Characterization of vascular endothelial growth factor's effect on the activation of protein kinase C, its isoforms, and endothelial cell growth
Characterization of vascular endothelial growth factor's effect on the activation of protein kinase C, its isoforms, and endothelial cell growth
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DOI：
10.1172/jci119006
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发表时间：
1996-11-01
影响因子：
15.9
通讯作者：
King, GL
中科院分区：
文献类型：
--
作者：
Xia, P;Aiello, LP;King, GL
血管内皮生长因子（Vascular endothelial growth factor，VEGF）是一种强有力的内皮细胞有丝分裂原，通过与酪氨酸激酶受体结合来介导其作用。VEGF诱导蛋白激酶C（PKC）激活的浓度和时间依赖性增加，在5 × 10（-）细胞浆和细胞膜组分中PKC亚型的免疫印迹分析表明，在VEGF刺激后，细胞膜组分中Ca 2+敏感的PKC亚型（α和β II）的含量增加，而PKC亚型δ和β II没有观察到变化。VEGF对PKC活性的刺激是通过激活磷脂酶C γ来实现的。这通过在牛主动脉内皮细胞中PLC γ酪氨酸磷酸化、[H-3]肌醇磷酸产生和[H-3]花生四烯酸标记的二酰基甘油形成的平行增加来证明。此外，VEGF使磷脂酰肌醇3-激酶活性增加2.1倍，这被渥曼青霉素抑制，有趣的是，Genistein，酪氨酸激酶抑制剂，GFX或H-7，PKC抑制剂，消除VEGF诱导的PKC激活和内皮细胞增殖，VEGF的促有丝分裂作用被PKC亚型β选择性抑制剂LY 33 -3531以浓度依赖性方式抑制。相反，反义PKC-α寡核苷酸增强VEGF刺激的细胞生长，同时PKC-α蛋白含量降低70%。因此，VEGF似乎部分通过激活PLC γ和PKC途径介导其促有丝分裂作用，主要涉及内皮细胞中PKC-β亚型的激活。
Vascular endothelial growth factor (VEGF) is a potent endothelial cell mitogen which mediates its effects by binding to tyrosine kinase receptors, We have characterized the VEGF-activated intracellular signal transduction pathway in bovine aortic endothelial cells and correlated this to its mitogenic effects, VEGF induced concentration- and time-dependent increases in protein kinase C (PKC) activation with a maximum of 2.2-fold above the basal level at 5 x 10(-10) M within 10 min as measured both by in situ and translocation assays, Immunoblotting analysis of PKC isoforms in cytosolic and membrane fractions indicated that after VEGF stimulation the content of Ca2+-sensitive PKC isoforms (alpha and beta II) was increased in the membrane fractions, whereas no changes were observed for PKC isoforms delta and epsilon. The stimulation of PKC activity by VEGF was preceded by the activation of phospholipase C gamma (PLC gamma), This was demonstrated by parallel increases in PLC gamma tyrosine phosphorylation, [H-3]inositol phosphate production, and [H-3]arachidonic acid-labeled diacylglycerol formation in bovine aortic endothelial cells, In addition, VEGF increased phosphatidylinositol 3-kinase activity 2.1-fold which was inhibited by wortmannin, a phosphatidylinositol 3-kinase inhibitor, without decreasing the VEGF-induced increase in PKC activity or endothelial cell growth, Interestingly, genistein, a tyrosine kinase inhibitor, and GFX or H-7, PKC inhibitors, abolished both VEGF-induced PKC activation and endothelial cell proliferation, VEGF's mitogenic effect was inhibited by a PKC isoform beta-selective inhibitor, LY33-3531, in a concentration-dependent manner. In contrast, antisense PKC-alpha oligonucleotides enhanced VEGF-stimulated cell growth with a simultaneous decrease of 70% in PKC-alpha protein content, Thus, VEGF appears to mediate its mitogenic effects partly through the activation of the PLC gamma and PKC pathway, involving predominately PKC-beta isoform activation in endothelial cells.