小凹蛋白1在血管紧张素Ⅱ诱导人脐静脉内皮细胞衰老中的变化

Caveolin-1 Expression Changes in Senescent Human Umbilical Vein Endothelial Cells Induced by Angiotensin Ⅱ

目的观察小凹蛋白1(caveolin-1,Cav-1)在血管紧张素Ⅱ(angiotensinⅡ,AngⅡ)诱导的人脐静脉内皮细胞(human umbilical vein endothelial cells,HUVECs)衰老中的变化。方法体外培养HUVECs,采用CCK-8法检测细胞存活率,分为对照组、AngⅡ诱导组(用终浓度为10-6 mol/L的AngⅡ干预)。以衰老相关β-半乳糖苷酶活性和细胞的增殖能力两种衰老标志物为主要观察指标。采用免疫化学染色方法检测衰老相关β-半乳糖苷酶的活性,流式细胞术检测细胞周期来反映细胞的增殖能力;利用Western印迹法分析AngⅡ诱导HUVECs 0、12、24、36、48h的Cav-1表达的时间效应关系。结果与对照组相比,10-6 mol/L AngⅡ诱导组存活的细胞数为对照组的(77.15±6.83)%;(81.80±0.92)%的细胞呈现β-半乳糖苷酶阳性染色,流式细胞仪检测细胞周期停滞于G0-G1[(89.4±3.4)%],证实细胞衰老;AngⅡ呈时间依赖性上调Cav-1表达。结论 AngⅡ可以诱导HUVECs衰老,其分子机制可能与上调衰老细胞Cav-1表达有关。

Objective To investigate the caveolin-1 expression changes in senescent human umbilical vein endothelial cells（HUVECs） induced by angiotensin Ⅱ（AngⅡ）.Methods HUVECs were cultured in vitro and the cell viability was observed by CCK-8.HUVECs were divided into control group and AngⅡ group（stimulated with AngⅡ with 10-6mol/L as the final concentration at 48 hours）.Two aging markers,i.e,β-galactosidase activity and cell proliferation activity were adopted as main observation indexes.β-galactosidase activity was measured by immunocytochemical method and cell proliferation activity was measured by flowcytometry.The expression of caveolin-1 was determined by Western bloting at 0,12,24,36,and 48 hours after AngⅡ intervention.Results The cell viability in the AngⅡ group was（77.15%±6.83）% of that in the control group.In AngⅡ group（81.80%±0.92）% cells had positive β-galactosidase staining,significantly higher than that in the control group,and the cell cycle was remained at G0/G1 [（89.4%±3.4）%].AngⅡ up-regulated the expression of caveolin-1 protein of HUVECs with time dependency.Conclusion AngⅡ can induce HUVEC senescence and the molecular mechanisms may relate with the up-regulated expressions of caveolin-1 protein.