曲美他嗪对棕榈酸诱导的H9C2心肌细胞脂毒性凋亡的影响  

The effects of trimetazidine on palmiticacid-induced lipotoxic apoptosis in H9C2 cardiomyocytes

作　　者：王凯[1] 张子钊 刘彤[1] WANG Kai;ZHANG Zizhao;LIU Tong(Department of Cardiology,The Second Hospital of Tianjin Medical University,Tianjin 300211,China)

机构地区：[1]天津医科大学第二医院心脏内科,天津300211

出　　处：《天津医科大学学报》2025年第1期30-35,共6页Journal of Tianjin Medical University

基　　金：天津市医学重点学科(专科)建设项目(TJYXZDX-029A)。

摘　　要：目的:建立棕榈酸(PA)诱导的H9C2细胞凋亡模型,验证曲美他嗪(TMZ)是否对H9C2细胞脂毒性凋亡产生保护作用及具体机制。方法:在体外培养的H9C2心肌细胞,当细胞密度为90%时分别给予不同时间(0、6、12、24 h)、不同浓度(0、50、100、150、200、300、350μmol/L)的PA刺激(n=3),建立脂毒性损伤模型,用CCK8检测细胞活性确定最佳刺激条件,蛋白印迹法证实细胞凋亡。实验共分6组,分别为对照组(CON)、PA(200μmol/L)组、CON+TMZ3(10μmol/L)组、PA(200μmol/L)+TMZ1(0.1μmol/L)组、PA(200μmol/L)+TMZ2(1μmol/L)组、PA(200μmol/L)+TMZ3(10μmol/L)组(n=3),用CCK8检测细胞活性、蛋白印迹测定凋亡相关蛋白表达量。后续为观察细胞内脂质含量变化,实验分为4组,分别为对照组(CON)、PA(200μmol/L)组、TMZ3(10μmol/L)组、PA(200μmol/L)+TMZ3组(n=3),通过油红O染色镜下观察。结果:与CON组相比,200μmol/L PA组及12 h组细胞活力明显下降(t=15.8、20.82,均P<0.05),使用CCK8检测并最终确定PA 200μmol/L、刺激12 h的条件下,H9C2细胞脂毒性损伤较大、存活细胞最多。与CON组相比,200μmol/L PA组Bax蛋白表达明显升高(t=3.201,P<0.05),150μmol/L PA组Bcl-2蛋白表达明显降低(t=2.479,P<0.05),蛋白检测证实细胞凋亡。与PA刺激组相比,PA+TMZ1组、PA+TMZ2组、PA+TMZ3组的细胞活力升高(F=420.1,均P<0.05)、PA+TMZ2组、PA+TMZ3组凋亡相关蛋白Bcl-2表达升高(t=6.028、8.952,均P<0.05)、Bax表达下降(t=3.392、4.275,均P<0.05),随着TMZ浓度的增加,Bax表达降低、Bcl-2表达增加(F=3.763、7.548,均P<0.05)。与CON组相比,PA组H9C2细胞内脂滴含量明显增多。与PA组相比,PA+TMZ3组H9C2细胞内脂滴含量明显减少。结论:棕榈酸刺激可诱导H9C2心肌细胞脂毒性凋亡,曲美他嗪可抑制棕榈酸诱导的H9C2细胞脂毒性凋亡。Objective:To establish a palmitic acid(PA)-induced apoptosis model of H9C2 cells to verify whether trimetazidine(TMZ)had a protective effect on lipotoxic apoptosis in H9C2 cells and its specific mechanism.Methods:H9C2 cardiomyocytes cultured in vitro were stimulated with PA for different time(0,6,12,24 h)and different concentrations(0,50,100,150,200,300,350μmol/L)(n=3)when the cell density was 90%to establish a lipotoxic injury model.CCK8 was used to detect cell viability to determine the optimal stimulation conditions,and Western blotting was used to confirm cell apoptosis.There were 6 groups in the experiment,including control group(CON),PA(200μmol/L),CON+TMZ3(10μmol/L),PA(200μmol/L)+TMZ1(0.1μmol/L),PA(200μmol/L)+TMZ2(1μmol/L),and PA(200μmol/L)+TMZ3(10μmol/L)(n=3).CCK8 was used to detect cell viability and Western blotting was used to determine the expression of apoptosis-related proteins.To observe the changes in intracellular lipid content,the experiment was divided into 4 groups,including control group(CON),PA(200μmol/L),TMZ3(10μmol/L),and PA(200μmol/L)+TMZ3(n=3),which were observed under the microscope by Oil Red O staining.Results:Compared with the CON group,the cell viability in the 200μmol/L PA group and the 12 h group was significantly decreased(t=15.8,20.82,both P<0.05).CCK8 was used to detect and finally determine that under the conditions of PA 200μmol/L and 12 h stimulation,H9C2 cells suffered greater lipotoxic damage and the most surviving cells.Compared with the CON group,the expression of Bax protein in the 200μmol/L PA group was significantly increased(t=3.201,P<0.05),and the expression of Bcl-2 protein in the 150μmol/L PA group was significantly decreased(t=2.479,P<0.05).Protein detection confirmed cell apoptosis.Compared with the PA stimulation group,the cell activity of the PA+TMZ1 group,PA+TMZ2 group,and PA+TMZ3 group increased(F=420.1,all P<0.05),the expression of apoptosis-related protein Bcl-2 increased(t=6.028,8.952,both P<0.05)and the expression of Bax decreased(t=3.392,

关 键 词：棕榈酸 凋亡 曲美他嗪 BAX/BCL-2 

分 类 号：R541.9[医药卫生—心血管疾病]