increase in fibroblast growth factor receptor-1 (FGFR-1) expression over a seven day course, with higher levels seen in lysates from nondenuded explants. Immunoreactive FGFR-1, detected in both endothelial cells and ISMCs of nondenuded explants, were not detected in ISMC of denuded explant sections. Quantitative ligand binding and in situ autoradiography demonstrated specific, high affinity binding of $\sp{125}$I-bFGF to receptors on intimal cells. FGF receptors 3 and 4 were also expressed in intimal lysates; however, their expression did not correlate with neointimal formation and was markedly less abundant than FGFR-1 expression. The addition of bFGF neutralizing antibodies which resulted in partial reduced bFGF did not decrease ISMC accumulation but increased FGFR-1 expression. When bFGF neutralizing antibodies were administered more frequently (every 24 hours), resulting in a marked reduction in bFGF, neointimal formation was reduced despite increased FGFR-1 protein. Thus, bFGF mediates neointimal formation and appears to reciprocally regulate FGFR-1 in porcine aortic explants.Ph.D