VEGF-A and αVβ3 integrin synergistically rescue angiogenesis via N-Ras and PI3-K signaling in human microvascular endothelial cells
VEGF-A and αVβ3 integrin synergistically rescue angiogenesis via N-Ras and PI3-K signaling in human microvascular endothelial cells
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DOI：
10.1096/fj.02-1171fje
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发表时间：
2003-08-01
期刊：
影响因子：
4.8
通讯作者：
Velazquez, OC
中科院分区：
文献类型：
--
作者：
Liu, ZJ;Snyder, R;Velazquez, OC
我们最近发现，正常的成纤维细胞介导的毛细血管样分化的人微血管内皮细胞（HMVEC）的三维血管生成模型。在这里，我们发现VEGF-A和α（v）β（3）整合素的协同作用在成纤维细胞介导的血管生成中是至关重要的，因为在低血清条件下，成纤维细胞产生的VEGF和HMVEC中β（3）整合素表达的增强可以挽救毛细血管样内皮分化。为了研究下游信号传导机制，我们比较了N-Ras和Rho/Rac/Cdc 42以及磷脂酰肌醇3-激酶（PI 3-K）和Akt参与毛细血管样网络形成的情况。N-Ras的显性负突变体（N-Ras（N17））抑制了网络形成，而Rho/Rac/Cdc 42的突变体不抑制网络形成。组成型活性形式的PI 3-K的过表达挽救了网络形成，其被显性负性β（3）整联蛋白抑制;然而，活性形式的Akt未能挽救抑制，但诱导了HMVEC的表型变化。此外，PI 3-K是N-Ras的下游靶点，因为它可以与N-Ras免疫共沉淀，并且其活性形式可以挽救N-Ras（N17）的抑制作用。因此，我们的数据表明VEGF-A和alpha（v）beta（3）对成纤维细胞介导的微血管网络形成的协同作用存在N-Ras和PI 3-K依赖性但Rho/Rac/Cdc 42和Akt独立性的信号传导机制。
We recently showed that normal fibroblasts mediate capillary-like differentiation of human microvascular endothelial cells (HMVEC) in a 3-D angiogenesis model. Here, we show that a collaborative effect of VEGF-A and alpha(v)beta(3) integrin is critical in fibroblast-mediated angiogenesis because enhancement of both VEGF production by fibroblasts and beta(3) integrin expression in HMVEC can rescue capillary-like endothelial differentiation under reduced serum conditions. To investigate the downstream signaling mechanisms, we compared N-Ras and Rho/Rac/Cdc42, as well as phosphatidylinositol 3-kinase (PI3-K) and Akt, for their involvement in the capillary-like network formation. The dominant-negative mutant of N-Ras (N-Ras(N17)), but not the mutants of Rho/Rac/Cdc42, suppressed network formation. Overexpression of a constitutively active form of PI3-K rescued the network formation, which was inhibited by a dominant-negative beta(3) integrin; however, an active form of Akt failed to rescue the inhibition but induced a phenotypic change in HMVEC. Moreover, PI3-K is a downstream target of N-Ras because it could be coimmunoprecipitated with N-Ras, and its active form could rescue the inhibitory effect of N-Ras (N17). Thus, our data indicate the existence of N-Ras- and PI3-K-dependent but Rho/Rac/Cdc42- and Akt-independent signaling mechanisms for the synergistic effect of VEGF-A and alpha(v)beta(3) on fibroblast-mediated microvascular network formation.