血管紧张素Ⅱ对内皮细胞致凋亡效应及血脂康的保护作用

Proapoptotic Effect of AngiotensinⅡ on Cultured Endothelial Cells and Protective Effect of Xuezhikang

目的观察血管紧张素Ⅱ(AngⅡ)对体外培养的人脐静脉内皮细胞(HUVECs)增殖与凋亡的影响及血脂康对AngⅡ此效应的干预作用,探讨AngⅡ的致动脉粥样硬化作用及血脂康调脂之外的内皮保护作用。方法将体外培养内皮细胞株ECV304进行实验分组:空白对照组;AngⅡ诱导组,培养基中AngⅡ终浓度分别为10-8、10-7、10-6、10-5、10-4mol/L,与细胞共孵育18h。利用三磷酸腺苷生物素发光法(ATP法)检测AngⅡ对HU VECs生长增殖的影响,用电子显微镜观察AngⅡ(10-4mol/L)诱导后内皮细胞的超微结构特点,用流式细胞仪检测AngⅡ作用后内皮细胞凋亡率的变化,采用比色法检验AngⅡ(10-4mol/L)诱导后内皮细胞Caspase3活性的改变;血脂康干预组,培养基中AngⅡ浓度为10-4mol/L,同时加入血脂康500ng/ml与细胞共培养18h,检测该组内皮细胞凋亡率和Caspase3活性的变化。结果与对照组比较,不同浓度(10-8～10-4mol/L)的AngⅡ均能抑制内皮细胞生长增殖(P<0.05),且其作用呈剂量依赖性;不同浓度(10-7～10-4mol/L)的AngⅡ均可显著诱导内皮细胞凋亡(P<0.05),同时Caspase3的活性在AngⅡ(10-4mol/L)作用后明显增加(P<0.01);透射电镜下可见AngⅡ诱导后内皮细胞呈明显凋亡改变;血脂康(500ng/ml)可抑制AngⅡ(10-4mol/L)所诱导的内皮细胞凋亡(P<0.05)。

Objective To clarify whether AngⅡ induce apoptosis of cultured HUVECs,and to investigate the protective effect of Xuezhikang. To elucidate the proatherogenic effect of AngⅡ and the non-lipid mechanisms of Xuezhikang.Methods Cultured ECV304 cells were randomly divided into ①、②and③ groups. Group ①: control ; Group ②: Cells were stimulated with AngⅡ(10 -8、10 -7、10 -6、10 -5、10 -4 mol/L)for 18 h;Group ③: Cells were exposed to AngⅡ(10 -4mol/L) and interfered with Xuezhikang(500 ng/ml) for 18h.The growth inhibition of cells induced by AngⅡ(10 -8、10 -7、10 -6、10 -5、10 -4 mol/L)was assessed by ATP-biolum inesence Assay. The change of apoptosis of HUVECs was observed by electron microscope. The activity of Caspase-3 was assayed by spectrophotometry . Flow cytometry was used to measure the apoptosis ratio induced by AngⅡin different concentrations ,and to determine the effects of Xuezhikang .Results Compared with controls , HUVECs were inhibited by different concentrations (10 -8～10 -4 mol/L)of AngⅡ(P<0.05). These alteration was in a dose-dependent manner;Apoptosis ratio was induced by AngⅡ(10 -7～10 -4 mol/L)with increase of concentrations (P<0.05).The activity of Caspase-3 was significantly higher in the AngⅡ(10 -4 mol/L)treatment group than that in the control group(P<0.01);The change of apoptosis of HUVECs may be observed by electron microscope;A desease in apoptosis cells were observed when Xuezhikang(500 ng/ml)and AngⅡ(10 -4 mol/L) used jointly.Conclusion Apoptosis may be induced by AngⅡand the ratio increases with the rise of concentrations, and Xuezhikang may reduce apoptosis ratio of endothelial cells, which might be helpful to explore the pathogenesis of atherosclerosis and the non-lipid mechanisms of Xuezhikang.