THE 140-KILODALTON ANTIANGIOGENIC FRAGMENT OF THROMBOSPONDIN-1 BINDS TO BASIC FIBROBLAST GROWTH-FACTOR

Thrombospondin-1 (TSP) inhibits the angiogenic activity of basic fibro blast growth factor (bFGF), Here we address the hypothesis of a direct interaction between TSP and bFGF, Gel permeation chromatography and c ross-linking experiments demonstrated that bFGF binds to TSP in soluti on, bFGF also bound to immobilized TSP in a solid-phase assay, Binding was dose-dependent, with a K-d in the nanomolar range, and was inhibi ted by anti-TSP antibodies. The 140-kDa carboxyl-terminal fragment of TSP, but not the 25-kDa heparin-binding fragment, fully retained the b FGF binding capacity, Accordingly, binding was inhibited by monoclonal antibodies directed against this fragment. Heparin completely blocked bFGF binding to TSP and to the 140-kDa fragment, TSP and its 140-kDa fragment inhibited the binding of bFGF to endothelial cells at concent rations (greater than or equal to 100 nM) that inhibited endothelial c ell proliferation but not motility, Low-affinity binding was inhibited more than high-affinity binding (up to 76 and 41% inhibition, respect ively), and the inhibition was reversed by anti-TSP antibodies, Vitron ectin and transforming growth factor beta, potentially associated with TSP, did not affect bFGF binding to endothelial cells, Although TSP d id not affect the activation of the high-affinity receptors, it reduce d the long-term internalization of bFGF. We conclude that TSP binds to bFGF through a domain within its 140-kDa fragment, a mechanism that m ight affect bFGF interaction with endothelial cells, activity, and ass ociation with the extracellular matrix.