摘要
目的观察松果菊苷(ECH)对过氧化氢(H_2O_2)所致心肌细胞H9c2损伤的保护作用并探讨其机制。方法体外培养H9c2细胞系,采用MTT法检测H_2O_2对细胞存活率的影响,筛选最适作用浓度。将细胞分为5组,空白对照组、H_2O_2模型组(300μmol/L)、松果菊苷低剂量组(5 mg/L)、中剂量组(10 mg/L)、高剂量组(20 mg/L)。采用流式细胞术检测细胞凋亡率和线粒体膜电位(MMP);采用试剂盒检测培养基心肌激酶(CK)、乳酸脱氢酶(LDH)及丙二醛(MDA)释放量;采用蛋白免疫印记(WB)检测细胞质、细胞核中Nrf2蛋白表达水平。采用Nrf2抑制剂鸦胆苦醇预处理心肌细胞,检测细胞凋亡率和线粒体膜电位。结果 H_2O_2最适作用浓度为300μmol/L。与空白对照组相比,H_2O_2模型组心肌细胞凋亡率显著升高(P<0.05),MMP显著降低(P<0.05);与H_2O_2模型组比较,松果菊苷处理组心肌细胞凋亡率明显下降(P<0.05),MMP显著升高(P<0.05)。与空白对照组相比,H_2O_2模型组CK、LDH和MDA释放率明显升高(P<0.05);松果菊苷处理后CK、LDH和MDA释放率显著降低(P<0.05)。与空白对照组相比,H_2O_2模型组Nrf2蛋白核转移水平增加(P<0.05),与H_2O_2模型组比较,松果菊苷处理组Nrf2蛋白核转移水平进一步显著增加(P<0.05)。与松果菊苷处理组比较,鸦胆苦醇处理组心肌细胞凋亡率显著升高(P<0.05),MMP显著降低(P<0.05)。结论松果菊苷可以减轻H_2O_2对心肌细胞造成的损伤,推测其机制是通过促进Nrf2蛋白核移位激活Nrf2通路实现的。
Objective： To explore protective effect and mechanism of echinaeea （ECH） on H9c2 damage in cardiomyocytes induced by hydrogen peroxide（H2O2）. Methods： H9c2 cell line was cultured in vitro,and the in- fluence of H2O2 on cell viability was detected by MTF method,and the optimum concentration was screened. The cells were divided into five groups： the blank control group,H2O2 model group （300 mol/L）,ECH low-dose group （5 rag/L）,mid-dose group（10 mg/L） and high-dose group（20 mg/L）. Apoptosis rate and mitochondrial membrane potential （MMP） were measured by flow cytometry,and cardiac kinase （CK）,lactate dehydrogenase （LDH） and malondialdehyde （MDA） release in culture medium were measured by reagent kit. Nrf2 protein expression level in cytoplasm and nucleus were detected by protein immuno imprinting （WB）. Apoptosis rate and mitochondrial membrane potential were examined by pretreatment of cardiac muscle cell with Nrf2 inhibitor. Results： The opti- mal concentration of H2O2 was 300 mol/L. Compared with that in the blank control group,apoptosis rate of my- ocardial cells in H2O2 model group increased significantly （P〈 0.05 ）, and MMP decreased significantly （P 〈 0.05）. Compared with that in H2O2 model group, apoptotic rate of myocardial cells in ECH treatment group decreased significantly （P 〈 0.05）,and the MMP increased significantly （P 〈 0.05）. Compared with that in blank control group,the release rate of CK, LDH and MDA in H2O2 model group increased significantly （P〈 0.05）,and the rate of CK,LDH and MDA release was significantly lower （P〈 0.05）. Compared with that in blank control group,the nuclear transfer level of Nrf2 protein in H2O2 model group increased （P〈 0.05）. Compared with that in H2O2 model group,the level of nuclear transfer of Nrf2 protein in ECH treatment group significantly increased （P〈 0.05）.Compared with that in ECH treatment group,apoptosis rate of myocardial＇ cells was significantly higher in ECH tr
出处
《中国中医急症》
2017年第12期2078-2082,共5页
Journal of Emergency in Traditional Chinese Medicine
基金
广东省医学科学技术研究基金项目(A2014273)
