黄芪多糖对H_2O_2所致乳鼠心肌细胞氧化应激损伤的影响

Effects of Astraglus Polysaccharides on oxidative stress injury in neonatal rat cadiocytes induced by H_2O_2

目的研究黄芪多糖对H_2O_2所致乳鼠心肌细胞氧化应激损伤的影响,并探讨其可能作用机制。方法分离并培养乳鼠心肌细胞72 h后分为空白对照组、H_2O_2组、黄芪多糖20μmol/L+H_2O_2组、黄芪多糖40μmol/L+H_2O_2组、黄芪多糖80μmol/L+H_2O_2组,每组设10个复孔。每组给予相应干预24 h后,采用MTT法检测细胞存活率,采用紫外-可见分光光度计检测细胞中抗氧化酶[超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)、过氧化氢酶(CAT)]活性,采用氧敏感荧光探针DCFH-DA检测氧自由基(ROS)含量,采用全自动生化分析仪检测培养液中心肌酶[谷草转氨酶(AST)、磷酸肌酸激酶(CPK)、乳酸脱氢酶(LDH)]和丙二醛(MDA)含量,通过流式细胞仪检测细胞凋亡状况并计算凋亡率,采用免疫印迹法检测细胞核因子-κB(NF-κB)蛋白表达情况并进行半定量分析。结果与H_2O_2组比较,黄芪多糖40μmol/L+H_2O_2组和黄芪多糖80μmol/L+H_2O_2组细胞存活率及细胞中SOD、GSH-Px、CAT活性显著升高(P均<0.05),ROS含量显著降低(P<0.05),培养液中AST、CPK、LDH和MDA含量及细胞凋亡率、NF-k B蛋白表达量均显著降低(P均<0.05);黄芪多糖80μmol/L+H_2O_2组细胞存活率及细胞中SOD、GSH-Px活性均显著高于黄芪多糖40μmol/L+H_2O_2组(P均<0.05),CPK、MDA含量及细胞凋亡率、细胞中NF-κB蛋白表达量均显著低于黄芪多糖40μmol/L+H_2O_2组(P均<0.05)。结论黄芪多糖可能通过提高心肌细胞存活率、改善抗氧化酶活性、提高ROS清除能力、降低心肌酶含量、抑制细胞凋亡、下调NF-k B蛋白表达而对H_2O_2诱导乳鼠心肌细胞氧化应激损伤起保护作用,且高剂量效果更好。

Objective It is to investigate the effects of Astraglus Polysaceharides （APS） on oxidative stress injury of neo- natal rat eadiocytes induced by H2 Oz. Methods Cadiocytes of neonate rat was separated and cultivated for 72 hours and then were divided into six groups： normal control group, H202 group, 20 μmol/L APS ＋ H202 group, 40 μmol/L APS ＋ H202 group, 80 txmol/L APS ＋ H202 group, each group had 10 multiple pores. After intervention for twelve hours with respective drugs, the survival rate was detected by MTT; the activity of SOD, CAT, GSH -Px in eardiomyocytes were determined by ul- traviolet-visible spectrophotometer; and the content of ROS was detected by fluorescence probe technique, the content of AST, CPK, LDH and the content of MDA in culture medium were detected by automatic biochemistry analyzer; the cardiomyocytes apoptosis was observed by flow eytometry and the apoptosis rate was calculated; the expression of NF - KB proten was detected by Western blot and Semi-quantitative analysized. Results Compared with H202 group, the survival rate in APS（40, 80 ~mol/L） ＋ H202 groups were significantly increased; the activity of SOD, CAT in cardiomyocytes were significantly increased and the content of ROS were significantly decreased; the content of AST, CPK, LDH and MDA in culture medium were signif- icantly decreased; the cardiomyocytes apoptosis were improved and the apoptosis rate were significantly decreased, the expres- sion of NF - KB protein was significantly down-regulated, all of the difference were significant（ P 〈 0.05, P 〈 0. 01 ）. Conclu- sion APS had inhibitive effects on oxidative stress of neonatal rat cadiocytes induced by H202 , which perhaps related to its effects of improving the activity of antioxidase, enhancing the free radical scavenging ability, down-regulating the expression of NF - ~B protein, depressing apoptosis.