Effects Of CLDN6 On Invasion,Migration And Apoptosis Of Hepatocellular Carcinoma By Regulating JAK2/STAT3 Signaling Pathway

Background : Claudins(CLDNs)as a scaffold protein with tight junction between epithelial and endothelial cells,is highly expressed in hepatocellular carcinoma.The treatment of liver cancer has come a long way over the years,but metastasis is still one of the factors leading to poor prognosis.Claudins is considered as a potential new therapeutic target for patients with HCC,so it is vitial to study the mechanism of Claudin family in hepatocellular carcinoma cells.Methods:1.Downregulation and overexpression of CLDN6 in hepatocellular carcinoma were achieved by si RNA and plasmid transfection.2.q RT-PCR and immunofluorescence were used to know whether the transfection was successful,and the change of CLDN6 protein expression level was determined according to the results of Western Blot.3.Transwell assay and Wound healing assay were used to determine the changes of cell invasion and migration ability.4.CCK-8 assay was used to determine if the cell proliferation was significantly inhibited.5.Flow cytometry was used to obtain the apoptosis rate of cells after CLDN6 knockdown.6.The expressions of related proteins(JAK2,STAT3,p-JAK2,p-STAT3)and apoptosis-related proteins(cleaved caspase-3,Bax,Bcl-2)in the signaling pathway were detected by Western Blot.Results:1.After transfection,the expression of CLDN6 gene was changed by q RT-PCR,immunofluorescence and Western Blot,indicating that the cell model had been successfully constructed.2.From Transwell test and Wound healing assay we know that the invasion and migration ability of cells was decreased because of the downregulation of CLDN6 expression.3.CCK-8 results showed that cell proliferation was significantly inhibited,and the apoptosis rate was significantly increased by flow cytometry.4.Western Blot results showed that the protein expressions of p-JAK2 and p-STAT3 decreased,and the ratio of p-JAK2/JAK2 and p-STAT3/STAT3 also decreased correspondingly,which indicated that the JAK2/STAT3 signaling pathway was inhibited.5.Western Blot results showed that the expression of anti-apoptotic protein Bcl-2 was decreased,cleaved caspase-3 and Bax were increased,and knockdown of CLDN6 promoted apoptosis.6.When CLDN6 was overexpressed,the invasion and migration ability of cells were enhanced,and apoptosis is inhibited,while the influence of CLDN6 overexpression on cells is weakened after the use of pathway inhibitors.Conclusions : Knocking down CLDN6 can inhibit the invasion and migration of hepatocellular carcinoma cells and promote apoptosis by down-regulating JAK2/STAT3 signaling pathway.