高脂通过线粒体通路诱导H9c2心肌细胞损伤
|
摘要
目的探讨高脂通过线粒体凋亡通路对H9c2心肌细胞的损伤作用。方法棕榈酸(0～0.4 mmol·L~(-1))刺激H9c2心肌细胞24 h和0.2 mmol·L~(-1)棕榈酸刺激H9c2心肌细胞(0～48 h);MTT法评估细胞生长状态;活性氧试剂盒检测细胞内活性氧水平;凋亡试剂盒检测细胞凋亡情况;线粒体膜电位试剂盒检测线粒体膜电位变化;免疫印迹法检测细胞内线粒体凋亡相关蛋白Cyt-C、Cleaved casepase-3、Bax、Bcl-2表达水平。结果棕榈酸刺激H9c2心肌细胞24 h,棕榈酸0.2、0.4 mmol·L~(-1)组细胞增殖率均出现明显下降;细胞内活性氧水平逐渐升高,线粒体膜电位下降,细胞凋亡增加。棕榈酸(0.2 mmol·L~(-1))刺激H9c2心肌细胞24、36、48 h均引起细胞增殖率明显下降。棕榈酸(0.4 mmol·L~(-1))刺激H9c2心肌24 h,线粒体相关蛋白Cyt-C、Cleaved casepase-3、Bax表达均明显增高(P <0. 05),而Bcl-2明显降低(P <0. 05)。结论线粒体凋亡信号通路可能对高脂所致H9c2心肌细胞损伤起重要作用。
Aim To explore the role of mitochondrial apoptosis pathway in high fatty acid induced injury in H9c2 cardiomyocytes. Methods Cardiomyocytes were exposed to different concentrations of palmitic acid(PA) at 0～0.4 mmol·L~(-1) for 24 h and different time points(0～48 h) of PA 0.2 mmol·L~(-1). Cell viability was measured by MTT; the intracellular reactive oxygen species(ROS) was detected by ROS kit; the cells were detected by apoptosis kit; the cell mitochondrial membrane potential changes were detected by mitochondrial membrane potential kit, and the protein expressions of mitochondrial apoptosis pathway such as Cyt-C, Cleaved caspase-3, Bax, and Bcl-2 were detected by Western blot. Results When the cells were stimulated with PA for 24 h, the cell proliferation rates of 0.2 and 0.4 mmol·L~(-1) PA groups significantly decreased. The level of ROS increased gradually, the cell mitochondrial membrane potential decreased and the cell apoptosis increased. When the cells were stimulated with PA(0.2 mmol·L~(-1)) for 24 h, 36 h and 48 h, and all of the cell proliferation rates showed significant decline. Cardiomyocytes exposed to PA(0.4 mmol·L~(-1)) for 24 h showed an increase in the expression of mitochondrial related proteins(Cyt-C, Cleaved caspase-3 and Bax)(P<0.05), while Bcl-2 expression was significantly reduced(P<0.05). Conclusion Mitochondrial apoptosis signaling pathway might play an important role in high fatty acid induced H9c2 myocardial injury.
Aim To explore the role of mitochondrial apoptosis pathway in high fatty acid induced injury in H9c2 cardiomyocytes. Methods Cardiomyocytes were exposed to different concentrations of palmitic acid(PA) at 0～0.4 mmol·L~(-1) for 24 h and different time points(0～48 h) of PA 0.2 mmol·L~(-1). Cell viability was measured by MTT; the intracellular reactive oxygen species(ROS) was detected by ROS kit; the cells were detected by apoptosis kit; the cell mitochondrial membrane potential changes were detected by mitochondrial membrane potential kit, and the protein expressions of mitochondrial apoptosis pathway such as Cyt-C, Cleaved caspase-3, Bax, and Bcl-2 were detected by Western blot. Results When the cells were stimulated with PA for 24 h, the cell proliferation rates of 0.2 and 0.4 mmol·L~(-1) PA groups significantly decreased. The level of ROS increased gradually, the cell mitochondrial membrane potential decreased and the cell apoptosis increased. When the cells were stimulated with PA(0.2 mmol·L~(-1)) for 24 h, 36 h and 48 h, and all of the cell proliferation rates showed significant decline. Cardiomyocytes exposed to PA(0.4 mmol·L~(-1)) for 24 h showed an increase in the expression of mitochondrial related proteins(Cyt-C, Cleaved caspase-3 and Bax)(P<0.05), while Bcl-2 expression was significantly reduced(P<0.05). Conclusion Mitochondrial apoptosis signaling pathway might play an important role in high fatty acid induced H9c2 myocardial injury.
引文
[1] International Diabetes Federation. IDF Diabetes ATLAS (8th edition)[M].Belgium:International Diabetes Federation,Brussels, 2017.
[2] Wang L, Gao P, Zhang M, et al. Prevalence and ethnic pattern of diabetes and prediabetes in China in 2013[J]. JAMA, 2017,317:2515-23.
[3] Wang Z V, Hill J A. Diabetic cardiomyopathy:catabolism driving metabolism[J]. Circulation, 2015,131(9):771-3.
[4] 陈丹,吴基良,李晶.内质网应激信号通路在高脂诱导的心肌细胞损伤中的作用[J].中国药理学通报,2017,33(7):966-71.[4] Chen D,Wu J L,Li J.Role of endoplasmic reticulum stress in high fatty acid induced injury in cardiomyocytes[J].Chin Pharmacol Bull, 2017,33(7):966-71.
