CircCDR1as调节miR-671-5p/CBX4轴对NSCLC细胞生物学行为的影响  

Effects of CircCDR1as on the biological behavior of NSCLC cells by regulating the miR-671-5p/CBX4 axis

作　　者：杜文峰[1] 周玲 李琪[1] DU Wenfeng;ZHOU Ling;LI Qi(Department of Respiratory and Critical Care,Puren Hospital Affiliated to Wuhan University of Science and Technology,Wuhan 430081,China)

机构地区：[1]武汉科技大学附属普仁医院呼吸与危重症科,430081

出　　处：《天津医药》2023年第6期573-579,共7页Tianjin Medical Journal

基　　金：2020年度中国金属学会冶金安全与健康分会健康卫生科研项目(jkws202005)。

摘　　要：目的探讨环状RNA(CircRNA)反义小脑变性相关蛋白1转录本(CircCDR1as)通过调节miR-671-5p/染色盒同源物4(CBX4)轴对非小细胞肺癌(NSCLC)细胞增殖、凋亡、迁移和侵袭的影响。方法体外培养人NSCLC细胞株(NCI-H524、NCI-H1734、Calu-3和A549)和人支气管上皮样细胞(HBE)。A549细胞分成Control组、si-NC组、si-CircCDR1as组、si-CircCDR1as+anti-miR-NC组、si-CircCDR1as+anti-miR-671-5p组、pcDNA组、CircCDR1as组、miR-NC组、miR-671-5p组、miR-671-5p+pcDNA组、miR-671-5p+CBX4组、anti-miR-NC组和anti-miR-671-5p组。采用实时定量聚合酶链反应(qRT-PCR)检测CircCDR1as、miR-671-5p和CBX4 mRNA表达。Western blot检测CBX4蛋白表达。分别采用四甲基偶氮唑盐(MTT)、克隆形成实验、流式细胞术和Transwell检测细胞活力、增殖、凋亡、迁移和侵袭。通过双荧光素酶报告基因检测验证miR-671-5p与CircCDR1as或CBX4之间的靶向关系。结果与HBE细胞相比,CircCDR1as和CBX4在4种NSCLC细胞中表达升高,miR-671-5p表达降低(P<0.05)。与si-NC组比较,si-CircCDR1as组A549细胞活力和克隆形成数显著降低,细胞迁移和侵袭数目减少,凋亡率增高(P<0.05)。miR-671-5p作为CircCDR1as的靶点,其下调减弱了CircCDR1as沉默对NSCLC进展的调控作用(P<0.05)。miR-671-5p靶向CBX4在体外抑制NSCLC的恶性进展(P<0.05)。沉默CircCDR1as可通过上调miR-671-5p水平降低CBX4的表达(P<0.05)。结论CircCDR1as沉默可通过上调miR-671-5p和下调CBX4表达来抑制细胞增殖、迁移和侵袭,诱导细胞凋亡。Objective To investigate effects of CircRNA antisense cerebellar degeneration-related protein 1 transcript(CircCDR1as)on the proliferation,apoptosis,migration and invasion of non-small cell lung cancer(NSCLC)cells by regulating miR-671-5p/chromobox 4(CBX4)axis.Methods Human NSCLC cell lines(NCI-H524,NCI-H1734,Calu-3 and A549)and human bronchial epithelioid cells(HBE)were cultured in vitro.A549 cells were randomly separated into the control group,the si-NC group,the si-CircCDR1as group,the si-CircCDR1as+anti-miR-NC group,the si-CircCDR1as+anti-miR-671-5p group,the pcDNA group,the CircCDR1as group,the miR-NC group,the miR-671-5p group,the miR-671-5p+pcDNA group,the miR-671-5p+CBX4 group,the anti-miR-NC group and the anti-miR-671-5p group.The expression levels of CircCDR1as,miR-671-5p and CBX4 mRNA were measured by quantitative real-time polymerase chain reaction(qRT-PCR).The protein expression of CBX4 was detected by Western blot assay.The cell viability,proliferation,apoptosis,migration and invasion were detected by tetramethylazolium salt(MTT),clonogenic assay,flow cytometry and Transwell,respectively.The targeting relationship between miR-671-5p and CircCDR1as or CBX4 was verified by dual-luciferase reporter assay.Results Compared with HBE cells,expression levels of CircCDR1as and CBX4 were increased and the expression of miR-671-5p was decreased in the four NSCLC cell groups(P<0.05).Compared with the si-NC group,the viability and the number of colonies of A549 cells were obviously decreased in the si-CircCDR1as group,the numbers of cell migration and invasion were decreased,and the apoptosis rate was increased(P<0.05).The down regulation of miR-671-5p,a target of CircCDR1as,attenuated the regulatory effect of CircCDR1as silencing on NSCLC progression(P<0.05).miR-671-5p targeting CBX4 inhibited the malignant progression of NSCLC in vitro(P<0.05).The down regulation of CircCDR1as could reduce the expression of CBX4 by up-regulating the level of miR-671-5p(P<0.05).Conclusion CircCDR1as silencing can inhibit cell p

关 键 词：癌 非小细胞肺 A549细胞 CircRNA反义小脑变性相关蛋白1转录本 miR-671-5p 染色盒同源物4 

分 类 号：R734.2[医药卫生—肿瘤]