Liver Protection And Antiviral Effect And Immunological Mechanism Of The Main Active Ingredients Of Sophora Tonkinensis Gagnep

HBV infection is an urgent social problem which causing high risk for developing liver cirrhosis and hepatocellular carcinoma and leads to high mortality each year.The pathogenesis of HBV is very complicated.Current studies have shown that hepatitis B virus does not directly cause liver cell damage.During the antiviral immunity,T lymphocytes are the important cell regulating immune response.Some research have pointed out that CD4 T lymphocyte function deficiency or defect could lead to immune tolerance.Therefore,study of the immune adjustment especially the immunomodulatory effects of CD4 T cells provides a new direction and strategy for the control of HBV infection.In our country,traditional Chinese medicine has been widely used in the treatment of hepatitis for many years.Sophora tonkinensis Gagnep is a traditional Chinese medicine,the main function of it is qingrejiedu.It has been used to treat pharyngitis jaundice and arrhythmia.A lot of literatures reports that Oxymatrine possessing a wide range of pharmacological effects,such as antivirus,immunity regulation and amelioration of inflammation in several animal inflammation models.According to recent researches,Oxymatrine could inhibit the replication of HBV and the expression of HBsAg and HBeAg in HepG2.2.15 cells and transgenic mice.Oxymatrine was confirmed to reduce the serum levels of HBeAg and HBV DNA which is similar to the interferon-?.Sophocarpine has been shown to against Coxsackie B virus and regulate the immune.In the current study,a HBV persistence murine model was used to investigate the efficiency of Oxymatrine and its immune mechanism leading toHBV clearance.A T cell-dependent hepatitis mouse model was used to examine the protective effects of Sophocarpine to expand the clinical application of the active ingredients of Sophora tonkinensis Gagnep.Part ? The antiviral effect and immune mechanism of Oxymatrine in hydrodynamic injection HBV mouse modelObjective:To investigate the antiviral effect and immunological mechanism of Oxymatrine in hydrodynamic injection HBV mouse model.Method:1.The model were achieved by quickly injected with saline which equivalent to 10% of the mouse body weight and contains plasmid DNA.2.The efficacy of OMT were evaluated by the levels of HBV DNA,HBs Ag,HBeAg and HBcAg.3.The expression of serum IFN-? and TNF-? were measured by ELISA.The lymphocyte of mice were separated and measured by Flow cytometric to explore the immunological mechanism of Oxymatrine.Result:1.ETV showed tremendous suppression effect on HBV infection in the indicated time point.And OMT at 20 mg·kg^-1 also reduced the HBV titers compared with the model group that still carried relatively high level of DNA copies at the 3rd and 6th week.However,OMT at 2.2 mg·kg^-1 and 6.7 mg·kg^-1had no effect even with long-term administration.2.Although ETV was able to reduce the level of serum HBV DNA in transfected mice,it had very limited impact on serum HBsAg.OMT administrated intraperitoneally was beneficial for the control of HBV infection and it produced obvious reduction in HBeAg and HBcAg.3.ELISA showed that OMT of 6.7 and 20 mg·kg^-1 were capable to promote the expression of IFN-? while it displayed no influence on TNF-?.4.Flow cytometric showed that after OMT treatment the expression of IFN-? produced by CD4+ T cells were increased in a dose-dependent mannerwhile it had no influence on IL-4.Conclusion:1.OMT was able to reduce the level of serum HBV DNA in infected mice,while efficacy is relatively weak.2.OMT was beneficial for the control of HBV infection and it produced obvious reduction on HBsAg,HBeAg and HBcAg.The efficacy of OMT was better than ETV.3.The efficacy of OMT responsed to HBV clearance was related to the high production of IFN-? produced by CD4+ T cells.Part ? The protective effect and immune mechanism of Sophocarpine on ConA-induced hepatitisObjective:The aim of this study was to determine the protective effects and immunological mechanisms of Sophocarpine on ConA-induced hepatitis.Method:1.BALB/c mice were pretreated with Sophocarpine or Bicyclol for 5consecutive days,control group were treated with saline.Thirty minutes after the final administration,the mice were injected with 15 mg·kg^-1 of ConA intravenously except the control group.2.The hepatoprotective effect of Sophocarpine was evaluated by serum ALT,AST,TBIL,HE and TUNEL staining on ConA induced hepatitis.3.The immunological mechanism was investigated by ELISA,RT-PCR,flow cytometry and Western Blotting.Result:1.Compared with the normal group,the ALT,AST and TBIL in the ConA group were significantly increased.Administration of Sophocarpine inhibited the elevation of ALT,ASTand TBIL compared with the ConA group.Sophocarpine inhibited ConA-induced liver inflammation and cell death demonstrated by HE and TUNEL staining.2.RT-PCR showed that Sophocarpine effectively suppressed the mRNAexpression of chemokines and adhesion molecules such as MIP^-1?,CXCL10 and ICAM^-1.3.ELISA showed that Sophocarpine reduced the expression of IFN-? and TNF-?.Sophocarpine reduced the expression of IFN-? in a dose-dependent manner.4.Flow cytometry showed that Sophocarpine decrease the proportion of IFN-?.5.Further study showed that Sophocarpine via overexpression SOCS1 inhibited the activation of STAT1.Conclusion:1.Sophocarpine could alleviate ConA-induced liver injury in mice.2.The expression of chemokines and adhesion molecules such as MIP^-1?,CXCL10 and ICAM^-1were reduced in Sophocarpine pretreated group.3.Sophocarpine decrease the expression of IFN-? in serum and splenic lymphocyte.4.Sophocarpine could up-regulate the expression of SOCS1 and inhibit the activation of STAT1 to protect mice from ConA induced hepatitis.5.The hepatoprotective effect of Sophocarpine was associated with its inhibition of the IFN-?/STAT1 signaling pathway.