摘要
目的 探讨微小RNA-338-3p(miR-338-3p)对表皮生长因子受体酪氨酸激酶抑制剂(EGFR-TKI)耐药肺癌细胞株PC-9/GR中程序性死亡配体1(PD-L1)表达和细胞凋亡的影响及相关机制。方法 体外培养PC-9细胞、PC-9/GR细胞,采用0、0.25、0.50、1.00、2.00、4.00、8.00μmol/L吉非替尼处理确定用药浓度;实时荧光定量聚合酶链反应(qRT-PCR)检测细胞中miR-338-3p表达。将PC-9/GR细胞分为对照组、miR-338-3p NC组(转染miR-338-3p NC)、miR-338-3p组(转染miR-338-3p模拟物)和信号转导和转录激活因子1(STAT1)抑制剂组(转染miR-338-3p模拟物+100μmol/L氟达拉滨)。4组均添加1.00μmol/L吉非替尼,转染后qRT-PCR检测细胞中miR-338-3p表达;MTT法检测细胞增殖;流式细胞仪检测细胞凋亡;蛋白印迹(WB)法STAT1、p-STAT1、PD-L1、Bcl-2、Bax蛋白表达。结果 当吉非替尼浓度升高至1.00μmol/L时,PC-9细胞与PC-9/GR细胞增殖率比较,差异有统计学意义(P<0.05)。与PC-9细胞相比,PC-9/GR细胞中miR-338-3p表达水平降低,差异有统计学意义(P<0.05)。转染后,qRT-PCR结果显示,miR-338-3p组、STAT1抑制剂组的miR-338-3p表达水平均高于miR-338-3p NC组、对照组,差异有统计学意义(P>0.05)。与miR-338-3p NC组相比,miR-338-3p组PC-9/GR细胞增殖率及细胞中PD-L1、Bcl-2蛋白表达水平降低,凋亡率及细胞中p-STAT1/STAT1、Bax蛋白表达水平升高,差异有统计学意义(P<0.05);与miR-338-3p组相比,STAT1抑制剂组PC-9/GR细胞增殖率及细胞中PD-L1、Bcl-2蛋白表达水平升高,凋亡率及细胞中p-STAT1/STAT1、Bax蛋白表达水平降低,差异有统计学意义(P<0.05)。结论 miR-338-3p可抑制EGFR-TKI耐药肺癌细胞株PC-9/GR细胞中PD-L1表达,并诱导细胞凋亡,其作用机制可能与激活STAT1有关。
Objective To investigate the effects and related mechanisms of miR-338-3 p on the expression of programmed death ligand 1(PD-L1) and apoptosis of epidermal growth factor receptor-tyrosine kinase inhibitor(EGFR-TKI)-resistant lung cancer cell line PC-9/GR cells. Methods PC-9 cells and PC-9/GR cells were cultured in vitro, and treated with 0, 0.25, 0.50, 1.00, 2.00, 4.00 and 8.00 μmol/L gefitinib to determine the drug concentration;quantitative real-time polymerase chain reaction(qRT-PCR) method was used to detect the expression of miR-338-3 p in cells. PC-9/GR cells were divided into control group,mi R-338-3 p NC group( transfected with mi R-338-3 p NC),mi R-338-3 p group( transfected with mi R-338-3 p mimic) and signal transducer and activator of transcription 1( STAT1) inhibitor group( transfected mi R-338-3 p mimic + 100 μmol/L fludarabine). All four groups were added with 1. 00 μmol/L gefitinib,and q RT-PCR method was used to detect the expression of mi R-338-3 p in the cells after transfection;MTT method was used to detect cell proliferation;flow cytometry was used to detect cell apoptosis;western blot( WB) method was used to detect the expression of STAT1,p-STAT1,PD-L1,Bcl-2 and Bax proteins. Results When the concentration of gefitinib was increased to 1. 00 μmol/L,the differences in the proliferation rates of PC-9 cells and PC-9/GR cells were statistically significant( P < 0. 05). Compared with PC-9 cells,the expression level of mi R-338-3 p in PC-9/GR cells was significantly reduced( P < 0. 05). The results of q RT-PCR after transfection showed that the expression levels of mi R-338-3 p in the mi R-338-3 p group and the STAT1 inhibitor group were significantly higher than those in the mi R-338-3 p NC group and the control group( P > 0. 05). Compared with the mi R-338-3 p NC group,the PC-9/GR cell proliferation rate,the expression levels of PD-L1 and Bcl-2 proteins in the mi R-338-3 p group were significantly reduced,the apoptosis rate and expression levels of p-STAT1/STAT1 and Bax proteins in cells wer
作者
朱洪宇
史志敏
ZHU Hongyu;SHI Zhimin(Department of Radiotherapy,Suzhou Science and Technology City Hospital Affiliated to Nanjing Medical University,Suzhou,Jiangsu,215010)
出处
《实用临床医药杂志》
CAS
2022年第4期100-105,共6页
Journal of Clinical Medicine in Practice
