AngⅡ诱导人脐静脉内皮细胞建立高血压损伤模型

Establishment of Angiotensin Ⅱ-Induced Model in Human Umbilical Vein Endothelial Cells

目的:采用高血压发病因子血管紧张素Ⅱ(angiotensinⅡ,AngⅡ)诱导人脐静脉内皮细胞(human umbilical vein endothelial cells,HUVECs)建立高血压损伤模型。方法:使用不同浓度(0.01、0.10、1.00μmol/L和10.00μmol/L)AngⅡ诱导HUVECs不同时间(3、6、9、12 h和24 h),通过考察HUVECs形态、活性、功能、微观结构及凋亡程度来评价HUVECs损伤程度,同时采用交叉设计方差分析和多重比较方法得到AngⅡ最适诱导浓度和最佳诱导时间。结果:AngⅡ呈浓度依赖式诱导HUVECs损伤,最佳诱导条件为1.00μmol/L AngⅡ诱导12 h,此时HUVECs活性为44.85%、NO含量为43.57μmol/L、总一氧化氮合酶活力为6.99 U/mg pro、内皮型一氧化氮合酶活力为1.89 U/mg pro、丙二醛含量为7.46 nmol/mL、超氧化物歧化酶活力为27.29 U/mg pro、细胞凋亡率为41.5%,微观结构显示核膜皱缩,核仁消失,胞浆内出现大量空泡状结构,线粒体或细小或肿胀,粗面内质网扩张数目较少,部分核糖体丢失,平面内质网扩张,但细胞未解体,仍然保持细胞结构。结论:1.00μmol/L AngⅡ诱导12 h可以成功诱导HUVECs建立高血压损伤模型。

Objective:This research was aimed to establish a hypertensive damage model in human umbilical vein endothelialcells(HUVECs)induced by angiotensin(Ang)Ⅱ.Methods:HUVECs were induced with different concentrations(0.01,0.10,1.00and10.00μmol/L)of AngⅡfor different times(3,6,9,12and24h),respectively.The injury degree of HUVECswas evaluated by cellular morphology,viability,function,microstructure,and apoptosis.The optimal concentration and timewere obtained by using variance analysis and multiple comparison tests.Results:AngⅡinduced injury in HUVECs in adose-and time-dependent manner.The optimal AngⅡconcentration and induction time were respectively1.00μmol/L,and12h.After12h induction by1μmol/L AngⅡinduced,the cell viability was44.85%,nitric monoxide(NO)content was43.57μmol/L,total nitric oxide synthase(TNOS)activity was6.99U/mg pro,endothelial nitric oxide synthase(eNOS)activity was1.89U/mg pro,malonaldehyde(MDA)content was7.46nmol/mL,superoxide dismutase(SOD)activity was27.29U/mg pro,and apoptosis percentage was41.5%.The microstructure showed that the nuclear membrane shrivel andnucleolus disappeared,and a large number of air bubbles appeared inside the cytoplasm.Small and swollen mitochondria,only a small number of expanded rough endoplasmic reticula,partial loss of ribosome,and endoplasmic reticulum expansionwere observed,but the cells did not collapse,and still maintained their normal structure.Conclusion:Induction with1.00μmol/L AngⅡfor12h allows successful establishment of a hypertensive damage model in HUVECs.