荭草花醇提物对H2O2诱导的H9c2细胞氧化损伤的保护作用机制研究

摘要

研究荭草花醇提物对H9c2心肌细胞氧化损伤的保护作用机制.采用200 μmol/L H2O2作用H9c2细胞0.5h,建立H9c2细胞氧化损伤模型.将细胞分为正常对照组、H9c2细胞氧化损伤模型组、不同浓度荭草花醇提物(20、40、80 μg/mL)预处理组.采用MTS法检测细胞存活率;生化试剂盒检测细胞乳酸脱氢酶(LDH)释放量,细胞内丙二醛(MDA)含量及超氧化物歧化酶(SOD)、过氧化氢酶(CAT)活力;Western blot测定cleaved-caspase-3、Bcl-2、Bax、p-AKT和AKT的表达.与模型组比较,荭草花醇提物能明显增加心肌细胞存活率,降低LDH释放量和MDA含量,提高SOD及CAT活性,并呈剂量依赖性抑制H2O2诱导的氧化应激损伤.荭草花醇提物下调cleaved caspase-3和Bax的蛋白水平,上调Bcl-2的表达,减少心肌细胞凋亡;增加细胞中p-AKT的表达,且这种表达可被PI3 Ks抑制剂LY294002抵消.表明荭草花醇提物可减轻H2O2诱导的H9c2心肌细胞氧化应激损伤,其机制可能与减少细胞凋亡,平衡氧化应激产物有关,且部分依赖于磷脂酰肌醇3-激酶(PI3 K/Akt)通路.%To investigate the protective effect of Polygonum orientale Flower Ethanol Extracton H2O2 induced Oxidative damage in H9c2 Cells.200 μ mol/L H2O2 was used to induce the oxidativestress damage model in H9c2 cells for 0.5 h in vitro.The H9c2 cells were randomly divided into control group,the model group(200 μmol/L H2O2)and the Polygonum orientale Flower ethanol Extract different groups(20、40、80 μg/mL).Cellviability was determined by MTS assay.Thecontentoflactate dehydrogenase (LDH),malondialdehyde (MDA) and the activities of superoxidedimutase (SOD)and catalase (CAT) were detected by biochemical kits.The protein levels of caspase-3,Bax,Bcl-2,p-AKT and AKT were detected by Western blot.Compared with model group,Polygonum orientale Flower Ethanol Extract pre-treatment improved cell viabilityand the activities of SOD and CAT in a dose-dependent manner,and attenuated leakage of LDH and content of MDA.Polygonum orientale Flower Ethanol Extract depressed myocardial apoptosis by down-regulating pro-apoptotic protein cleavedcaspase-3 and Bax,up-regulating apoptosis inhibitory protein Bcl-2 and increasing the protein level of p-AKT.LY294002 could offsets such an effect of Polygonum orientale Flower Ethanol Extract.In summary,Polygonum orientale Flower Ethanol Extract showed a protective effect on H2O2 induced injury in H9c2 cells.This protection may result from inhibiting myocardial oxidative apoptosis,balancing the oxidative stress and maybe related with the PI3K/Akt signaling pathway.