Effect And Mechanism Of Angiotensin(1-7) On Cultured Human Umbilical Vein Endothelial Cells Oxitadive Stress Induced By Angiotensin Ⅱ

Objective:This experiment establish the oxidative stress model by angiotensin-Ⅱinducing human umbilical vein endothelial cells (HUVECs),and explore the influence and mechanism of Ang-(1-7) on oxidative stress injury induced by Ang-Ⅱto HUVECs.Methods:We culture HUVECs in vitro, the 2 to 5 generation cells were used in our study. Experiment was divided into:(1) control group:no intervention factor; (2) Ang-(1-7) Group:add 10-6mol/L of Ang-(1-7); (3) A-779 Group:join the 10-6mol/L of the A-779; (4) AngⅡgroup: add 10-6mol/L of AngⅡ; (5) AngⅡ+Ang-(1-7) Group:Add 10-6mol/L of AngⅡon the basis of, different concentrations of Ang-(1-7) (10-6～10-9mol/L); (6) AngⅡ+Ang-(1-7)+ A-779 Group:a 10-6mol/L of A-779 pretreatment, after 30min, and then a final concentration of 10-6mol/LAng-(1-7) pretreatment 30min, finally adding a final concentration of 10-6mol/LAngⅡ. Cells were collected 16 hours after the intervention and the culture medium. Cells by flow cytometry the fluorescence intensity of ROS, using xanthine oxidase method of superoxide dismutase (SOD) of the activity, with thiobarbituric acid (Thibabituric Acid TBA) Determination of MDA content.Results1. Cell morphology:fluorescent microscope, AngⅡcells compared with the control group markedly enhanced green fluorescence intensity, in AngⅡbasis of the Ang-(1-7), the green fluorescence intensity compared with AngⅡsignificantly reduced compared, Ang-(1-7) and A-779 group compare with the control group showed no significant difference.2. Flow cytometry results of flow cytometry analysis:Ang-(1-7) (10-6 mol/L) and A-779 (10-6 mol/L) group compared with control group (38.9±3.5,39.2±2.8 and 39.3±2.2, P> 0.05), not significant, suggesting that individual to join Ang-(1-7) or A-779 on endothelial cells did not affect ROS; AngⅡ(10-6 mol/L) group compared with the control group (98.9±4.5 and 39.3±2.2, P<0.05), prompted AngⅡinduced increase in intracellular ROS generation; AngⅡ+ Ang-(1-7) group compared with AngⅡgroup (43.7±2.3 and 98.9±4.5, P<0.05), prompted Ang-(1-7) inhibited AngⅡrole in promoting cell ROS generation; AngⅡ+Ang-(1-7)+ A-779 group compared with AngⅡ+Ang-(1-7) group (78.2±4.3 and 43.7±2.3, P<0.05), prompted the role of Ang-(1-7) is blocked by A-779, Ang-(1-7) on the antagonism of AngⅡ MAS may be mediated;the different concentration of Ang-(1-7) groups (77.5±2.7,68.1±2.5,55.2±3.2,43.7±2.3,P<0.05), prompted Ang-(1-7) in a dose-dependent inhibition of AngⅡin promoting the role of ROS generation.3. Ang-(1-7) (10-6 mol/L) and A-779 (10-6 mol/L) group compared with control group (34.2±1.23,34.8±1.03,34.5±1.27,P> 0.05), no statistically significant, suggesting that individual to join Ang-(1-7) or A-779 on endothelial cells did not affect SOD activity; AngⅡ(10-6 mol/L) group compared with control group (12.4±1.01 and 34.5±1.27,P<0.05), AngⅡinduced cells prompted SOD activity decreased; AngⅡ+Ang-(1-7) group compared with AngⅡgroup (30.9±0.79 and 12.4±1.01.P<0.05), prompted Ang-(1-7) inhibited Ang II SOD activity decrease of the role of induced cells; AngⅡ+Ang-(1-7)+A-779 group compared with AngⅡ+Ang-(1-7) group (19.2±0.87 and 30.9±0.79,P< 0.05), suggesting that the role of Ang-(1-7) was blocked by A-779, Ang-(1-7) on the antagonism of AngⅡMAS may be mediated; the different concentrations Ang-(1-7)(17.3±0.78,22.9±0.87,27.3±0.67,30.9±0.79,P<0.05), prompted Ang-(1-7) in a dose-dependent inhibition of AngⅡSOD activity decrease caused by the role of intracellular.4.The content of MDA:the Ang-(1-7) (10-6 mol/L) or the A-779 (10-6 mol/L) group compared with control group (11.02±0.33,10.09±0.41 and 10.08±0.32, P> 0.05), no statistically significant, suggesting that individual to join Ang-(1-7) or A-779 on endothelial cells did not affect MDA formation; AngⅡ(10-6 mol/L) group compared with control group (45.9±0.59,10.08±0.32,P<0.05), AngⅡprompted increased production of endothelial cells MDA; AngⅡ+Ang-(1-7) group compared with AngⅡgroup (18.9±0.43,45.9±0.59,P<0.05), prompted Ang-(1-7) inhibited AngⅡin promoting the role of endothelial cells MDA formation; AngⅡ+Ang-(1-7)+A-779 group compared with AngⅡ+Ang-(1-7) group (44.8±0.62,18.9±0.43,P< 0.05), suggesting that the role of Ang-(1-7) was blocked by A-779, Ang-(1-7) on the antagonism of AngⅡMAS may be mediated;the different concentrations of Ang-(1-7) (38.5±0.54,32.1±0.51,24.8±0.47,17.9±0.43,P<0.05), prompted Ang-(1-7) in a dose-dependent inhibition of MDA formation induced by AngⅡin the role.Conclusion:1. Ang II can induce human umbilical vein endothelial cells (HUVECs) oxidative stress.2. Ang-(1-7) in a dose-dependent inhibition of Ang II induced oxidative stress in HUVECs3. A-779 can block Ang-(1-7) inhibited Ang II induced oxidative stress in HUVECs, suggesting that Ang-(1-7) the role is mediated by specific Mas...