The alpha(V) beta(3) integrin plays a fundamental role during the angiogenesis process by inhibiting endothelial cell apoptosis. However, the mechanism of inhibition is unknown. In this report, we show that integrin mediated cell survival involves regulation of nuclear factor-kappa B (NF-kappa B) activity. Different extracellular matrix molecules were able to protect rat aorta-derived endothelial cells from apoptosis induced by serum withdrawal. Osteopontin and beta(3) integrin ligation rapidly increased NF-kappa B activity as measured by gel shift and reporter activity. The p65 and p50 subunits were present in the shifted complex. In contrast, collagen type I (a beta(1)-integrin ligand) did not induce NF-kappa B activity. The alpha(V) beta(3) integrin was most important for osteopontin-mediated NF-kappa B induction and survival, since adding a neutralizing anti-beta(3) integrin antibody blocked NF-kappa B activity and induced endothelial cell death when cells were plated on osteopontin. NF-kappa B was required for osteopontin-and vitronectin-induced survival since inhibition of NF-kappa B activity with nonphosphorylatable I kappa B completely blocked the protective effect of osteopontin and vitronectin. In contrast, NF-kappa B was not required for fibronectin, laminin, and collagen type I-induced survival. Activation of NF-kappa B by osteopontin depended on the small GTP-binding protein Ras and the tyrosine kinase Src, since NF-kappa B reporter activity was inhibited by Ras and Src dominant-negative mutants. In contrast, inhibition of MEK and PI3-kinase did not affect osteopontin-induced NF-kappa B activation. These studies identify NF-kappa B as an important signaling molecule in alpha(V) beta(3) integrin-mediated endothelial cell survival.
