摘要
目的探讨姜黄素对Hep G2肝癌细胞系增殖和凋亡的影响及机制。方法体外培养Hep G2肝癌细胞系,加入终浓度5、10、20μg/m L的姜黄素培养细胞24、48 h,观察细胞生长状态,并利用CCK-8试剂盒检测细胞增殖情况,选择最佳药物浓度和作用时间,再分为对照组、姜黄素组以及二甲基亚砜(DMSO)组进行实验。利用DCFH-DA荧光染色检测各组细胞ROS水平,通过流式细胞仪分析各组细胞凋亡情况,Western blot检测各组细胞Cleaved caspase-3、细胞色素C(cytochrome C,Cyt C)、Yes相关蛋白(yes-associated protein,YAP)的表达情况。结果 CCK-8试剂盒检测显示,5、10、20μg/m L姜黄素培养细胞24 h,细胞增殖抑制率分别为(15.9±1.3%)、(49.7±1.8)%、(63.7±1.7)%;培养细胞48 h后增殖抑制率分别为(27.2±1.1)%、(63.3±2.0)%、(70.5±2.3)%,本研究选取10μg/m L姜黄素作用48 h进行后续实验。与对照组相比,姜黄素组细胞ROS水平明显增加,凋亡率增加到26.74%,凋亡相关蛋白Cleaved caspase-3、Cyt C明显上调,YAP蛋白含量显著降低(P<0.05);而DMSO组各项指标均无明显变化(P>0.05)。结论姜黄素能抑制Hep G2肝癌细胞系的增殖,诱导凋亡,其机制可能与姜黄素下调YAP表达,进而促进氧化应激有关。
Objective To investigate the effect and curcumin on proliferation and apoptosis of HepG2 cell lines and its mechanism.Methods HepG2 cells were cultured in vitr o at 37℃with curcumin of different final concentrations(5,10,20μg/mL)for 24 and 48 hours,with the growing status of HepG2 cells observed.CCK-8 kit was used to detect cell proliferation.The following tests were implemented in the control group,curcumin group and DMSO group after determining the optimal drug concentration and action time.DCFH-DA fluorescence staining was adopted to detect the intracellular ROS level.The apoptosis status of cells was demonstrated via the flow cytometry(FCM).Western blot analysis was performed to analyze the expression of Cleaved caspase-3,Cytochrome C(Cyt C)and Yes-associated protein(YAP).Results CCK-8 assay indicated that the cellular proliferation inhibition rates were(15.9±1.3%)%,(49.7±1.8)%,(63.7±1.7)%respectively when cells were cultured in 5,10,20μg/mL of curcumin for 24 h,and(27.2±1.1)%,(63.3±2.0)%,(70.5±2.3)%respectively for 48 h.Therefore,cells cultured in 10μg/mL of curcumin for 48 h were selected to proceed the follow-up experiments.Compared with the control group,in curcumin group,ROS level markedly increased,apoptosis rate increased to 26.74%,and the expression level of proteins associated with apoptosis including Cleaved caspase-3 and Cyt C was up-regulated obviously,while the content of YAP was down-regulated remarkably(P<0.05).However,the results manifested that no significant changes of ROS level,apoptosis rate and protein expression were observed in the DMSO group(P>0.05).Conclusion Curcumin can inhibit the proliferation of HepG2 cell line and induce apoptosis,and its mechanism may be related to curcumin s down-regulating the expression of YAP,thereby promoting oxidative stress.
作者
于洪丹
张舒文
谷学静
Yu Hongdan;Zhang Shuwen;Gu Xuejing(Liaoning Province Key Laboratory of Diabetic Perception Dysfunction,Jinzhou Medical University;Department of gastroenterology,the First Affiliated Hospital of Jinzhou Medical University;Department of Human Anatomy,Jinzhou Medical University,Jinzhou 121000 China)
出处
《锦州医科大学学报》
CAS
2020年第3期13-18,共6页
Journal of Jinzhou Medical University
基金
辽宁省自然科学基金,资助项目编号:2019-ZD-0807。
