Reactive oxygen species as downstream mediators of angiogenic signaling by vascular endothelial growth factor receptor-2/KDR
Reactive oxygen species as downstream mediators of angiogenic signaling by vascular endothelial growth factor receptor-2/KDR
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DOI：
10.1074/jbc.m107711200
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发表时间：
2002-02-01
影响因子：
4.8
通讯作者：
Galeotti, T
中科院分区：
文献类型：
--
作者：
Colavitti, R;Pani, G;Galeotti, T
最近的证据表明，活性氧（ROS）的参与有丝分裂级联启动的酪氨酸激酶受体的几个生长因子肽。我们已经询问血管内皮生长因子（VEGF）是否也利用ROS作为VEGF受体-2（VEGFR-2）/KDR受体下游的信使中间体，因为在新血管生成期间内皮细胞的增殖在生理上受氧调节，并且可能受其衍生物种类调节。在猪主动脉内皮细胞稳定表达人KDR，受体激活VEGF后，过氧化氢的细胞内生成的快速增加，所揭示的过氧化物敏感的探针二氯荧光素二乙酸酯。遗传学和药理学研究表明，这种氧化剂爆发需要作为上游事件的磷脂酰肌醇3-激酶和小GT3 Rac-1的激活，并且可能由脂氧合酶启动。有趣的是，响应于VEGF的ROS产生并没有被阻断，而是被内皮一氧化氮合酶抑制剂二苯碘鎓和N（G）甲基-L-精氨酸增强，排除了一氧化氮是在VEGF刺激的细胞中检测到的氧化剂种类的可能性。抑制KDR依赖的ROS产生减弱早期信号传导事件，包括受体自磷酸化和与磷脂酶C-γ-谷胱甘肽S-转移酶融合蛋白的结合。此外，过氧化氢酶、脂氧合酶抑制剂去甲二氢愈创木酸、合成ROS清除剂EUK-134和磷脂酰肌醇3-激酶抑制剂渥曼青霉素都能降低ERK磷酸化对VEGF的反应，抗氧化剂可阻止VEGF依赖性有丝分裂。最后，在几乎缺氧的环境中进行细胞培养和刺激，模拟ROS清除剂对受体和ERK磷酸化的作用，加强了ROS是由KDR启动的促有丝分裂信号级联的必要组分的观点。这些数据确定活性氧作为一类新的细胞内血管生成介质，并可能代表一个潜在的前提，为新的抗氧化剂为基础的抗血管生成疗法。
Recent evidence shows the involvement of reactive oxygen species (ROS) in the mitogenic cascade initiated by the tyrosine kinase receptors of several growth factor peptides. We have asked whether also the vascular endothelial growth factor (VEGF) utilizes ROS as messenger intermediates downstream of the VEGF receptor-2 (VEGFR-2)/KDR receptor given that the proliferation of endothelial cells during neoangiogenesis is physiologically regulated by oxygen and likely by its derivative species. In porcine aortic endothelial cells stably expressing human KDR, receptor activation by VEGF is followed by a rapid increase in the intracellular generation of hydrogen peroxide as revealed by the peroxide-sensitive probe dichlorofluorescein diacetate. Genetic and pharmacological studies suggest that such oxidant burst requires as upstream events the activation of phosphatidylinositol 3-kinase and the small GTPase Rac-1 and is likely initiated by lipoxygenases. Interestingly, ROS generation in response to VEGF is not blocked but rather potentiated by endothelial nitric-oxide synthase inhibitors diphenyleneiodonium and N(G)methyl-L-arginine, ruling out the possibility of nitric oxide being the oxidant species here detected in VEGF-stimulated cells. Inhibition of KDR-dependent generation of ROS attenuates early signaling events including receptor autophosphorylation and binding to a phospholipase C-gamma-glutathione S-transferase fusion protein. Moreover, catalase, the lipoxygenase inhibitor nordihydroguaiaretic acid, the synthetic ROS scavenger EUK-134, and phosphatidylinositol 3-kinase inhibitor wortmannin all reduce ERK phosphorylation in response to VEGF, and antioxidants prevent VEGF-dependent mitogenesis. Finally, cell culture and stimulation in a nearly anoxic environment mimic the effect of ROS scavenger on receptor and ERK phosphorylation, reinforcing the idea that ROS are necessary components of the mitogenic signaling cascade initiated by KDR. These data identify ROS as a new class of intracellular angiogenic mediators and may represent a potential premise for new antioxidant-based antiangiogenic therapies.