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目的:研究黄芪甲苷(astragaloside IV,As IV)对过氧化氢(H2O2)诱导的人脐静脉内皮细胞(HUVECs)保护作用并探讨其可能的作用机制。方法:在37℃,5%CO2,95%空气饱和湿度的培养箱内体外培养HUVECs,将细胞分为正常组,模型组(过氧化氢180μmol·L-1),黄芪甲苷不同剂量(25,50,100μmol·L-1)+过氧化氢(180μmol·L-1)组。MTT法测定各组细胞活力,二氢乙锭原位定性细胞内超氧阴离子含量,Western blot测定细胞内核因子(NF)-κB p65,NF-κB抑制蛋白α亚基(IκB-α),内皮型一氧化氮合酶(e NOS)蛋白表达,酶联免疫吸附测定法(ELISA)检测HUVECs外液肿瘤坏死因子-α(TNF-α)和白细胞介素-6(IL-6)的含量。结果:与正常组比较,模型(H2O2)组细胞活力明显降低,细胞中超氧阴离子明显升高,NF-κ`B p65蛋白表达升高,IκB-α蛋白表达降低,e NOS蛋白表达升高,细胞上清液中TNF-α和IL-6的含量升高(P<0.01)。与模型(H2O2)组相比,黄芪甲苷不同剂量组表现为细胞活力明显升高,细胞中超氧阴离子含量减少,e NOS,p65蛋白表达有不同程度的降低,IκB-α的蛋白含量增加,细胞上清液中TNF-α和IL-6的含量降低,且呈浓度依赖性(P<0.05)。结论:黄芪甲苷能改善H2O2诱导的HUVECs的损伤,其机制可能是通过降低超氧阴离子量,抑制e NOS的解耦联,减少其产生超氧阴离子,从而减轻氧化应激,降低NF-κB和相关炎症因子表达。
Objective: To investigate the protective effect of astragaloside IV (As IV) on human umbilical vein endothelial cells (HUVECs) induced by hydrogen peroxide (H2O2) and to explore its possible mechanism. Methods: HUVECs were cultured in vitro at 37 ℃ with 5% CO2 and 95% air saturated humidity. The cells were divided into normal group, model group (180μmol·L-1 H2O2), different doses of astragaloside , 50,100μmol·L-1) + hydrogen peroxide (180μmol·L-1) group. Cell viability was assayed by MTT assay, and the contents of superoxide anion (ROS) in cells were determined by in-situ ethidium bromide staining. The expressions of NF-κB p65, IκB-α, The expression of eNOS protein and TNF-α and IL-6 in HUVECs were detected by enzyme linked immunosorbent assay (ELISA). Results: Compared with the normal group, the viability of model group (H2O2) was significantly decreased, the superoxide anion concentration was significantly increased, the expression of NF-κB p65 protein was increased, the expression of IκB-α protein was decreased and the expression of e NOS protein was increased, The contents of TNF-α and IL-6 in the cell supernatant increased (P <0.01). Astragaloside at different doses showed significantly higher cell viability, decreased superoxide anion content, decreased eNOS and p65 protein expression, and increased the protein content of IκB-α compared with model group (H2O2) The contents of TNF-α and IL-6 in the cell supernatant decreased in a concentration-dependent manner (P <0.05). CONCLUSION: Astragaloside can improve the H2O2-induced HUVECs injury by reducing the amount of superoxide anion, inhibiting the decoupling of eNOS, reducing the production of superoxide anion, thereby reducing oxidative stress and decreasing the expression of NF-κB And related inflammatory factor expression.