EDN1基因5′上游AP-1顺式调控元件在同型半胱氨酸诱导HUVECs EDN1基因转录中的作用  

Effects of AP-1 element on endothelin-1 gene transcription in homocysteine-induced HUVECs

作　　者：段金虹[1] 徐海珊[1] 戴顺龄[1] 王小明[1] 吴云清[1] 张彦东[1] 程锦轩[1] 孙仁宇[1] 李良[2] 陈槐卿[2] 

机构地区：[1]中国医学科学院基础医学研究所,中国协和医科大学基础医学院,北京100005 [2]四川大学华西医学中心生物医学工程研究所,四川成都610041

出　　处：《中国病理生理杂志》2007年第9期1665-1670,共6页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.30170376)

摘　　要：目的:探讨EDN1基因5′上游AP-1顺式调控元件在同型半胱氨酸(Hcy)诱导HUVECs EDN1基因转录中的作用。方法:用荧光探针DCF检测HUVECs内活性氧的含量;用RT-PCR法检测EDN1mRNA的表达,用Western blotting法测定c-jun/AP-1蛋白的表达;同时用双抗夹心法测定HUVECs上清液中EDN1的分泌;利用重组质粒的瞬时转染技术观察了HUVECs的AP-1报告基因的活性。结果:Hcy能明显增加HUVECs ROS的生成,促进EDN1的分泌和提高EDN1mRNA的表达;瞬时转染分析实验结果显示Hcy能明显诱导质粒pGL3-EDN1-AP-1(-115/+135)荧光素酶的表达,但对质粒pGL3-EDN1-AP-1(-115/+135)的突变体pGL3-EDN1-AP-1MU(-115/+135)荧光素酶的基础表达及Hcy的诱导作用均明显降低。结论:推测Hcy可能改变HUVECs内的氧化还原状态,促进ROS的释放,激活核转录因子AP-1,通过EDN1基因中AP-1顺式作用元件调控该基因转录。AIM： The objective of this study was to observe the effects of AP - 1 element on endothelin - 1 （EDN1） gene transcription in homecysteine （Hey） -induced human umbilical vein endothelial cells（HUVECs）. METHODS： The redox level in Hey - induced HUVECs was determined by using the fluorescent product DCF, The influences of Hey on expressions of EDN1 mRNA and protein of c - jun/AP - 1 in HUVECs were measured by the methods of RT - PCR and Western blotting, respectively. The EDN1 secretion level of HUVECs induced by Hey was determined by enzymatic immunoassay. The transient transfection assay was performed and luciferase activity of transcriptional factor AP - 1 reporter gene induced by Hey was detected. RESULTS： The Hey significantly increased EDN1 secretion, EDN1 mRNA expression and oxidative stress in HUVECs. Hey remarkably induced the expression of pGL3 -EDN1 -AP- 1 （115/＋ 135 ） luciferase reporter gene plasmid. However, basic expression of mutation of pGL3 - EDN1 - AP - 1 （ - 115/＋ 135 ） luciferase repetter gene plasmid was downregnlated markedly in HUVECs induced by Hey. CONCLUSION： Redox - active and release of ROS are changed by Hey. Activated AP - 1 element plays an important role in EDNlgene transcription in HUVECs induced by Hey.

关 键 词：转录因子AP-1 高半胱氨酸 基因 EDN1 

分 类 号：R363[医药卫生—病理学]