灯盏乙素对血管内皮细胞抗氧化损伤作用及其分子机制探讨
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摘要
目的:研究灯盏乙素对血管内皮细胞(vascular endothelial cells,VEC)抗氧化损伤作用,并从分子水平探讨其作用机制。
方法:以过氧化氢(H_2O_2)诱导HUVEC损伤,体外模拟自由基损伤模型,采用MTT法以细胞存活率为指标,从细胞水平研究灯盏乙素对氧化损伤的HUVEC的保护作用,用原位分子杂交方法测定H_2O_2损伤HUVEC后超氧化物歧化酶(superoxide dismutase,SOD)、NADPH氧化酶(NADPH oxidase,NOX)的mRNA表达;用western blotting方法测定H_2O_2损伤HUVEC后SOD、NOX的蛋白表达。
结果:1 mmol/L H_2O_2作用4 h,可引起血管内皮细胞损伤,与正常对照组比较差异有统计学意义(P＜0.05);灯盏乙素可上调H_2O_2诱导的体外培养血管内皮细胞抗氧化酶SOD的mRNA及蛋白表达,并呈浓度依赖性,与模型组比较差异有统计学意义(P＜0.05)。灯盏乙素呈浓度依赖性下调H_2O_2诱导的体外培养血管内皮细胞氧化酶NOX的mRNA及蛋白表达,与模型组比较差异有统计学意义(P＜0.05)。
结论:灯盏乙素对H_2O_2氧化损伤的血管内皮细胞具有保护作用,其机制可能与提高血管内皮细胞抗氧化能力、下调氧化酶NOX的mRNA及蛋白表达和上调抗氧化酶SOD的mRNA及蛋白表达有关。
Objective:To investigate the protective effects of scutellarin on vascular endothelial cell injury induced by oxidative damage and to claim its mechanisms at the molecular level.
Methods:Peroxide hydrogen(H_2O_2) was used to induce injury of human umbilical vein endothelial cells(HUVEC) to duplicate the in-vitro free radical-injured model. The MTT assay was used to evaluate the cell survival rate in order to explore the protective effect of scutellarin against injury of vascular endothelial cells.On the above basis,the mRNA expression level of superoxide dismutase(SOD) and NADPH oxidase(NOX) was detected by the method of molucular hybridization in situ.The protein expression of SOD and NOX was assayed by the method of western blotting.
Results:After incubated with 1 mmol/L H_2O_2 for 4 h,the viability of HUVEC significantly decreased(P＜0.05 vs normal control group).Scutellarin could concentration-dependently up-regulate the expression of SOD in HUVEC injuried by H_2O_2(P＜0.05 vs model group);scutellarin could concentration-dependently down-regulate the expression of NOX in HUVEC injuried by H_2O_2(P＜0.05 vs model group).
Conclusion:Scutellarin showed obvious protective effects against vascular endothelial cells induced by oxidative stress and the probable mechanisms may be closely associated with its promoting the capability of antioxidases,down-regulating the expression of NOX protein and mRNA and up-regulating the expression of SOD protein and mRNA.
方法:以过氧化氢(H_2O_2)诱导HUVEC损伤,体外模拟自由基损伤模型,采用MTT法以细胞存活率为指标,从细胞水平研究灯盏乙素对氧化损伤的HUVEC的保护作用,用原位分子杂交方法测定H_2O_2损伤HUVEC后超氧化物歧化酶(superoxide dismutase,SOD)、NADPH氧化酶(NADPH oxidase,NOX)的mRNA表达;用western blotting方法测定H_2O_2损伤HUVEC后SOD、NOX的蛋白表达。
结果:1 mmol/L H_2O_2作用4 h,可引起血管内皮细胞损伤,与正常对照组比较差异有统计学意义(P＜0.05);灯盏乙素可上调H_2O_2诱导的体外培养血管内皮细胞抗氧化酶SOD的mRNA及蛋白表达,并呈浓度依赖性,与模型组比较差异有统计学意义(P＜0.05)。灯盏乙素呈浓度依赖性下调H_2O_2诱导的体外培养血管内皮细胞氧化酶NOX的mRNA及蛋白表达,与模型组比较差异有统计学意义(P＜0.05)。
结论:灯盏乙素对H_2O_2氧化损伤的血管内皮细胞具有保护作用,其机制可能与提高血管内皮细胞抗氧化能力、下调氧化酶NOX的mRNA及蛋白表达和上调抗氧化酶SOD的mRNA及蛋白表达有关。
Objective:To investigate the protective effects of scutellarin on vascular endothelial cell injury induced by oxidative damage and to claim its mechanisms at the molecular level.
Methods:Peroxide hydrogen(H_2O_2) was used to induce injury of human umbilical vein endothelial cells(HUVEC) to duplicate the in-vitro free radical-injured model. The MTT assay was used to evaluate the cell survival rate in order to explore the protective effect of scutellarin against injury of vascular endothelial cells.On the above basis,the mRNA expression level of superoxide dismutase(SOD) and NADPH oxidase(NOX) was detected by the method of molucular hybridization in situ.The protein expression of SOD and NOX was assayed by the method of western blotting.
Results:After incubated with 1 mmol/L H_2O_2 for 4 h,the viability of HUVEC significantly decreased(P＜0.05 vs normal control group).Scutellarin could concentration-dependently up-regulate the expression of SOD in HUVEC injuried by H_2O_2(P＜0.05 vs model group);scutellarin could concentration-dependently down-regulate the expression of NOX in HUVEC injuried by H_2O_2(P＜0.05 vs model group).
Conclusion:Scutellarin showed obvious protective effects against vascular endothelial cells induced by oxidative stress and the probable mechanisms may be closely associated with its promoting the capability of antioxidases,down-regulating the expression of NOX protein and mRNA and up-regulating the expression of SOD protein and mRNA.
引文
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