HBx Regulates The Invasion And Metastasis Of Hepatocellular Carcinoma By Upregulating LncRNA-ATB

Background and objective:Hepatocellular carcinoma(HCC)is the second most common cause of malignancy mortality in China,and chronic infection of hepatitis B virus(HBV)and its associated liver disease is one of the most common cause of high incidence of HCC.The pathology,invasion and metastasis of HBV-related liver cancer were stronger than non-HBV-related liver cancer,with hidden onset,rapid progress.Up to now,early detection rate and 5-year survival rate of HBV-related liver cancer are still very low.The main reason for poor prognosis and faster progress of HBV-related liver cancer,are intrahepatic,extrahepatic metastasis and tumor tissue invasion of large vessels.At present,understanding on how HBV promotes the occurrence,development and even metastasis of liver cancer is still not clear.Most of the studies suggest that HBx protein encoded by X gene in HBV genome plays a major role in the development of hepatocellular carcinoma,but there is a lack of understanding of HBx-induced hepatocellular carcinogenesis and the influence of hepatocyte growth,differentiation and migration.With more and more studies on the discovery of non-coding RNA’s regulation of gene expression that the oncogene,tumor suppressor gene dysfunction is due to abnormal control of non-coding RNA.The role of non-coding RNA in the development and progression of hepatocellular carcinoma is gradually being revealed,and it is also gradually accepted by researchers.As a kind of functional non-coding RNA,long noncoding RNA(lncRNA)has been shown to regulate the expression of many genes in the development and progression of hepatocellular carcinoma and affects the progression of liver cancer.However,the role of lncRNA in HBV-related liver cancer and regulation is still unclear.In order to explore the role of lncRNA in the development of HBV-related hepatocellular carcinoma,we selected a newly discovered IncRNA-ATB,which is closely related to the invasion and metastasis of HCC,explored the relationship between the expression of HBx and IncRNA-ATB.On the basis of this,we further explored the upstream and downstream relationship between HBx and IncRNA-ATB and possible regulatory mechanisms to detect the relationship between lncRNA-ATB and invasion.migration of HBx(+)hepatocellular carcinoma cells in order to clarify whether HBx can promote liver cancer invasion and metastasis through regulation of IncRNA-ATB expression,in order to improve the early diagnosis of HBV-related liver cancer to provide a theoretical basis.Methods:Part Ⅰ:Effect of HBx on IncRNA-ATB in Hepatocellular Carcinoma Cells and Its MechanismThe expression of lncRNA-ATB in HBx-HepG2 and HepG2 cells was detected by using lentivirus to construct HBx-HepG2,a hepatocarcinoma cell line stably expressing HBx.The levels of autophagy and TGF-β in two kinds of cells were measured.The changes of lncRNA-ATB and TGF-β were observed after induced autophagy.The correlation between autophagy,TGF-β and lncRNA-ATB was confirmed.The changes of autophagy,TGF-p and lncRNA-ATB in HBx-HepG2 were clarified by knockout of TGF-β.Part Ⅱ:Effect of lncRNA-ATB on the migration of HBx(+)hepatoma cellsCollect clinical specimens of liver cancer to detect whether the level of IncRNA-ATB in tissues was correlated with HBV infection and malignancy of hepatocellular carcinoma.The changes of migration and healing ability of HBx(+)hepatocellular carcinoma cells before and after knockout of TGF-β and lncRNA-ATB were compared according to the results of the first part,and whether HBx could promote the migration and healing of hepatocarcinoma cells by up-regulating the expression of lncRNA-ATB.Results:Part Ⅰ:The levels of IncRNA-ATB and TGF-β in HBx-HepG2 were higher than those in HepG2 cells after HBX transfection of HepG2 cells.After transfection of HBx,the autophagy level of HepG2 cells was significantly increased,and the contents of lncRNA-ATB and TGF-β were further increased after EBSS induced autophagy.The expression of lncRNA-ATB and the autophagy of HBx-HepG2 cells were inhibited after knockout of TGF-β,but the levels of TGF-β and lncRNA-ATB were induced by autophagy again,indicating that autophagy in HBx-HepG2 is located on the upstream of TGF-β.Part II:The level of lncRNA-ATB in hepatocellular carcinoma tissues was significantly correlated with HBV infection,and the content of lncRNA-ATB in HBV(+)hepatocellular carcinoma tissues was higher than that in HBV(-)tissues.In addition,higher expression of lncRNA-ATB was associated with a later TNM staging.Cell function test confirmed that invasion,migration ability of HBx(+)hepatoma cells are stronger than that of HBx(-)hepatoma cells,and knockout TGF-β or lncRNA-ATB can reduce the migration,healing ability of HBx(+)liver cancer cell.Conclusion:1.There was an increase of lncRNA-ATB in hepatocellular carcinoma with high expression of HBx.2.Highly expressed lncRNA-ATB patients with hepatocellular carcinoma had poor tumor staging.3.HBx increased the expression of lncRNA-ATB by promoting the autophagy of hepatocellular carcinoma cells and up-regulating the level of TGF-β.4.The high expression of lncRNA-ATB induced by the above mechanism promoted the migration and healing of hepatocarcinoma cells.