Both vascular endothelial growth factor receptors (VEGFR) and integrins are major regulators of VEGF-induced angiogenesis. Previous work has shown that ␤3 integrin can regulate negatively VEGFR2 expression. Here we show that ␤3 integrin can regulate negatively VEGF-mediated angiogenesis by limiting the interaction of the co-receptor NRP1 (neuropilin-1) with VEGFR2. In the presence of ␣v␤3 integrin, NRP1 contributed minimally to VEGF-induced angiogenic processes in vivo, ex vivo, and in vitro. Conversely, when ␤3 integrin expression is absent or low or its function is blocked with RGD-mimetic inhibitors, VEGF-mediated responses became NRP1-dependent. Indeed, combined inhibition of ␤3 integrin and NRP1 decreased VEGF-mediated angiogenic responses further than individual inhibition of these receptors. We also show that ␣v␤3 integrin can associate with NRP1 in a VEGF-dependent fashion. Our data suggest that ␤3 integrin may, in part, negatively regulate VEGF signaling by sequestering NRP1 and preventing it from interacting with VEGFR2.

Integrin ␣3␤1 is a major receptor for laminin. The expression levels of laminins-8 and -10 in the basement membrane surrounding blood vessels are known to change during tumor angiogenesis. Although some studies have suggested that certain ligands of ␣3␤1 can affect angiogenesis either positively or negatively, either a direct in vivo role for ␣3␤1 in this process or its mechanism of action in endothelial cells during angiogenesis is still unknown. Because the global genetic ablation of ␣3-integrin results in an early lethal phenotype, we have generated conditional-knockout mice where ␣3 is deleted specifically in endothelial cells (ec-␣3؊/؊). Here we show that ec-␣3؊/؊ mice are viable, fertile, and display enhanced tumor growth, elevated tumor angiogenesis, augmented hypoxia-induced retinal angiogenesis, and increased vascular endothelial growth factor (VEGF)-mediated neovascularization ex vivo and in vivo. Furthermore, our data provide a novel method by which an integrin may regulate angiogenesis. We show that ␣3␤1 is a positive regulator of endothelial-VEGF and that, surprisingly, the VEGF produced by endothelial cells can actually repress VEGF-receptor 2 (Flk-1) expression. These data, therefore, identify directly that endothelial ␣3␤1 negatively regulates pathological angiogenesis and implicate an unexpected role for low levels of endothelial-VEGF as an activator of neovascularization.