摘要
目的:探讨黄芪注射液对血管紧张素Ⅱ(AngⅡ)诱导的H9c2心肌细胞凋亡的抑制作用及其可能作用机制。方法:采用细胞计数试剂盒-8(CCK-8)方法筛选H9c2细胞培养的黄芪注射液浓度;H9c2细胞分为正常对照组、AngⅡ模型组、AngⅡ+黄芪注射液组。采用实时定量聚合酶链式反应(RT-PCR)法检测3组H9c2细胞电压依赖性阴离子通道1(VDAC1)、线粒体融合蛋白2(Mfn2)及分子伴侣葡萄糖调节蛋白75(GRP75)mRNA的表达;蛋白免疫印迹法(Western Blot)检测VDAC1、Mfn2、GRP75的蛋白表达;Fura-2 AM法测定细胞内Ca^(2+)浓度变化;用流式细胞仪检测各组细胞凋亡情况。结果:黄芪注射液在浓度为0.125%时对细胞的促增殖作用最为显著。与正常对照组比较,AngⅡ模型组Mfn2、GRP75 mRNA表达水平明显增加(P<0.05),AngⅡ+黄芪注射液组Mfn2、GRP75 mRNA表达低于AngⅡ模型组(P<0.05或P<0.01),而3组VDAC1 mRNA表达比较差异无统计学意义(P>0.05)。与正常对照组比较,AngⅡ模型组VDAC1、Mfn2和GRP75的蛋白表达明显增加(P<0.05或P<0.01),AngⅡ+黄芪注射液组VDAC1、Mfn2、GRP75的蛋白表达低于AngⅡ模型组(P<0.01)。与正常对照组比较,AngⅡ模型组细胞钙含量、细胞凋亡率明显增加(P<0.05或P<0.01),AngⅡ+黄芪注射液组细胞钙含量、细胞凋亡率低于AngⅡ模型组(P<0.01)。结论:黄芪注射液能够抑制血管紧张素Ⅱ诱导的H9c2心肌细胞凋亡,其机制可能与其调节VDAC1、Mfn2和GRP75等MAM结构蛋白表达和影响细胞内钙平衡有关。
Objective:To study the inhibitory effect of Astragalus Injection on H9c2 cardiomyocyte apoptosis induced by angiotensinⅡ(AngⅡ)and its possible mechanism.Methods:The CCK-8 method was used to screen the concentration of Astragalus Injection in H9c2 cell culture.H9c2 cells were divided into normal control group,AngⅡmodel group,and AngⅡ+Astragalus Injection group.Reverse transcription-polymerase chain reaction(RT-PCR)was used to detect the mRNA expression of voltage dependent anion channel 1(VDAC1),mitofusin 2(Mfn2),and glucose regulated protein 75(GRP75)in H9c2 cells.The protein expressions of VDAC1,Mfn2,and GRP75 were detected by Western Blot.The intracellular Ca^(2+)concentration was measured by Fura-2 AM method.Cell apoptosis was detected by flow cytometry.Results:The cell proliferation promoting effect was the most significant when the concentration of Astragalus Injection was 0.125%.Compared with control group,the mRNA expression levels of Mfn2 and GRP75 in AngⅡmodel group were significantly increased(P<0.05),and the mRNA expressions of Mfn2 and GRP75 in AngⅡ+Astragalus Injection group were lower than those in AngⅡmodel group(P<0.05 or P<0.01),but there was no significant difference in the mRNA expression level of VDAC1 in each group(P>0.05).Compared with normal control group,the protein expressions of VDAC1,Mfn2,and GRP75 in AngⅡmodel group were significantly increased(P<0.05 or P<0.01).The protein expressions of VDAC1,Mfn2,and GRP75 in AngⅡ+Astragalus Injection group were lower than those in AngⅡmodel group(P<0.01).Compared with normal control group,the calcium content and apoptosis rate of cells in AngⅡmodel group were significantly increased(P<0.05 or P<0.01).The calcium content and apoptosis rate of AngⅡ+Astragalus Injection group were lower than those of AngⅡmodel group(P<0.01).Conclusion:Astragalus Injection can inhibit the apoptosis of H9c2 cardiomyocytes induced by AngⅡ,and its mechanism may be related to the regulation of the expression of MAM structural proteins such as VDAC1,Mfn2,and GRP75,and the influence of intracellular calcium balance.
作者
程炜婷
张婷
金秋硕
马喆
吴爱明
薛程元
高永红
聂波
赵明镜
娄利霞
CHENG Weiting;ZHANG Ting;JIN Qiushuo;MA Zhe;WU Aiming;XUE Chengyuan;GAO Yonghong;NIE Bo;ZHAO Mingjing;LOU Lixia(Chinese Internal Medicine Laboratory of Ministry of Education and Beijing,Dongzhimen Hospital,Beijing University of Chinese Medicine Department of Cardiology,Beijing 100700,China)
出处
《中西医结合心脑血管病杂志》
2023年第1期40-45,共6页
Chinese Journal of Integrative Medicine on Cardio-Cerebrovascular Disease
基金
北京中医药大学新奥奖励基金课题(No.2017-XA-JLJJ-012)。
