摘要
目的研究曲格列酮的心肌细胞毒性特征,并从线粒体氧化应激和自噬角度探讨其潜在的毒作用机制。方法人源心肌细胞AC16给予不同浓度曲格列酮0~40μmol·L^-1孵育24 h。倒置显微镜观察细胞形态,CCK-8法检测细胞存活率,漏出法检测乳酸脱氢酶(LDH)释放量;荧光探针TMRM检测线粒体膜电位和CM-H2DCFDA检测全细胞活性氧的含量;Western印迹法检测微管相关蛋白Ⅱ/Ⅰ轻链3(LC3-Ⅱ/LC3-Ⅰ)比值和P62蛋白表达水平。结果与细胞对照组相比,曲格列酮可浓度依赖性地引起细胞质皱缩、细胞存活率下降(r=-0.928,P<0.05)和LDH释放量增加(r=0.746,P<0.05);曲格列酮10和20μmol·L^-1可明显降低细胞线粒体膜电位(P<0.05),增加全细胞活性氧含量(P<0.05);显著增加LC3-Ⅱ/LC3-Ⅰ比值,上调P62蛋白表达(P<0.05)。结论曲格列酮可引起心肌细胞损伤和线粒体功能障碍,其机制与线粒体氧化应激和自噬体降解受阻密切相关。
OBJECTIVE To investigate the effect of troglitazone-induced toxicity,and explore its mechanism related to mitochondrial oxidative stress and autophagy.METHODS AC16 cells were treated with different concentration of troglitazone 0-40μmol·L^-1 for 24 h.The morphological changes were observed under an inverted microscope,cell viability was examined by CCK-8 assay,whereas lactate dehydrogenase(LDH)leakage was estimated by assay kit.The mitochondrial membrane potential and intracellular reactive oxygen species(ROS)were detected by high content analysis using TMRM and CM-H2DCFDA fluorescent probes.The protein expressions of microtubule-associated protein Ⅰ/Ⅱ light chain 3(LC3-Ⅱ/LC3-Ⅰ)ration and P62 were detected by Western blotting.RESULTS Compared with cell control group,troglitazone induced morphological changes,reduction of cell viability(r=-0.928,P<0.05),and increase of LDH leakage(r=0.746,P<0.05)in a concentration-dependent manner.The mitochondrial membrane potential was decreased while intracellular ROS was increased in troglitazone 10 and 20μmol·L^-1 group(P<0.05).Troglitazone was found to significantly increase the ratio of LC3-Ⅱ/LC3-Ⅰ(P<0.05)and up-regulate P62 protein expressions(P<0.05).CONCLUSION Troglitazone can induce cytotoxicity and mitochondrial dysfunction,which is possibly related to mitochondrial oxidative stress and inhibited autophagosome degradation in human cardiomyocytes.
作者
李凤祥
张弛
唐瑶
李宇杰
张林聪
彭双清
郭家彬
康金森
LI Feng-xiang;ZHANG Chi;TANG Yao;LI Yu-jie;ZHANG Lin-cong;PENG Shuang-qing;GUO Jia-bin;KANG Jin-sen(School of Pharmacy,Xinjiang Medical University,Urumqi 830011,China;Center of Disease Prevention and Control,PLA,Beijing 100071,China)
出处
《中国药理学与毒理学杂志》
CAS
北大核心
2020年第1期24-29,共6页
Chinese Journal of Pharmacology and Toxicology
基金
新疆维吾尔自治区自然科学基金面上项目(2015211C006)
北京市科技新星项目(Z171100001117103)
国家自然科学基金(81430090)
联合利华国际合作项目(MA201500410)。
关键词
曲格列酮
心肌细胞
线粒体
氧化应激
自噬
troglitazone
human cardiomyocyte
mitochondria
oxidative stress
autophagy
