二甲双胍通过抑制p38MAPK信号通路逆转卵巢癌细胞对顺铂的耐药性  

Metformin reverses drug-resistance of ovarian cancer cells to cisplatin by inhibiting p38 mitogen-activated protein kinase signaling pathway

作　　者：彭铮[1] 史惠蓉[1] 谢娅[1] 王静璐 

机构地区：[1]郑州大学第一附属医院妇产科,河南郑州450052

出　　处：《肿瘤》2015年第2期129-138,共10页Tumor

基　　金：国家自然科学基金资助项目(编号:81202070)~~

摘　　要：目的:探讨二甲双胍是否能够逆转卵巢癌细胞对顺铂的耐药性,以及其可能的作用机制。方法:CCK-8法检测二甲双胍作用后卵巢癌细胞顺铂耐药株C13K和CP70细胞对顺铂的耐药性变化,以及转染针对切除修复交叉互补基因1(excision repair cross-complemention 1,ERCC1)的特异性小干扰RNA(small interfering RNA,si RNA)后C13K和CP70细胞耐药性的变化。实时荧光定量PCR法检测不同浓度二甲双胍作用后,C13K和CP70细胞中ERCC1基因表达的变化。蛋白质印迹法检测二甲双胍作用后C13K和CP70细胞中腺苷酸活化蛋白激酶(AMP-activated protein kinase,AMPK)信号通路的激活和p38丝裂原活化蛋白激酶(p38 mitogen-activated protein kinase,p38MAPK)信号通路的抑制情况,以及联合或不联合AMPK信号通路阻断剂Compound C作用后ERCC1蛋白的表达情况。同时,蛋白质印迹法检测p38MAPK信号通路抑制剂SB203580作用后耐药细胞中ERCC1蛋白表达的变化,以及二甲双胍作用联合p38MAPK si RNA转染前后耐药细胞中ERCC1蛋白的表达变化。结果:二甲双胍能够逆转C13K和CP70细胞对顺铂的耐药性(P<0.05)。二甲双胍作用能够降低C13K和CP70细胞中ERCC1 m RNA和蛋白的表达水平(P<0.05),并激活AMPK信号通路(P<0.05);而联合应用AMPK信号通路阻断剂Compound C可以阻断二甲双胍的这种作用(即ERCC1蛋白表达水平升高)(P<0.01)。p38MAPK信号通路抑制剂SB203580作用后,C13K和CP70细胞中ERCC1蛋白表达水平降低(P<0.05);而二甲双胍作用可降低p38MAPK的磷酸化水平(P<0.01),抑制p38MAPK信号通路。二甲双胍作用转染了p38MAPK si RNA的C13K和CP70细胞后,ERCC1蛋白的表达水平无明显变化(P>0.05)。结论:二甲双胍可能逆转卵巢癌顺铂耐药株对顺铂的耐药性,并且这一作用可能是通过抑制p38MAPK信号通路和降低细胞中ERCC1的表达而实现的。Objective： To investigate whether metform cells to cisplatin, and explore its underlying n can reverse drug-resistance of ovarian cancer mechanism. Methods： Cisplatin-resistant ovarian cancer cells C13K and CP70 were used in this study. The small interfering RNAs （siRNAs） targeting excision repair cross-complemention 1 （ERCC1） gene were synthesized and transfected into C13K and CP70 cells, respectively. The change of cisplatin-resistance of C13K and CP70 cells was detected by cell count kit-8 （CCK-8） assay after metformin treatment and ERCC1 siRNA transfection. The expressions of ERCC1 mRNA in C13K and CP70 cells treated with different concentrations of metformin were detected by real-time fluorescent quantitative-PCR. The activation of AMP-activated protein kinase （AMPK） and suppression of p38 mitogen-activated protein kinase （p38MAPK） in C13K and CP70 cells after metformin treatment were evaluated by Western blotting. Then the expression level of ERCC1 protein in C13K and CP70 cells treated with metformin （combined with Compound C, a blocking agent of AMPK signaling pathway, or p38MAPK siRNA transfection） or SB203580 （an inhibitor of p38MAPK signaling pathway） was detected by Western blotting. Results： Metformin could reverse drug-resistance of C13K and CP70 cells to cisplatin （P 〈 0.05）. Metformin reduced the expression levels of ERCC1 mRNA and protein in C13K and CP70 cells （P 〈 0.05） and activated the AMPK signaling pathway （P 〈 0.05）, but this effect of metformin was blocked by AMPK blocking agent Compound C （P 〈 0.01）. The p38MAPK inhibitor SB203580 suppressed the expression of ERCC1 protein in C13K and CP70 cells （P 〈 0.05）; while metformin could suppress the p38MAPK signaling pathway （P 〈 0.01）, but not reduce the expression level of ERCC1 protein in C13K and CP70 cells transfected with p38MAPK siRNA （P 〉 0.05）. Conclusion： Metformin may reverse the drug-resistance of cisplatin-resistant ovarian cancer cells to cisplatin. This

关 键 词：卵巢肿瘤 二甲双胍 抗药性 肿瘤 顺铂 DNA修复 P38丝裂原活化蛋白激酶类 切除修复交叉互补基因1 

分 类 号：R737.31[医药卫生—肿瘤]