【摘要】 目的:探讨Twist基因在SW480、HCT116和HT29结肠癌细胞系上皮-间质转化(EMT)中的作用,明确其对恶性肿瘤侵袭转移能力的影响。方法:利用重组质粒p Tracer-CMV/BSD-Twist和p Genesil1.2-Twist-shRNA转染SW480、HT29和HCT116;流式细胞术检测转染率;Real time-PCR和Western blot检测Twist、E-cadherin和Vimentin转录及蛋白表达水平;Transwell实验检测细胞的迁移和侵袭能力。结果:转染高表达Twist质粒后,各结肠癌细胞系中Twist和Vimentin表达水平显著升高(P<0.05),E-cadherin显著降低(P<0.05);转染低表达Twist质粒后,SW480和HT29中各指标均无显著变化(P>0.05),HCT116中Twist和Vimentin表达水平显著降低(P<0.05),E-cadherin无显著变化(P>0.05);Transwell实验表明抑制HCT116细胞系Twist表达后,其侵袭和迁移能力明显减弱(P<0.01)。结论:上调Twist表达可以促进EMT;抑制HCT116中Twist表达能减弱结肠癌细胞的侵袭和转移能力。更多还原

【Abstract】 Objective: To explore the role of Twist gene in epithelial-mesenchymal transition(EMT) in SW480, HCT116 and HT29, and then study the effect of Twist on invasion and metastasis of malignant tumors. Methods: Recombinant plasmids p Tracer-CMV/BSD-Twist and p Genesil1.2-Twist-shRNA were used to transfect SW480, HCT116 and HT29. Transfection efficacy of the plasmid in each cell line was confirmed by flowcytometry. The m RNA transcription level and protein expression level of Twist, E-cadherin and Vimentin were detected by RT-PCR and western blot, respectively. The migration and invasive analysis was done by transwell assay. Results: After transfected by recombinant high-expressed twist plasmid, the m RNA and protein expression levels of Twist and Vimentin increased significantly(P<0.05), whereas E-cadherin was inhibited(P<0.05). After transfected by recombinant low-expressed twist plasmid, all the parameters in SW480 and HT29 cell lines were statistically insignificant(P>0.05), the m RNA and protein levels of Twist and Vimentin were prominently inhibited in HCT116 cell line(P <0.05) and the level of E-cadherin was not statistically significant(P >0.05). Significant reduction of invasion and migration was observed in transwell assay after inhibiting the Twist expression in HCT116 cell line(P <0.01).Conclusion: Up-regulation of Twist gene expression can promote the EMT and inhibiting the Twist gene expression can lessen the migration and invasion of HCT116.更多还原