摘要
目的探讨培美曲塞(Pem)通过PI3K/AKT通路对人肺腺癌HCC827细胞株PD-L1表达的影响及其可能的作用机制。方法分别采用不同浓度的Pem(0、0.01、0.1、1、10、100μmol/L)在不同时间下(12、24、48h)处理HCC827细胞,CCK-8法检测细胞的增殖抑制作用及确定半数抑制浓度(IC50)作为后续实验的药物浓度,选最佳作用时间作为后续实验的干预时间;流式细胞术检测最佳时间下不同浓度的Pem对PD-L1的表达影响;试验分组:对照组(加入培养液)和试验组(IC50pem与PI3K/AKT通路激活剂740-YP分别单用或联用),Western Blotting检测各组AKT、P-AKT蛋白表达情况;流式细胞术检测最佳时间下各组对HCC827细胞株PD-L1表达的影响。结果与对照组比较,Pem呈浓度依赖性抑制HCC827细胞增殖(P<0.05),各组差异均有统计学意义(均P<0.05),培养曲塞48h组抑制作用强于12 h组(P<0.05),与24h组抑制率无明显差别(P>0.05)。随着药物浓度增加,各实验组细胞PD-L1表达逐渐下降,显著低于对照组(P<0.05);与对照组比较,Pem组可明显下调P-AKT/GAPDH值以及PD-L1的表达(P<0.05),该作用可被740-YP显著逆转(P<0.05),各组AKT/GAPDH值差异均无统计学意义(P>0.05)。与对照组相比,PD-L1表达量在740-YP组上调(P<0.05),在IC50pem组、IC50pem+740-YP组中表达量降低P<0.05),且IC50pem组的PD-L1表达量低于IC50pem+740-YP组(P<0.05)。结论Pem可下调HCC827细胞PD-L1的表达,其作用机制可能与抑制PI3K/AKT信号通路有关。
Objective To investigate the effect of pemetrexed on the expression of PD-L1 in human lung adenocarcinoma cell line hcc827 and its possible mechanism.Methods Hcc827 cells were treated with different concentrations of PEM(0,0.01,0.1,1,10,100μmol/L)at different time(12,24,48 h).CCK-8 method was used to detect the proliferation inhibition of hcc827 cells.The IC50 was determined as the drug concentration in the follow-up experiment,and the optimal action time was selected as the intervention time.Flow cytometry was used to detect the effect of different concentrations of PEM on PD-L1 The expression of the protein was influenced by the expression;Experimental group:control group(adding culture medium)and experimental group(IC50 PEM and PI3K/Akt pathway activator 740y-p were used alone or in combination),The expression of Akt and p-Akt protein in each groμp was detected by Western blotting;the effect of each group on the expression of PD-L1 in hcc827 cell line was detected by flow cytometry.Results Compared with the control group,PEM inhibited the proliferation of hcc827 cells in a concentration dependent manner(P<0.05),and the difference was statistically significant(P<0.05).The inhibitory effect of pemetrexed in 48h group was stronger than that in 12h group(P<0.05),and there was no significant difference between pemetrexed and 24h group.With the increase of drug concentration,the expression of PD-L1 on the cell surface of each experimental group decreased gradually,which was significantly lower than that of the control group(P<0.05).Compared with the control group,the expression of p-Akt/GAPDH in the PEM group was significantly decreased.There was no significant difference in Akt/GAPDH between the two groups(P>0.05).Compared with the control group,the expression of PD-L1 increased in 740y-p group(P<0.05),decreased in IC50 PEM group and IC50 PEM+740y-p group(P<0.05),and the expression of PD-L1 in IC50 PEM group was lower than that in IC50 PEM+740y-p group(P<0.05).Conclusion PEM can down reulate the expression of PD-L1 in hcc827 cells which may be related to the inhibition of PI3k/Akt signaling pathway.
作者
王洁
刘单
邓述恺
WANG Jie;LIU Dan;DENG Shukai(Department of Respiratory and Critical Care Medicine,The Affiliated Hospital of Southwest Medical University,Luzhou 646000,Sichuan,China)
出处
《西部医学》
2021年第9期1290-1295,共6页
Medical Journal of West China
