CTRP6对阿霉素诱导的心肌细胞损伤的保护作用  

Protective Effect of CTRP6 on Doxorubicin-induced Cardiomyocyte Injury

作　　者：黄荣 孔春燕 沈鐲钰 黄丹[1] 赵淑红 马振国[1] HUANG Rong;KONG Chunyan;SHEN Zhuoyu(Department of Cardiology,Renmin Hospital of Wuhan University,Hubei Key Laboratory of Metabolism and Chronic Diseases,Hubei 430060,China)

机构地区：[1]武汉大学人民医院心内科、代谢与相关慢病湖北省重点实验室,430060

出　　处：《医学研究杂志》2024年第6期23-28,共6页Journal of Medical Research

基　　金：国家自然科学基金青年科学基金资助项目(81700254);湖北省青年拔尖人才项目。

摘　　要：目的探讨C1q/肿瘤坏死因子相关蛋白6(C1q/tumour necrosis factor-related protein 6,CTRP6)对阿霉素(doxorubicin,DOX)诱导的心肌细胞凋亡及氧化应激损伤的影响及其相关作用机制。方法建立H9C2心肌细胞损伤模型,设置对照组(NS组)、DOX(1μmol/L)组和DOX(1μmol/L)+浓度梯度CTRP6组(CTRP6浓度分别为0.5、1、3、5μg/ml),培养24h后检测心肌细胞生存率,结果表明,CTRP6浓度为3μg/ml时,心肌细胞的生存率提高最显著,因此后续实验CTRP6浓度选择3μg/ml。后续H9C2心肌细胞实验分组为对照组(NS组)、CTRP6组、DOX(1μmol/L)组和DOX(1μmol/L)+CTRP6(3μg/ml)组。荧光实时定量聚合酶链反应(real-time quantitative polymerase chain reaction,RT-qPCR)和Western blot法检测DOX刺激后CTRP6转录及翻译水平改变,Tunel法检测细胞凋亡水平,使用酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)试剂盒检测细胞谷胱甘肽过氧化物酶(glutathione peroxidase,GSH-Px)活性、超氧化物歧化酶(superoxide dismutase,SOD)活性和丙二醛(malondialdehyde,MDA)含量,Western blot法检测脂联素受体1(adiponectin receptor 1,AdipoR1)蛋白水平改变,检测蛋白激酶B/糖原合成酶激酶-3(protein kinase B/glycogen synthase kinase-3β,Akt/GSK-3β)信号通路蛋白表达改变情况。结果与对照组(NC组)比较,DOX组细胞CTRP6蛋白表达水平及mRNA水平显著降低(分别降低46.26%及67.74%,P<0.05);与DOX组比较,DOX+CTRP6组CTRP6蛋白表达水平显著提高(P<0.05);加入不同浓度梯度的CTRP6蛋白,其中DOX+3μg/ml CTRP6处理组心肌细胞存活率显著提高了26.48%(P<0.05);Tunel染色显示凋亡减少,Bcl-2表达升高;抗氧化分子GSH-Px及SOD活性分别升高20.49%及36.89%,MDA水平受到显著抑制(降低47.09%,P<0.05);AdipoR1蛋白水平显著提高(P<0.05);Akt/GSK-3β信号通路蛋白磷酸化水平显著升高。结论CTRP6改善DOX诱导的心肌细胞凋亡及氧化应激损伤,可能是通过Akt/GSK-3β信号通路发挥的保护作用。Objective To explore the impact of C1q/tumour necrosis factor-related protein 6(CTRP6)on doxorubicin(DOX)-induced apoptosis and oxidative stress injury of cardiomyocytes,along with its associated mechanism of action.Methods The H9C2 cardiomyocyte injury model was established by dividing the subjects into different groups.These groups included the control group(NS group),the DOX(1μmol/L)group,and the DOX(1μmol/L)group supplemented with CTRP6 at concentration gradients of 0.5μg/ml,1μg/ml,3μg/ml,and 5μg/ml,respectively.After 24hours of culture,the survival rate of the cardiomyocytes was measured,the results showed that the highest increase in cardiomyocyte survival rate was observed at a CTRP6 concentration of 3μg/ml,which was selected as the optimal concentration.Subsequently,the experimental groups of H9C2 cardiomyocytes consisted of the control group(NS group),the CTRP6 group,the DOX(1μmol/L)group,and the DOX(1μmol/L)group supplemented with CTRP6(3μg/ml)group.The impact of DOX stimulation on the transcriptional and translational levels of CTRP6 was assessed using fluorescence real-time quantitative polymerase chain reaction(RT-qPCR)and Western blot assay.Apoptosis levels were measured using Tunel assay,while enzyme-linked immunosorbent assay(ELISA)kits were was used to detect the activity of cellular total glutathione peroxidase(GSH-Px)and superoxide dismutase(SOD),as well as the levels of malondialdehyde(MDA).Western blot assay was used to detect the any alterations in the levels of AdipoR1 protein and the protein expression of the protein kinase B/glycogen synthase kinase-3β(Akt/GSK-3β)signaling pathway.Results Compared with the control group(NS group),the expression levels of CTRP6 protein and mRNA were significantly decreased in the DOX group(46.26%and 67.74%reduction,respectively,P<0.05).In contrast,compared with DOX group,the DOX+CTRP6group showed a significant increase in CTRP6 protein expression levels(P<0.05).Among the different concentrations of CTRP6 added to the DOX+CTRP6group,the survival

关 键 词：C1q/肿瘤坏死因子相关蛋白6 凋亡 氧化应激 阿霉素 

分 类 号：R541[医药卫生—心血管疾病]