顺铂通过激活细胞外信号调节激酶(ERK)通路促进A549人肺腺癌细胞PD-L1表达

Cisplatin promotes PD-L1 expression in A549 human lung adenocarcinoma cells via activating the ERK pathway

目的探讨顺铂(DDP)对人肺腺癌A549细胞的程序性死亡蛋白1配体1(PD-L1)表达的影响及其可能的机制。方法体外培养人肺腺癌A549细胞,采用(0、0.5、1、2、4、8)mg/L DDP处理24、36、48 h,CCK-8法检测细胞增殖抑制率,并计算其半数抑制浓度(IC_(50)),选择最适抑制时间48 h及其IC_(50)用于后续实验。后续将A549细胞分为4组:空白对照组、IC_(50)的DDP组、20μmol/L的丝裂原活化蛋白激酶激酶抑制剂PD98059组、IC_(50)的DDP联合20μmol/L PD98059组。培养48 h,Western blot法检测细胞外信号调节激酶(ERK)、磷酸化的ERK(p-ERK)蛋白表达,流式细胞术检测PD-L1表达。结果与对照组相比,DDP处理的A549细胞PD-L1表达均上调,且呈剂量依赖性,选择最适作用时间点为48h,其IC_(50)值为3.586 mg/L。与对照组相比,DDP处理组p-ERK、PD-L1表达增加,PD98059组p-ERK表达降低;DDP联合PD98059组较DDP组的p-ERK、PD-L1的表达降低,较PD98059组的p-ERK、PD-L1表达增加。结论DDP激活ERK信号通路上调A549细胞PD-L1表达。

Objective To investigate the effect of cisplatin(DDP)on the expression of programmed death 1 ligand 1(PD-L1)in human lung adenocarcinoma A549 cells and its possible mechanism.Methods Human lung adenocarcinoma A549 cells were cultured in vitro and treated with(0,0.5,1,2,4,8)mg/L of DDP for 24,36,48 hours.CCK-8 assay was used to detect the cell proliferation inhibition rate and the half maximal inhibitory concentration(IC_(50))was calculated.The optimal inhibition time of 48 hours and its IC_(50)were selected for subsequent experiments.A549 cells were then randomized into four groups:blank control group,group with DDP(IC_(50)),group with 20μmol/L of mitogen-activated protein kinase kinase inhibitor PD98059,and group with DDP(IC_(50))combined with 20μmol/L of PD98059.The cells were cultured for 48 hours.The expression of ERK and p-ERK were detected by Western blotting,and PD-L1 expression was detected by flow cytometry.Results Compared with that in the control group,the expression of PD-L1 in A549 cells treated with DDP was up-regulated in a dose-dependent manner.The optimal time was 48 hours and the IC_(50)was 3.586 mg/L.Also compared with that in the control group,the expression of p-ERK and PD-L1 in the DDP treatment group increased,and the expression of p-ERK in the group with PD98059 decreased.The expressions of p-ERK and PD-L1 in the group with DDP combined with PD98059 were lower than those in the group with DDP,but higher than those in the group with PD98059.Conclusion DDP up-regulates the expression of PD-L1 in A549 cells by activating the ERK signaling pathway.