Expression Of ZWINT In Hepatocellular Carcinoma And Its Effect On Biological Function Of Hepatoma Cell

Objective:Hepatocellular carcinoma(HCC)is a kind of malignant tumor.And this cancer has high mortality and its prognosis is very pooor.The purpose of this study was to explore the expression level of ZWINT in HCC through database,and to explore the effects of ZWINT on various biological behaviors of hepatoma cells and the possible mechanism of action by a series of experiments.Methods:1.To investigate the expression of ZWINT in cancer and paracancer tissues of liver cancer patients through public database.Using TIMER database and GEPIA database,the expression of ZWINT in HCC patients was investigated.UALCAN database was also used by us,which,can tell us the expression of ZWINT in HCC patients with liver cancer under various clinicopathological characteristics.Kaplan-Meier Plotter database analyzed the relationship between ZWINT and overall survival(OS)and relapse-free survival(DFS).The correlation between ZWINT expression and immune cell infiltration in HCC was analyzed by TIMER database.The Linked Omics database was used for gene enrichment analysis of ZWINT to identify its function and related signaling pathways.2.The expression of ZWINT was subsequently verified in hepatoma cell lines and normal liver cells.Hepatoma cell SUN-449 with relatively high expression of ZWINT were selected for knockdown using small interfering RNA and divided into Control group(Control),negative control group(Si-NC)and ZWINT knockdown group(Si-ZWINT).Hepatoma cell Huh-7 with relatively low expression of ZWINT were selected,and lentivirus was used to construct overexpressed stable strains,which were divided into no-load control group(Vector)and overexpression group(Oe-ZWINT).The transfection efficiency was verified by Western Blot,and the follow-up experiments were conducted on these basis.CCK-8 assay and EDU assay were used to detect the proliferation of HCC cells in each group.The metastasis of HCC cells in each group were detected by wound healing test and Transwell test.The cell cycle and apoptosis of HCC cells in each group were detected by flow cytometry.Western Blotting(WB)was used to detect the effect of ZWINT on the expression of EMT-related proteins.3.The effect of ZWINT on HCC proliferation was investigated in vivo by subcutaneous tumorigenesis in nude mice.Results:1.Public databases showed that ZWINT was highly expressed in HCC;ZWINT was statistically significant in the P53 mutation subgroup.Patients with high expression of ZWINT had poor postoperative OS and DFS.ZWINT was correlated with the infiltration degree of B cells,CD4~+T cells,CD8~+T cells,neutrophils,dendritic cells,macrophages and other immune cells in HCC.Gene enrichment analysis showed that ZWINT was highly enriched in biological processes and signaling pathways such as DNA replication,cell division and cell cycle checkpoint.2.In vitro experiments:CCK-8 and EDU experiments showed that compared with Control group and Si-NC group,the proliferative ability of Si-ZWINT group was decreased,while compared with Vector group,the proliferative ability of Oe-ZWINT group was improved(P<0.05).The wound healing experiment and Transwell experiment showed that the metastasis ability of Si-ZWINT group was significantly weaker comparing with the Control group and the Si-NC group,while the cell metastasis ability of Oe-ZWINT group was enhanced compared with Vector group(P<0.05).Flow cytometry showed that the apoptosis rate in the Si-ZWINT group was significantly higher than that in the Control and Si-NC groups,and the proportion of cells blocked in G1 phase was increased,while the apoptosis rate in the Oe-ZWINT group was lower than that in the Vector group,and the cell cycle arrest was reduced in G1 phase(P<0.05).WB results demonstrated that,compared with the Control group(Control group)and the negative control group(Si-NC group),the expression of E-cadherin was higher in the knockdown group,while the expression of Vimentin and N-cadherin was lower.The expression levels of E-cadherin in Oe-ZWINT group were lower,while the expression levels of Vimentin and N-cadherin were higher(P<0.05).3.The results of nude mice experiment indicated that:compared with the control group,the tumor growth rate in the knock down group was significantly slowed down(P<0.05),and the tumor weight was significantly lighter(P<0.05).All these figures indicated that the growth of transplanted tumors in the knock down group was significantly inhibited.Conclusion:ZWINT gene is highly expressed in HCC patients,and high expression of ZWINT is associated with poor survival prognosis.They are positively correlated.In vitro experiments have verified that ZWINT can promote the proliferation and metastasis of HCC cells,while inhibit their apoptosis,and promote the progression of HCC by promoting the occurrence of EMT.ZWINT promoted the growth of hepatoma cell in nude mice.