摘要
目的 以内皮细胞产生NO的关键酶———eNOS(内皮一氧化氮合酶 )为研究目标 ,探讨外源性内皮源性超极化因子EDHF(EETs)对内皮细胞合成NO的影响。方法 在原代培养 3～ 4代以内的牛主动脉内皮细胞中 ,分别加入不同浓度 (5 0～ 2 0 0nmol·L-1)的 8,9 EET、11,12 EET、14 ,15 EET ,作用 1h后用不同的方法收获细胞。用WesternBlot以及NorthernBlot方法检测EETs对eNOS基因表达的影响 ;同时通过检测L [3 H] 精氨酸转化为L [3 H] 瓜氨酸的量研究EETs对NOS活性的影响。结果 显示 8,9 EET、11,12 EET、14 ,15 EET均呈浓度依赖性地增加eNOS蛋白质的表达 ,并提高eNOSmRNA表达水平以及NOS酶活性。结论 外源性EDHF对eNOS基因表达是一种正反馈调节作用 ,从而能够促进内皮细胞NO的产生 。
AIM In addition to NO and PGI 2, endothelium derived hyperpolarizing factor(EDHF) is a third endothelium derived vasodilator. It has been suggested that epoxyeicosatrienoic acids (EETs), arachidonic acid cytochrome P450 epoxygenase metabolites, are EDHF. Previous studies reported that NO attenuated the release of EDHF. However, little is known about the effect of EDHF on nitric oxide synthesis. Therefore, we studied the effect of EDHF (EETs) on eNOS gene expression and NOS activity in the cultured bovine endothelial cells. METHODS Bovine aortic endothelial cells were isolated and cultured. The third or fourth passages of the endothelial cells were treated with physiologically relevant concentrations (50 nmol·L -1 , 200 nmol·L -1 ) of , , EET for 1 hour and then the treated cells were harvested to analyze eNOS gene expression by Western blotting and Northern blotting. The eNOS activity was determined by measuring the conversion of L arginine to L citrulline. RESULTS Various EETs all significantly up regulated the eNOS protein expression in a concentration dependent manner, and increased eNOS mRNA expression levels and eNOS activity. CONCLUSIONS Each of the EET regioisomers induced a significant increase in eNOS expression and eNOS activity. These results indicated a positive feedback regulation of exogenous EDHF on eNOS. So EDHF may increase the synthesis of NO in endothelial cells, which suggests a feasibility to prevent and treat cardiovascular diseases by promoting epoxygenase expression.
出处
《中国药理学通报》
CAS
CSCD
北大核心
2003年第9期1003-1008,共6页
Chinese Pharmacological Bulletin
基金
国家自然科学基金资助课题 No 3 9870 3 0 7
关键词
内皮源性超极化因子
EETS
内皮一氧化氮合酶
内皮细胞
endothelium derived hyperpolarizing factor
EETs
endothelial nitric oxide synthase
endothelial cell
