Basic fibroblast growth factor (bFGF) has been previously shown to be mitogenic for vascular smooth muscle cells (VSMCs) in vivo, but only after vascular injury. We show in the present study that the regulation of bFGF-stimulated VSMC proliferation, by vascular cell-secreted heparin-like compounds, correlates with inhibition of bFGF binding to cell-associated heparan sulfate proteoglycans. The stimulation of cultured VSMC proliferation by bFGF was markedly reduced when these cells were cocultured with confluent endothelial cells or confluent VSMCs (100.8±8.4% and 55.6±2.3% inhibition, respectively) or with conditioned media from these two cell types. Balb/c3T3 fibroblasts had no statistically significant effect on bFGF-stimulated VSMC proliferation. Vascular cell-conditioned media also inhibited bFGF binding to heparan sulfate proteoglycans on VSMCs, and the inhibition of binding correlated linearly with the inhibition of proliferation after a critical amount of binding was inhibited (44%) (r=.952, P<.0001). Heparinase or heparitinase treatment of conditioned media removed the bFGF-inhibitory effects, presumably by degrading heparin-like compounds. Indeed, heparin itself mimicked the inhibitory effects of conditioned media on bFGF-mediated proliferation and binding to heparan sulfate proteoglycans. These results suggest a bFGF regulatory role for vascular cell-produced heparin-like compounds, linking the mitogenic effects with binding to heparan sulfate proteoglycans for this heparin-binding growth factor.