Effects Of Silencing CRAF Gene Expression On Proliferation, Metastasis And Invasion Of Hepatocellular Carcinoma Cells

ObjectiveTo investigate the expression of CRAF in human liver cancer and to investigate the effect of CRAF expression on the proliferation,invasion and migration of hepatocellular carcinoma cells in order to elucidate the mechanism of CRAF promoting the proliferation,invasion and migration of hepatoma cells.It provides a choice scheme for the personalized treatment of hepatocellular carcinoma and provides a new idea for the potential target of clinical targeted therapy.Methods1.Immunohistochemistry and Western blot were used to detect the expression of CRAF in hepatocellular carcinoma and its adjacent tissues.2.The expression of CRAF in HepG2,QGY7703 and SMMC-7721 hepatocellular carcinoma cells was analyzed by Western blotting,and the suitable cell lines were selected.3.The transfected shRNA was transfected into SMMC-7721 hepatocellular carcinoma cells to down-regulate the expression of CRAF.Western blotting was used to detect the expression of CRAF to determine the effect of downregulation.4.The MTT assay was used to detect the effect of silencing CRAF on the proliferation of hepatoma cells in vitro.5.The cell scratch assay was used to detect the effect of CRAF on the migration of liver cancer cells in vitro.The effect of CRAF on the invasion of liver cancer cells in vitro was detected by Transwell assay.6.The effect of silencing CRAF on MMP-2,MMP-9 was analyzed by Western blotting.ResultsWestern Blot results show that SMMC7721 is highly expressed in CRAF hepatoma cells and can be used in subsequent experiments.Transfection of shRNA downregulated CRAF in SMMC-7721 hepatoma cells.MTT assay and clonogenic assay in vitro showed that the proliferation and clonogenic capacity of HCC cells decreased compared with the control group(p <0.05).The cell scratch assay and the transwell assay The experimental results showed that the ability of migration and invasion of HCC cells decreased compared with the control group(p <0.05).The results of Western Blot showed that the expression of MMP-2 and MMP-9 in HCC cells decreased significantly compared with the control group(p <0.05)Conclusion1.CRAF is highly expressed in hepatocellular carcinoma cell line SMMC-7721.2.The silencing of CRAF by shRNA can inhibit the proliferation,migration and invasion of hepatocellular carcinoma cells.3.Silencing CRAF can down-regulate the expression of MMP-2,MMP-9 and other proteins related to invasion and migration.