摘要
目的:探讨miRNA-338(miR-338)通过靶向调控谷胱甘肽过氧化物酶4(GPX4)表达在非小细胞肺癌(non-small cell lung cancer,NSCLC)增殖中的作用及机制研究。方法:通过实时定量聚合酶链式反应(qRT-PCR)检测非小细胞肺癌组织、癌旁、细胞系及对照细胞系中miR-338及GPX4 mRNA的表达水平;通过Western Blot检测非小细胞肺癌组织、癌旁、细胞系及对照细胞系中GPX4蛋白水平;通过荧光素酶报告基因时间验证miR-338直接靶向调节GPX4的表达;通过CCK-8探索miR-338是否通过调节铁死亡影响肿瘤细胞增殖;通过试剂盒检测细胞脂质氧化和活性氧水平。结果:NSCLC组织和细胞系中miR-338表达水平低于癌旁组织和对照细胞系;NSCLC组织和细胞系中GPX4 mRNA及蛋白表达水平高于癌旁组织和对照细胞系;Starbase软件分析发现GPX4 mRNA序列中含有miR-338特异作用位点,荧光素酶报告基因实验结果证实miR-338直接靶向调节GPX4表达;过表达miR-338提高肿瘤细胞中脂质氧化及活性氧水平,并抑制肿瘤细胞增殖;而铁死亡抑制剂预处理可以逆转miR-338的抑癌作用。结论:miR-338通过负向调控GPX4表达进而促进肿瘤细胞铁死亡,最终抑制NSCLC细胞增殖。
Objective:To explore the effect and molecular mechanism of miR-338 on proliferation ability of non-small cell lung cancer(NSCLC)by targeting GPX4.Methods:The expression of miR-338 and GPX4 mRNA level in NSCLC or control tissues and cell lines was detected by qRT-PCR.Western blot was used to explore the protein expression of GPX4 in NSCLC or control tissues and cell lines.Luciferase reporter assay was employed to verify that miR-338 directly targeted and negatively modulated GPX4.CCK8 assay was used to explore the effect of miR-338 on proliferation ability of NSCLC cells.ROS and lipid oxidation assay kit were utilized to measure the ROS and lipid oxidation level.Results:miR-338 was obviously decreased in NSCLC tissue and cell lines compared with normal tissues and cell line.The mRNA and protein level of GPX4 was highly expressed in NSCLC tissues and cell lines compared with normal tissue and cell line.Analysis in Starbase software indicated that the mRNA sequence of GPX4 harbored a potential binding motif of miR-338.Luciferase reporter assay result showed that miR-338 directly bound to and regulated GPX4.Functionally,overexpression of miR-338 induced the increase of ROS and lipid oxidation level which suppressed the proliferation of NSCLC cells.While pretreatment with ferroptosis inhibitor reversed the inhibitory effect of miR-338.Conclusion:miR-338 promoted the ferroptosis and inhibited the proliferation ability of NSCLC cells via negatively regulating GPX4 expression.
作者
张伟
彭燕琪
费思佳
刘洋
黄盼
田艳妮
石静
王继钊
梁华
何建军
许静
ZHANG Wei;PENG Yanqi;FEI Sijia;LIU Yang;HUANG Pan;TIAN Yanni;SHI Jing;WANG Jizhao;LIANG Hua;HE Jianjun;XU Jing(Department of Breast Surgery,the First Affiliated Hospital of Xi'an Jiaotong University,Shanxi Xi'an 710061,China;Department of Geriatric Endocrinology,the First Affiliated Hospital of Xi'an Jiaotong University,Shanxi Xi'an 710061,China;Department of Thoracic Surgery,the First Affiliated Hospital of Xi'an Jiaotong University,Shanxi Xi'an 710061,China;Department of Pathology,the First Affiliated Hospital of Xi'an Jiaotong University,Shanxi Xi'an 710061,China)
出处
《现代肿瘤医学》
CAS
2020年第23期4041-4045,共5页
Journal of Modern Oncology
基金
陕西省自然科学基础研究计划一般项目(面上)(编号:2020JM-393)
西安交通大学第一附属医院教改项目(编号:JG20190312)。
