LINC00628 Suppresses Migration And Invasion Of Hepatocellular Carcinoma By Its Conserved Region Interacting With The Promoter Of VEGFA

ObjectiveTo investigate the role and molecule mechanisms of LINC00628 in the process of HCC.Methods1.RT-qPCR was used to detect the expression level of LINC00628 in HCC tumor tissues and adjacent tissues,normal hepatocytes and several HCC cell lines.2.Overexpression and interference vectors of LINC00628 were constructed.The effects of LINC00628 on the biological function of HCC cells were examined by colony formation assay,MTS,transwell assay and wound healing assay in vitro.3.RT-qPCR and Western blot were used to find the target gene of LINC00628.The target gene overexpression vector was constructed and used wound healing assay and transwell assay to determine the effect of target gene on HCC migration and invasion.4.Different length fragments of VEGFA promoter were cloned.Dual luciferase reporter assay was employed to analyze the effect of LINC00628 on the promoter of VEGFA.5.Bioinformatics website was used to analyze the conserved sequence of LINC00628.Stable cell lines of overexpressing the conserved sequence were established to test their impacts on the migration and invasion capacity of HCC cells by wound healing assay and transwell assay.6.Western blot assay was performed to evaluate the influence of the conserved sequence on the expression of VEGFA.Dual luciferase reporter assay was used to testify the relationship of the conserved sequence and the VEGFA promoter activity.7.Nude mice model of hepatic metastasis with LINC00628 overexpression was constructed.The H&E staining,RT-qPCR,western blot and immunohistochemistry assays were used to verify the effect of LINC00628 on the metastasis of HCC and the expression of related molecules.Results1.LINC00628 was frequently downregulated in HCC tumor tissues and HCC cell lines.2.LINC00628 suppressed HCC cell migration and invasion in vitro.3.LINC00628 inhibited the expression of VEGFA.4.VEGFA could promote HCC cell migration and invasion in vitro.5.LINC00628 repressed VEGFA expression by inhibiting its promoter activity.6.The conserved sequence of LINC00628 was essential for repressing migration and invasion of HCC cells.7.The conserved sequence element of LINC00628 reduced VEGFA expression and its promoter activity.8.LINC00628 suppressed the metastasis of HCC cells and the expression of VEGFA in vivo.ConclusionLINC00628 can inhibit the migration and invasion of HCC cells through the interaction between its conserved sequence and the promoter of VEGFA.