Effect Of Targeted SiRNA Silencing ZFX Gene On Biological Function Of Intrahepatic Cholangiocarcinoma Cells

ObjectiveThe ZFX gene siRNA fragment was constructed and transfected into the intrahepatic cholangiocarcinoma cell line HCCC-9810.Observe changes in the expression level of ZFX in intrahepatic cholangiocarcinoma cells after transfection.By down-regulating the expression level of ZFX in HCCC-9810 cells,the effects on the proliferation,migration,invasion,and cloning of the intrahepatic cholangiocarcinoma cells were observed and preliminarily clarify the role of ZFX gene in the occurrence and development of intrahepatic cholangiocarcinoma.MethodsThe ZFX gene siRNA sequence reported in the literature was synthesized and transfected into the intrahepatic cholangiocarcinoma cell line HCCC-9810 by lipofectamine 2000.According to the fragment of transfected siRNA,the experiment was divided into three groups: blank(Cell),siRNA containing nonsense sequence(NC-siRNA),and siRNA transfected with ZFX gene(ZFX-siRNA).The mRNA expression level of ZFX gene was detected by RT-PCR,and the level of ZFX gene protein was detected by Western blot.Finally,we used the soft lipid agarose colony formation assay,CCK-8 proliferation assay,and Transwell assay to detect the colony-forming ability,cell proliferation ability,cell migration and metastasis ability of the HCCC-9810 cells after transfection.ResultsAfter siRNA was transfected into HCCC-98110 cell line for 72 h,the results of RT-PCR and Western blot showed that ZFX-siRNA effectively reduced ZFX mRNA and protein levels in HCCC-9810 cell line compared with Cell and NC-siRNA groups(P<0.05),but there was no significant difference between Cell and siRNA-NC groups(P>0.05).It was proved that ZFX-siRNA had significant silencing efficiency on ZFX gene in HCCC-9810 cells(65.9%).Clone formation experiments showed that after the silencing of ZFX,the clone formation ability of the cell line was significantly reduced(P=0.019).The CCK-8 proliferation experiment demonstrated that the cell proliferation ability of the experimental group was significantly decreased on the second day(t=5.366 P=0.0058).Transwell experiments showed that the ability of cell migration and metastasis in Cell and siRNA-NC groups was significantly higher than that in siRNA-ZFX group(P=0.001,P=0.002).There was no significant difference between Cell and siRNA-NC groups in each group(P=0.1569,P=0.0649).ConclusionIn this experiment,ZFX-siRNA successfully reduced the expression of ZFX gene in intrahepatic cholangiocarcinoma cell line HCCC-9810,demonstrating that the decreased expression of ZFX gene can inhibit the proliferation,clone,migration and metastasis of intrahepatic cholangiocarcinoma cells.It has been initially clarified that ZFX plays an important role in the development of intrahepatic cholangiocarcinoma HCCC-9810.