肉豆蔻-8散提取物对过氧化氢诱导心肌细胞损伤的影响

Effects of Roudoukou-8 San against hydrogen peroxide-induced injury of cardiomyocyte

研究肉豆蔻-8散提取物对过氧化氢(H_2O_2)诱导乳鼠心肌细胞损伤的影响,并探讨其作用机制。分离并培养大鼠乳鼠心肌细胞,建立H_2O_2诱导心肌细胞损伤模型。倒置显微镜下观察肉豆蔻-8散提取物对心肌细胞形态学的影响;采用MTT法检测心肌细胞活力;使用全自动生化分析仪检测细胞培养液中乳酸脱氢酶(LDH)、磷酸肌酸激酶(CK)、天冬氨酸氨基转移酶(AST)的含量;采用试剂盒法检测细胞中丙二醛(MDA)、超氧化物歧化酶(SOD)、一氧化氮(NO)的含量;采用Hoechst荧光细胞核染色观察心肌细胞凋亡形态;使用流式细胞仪测定心肌细胞凋亡率。100μmol/L H_2O_2作用2 h能明显造成约50%心肌细胞损伤。H_2O_2模型组出现细胞间隙增大、细胞数减少、细胞胞浆空泡等明显损伤现象。与H_2O_2模型组比较,肉豆蔻-8散提取物各剂量组,心肌细胞形态不同程度好转;能明显降低H_2O_2损伤心肌细胞培养液中LDH、CK、AST含量;显著降低H_2O_2损伤心肌细胞内MDA和NO含量,并提高SOD活力,能明显抑制H_2O_2损伤心肌细胞凋亡。通过改善细胞生存状态、提高细胞活力、降低氧化应激损伤、抑制炎症反应、抑制细胞凋亡,肉豆蔻-8散提取物可改善H_2O_2诱导心肌细胞损伤状态,从而对H_2O_2损伤的心肌细胞起到保护作用。

To study the effects of Roudoukou-8 San against hydrogen peroxide-induced cardiomyocyte in neonatal rats and to explore its mechanism.Cardiomyocytes were isolated and cultivated by neonatal rats and the injure models were established by H 2O 2.Methyl thiazolyl tetrazolium(MTT)assay was used to detect the protective effects of Roudoukou-8 San on H 2O 2-induced cardiomyocyte.The effects of Roudoukou-8 San on myocardium morphology were observed under inverted microscope.The contents of lactate dehydrogenase(LDH),creatine kinase(CK)and aspartate aminotransferase(AST)in cell culture medium were determined by Automatic biochemical instrument;The levels of malonydialdehyde(MDA),superoxide dismutase(SOD)and NO in the cells were detected by kit method.The apoptotic morphology of cardiomyocytes was observed by Hoechst fluorescence staining.Cell apoptosis were measured by Annexin V and PI double staining and flow cytomety.100μmol/L H 2O 2 for 2 h could cause about 50%of myocardial injury.In the inverted optical microscope,H 2O 2 model group showed increased cell gap,decreased cell count,cell cytoplasmic vacuoles and other obvious damage.Roudoukou-8 San protected cell from H 2O 2-induced morphlogical improved in different degrees,reduced the release of LDH,CK and AST content,reduced the content of MDA,NO in myocardial cells significantly and increased the activity of SOD.Roudoukou-8 San energy significantly inhibited H 2O 2 damage myocardial cell apoptosis.Our study suggested that Roudoukou-8 San can protect cardiomyocyte from H 2O 2-induced injury by improving the cell viability,reducing oxidative stress injury,inhibiting inflammatory reaction and inhibiting cell apoptosis.