[5] Katunga L A, Gudimella P, Efird J T, et al. Obesity in a model of gpx4 haploinsufficiency uncovers a causal role for lipid-derived aldehydes in human metabolic disease and cardiomyopathy[J]. Mol Metab, 2015,4(6):493-506.
[6] 俞牧雨,米日阿依·阿里木江,刘威,等.小檗碱通过抑制脂肪酸摄取降低小鼠原代肝细胞脂质沉积[J].中国药理学通报,2018,34(3):337-42.[6] Yu M Y,Miriayi A J,Liu W, et al. Berberine ameliorates lipid accumulation of mouse primary hepatocytes by decreasing fatty acids uptake[J].Chin Pharmacol Bull,2018,34(3):337-42.
[7] Carpentier A C. Abnormal myocardial dietary fatty acid metabolism and diabetic cardiomyopathy[J]. Can J Cardiol,2018,34(5):605-14.
[8] Li Q, Yu Q, Na R, et al. Omega3 polyunsaturated fatty acids prevent murine dilated cardiomyopathy by reducing oxidative stress and cardiomyocyte apoptosis[J]. Exp Ther Med, 2017,14(6):6152-8.
[9] Liu H, Peng H, Chen S, et al. S1PR2 antagonist protects endothelial cells against high glucose-induced mitochondrial apoptosis through the Akt/GSK-3β signaling pathway[J]. Biochem Biophys Res Commun, 2017,490(3):1119-24.
[10] Chen X, Bian M, Zhang C, et al. Dihydroartemisinin inhibits ER stress-mediated mitochondrial pathway to attenuate hepatocyte lipoapoptosis via blocking the activation of the PI3K/Akt pathway[J]. Biomed Pharmacother, 2018, 97:975-84.
[11] Fan X Y, Liu Y J, Chen K, et al. Organic arsenicals target thioredoxin reductase followed by oxidative stress and mitochondrial dysfunction resulting in apoptosis[J]. Eur J Med Chem, 2018,143:1090-102.
[12] Sun X, Mao Y, Dai P, et al. Mitochondrial dysfunction is involved in the aggravation of periodontitis by diabetes[J]. J Clin Periodontol, 2017, 44:463-71.
[13] Iurlaro R, Muňoz‐Pinedo C. Cell death induced by endoplasmic reticulum stress[J]. FEBS J, 2016,283(14):2640-52.
[2] Wang L, Gao P, Zhang M, et al. Prevalence and ethnic pattern of diabetes and prediabetes in China in 2013[J]. JAMA, 2017,317:2515-23.
[3] Wang Z V, Hill J A. Diabetic cardiomyopathy:catabolism driving metabolism[J]. Circulation, 2015,131(9):771-3.
[4] 陈丹,吴基良,李晶.内质网应激信号通路在高脂诱导的心肌细胞损伤中的作用[J].中国药理学通报,2017,33(7):966-71.[4] Chen D,Wu J L,Li J.Role of endoplasmic reticulum stress in high fatty acid induced injury in cardiomyocytes[J].Chin Pharmacol Bull, 2017,33(7):966-71.
[5] Katunga L A, Gudimella P, Efird J T, et al. Obesity in a model of gpx4 haploinsufficiency uncovers a causal role for lipid-derived aldehydes in human metabolic disease and cardiomyopathy[J]. Mol Metab, 2015,4(6):493-506.
[6] 俞牧雨,米日阿依·阿里木江,刘威,等.小檗碱通过抑制脂肪酸摄取降低小鼠原代肝细胞脂质沉积[J].中国药理学通报,2018,34(3):337-42.[6] Yu M Y,Miriayi A J,Liu W, et al. Berberine ameliorates lipid accumulation of mouse primary hepatocytes by decreasing fatty acids uptake[J].Chin Pharmacol Bull,2018,34(3):337-42.
[7] Carpentier A C. Abnormal myocardial dietary fatty acid metabolism and diabetic cardiomyopathy[J]. Can J Cardiol,2018,34(5):605-14.
[8] Li Q, Yu Q, Na R, et al. Omega3 polyunsaturated fatty acids prevent murine dilated cardiomyopathy by reducing oxidative stress and cardiomyocyte apoptosis[J]. Exp Ther Med, 2017,14(6):6152-8.
[9] Liu H, Peng H, Chen S, et al. S1PR2 antagonist protects endothelial cells against high glucose-induced mitochondrial apoptosis through the Akt/GSK-3β signaling pathway[J]. Biochem Biophys Res Commun, 2017,490(3):1119-24.
[10] Chen X, Bian M, Zhang C, et al. Dihydroartemisinin inhibits ER stress-mediated mitochondrial pathway to attenuate hepatocyte lipoapoptosis via blocking the activation of the PI3K/Akt pathway[J]. Biomed Pharmacother, 2018, 97:975-84.
[11] Fan X Y, Liu Y J, Chen K, et al. Organic arsenicals target thioredoxin reductase followed by oxidative stress and mitochondrial dysfunction resulting in apoptosis[J]. Eur J Med Chem, 2018,143:1090-102.
[12] Sun X, Mao Y, Dai P, et al. Mitochondrial dysfunction is involved in the aggravation of periodontitis by diabetes[J]. J Clin Periodontol, 2017, 44:463-71.
[13] Iurlaro R, Muňoz‐Pinedo C. Cell death induced by endoplasmic reticulum stress[J]. FEBS J, 2016,283(14):2640-52.