ACE2对脂多糖损伤人脐静脉内皮细胞的抗黏附作用  

Anti-adhesion effect of angiotensin-converting enzyme 2 on human umbilical vein endothelial cells induced by lipopolysaccharide

作　　者：何芸[1] 袁颖琳 张琴[1] 杨发健 杨俊霞[1] 

机构地区：[1]重庆医科大学药理教研室,重庆市生物化学与分子药理重点实验室,重庆400016

出　　处：《第三军医大学学报》2015年第13期1302-1307,共6页Journal of Third Military Medical University

基　　金：国家自然科学基金青年科学基金项目(30600812)~~

摘　　要：目的观察脂多糖(lipopolysaccharide,LPS)诱导血管内皮细胞损伤后细胞黏附性改变与血管紧张素转化酶2(angiotensin-converting enzyme 2,ACE2)的表达变化的关系,探讨ACE2对血管内皮细胞损伤所致黏附性增加的对抗作用。方法分别用不同浓度(0、10、100、1 000 ng/m L)LPS诱导人脐静脉内皮细胞(umbilical vein endothelial cell,HUVEC)致其损伤,人单核细胞白血病细胞(human acute monocytic leukemia cell line,THP-1)黏附实验观察HUVEC的黏附性改变。本实验设置4个实验组,分别为未经任何处理的空白对照组、1 000 ng/m L LPS处理6 h的模型组、以10μmol/L氯沙坦钾预处理模型组的氯沙坦钾组及以10μmol/L A779预处理空白对照组的A779组(n=3)。Western blot及RT-PCR法分别检测ACE2的蛋白和基因表达,ELISA法检测细胞上清中血管紧张素Ⅱ(AngⅡ)、血管紧张素1-7(Ang1-7)及细胞间黏附分子1(ICAM-1)的含量。结果单核细胞黏附实验可见黏附在HUVEC上的THP-1呈绿色荧光,黏附细胞数呈LPS诱导浓度及时间依赖性显著增加(P<0.01);Western blot结果显示,LPS诱导HUVEC损伤模型组中ACE2蛋白表达水平较空白对照组显著减少(37.33±3.50)%(P<0.01);RT-PCR结果显示,模型组中ACE2 mRNA表达水平较空白对照组显著减少(26.31±2.19)%(P<0.01);ELISA结果显示,模型组较空白对照组细胞上清Ang1-7显著降低(P<0.05),AngⅡ及ICAM-1均显著升高(P<0.01),氯沙坦钾处理组较模型组ICAM-1显著降低(P<0.01),A779处理组较空白对照组ICAM-1显著升高(P<0.05)。结论 LPS诱导损伤HUVEC细胞后其黏附性增加与ACE2表达下调密切相关,提示ACE2具有一定抗黏附作用。Objective To investigate the relationship of the changes of cell adhesion with the expression of angiotensin-converting enzyme 2（ ACE2） in vascular endothelial cells after LPS injury,and to determine the antagonism of ACE2 on increasing adhesion caused by vascular endothelial injury. Methods After human umbilical vein endothelial cells（ HUVECs） were injured by LPS at different concentrations（ 0,10,100 and 1 000 ng / m L）,the cells adhesion was observed through human acute monocytic leukemia THP-1cells adhesion experiment. The HUVECs were assigned into 4 groups,including untreated blank control cells,injured cells（ induced by 1 000 ng / m L LPS for 6 h）,injured cells with pretreatment of 10 μmol / L losartan potassium,and blank control cells with pretreatment of 10 μmol / L A779 for 0. 5 h. Expression of ACE2 at protein and mRNA levels was detected by Western blotting and RT-PCR respectively,and the contents of AngⅡ,Ang1-7 and ICAM-1 in the cultural supernatants were measured by ELISA. Results THP-1 adhered HUVEC showed green fluorescence,and the number of adhesion cells was increased significantly in a dose- and timedependent manner（ P〈0. 01）. Compared with blank control group,Western blot results showed that the protein expression of ACE2 was reduced significantly in the injured cells by（ 37. 33 ± 3. 50） %（ P〈0. 01）,RT-PCR results showed that its mRNA expression was decreased obviously by（ 26. 31 ± 2. 19） %（ P〈0. 01）.And ELISA results showed that the content of Ang1-7 was decreased significantly（ P〈0. 05）,but those of AngⅡ and ICAM-1 increased significantly in the injured cells than the blank control group（ P〈0. 01）. The content of ICAM-1was obviously lower in the losartan potassium pretreated cells than the injured cells（ P〈0. 01）,and higher in the A779 pretreated blank cells than the blank control（ P〈0. 05）. Conclusion LPS treatment induces cell adhesion in HUVECs,which is closely associated with down-regulation of ACE2. Our results indi

关 键 词：脂多糖 ACE2 脐静脉内皮细胞 单核细胞 

分 类 号：R543[医药卫生—心血管疾病] R969[医药卫生—内科学]