摘要
目的:探讨姜黄素改善阿霉素(DOX)诱导的心肌H9c2细胞毒性的作用,并阐明其作用机制。方法:采用DOX处理H9c2细胞建立心肌细胞毒性模型。将H9c2细胞分为正常组、DOX组、DOX+姜黄素组和DOX+姜黄素+沉默信息调节蛋白3(SIRT3)抑制剂3-TYP+DOX组(DOX+姜黄素+3-TYP组)。24 h后观察各组细胞形态表现,CCK-8法检测各组细胞活性,TUNEL染色检测各组细胞凋亡率,酶联免疫吸附试验(ELISA)法检测各组细胞中过氧化氢酶(CAT)和超氧化物歧化酶(SOD)活性及丙二醛(MDA)水平,2,7-二氯荧光素二乙酸酯(DCFH-DA)染色检测各组细胞中活性氧(ROS)水平,Western blotting法检测各组细胞中B细胞淋巴瘤-2(Bcl-2)、Bcl-2相关X蛋白(Bax)、还原型烟酰胺腺嘌呤二核苷酸磷酸(NADPH)氧化酶2(NOX2)、NADPH氧化酶4(NOX4)、SIRT3、乙酰化超氧化物歧化酶2(Ac-SOD2)和超氧化物歧化酶2(SOD2)蛋白表达水平。结果:与对照组比较,DOX组H9c2细胞肿胀,细胞活性降低(P<0.05),细胞中CAT和SOD活性降低(P<0.05),MDA和ROS水平升高(P<0.05),NOX2和NOX4蛋白表达水平升高(P<0.05),Bcl-2/Bax比值和SIRT3蛋白表达水平降低(P<0.05),SOD和Ac-SOD2蛋白表达水平升高(P<0.05);与DOX组比较,DOX+姜黄素组H9c2细胞形态改善,细胞活性升高(P<0.05),细胞中CAT和SOD活性升高(P<0.05),MDA和ROS水平降低(P<0.05),NOX2和NOX4蛋白表达水平降低(P<0.05),Bcl-2/Bax比值和SIRT3蛋白表达水平升高(P<0.05),SOD2和AcSOD2蛋白表达水平降低(P<0.05);与DOX+姜黄素组比较,DOX+姜黄素+3-TYP组细胞肿胀,细胞密度和细胞活性明显降低(P<0.05),细胞凋亡率明显升高(P<0.05),细胞中CAT和SOD活性降低(P<0.05),MDA水平和ROS水平明显升高(P<0.05),NOX2和NOX 4蛋白表达水平升高(P<0.05),Bcl-2/Bax比值和SIRT3蛋白表达水平降低(P<0.05),SOD2和Ac-SOD2蛋白表达水平升高(P<0.05)。结论:姜黄素通过激活SIRT3/SOD2信号抑制氧化应激和细胞凋亡,改善细胞活性,减轻DOX引起的心肌H9c2细胞毒性。
Objective:To discuss the effect of curcumin on ameliorating doxorubicin(DOX)-induced cytotoxicity in the H9c2 cardiomyocytes,and to clarify its mechanism.Methods:The H9c2 cardiomyocytes were treated with DOX to establish the cardiotoxicity model.The H9c2 cells were divided into normal group,DOX group,DOX+curcumin group,and DOX+curcumin+silent information regulator 3(SIRT3)inhibitor-3(3-TYP)group.After 24 h,the morphology of the cells in various groups were observed;CCK-8 method was used to detect the viabilities of the cells in various groups;TUNEL staining was used to detect the apoptotic rates of the cells in various groups;enzyme-linked immunosorbent assay(ELISA)method was used to detect the activities of catalase(CAT)and superoxide dismutase(SOD)and the levels of malondialdehyde(MDA)in the cells in various groups;2,7-dichlorofluorescein diacetate(DCFH-DA)staining was used to detect the levels of reactive oxygen species(ROS)in the cells in various groups;Western blotting method was used to detect the expression levels of B-cell lymphoma 2(Bcl-2),Bcl-2-associated X protein(Bax),nicotinamide adenine dinucleotide phosphate(NADPH)oxidase 2(NOX2),NADPH oxidase 4(NOX4),SIRT3,acetylated superoxide dismutase 2(Ac-SOD2),and superoxide dismutase 2(SOD2)proteins in the cells in various groups.Results:Compared with normal group,the H9c2 cells in DOX group exhibited swelling,the activity of the cells was decreased(P<0.05),the CAT and SOD activities were decreased(P<0.05),the MDA and ROS levels were increased(P<0.05),the expression levels of NOX2,and NOX4 proteins were increased(P<0.05),the ratio of Bcl-2/Bax and the expression level of SIRT3 protein were decreased(P<0.05),and the expression levels of SOD2 and Ac-SOD2 proteins were increased(P<0.05).Compared with DOX group,the H9c2 cells in DOX+curcumin group showed improved morphology,the activity of the cells was increased(P<0.05),the CAT and SOD activities were increased(P<0.05),the MDA and ROS levels were decreased(P<0.05),the expression levels of NOX2,and NOX4 proteins were decreased(P<0.05),the ratio of Bcl-2/Bax and the expression level of SIRT3 protein were increased(P<0.05),and the expression levels of SOD2 and Ac-SOD2 proteins were decreased(P<0.05).Compared with DOX+curcumin group,the cells in DOX+curcumin+3-TYP group exhibited swelling,the activity of the cells was decreased(P<0.05),the apoptotic rate of the cells was significantly increased(P<0.05),the CAT and SOD activities were decreased(P<0.05),the MDA and ROS levels were significantly increased(P<0.05),the expression levels of NOX2 and NOX4 proteins were increased(P<0.05),the ratio of Bcl-2/Bax and the expression level of SIRT3 protein were decreased(P<0.05),and the expression levels of SOD2 and Ac-SOD2 proteins were increased(P<0.05).Conclusion:Curcumin suppresses the oxidative stress and apoptosis by activating the SIRT3/SOD2 signaling pathway,improving the cell activity and alleviating the DOX-induced cytotoxicity in the H9c2 cells.
作者
熊凤梅
蔡玉香
刘卓
孙娜
李洋
XIONG Fengmei;CAI Yuxiang;LIU Zhuo;SUN Na;LI Yang(Department of Pharmacy,Children’s Hospital,Xi’an City,Shaanxi Province,Xi’an 710003,China;Department of Emergency,Children’s Hospital,Xi’an City,Shaanxi Province,Xi’an 710003,China;Institute of Basic Medicine,School of Basic Medical Sciences,Xi’an Medical University,Xi’an 710021,China)
出处
《吉林大学学报（医学版）》
CAS
CSCD
北大核心
2024年第5期1339-1347,共9页
Journal of Jilin University：Medicine Edition
基金
陕西省科技厅重点研发计划项目(2023-YBSF-530)
陕西省西安市儿童医院院级课题资助项目(2020C01)。
关键词
姜黄素
阿霉素
沉默信息调节蛋白
氧化应激
心肌毒性
Curcumin
Doxorubicin
Silent information regulator 3
Oxidative stress
Cardiotoxicity
