摘要
目的探讨沉默真核翻译起始因子5A-2(eIF5A-2)对人肺癌细胞株A549/DDP耐药及自噬的影响。方法体外培养人肺癌细胞株A549、A549/DDP,转染细胞A549/DDP,细胞分为eIF5A-2-SiRNA组、阴性对照组(NC组)及空白对照组,另以A549细胞为正常对照组。病毒感染72 h后,采用实时荧光定量PCR(qRT-PCR)检测细胞中eIF5A-2 mRNA表达水平,MTT法检测细胞DDP作用下A549/DDP细胞的耐药性,流式细胞术检测细胞的凋亡率,单酰戊二胺(MDC)染色检测细胞自噬水平,蛋白免疫印记(Western blot)法检测eIF5A-2、LC3Ⅱ及Beclin-1蛋白相对表达水平。结果与正常对照组相比,空白对照组、NC组A549/DDP细胞eIF5A-2 mRNA相对表达水平显著升高(均P<0.05);与空白对照组、NC组相比,eIF5A-2-SiRNA组细胞凋亡率显著升高,细胞IC50、自噬小体数目、eIF5A-2、LC3Ⅱ及Beclin-1蛋白相对表达水平显著降低(均P<0.05)。结论eIF5A-2下调可降低人肺癌细胞株A549/DDP耐药性,可能与下调LC3Ⅱ、Beclin-1表达,抑制自噬作用有关。
Objective To investigate the effects of silencing eukaryotic translation initiation factor 5A-2(eIF5A-2)on drug resistance and autophagy of human lung cancer cell line A549/DDP.Methods Human lung cancer cell lines A549 and A549/DDP were cultured in vitro.The A549/DDP cells were transfected,and divided into eIF5A-2-SiRNA group,negative control group and blank control group,A549 cells were used as the control group.The expression level of eIF5A-2 mRNA in cells was detected by real-time fluorescent quantitative PCR(qRT-PCR).The drug resistance of A549/DDP cells was detected by MTT.The apoptosis rate was detected by flow cytometry.The autophagy was detected by monodansyl cadaverine(MDC)staining.The relative expression levels of eIF5A-2,LC3Ⅱand Beclin-1 proteins were detected by Western blot.Results Compared with the normal control group,the relative expression levels of eIF5A-2 mRNA in A549/DDP cells in the blank control group and NC group were significantly higher(P<0.05).Compared with the control group and NC group,the apoptosis rate of eIF5A-2-SiRNA group was significantly higher(P<0.05),the IC50,number of autophagy bodies,relative expression levels of eIF5A-2,LC3Ⅱand Beclin-1 proteins decreased significantly(P<0.05).Conclusion The down-regulation of eIF5A-2 can reduce the drug resistance of human lung cancer cell line A549/DDP,which may be related to the down-regulation of LC3Ⅱand Beclin-1 expressions and the inhibition of autophagy.
作者
李星
刘东升
杜钢
LI Xing;LIU Dongsheng;DU Gang(Department of Radiotherapy, Bayannaoer Hospital, Bayan Nur 015000, Inner Mongolia, China)
出处
《西部医学》
2021年第4期493-497,共5页
Medical Journal of West China
基金
内蒙古自治区卫计委科研计划项目(201701118)。
关键词
真核翻译起始因子5A-2
肺癌
耐药
自噬
Eukaryotic translation initiation factor 5A-2
Lung cancer
Drug resistance
Autophagy
