敲低Ⅰ型胶原α1链基因抑制鼻咽癌细胞增殖、迁移和侵袭的研究

Inhibition of proliferation,migration and invasion of nasopharyngeal carcinoma cells by knocking down COL1A1 gene

目的研究Ⅰ型胶原α1链(COL1A1)基因对鼻咽癌细胞增殖、侵袭、迁移的影响。方法该研究于2017年9月至2018年12月完成,实时定量PCR(qPCR)和蛋白质印迹法(Western blotting)检测COL1A1在鼻咽癌(C666-1、CNE1、CNE2)细胞系中的表达量;CNE1细胞分m为对照组、阴性对照(si-NC组)、敲低COL1A1(si-COL1A1)组,通过Lipofectamine 2000将siRNA COL1A1质粒转染至鼻咽癌CNE1细胞中,对照组细胞正常培养,si-NC组细胞转染阴性对照;qPCR和Western blotting检测转染效果,噻唑蓝(MTT)、Transwell实验分别检测细胞增殖、侵袭、迁移;Western blotting检测钙黏蛋白E(E-cadherin)、波形蛋白(Vimentin)、钙黏蛋白N(Ncadherin)水平。结果 COL1A1 mRNA[(3.565±0.341)(4.253±0.410)(3.968±0.378)]和蛋白表达量[(2.214±0.216)(3.152±0.305)(2.519±0.221)]在鼻咽癌(C666-1、CNE1、CNE2)细胞系中表达量增加(P<0.05);CNE1细胞转染siRNA COL1A1质粒能显著下调COL1A1 mRNA(0.352±0.045)和蛋白表达量(0.245±0.232)(P<0.05);敲低COL1A1的表达量抑制CNE1细胞增殖(0.536±0.059)(0.665±0.067)(0.893±0.081)(P<0.05),降低其侵袭(52.895±5.635)、迁移(113.254±13.279)能力(P<0.05),上调E-cadherin蛋白水平(2.152±0.223)(P<0.05),下调Vimentin(0.362±0.045)、N-cadherin(0.423±0.043)蛋白水平(P<0.05)。结论 COL1A1可能通过抑制上皮细胞间充质化(EMT)促进鼻咽癌细胞增殖、侵袭、迁移。

Objective To investigate the effect of collagen type 1 alphα 1(COL1A1) gene on proliferation,invasion and migration of nasopharyngeal carcinoma cells.Methods The study was completed from September 2017 to December 2018.Quantitative polymerase chain reaction(qPCR) and Western blotting were used to detect the expression of COL1A1 in nasopharyngeal cancer cell lines(C666-1,CNE1,CNE2).CNE1 cells were randomly assigned into control group,negative control(si-NC) group and knockdown COL1A1(siCOL1A1) group.The siRNA COL1A1 plasmid was transfected into CNE1 cells by Lipofectamine 2000,and the control group were cultured normally,while those in si-NC group were transfected into negative control group.The transfection effect was detected by qRTPCR and Western blotting.The cells proliferation,invasion and migration were detected by MTT and Transwell assay.Western blotting was used to detect the expression of E-cadherin,Vimentin,N-cadherin.Results The expression of COL1A1 mRNA [(3.565±0.341),(4.253±0.410),(3.968±0.378)] and protein expressions [(2.214±0.216),(3.152±0.305),(2.519±0.221)] was significantly increased in nasopharyngeal cancer cell lines(C666-1,CNE1,CNE2)(P<0.05).The siRNA COL1A1 plasmid could down-regulate the expression of COL1A1(0.352±0.045) gene and(0.245±0.232) protein(P<0.05).Knocking down the expression of COL1A1 inhibited the proliferation[(0.536±0.059),(0.665±0.067),(0.893±0.081)] of CNE1 cells(P<0.05),decreased the invasion(52.895±5.635) and migration(113.254±13.279) ability(P<0.05),increased the level of E-cadherin(2.152±0.223) protein(P<0.05),and decreased the levels of Vimentin(0.362±0.045) and N-cadherin(0.423±0.043) protein(P<0.05).Conclusion COL1A1 may promote the proliferation,invasion and migration of nasopharyngeal carcinoma cells by inhibiting epithelial-mesenchymal transition(EMT).