miR-151a-5p对结直肠癌细胞增殖和转移的影响及其机制探讨

Effect of miR-151a-5p on proliferation and metastasis of colorectal cancer cells and its mechanism

目的:研究miR-151a-5p在结直肠癌(colorectal cancer,CRC)细胞中的生物学功能及其潜在机制。方法:采用qRT-PCR检测miR-151a-5p在不同CRC细胞株中的表达情况;使用Lipofectamine^(TM)2000将miR-151a-5p mimics/inhibitor和对照寡核苷酸分别转染至HCT116和SW480细胞中;通过CCK-8、细胞克隆形成实验和Transwell侵袭与迁移实验以及蛋白免疫印迹实验证明miR-151a-5p过表达组和敲低组对CRC细胞功能的影响及机制。结果:qRT-PCR结果显示,miR-151a-5p在HCT116细胞中低表达,在SW480细胞中高表达;CCK-8和细胞克隆形成实验结果显示,HCT116细胞过表达miR-151a-5p后细胞增殖能力明显高于对照组,而SW480细胞敲低miR-151a-5p后结果相反;Transwell实验结果显示,与相应的对照组相比,HCT116-mimics组细胞迁移和侵袭能力均增强,而SW480-inhibitor组细胞迁移和侵袭能力均降低;蛋白免疫印迹结果显示,与对照组相比,HCT116-mimics组E-cadherin和ZO-1蛋白表达水平降低,N-cadherin、Vimentin、β-Catenin、MMP2、MMP9蛋白和TGF-β/Smad信号通路相关蛋白表达水平升高,而SW480-inhibitor组呈相反结果。结论:miR-151a-5能够促进CRC细胞的增殖、侵袭和迁移,其机制可能与促进上皮-间充质转化(epithelial-mesenchymal transition,EMT)进程相关,而TGF-β/Smad信号通路相关蛋白在EMT发生进程中出现改变,提示TGF-β/Smad信号通路可能参与了该表型的调控。

Objective:To investigate the biological function and mechanism of miR-151a-5p in colorectal cancer(CRC)cells.Methods:The expression of miR-151a-5p in colorectal cancer cells was tested by qRT-PCR,and miR-151a-5p mimics/inhibitor small interfering and control oligonucleotides were transfected into HCT116 and SW480 cells respectively.The biological function of over-expressed and knockdown miR-151a-5p on CRC cells were verified by CCK-8,cell colony formation,Transwell invasion and migration experiment and Western blot experiment.Results:The results of qRT-PCR were showed that the expression level of miR-151a-5p was lower in HCT116 cells and higher in SW480 cells.CCK-8 assay and clonal formation assay showed that compared with the control group,the proliferation ability of HCT116 cells increased significantly after overexpression of miR-151a-5p,which the result of SW480 cells knocking down miR-151a-5p were opposite.Transwell assay showed that compared with the corresponding control group,the ability of cell migration and invasion in HCT116-mimics group was enhanced.However,the ability of cell migration and invasion decreased in SW480-inhibitor group.Western blot results showed that the protein expression levels of E-cadherin and ZO-1 were reduced and the expression levels of N-cadherin,Vimentin,β-Catenin,MMP2,MMP9 protein and TGF-β/Smad signaling pathway related protein were increased in the HCT116-mimics group compared to the control group,while the opposite result was observed in the SW480-inhibitor group.Conclusion:miR-151a-5p promotes the proliferation,migration,and invasion of CRC cells,and the mechanism may be related to the promotion of EMT process.The change of TGF-β/Smad signaling pathway protein during EMT suggests that TGF-β/Smad signaling pathway may be involved in regulating this phenotype.