N-terminal domain of soluble epoxide hydrolase negatively regulates the VEGF-mediated activation of endothelial nitric oxide synthase
N-terminal domain of soluble epoxide hydrolase negatively regulates the VEGF-mediated activation of endothelial nitric oxide synthase
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DOI：
10.1093/cvr/cvr267
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发表时间：
2012-01-01
影响因子：
10.8
通讯作者：
Lee, Tzong-Shyuan
中科院分区：
文献类型：
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作者：
Hou, Hsin-Han;Hammock, Bruce D.;Lee, Tzong-Shyuan
目的哺乳动物可溶性环氧化物水解酶(SEH)具有环氧化物水解酶和磷酸酶结构域。SEH水解酶活性在内皮细胞(ECs)环氧二十碳三烯酸(EETs)代谢和内皮型一氧化氮合酶(ENOS)激活中的作用已被明确。然而，关于sEH磷酸酶活性在eNOS激活中的作用还知之甚少。方法和结果sEH基因敲除(sEH(-/-))小鼠主动脉eNOS磷酸化水平明显高于野生型小鼠。在内皮细胞中，药物抑制sEH磷酸酶或过度表达sEH并结合无活性的磷酸酶结构域可促进血管内皮生长因子(VEGF)诱导的NO生成和eNOS磷酸化。相反，在Ser617、Ser635和Ser1179上过表达sEH的磷酸酶结构域可阻止血管内皮生长因子介导的NO产生和eNOS的磷酸化。此外，经血管内皮生长因子处理后，sEH瞬时酪氨酸磷酸化的c-Src激酶依赖性增加，并形成sEH-eNOS复合体，该复合体可被c-Src激酶抑制剂PP1或c-Src显性阴性突变体K298M所消除。通过检测小鼠血管内皮细胞的管状结构和Matrigel栓的新生血管，我们还证明了sEH的磷酸酶结构域在血管生成中起着关键作用。结论除了环氧化物水解酶活性外，sEH的磷酸酶活性在调节eNOS活性和NO介导的EC功能中起着关键作用。
Aims The mammalian soluble epoxide hydrolase (sEH) has both an epoxide hydrolase and a phosphatase domain. The role of sEH hydrolase activity in the metabolism of epoxyeicosatrienoic acids (EETs) and the activation of endothelial nitric oxide synthase (eNOS) in endothelial cells (ECs) has been well defined. However, far less is known about the role of sEH phosphatase activity in eNOS activation. In the present study, we investigated whether the phosphatase domain of sEH was involved in the eNOS activation in ECs.Methods and results The level of eNOS phosphorylation in aortas is higher in the sEH knockout (sEH(-/-)) mice than in wild-type mice. In ECs, pharmacological inhibition of sEH phosphatase or overexpressing sEH with an inactive phosphatase domain enhanced vascular endothelial growth factor (VEGF)-induced NO production and eNOS phosphorylation. In contrast, overexpressing the phosphatase domain of sEH prevented the VEGF-mediated NO production and eNOS phosphorylation at Ser617, Ser635, and Ser1179. Additionally, treatment with VEGF induced a c-Src kinase-dependent increase in transient tyrosine phosphorylation of sEH and the formation of a sEH-eNOS complex, which was abolished by treatment with a c-Src kinase inhibitor, PP1, or the c-Src dominant-negative mutant K298M. We also demonstrated that the phosphatase domain of sEH played a key role in VEGF-induced angiogenesis by detecting the tube formation in ECs and neovascularization in Matrigel plugs in mice.Conclusion In addition to epoxide hydrolase activity, phosphatase activity of sEH plays a pivotal role in the regulation of eNOS activity and NO-mediated EC functions.