绞股蓝皂苷LI与顺铂联用对食管癌EC109细胞生长的抑制作用  

Inhibitory effect of gypenoside LI combined with cisplatin on growth of esophageal cancer EC109 cells

作　　者：赵敏彤 齐彦爽 郝林瑶 纪小桐 朴香兰 ZHAO Min-tong;QI Yan-shuang;HAO Lin-yao;JI Xiao-tong;PIAO Xiang-lan(School of Pharmacy,Minzu University of China,Beijing 100081,China)

机构地区：[1]中央民族大学药学院,北京100081

出　　处：《中国药理学与毒理学杂志》2022年第10期746-753,共8页Chinese Journal of Pharmacology and Toxicology

基　　金：北京市大学生创新训练计划项目(BEIJ2020110030)。

摘　　要：目的研究绞股蓝皂苷LI(Gyp LI)与顺铂(DDP)联用对食管癌EC109细胞存活的抑制作用,以及对细胞凋亡、周期和迁移的影响。方法(1)将Gyp LI 0~200μmol·L^(-1)、DDP 0~800μmol·L^(-1)、Gyp LI3.125μmol·L^(-1)+DDP 0~200μmol·L^(-1)分别作用EC109细胞24 h,MTT法检测细胞存活率并计算半抑制浓度(IC_(50))值,利用CompuSyn软件计算联合指数(CI)。(2)将EC109细胞分为细胞对照组、Gyp LI60μmol·L^(-1)组、DDP 35μmol·L^(-1)组及Gyp LI 3.125μmol·L^(-1)和DDP 10μmol·L^(-1)联用组,给药24 h后,显微镜下观察细胞形态,Annexin V-FITC/PI双染、PI/RNase染色和流式细胞术检测细胞凋亡率和细胞周期,采用划痕实验检测细胞迁移,Western印迹法检测促凋亡因子Bax、细胞色素c、细胞周期蛋白A、细胞周期蛋白D1和基质金属蛋白酶9(MMP-9)蛋白表达。结果(1)Gyp LI和DDP单用组EC109细胞存活率的IC_(50)值分别为(55.34±3.52)μmol·L^(-1)和(37.48±2.99)μmol·L^(-1),Gyp LI 3.125μmol·L^(-1)与不同浓度DDP联用时,IC_(50)值降至(8.05±5.34)μmol·L^(-1),CI值均<1,表明两药联用具有协同作用。(2)Gyp LI与DDP联用时细胞出现生长缓慢、形态不规则、细胞脱落等特征。与细胞对照组比较,Gyp LI 60μmol·L^(-1)组、DDP35μmol·L^(-1)组和联用组的细胞凋亡率从(1.30±0.08)%分别上升至(4.87±0.04)%,(6.44±0.37)%和(9.12±0.20)%(P<0.01),联用组细胞凋亡率比DDP单用组显著升高(P<0.01)。Gyp LI组、DDP组和联用组细胞周期分别阻滞于S期,G_(2)/M期和G_(0)/G_(1)期。与细胞对照组比较,Gyp LI 60μmol·L^(-1)组、DDP 35μmol·L^(-1)组和联用组细胞迁移率从(29.54±5.56)%分别降至(15.30±3.64)%,(13.61±0.06)%和(5.95±0.56)%(P<0.01)。Western印迹法结果显示,与细胞对照组相比,DDP 35μmol·L^(-1)组和联用组促凋亡因子Bax表达水平均显著升高(P<0.01),Gyp LI 60μmol·L^(-1)组、DDP 35μmol·L^(-1)组和联用组细胞色素c表达水平均显著升高(P<0.01),其中联用组Bax和细OBJECTIVE To investigate the inhibitory effect of gypenoside LI(Gyp LI)combined with cisplatin(DDP)on EC109 cells and the effect on cell apoptosis,cycle and migration.METHODS(1)EC109 cells were treated with Gyp LI 0~200μmol·L^(-1),DDP 0~800μmol·L^(-1)and Gyp LI 3.125μmol·L^(-1)+DDP 0~200μmol·L^(-1)for 24 h,respectively.Cell viability was detected by MTT assay to calculate half maximal inhibitory concentration(IC_(50))values.CompuSyn software was used to calculate the combination index(CI).(2)EC109 cels were treated with Gyp LI 60μmol·L^(-1),DDP 35μmol·L^(-1)or Gyp LI 3.125μmol·L^(-1)and DDP 10μmol·L^(-1)combination for 24 h.Cell morphology was observed under a microscope,while the apoptotic rate and cell cycle were investigated by Annexin V-FITC/PI staining,PI/RNase staining and flow cytometry,respectively.The cell migration ability was detected by scratch test.The expression levels of Bax,cytochrome c,cyclin A,cyclin D1 and matrix metalloproteinase-9(MMP-9)were detected by Western blotting.RESULTS(1)The IC_(50)values of Gyp LI and DDP on EC109 cells were55.34±3.52 and(37.48±2.99)μmol·L^(-1),respectively.When Gyp LI 3.125μmol·L^(-1)was combined with DDP of different concentrations,the IC_(50)value was decreased to(8.05±5.34)μmol·L^(-1)and the CI values were<1,suggesting that the combination of the two drugs had a synergistic effect.(2)When Gyp LI was combinated with DDP,the cells grew slowly,shed,showed irregular morphology.Compared with the cell control group,the apoptotic rate was increased from(1.30±0.08)%to(4.87±0.04)%,(6.44±0.37)%and(9.12±0.20)%,respectively(P<0.01).The apoptotic rate of the combination group was significantly higher than that of the DDP group(P<0.01).Gyp LI,DDP and the combination group arrested cell cycle at S,G2/M and G0/G1phase,respectively.Compared with the cell control group,the cell migration rates of Gyp LI,DDP and the combination group was decreased from(29.54±5.56)%to(15.30±3.64)%,(13.61±0.06)%and(5.95±0.56)%,respectively(P<0.01).Western blotting re

关 键 词：绞股蓝皂苷LI EC109细胞 顺铂 联合用药 细胞凋亡 细胞周期 

分 类 号：R285.5[医药卫生—中药学] R979.1[医药卫生—中医学]