NF- Kappa B Mediates alpha v beta 3 Integrin-induced Endothelial Cell Survival

The alpha sub(v) beta sub(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 sub(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 sub(1)-integrin ligand) did not induce NF- Kappa B activity. The alpha sub(v) beta sub(3) integrin was most important for osteopontin-mediated NF- Kappa B induction and survival, since adding a neutralizing anti- beta sub(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 sub(v) beta sub(3) integrin-mediated endothelial cell survival.