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Biological role of EET in induction of vascular endotberial growth factor underr hypoxia

EET在缺氧条件下诱导血管内膜生长因子的生物学作用

基本信息

批准号：
16590129
负责人：
IMAOKA Susumu
金额：
$ 2.24万
依托单位：
Kwansei Gakuin University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2005
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-16590129/
关键词：
hypoxia VEGF EET HUAEC cytochrome P450 ヒト臍帯動脈内皮細胞(HUAEC)チトクロームP450 EDHF 血管内皮細胞

项目摘要

Cytochrome P450 (P450) catalyze the NADPH-dependent oxidation of arachidonic acid to several unique eicosanoids such as epoxyeicosatrienoic acid (EET) which has four isomers, 5,6-EET, 8,9-EET, 11,12-EET, and 14,15-EET. A particular interest in the epoxygenase reaction has developed because of the potent biological activities (modulation of vascular tone and anti-inflammatory activity etc.) attributed to the EETs. I focused on a new biological function of EETs which induce vascular endotherial growth factor (VEGF) under hypoxia. In this study, human hepatoma cell, Hep3B and human umbilical artery endotherial cell (HUAEC) were used under normoxic and hypoxic conditions. An inhibitor for phospholipase A2 which supplies arachidonic acid from plasma membrane, MAFP, inhibited the VEGF induction in HUAEC under hypoxia. NADPH-P450 reductase (NPR) supplies electrons to P450 and is necessary for P450 to oxidize arachidonic acid. An inhibitor for NPR, DPIC, inhibited the VEGF induction. These results suggest that arachidonic acid and P450 are important for the VEGF induction in HUAEC under hypoxia. The same effects on Hep3B cells were also observed. In Hep3B cells, inhibitors for P450s, sulfaphenazol (an inhibitor for CYP2C8 and CYP2C9) and ketoconazole (an inhibitor for CYP3A) inhibited hypoxia response (erythropoietin induction) efficiently. Addition of 11,12-EET or 14,15-EET induced VEGF in HUAEC under hypoxia but hydration products, 11,12-DHETE and 14,15-DHETE did not induce VEGF. Hypoxia-inducible factor (HIF-1) is necessary for induction of VEGF under hypoxia. 11,12-EET and 14,15-EET did not change the levels of HIF-1. In conclusion, EETs produced by P450s play an important role in hypoxic response of cells. EETs induced VEGF independent on HIF-1 which is an important factor of hypoxic response.

细胞色素P450 （P450）催化花生四烯酸nadph依赖的氧化生成几种独特的二十烷酸，如环氧二碳三烯酸（EET），它有四种异构体，5,6-EET, 8,9-EET, 11,12-EET和14,15-EET。由于eet具有强大的生物活性（调节血管张力和抗炎活性等），人们对环氧合酶反应产生了特别的兴趣。重点研究了EETs在缺氧条件下诱导血管内皮生长因子（VEGF）的新生物学功能。本研究使用人肝癌细胞Hep3B和人脐动脉内皮细胞（huec）在常氧和缺氧条件下进行实验。从质膜供给花生四烯酸的磷脂酶A2抑制剂MAFP可抑制缺氧条件下huec对VEGF的诱导。NADPH-P450还原酶（NPR）为P450提供电子，是P450氧化花生四烯酸所必需的。一种NPR抑制剂DPIC抑制了VEGF的诱导。上述结果提示，花生四烯酸和P450在缺氧条件下对血管内皮生长因子的诱导有重要作用。对Hep3B细胞也观察到相同的作用。在Hep3B细胞中，p450的抑制剂、磺胺苯唑（CYP2C8和CYP2C9的抑制剂）和酮康唑（CYP3A的抑制剂）有效地抑制缺氧反应（促红细胞生成素诱导）。在缺氧条件下，添加11,12- eet或14,15- eet可诱导小鼠血管内皮生长因子（VEGF），而水化产物11,12- dhete和14,15- dhete对VEGF无诱导作用。缺氧诱导因子（HIF-1）是缺氧条件下VEGF诱导的必需因子。11,12- eet和14,15- eet没有改变HIF-1的水平。综上所述，p450产生的eet在细胞缺氧反应中起重要作用。EETs诱导的VEGF不依赖于HIF-1， HIF-1是缺氧反应的重要因子。