Construction And Expression Of Live Attenuated Hepatitis B Virus Strains

Hepatitis B virus(HBV)is a DNA enveloped virus belonging to the hepatotropic DNA virus family.After the virus infects the human body,it will be in an acute infection stage in the early stage,and the body’s immune system will clear it.HBV infection changes from acute to chronic,accounting for about one tenth of adults.Chronic infection,on the other hand,produces low levels of inflammation,leading to chronic hepatitis,which can eventually lead to cirrhosis and liver cancer.Due to the particularity of hepatitis B virus infection,it has a completely different life cycle compared with other viruses,and this feature makes the complete elimination of the virus particularly difficult,and there is currently no clinical cure.At present,the mainstream drug for the clinical treatment of HBV is type I interferon.Interferon can directly exert its antiviral effect by inducing the expression of interferon-stimulated genes(ISGs),and on the other hand,it can also exert its antiviral effect indirectly by regulating the body’s immune system.Antiviral activity.But the current treatment effect is limited.The fundamental reason for the difficulty in clearing HBV is that ccc DNA cannot be completely cleared,and a sufficient level of cellular immunity must be activated to kill all infected cells to completely clear the HBV genome reservoir.Therefore,it is particularly important to study HBV-related cellular immune activation.The purpose of this study was to explore HBV therapeutic vaccines.The construction and expression of live attenuated HBV virus strains in liver cancer cells and non-hepatocytes were studied.The results are as follows:In this study,the construction of live attenuated HBV virus strains was initially discussed at the cellular level.The results showed that in human hepatoma cell Huh7,which supports HBV replication,after transfection of p HBV1.3ST1 with premature termination of S region,the cells could not secrete HBV particles,and HBV replication intermediates would accumulate in the cells.When the HBs Ag plasmid was re-expressed,the intracellular accumulation of HBV pg RNA,total DNA and ccc DNA disappeared and returned to the original normal level.At this time,HBs Ag and HBV rc DNA could be detected in the cell supernatant.This indicated that the virus was normally secreted to the outside of the cell,and the synthesis of HBV complete particles was not affected.The virus produced in this way has the same structural characteristics as natural HBV,and due to the modification of its viral genome,when used for infection,it cannot be released from the infected cell to the outside of the cell to cause persistent infection,which can be used as a Live attenuated virus strains were used.The second part of this study co-expressed multiple liver-enriched transcription factors and HBV 1.3-fold replicon in non-hepatocyte HEK293 T cells with a completely blank HBV replication background,and found that the liver-enriched transcription factor HNF4α could enhance HBV transcription and viral activity.The main manifestations were the increase of HBV pg RNA level in cells,the increase of HBe Ag expression in the supernatant of cells,and the increase of empty capsid virus particles and HBV intact virus particles in the supernatant.The results of this study provide a reference method for the development and production of HBV therapeutic vaccines,and provide new ideas for the treatment of chronic HBV infection.