川芎嗪对大鼠心肌细胞氧糖剥夺/复氧损伤的保护作用及机制研究

摘要

目的 研究川芎嗪(tetramethylpyrazine)对心肌缺血再灌注损伤的保护作用,并探究其作用机制.方法 原代培养大鼠心肌细胞,建立心肌细胞氧糖剥夺/复氧损伤模型:MTT方法 测定细胞存活率;紫外分光光度法测定细胞培养基上清液中LDH、CK、NO水平和心肌细胞内SOD、MDA水平;Western blot方法 检测细胞内eNOS和iNOS蛋白的表达水平.结果 川芎嗪能够显著改善大鼠心肌细胞氧糖剥夺/复氧损伤心肌细胞搏动频率,降低LDH、CK及NO释放水平,抑制MDA表达,提高SOD活性,下调细胞内iNOS蛋白表达水平,上调eNOS蛋白表达水平.结论 川芎嗪能够显著抑制氧化应激反应,从而增强心肌细胞对于过量氧自由基损伤的耐受能力,保持心肌细胞膜的完整性和保护心肌组织的正常生理功能.川芎嗪能够平衡eNOS/iNOS两种蛋白的表达,维持体内NO正常浓度,防止过量NO所造成的心肌组织损伤.%Objective This study explored the protective effect of tetramethylpyrazine on myocardial ischemia/reperfusion injury via inhibiting oxidative stress. Methods Primary cultured neonatal myocytes were applied to explore the anti-ischemia/reperfusion injury property in vitro. The survival viability of myocytes was determined by MTT; enzyme activities such as lactate dehydrogenase, creatine kinase, superoxide dismutase, malondialdehyde, and nitric oxide were analyzed with assay kits; inducible nitricoxide synthase and endothelial nitric oxide synthase expressions were determined by Westernblot. Results Tetramethylpyrazine significantly improved the beating frequencies of myocytes after oxygen-glucose deprivation/reoxygenation procedure, decreased lactate dehydrogenase, creatine kinase, and malondialdehyde levels and enhanced superoxide dismutase activity.Tetramethylpyrazine also inhibited excessive production of nitric oxide through downregulating inducible nitric oxide synthase as well as upregulating endothelial nitric oxide synthase during ischemia/reperfusion injury. Conclusion Tetramethylpyrazine could significantly improve the oxidative-stress tolerance of myocytes to keep cell membrane integrity and protect the myocardial tissue of normal physiological function via an antioxidant effect and by restoring the balance between inducible nitric oxide synthase and endothelial nitric oxide synthase, while inhibiting the generation of cytotoxic concentrations of NO.