Construction And Study On Hilar Cholangiocarcinoma QBC939 Cell Lines By SiRNA Silencing Vimentin Gene

Objective: 1.Construction of three groups of different nucleic acid sequence Small interference RNA(Small interfering RNA,si RNA)(si VIM-1,si VIM-2,si VIM-3)and the negative control sequence(si-Ctrl)model;2.The expression of Vimentin(VIM)in hilar bile duct cancer cell line QBC939 and normal bile duct cell line HIBEpi C were detected;3.To investigate the effect of RNA interference on the proliferation,invasion,migration and apoptosis of hilar cholangiocarcinoma cell line QBC939 after inhibiting the expression of VIM.Methods: 1.GEPLA database website(http://gepia.cancer-pku.cn/)was used to analyze the expression of VIM in cholangiocarcinoma and other tumors.2.The expression of VIM in hilar cholangiocarcino ma cell line QBC939 and normal cholangiocarcinoma cell line HIBEpi C were detected by RT-PCR and q RT-PCR.3.Three groups of Small interfering RNA(si RNA)with different sequences of nucleic acids(si VIM-1,si VIM-2,si VIM-3)and their negative control sequences(si-Ctrl)were designed and synthesized.The experimental group(transfected with si VIM-1,si VIM-2,si VIM-3)and the control group(transfected with si-Ctrl)were set up,and the group of si RNA with the best inhibitory effect was screened by RT-PCR and q RT-PCR for subsequent experiments.4.A group of si RNA and si-Ctrl with the best inhibitory effect were transfected into hilar cholangiocarcinoma cell line QBC939 by liposome method,and the expression of VIM at gene and protein levels was detected by RT-PCR,q RT-PCR and Western-blot.5.The experimental group(si RNA with the best inhibitory effect)and the control group(si-Ctrl transfection)were set up.Cell proliferation,invasion and migration were detected by Incu Cyte ZOOM system,and cell apoptosis was detected by flow cytometry.Results: 1.The expression of VIM in cholangiocarcinoma was higher than that in normal tissues(P<0.05)and no significant difference was found in different stages of cholangiocarcinoma(P>0.05)with GEPLA database website.2.Compared with normal bile duct cell line HIBEpi C,VIM expression was increased in hilar cholangiocarcinoma cell line QBC939(P < 0.05).3.Three groups of si RNA(si VIM-1,si VIM-2,si VIM-3)and negative control sequence(si-ctrl)models with different nucleic acid sequences were successfully designed and synthesized,among which si VIM-2 had better inhibitory effect than si VIM-1 and si VIM-3(P<0.05).4.si VIM-2 and si-Ctrl were successfully transfected into hilar cholangiocarcinoma cell line QBC939.Compared with the control group,the expression of VIM gene and protein was lower in the experimental group(P<0.05).5.Compared with the control group,the proliferation ability of cells in the experimental group was slower(P<0.05),invasion and migration ability was decreased(P < 0.05),and apoptosis increased(P<0.05).Conclusion: 1.The expression level of VIM in hilar cholangiocarcinoma cell line QBC939 was significantly higher than that in normal cholangiocarcinoma cell line HIBEpi C,suggesting that VIM may be related to the occurrence and development of hilar cholangiocarcinoma.2.si VIM-2 was successfully transferred into hilar cholangiocarcinoma cell line QBC939.After silencing VIM,QBC939 cell proliferation slowed,migration and invasion ability decreased,and apoptosis increased.3.The biological function of VIM is complex,which is related to the poor prognosis of tumor,and may become a major indicator of tumor detection.The expression of VIM is mostly related to the EMT process and promotes tumor invasion and metastasis through EMT.It is of great significance to further study the molecular mechanism,prognosis and targeted therapy of VIM in the occurrence and development of various cancers,and it is expected to become a new target of anti-tumor therapy in the future.