长链非编码RNA基因CCAT2抑制TGF-β信号通路促进非小细胞肺癌细胞的增殖  

Long non-coding RNA CCAT2 promotes proliferation of non-small cell lung cancer cells by inhibiting transforming growth factor-β pathway

作　　者：张婧婧 化瑞芳 姬颖华[2] 张敏[2] 李伟伟[2] 杨庆辉[2] 王瑾 于海川 路平[2] 王向鹏 

机构地区：[1]河南省新乡医学院医学检验学院和分子诊断与医学检验技术河南省协同创新中心,河南新乡453003 [2]河南省新乡医学院第一附属医院肿瘤科,河南卫辉453100

出　　处：《第三军医大学学报》2018年第2期122-129,共8页Journal of Third Military Medical University

基　　金：国家自然科学基金面上项目(31301135);河南省自然科学基金(162300410211);河南省医学科技攻关计划(201203068)~~

摘　　要：目的通过体内体外实验探讨结肠癌相关转录本2(colon cancer-associated transcript 2,CCAT2)对非小细胞肺癌(non-small cell lung cancer,NSCLC)细胞增殖的影响及其调控机制。方法利用shRNA干扰NSCLC细胞(A549和H1299)CCAT2表达。实时定量PCR(quantitative real-time reverse transcription PCR,qRT-PCR)检测CCAT2表达。肿瘤成球实验和CCK-8实验检测细胞增殖。蛋白印记法检测细胞增殖标记蛋白-细胞增殖核抗原-67(antigen identified by monoclonal antibody,Ki-67)和增殖细胞核抗原(proliferating Cell Nuclear Antigen,PCNA),TGF-β信号通路相关蛋白转化生长因子(transforming growth factor beta,TGF-β),细胞信号转导分子Smad2(suppressor of mothers against decapentaplegic 2)和α-平滑肌肌动蛋白(alpha smooth muscle actin,α-SMA)的表达。结果 NSCLC细胞系(A549、H1299、H1650和H358)中CCAT2的表达显著高于正常肺脏细胞系BEAS-2B(P<0.01)。与对照组相比,转染CCAT2 shRNA后A549和H1299细胞中CCAT2的表达显著降低(P<0.01)。A549和H1299细胞中CCAT2 shRNA组细胞增殖倍数明显低于对照组(P<0.05)。与对照组相比,A549和H1299细胞中CCAT2 shRNA组细胞增殖标记蛋白Ki-67和PCNA表达显著降低(P<0.01),TGF-β信号通路相关蛋白TGF-β、Smad2和α-SMA表达显著升高(P<0.01)。与对照组相比,CCAT2 shRNA组肿瘤体积显著变小(P<0.01),肿瘤组织中TGF-β信号通路相关蛋白TGF-β、Smad2和α-SMA表达显著升高(P<0.01)。结论 CCAT2可通过抑制TGF-β信号通路促进NSCLC细胞增殖和肿瘤生长。ObjectiveTo explore the role of long noncoding RNA （lncRNA） colon cancerassociated transcript 2 （CCAT2） in regulating the proliferation of nonsmall cell lung cancer （NSCLC） cells. MethodsThe expression of CCAT2 in NSCLC cell lines A549 and H1299 was silenced by transfecting the cells with a short hairpin RNA （shRNA） targeting CCAT2. CCAT2 expression in the transfected cells was detected with quantitative realtime PCR （qRTPCR）, and the cell proliferation was analyzed with CCK8 assay and tumor cell sphere formation assay. The expression of antigen identified by monoclonal antibody Ki67, proliferating cell nuclear antigen （PCNA）, transforming growth factor beta （TGFβ）, Smad2 and alpha smooth muscle actin （αSMA） in the cells were detected with Western blotting. ResultsNSCLC cell lines A549, H1299, H1650 and H358 expressed significantly higher levels of CCAT2 than the normal lung cell line BEAS2B （P〈0.01）. Compared with the control cells, A549 and H1299 cells transfected with CCAT2 shRNA showed obviously decreased expression of CCAT2 （P〈0.01） and significantly lowered proliferation rates （P〈0.01）. Transfection with CCAT2 shRNA resulted in significantly lowered expression of Ki67 and PCNA but obviously enhanced expression of TGFβ pathway related proteins TGFβ, Smad2 and αSMA （P〈0.01）. In mouse models bearing A549 cell xenografts, the tumor volume was significantly smaller in CCAT2 shRNA group than in the control group, and the expression of TGFβ, Smad2 and αSMA in the tumor tissues were significantly higher in CCAT2 shRNA group （P〈0.01）. ConclusionCCAT2 promotes the proliferation of NSCLC cells by inhibiting TGFβ pathway both in vitro and in vivo.

关 键 词：非小细胞肺癌 结肠癌相关转录本2 增殖 TGF-Β信号通路 

分 类 号：R394.3[医药卫生—医学遗传学] R730.23[医药卫生—基础医学]