| ObjectsHepatitis B virus(HBV) infection is easy to chronic and can progress to cirrhosis and liver cancer. Currently, there are two different treatment strategies for CHB patients:Pegylated interferon(PEG-IFN) or nucleot(s)ide analogues(NUC). This study mainly investigated the host immune relevant factors affecting the hepatitis B antiviral efficacy, the main contents include:1. To approach the clinical effect of nucleoside analog and peg-IFNα or ordinary IFNα antiviral therapy in patients with chronic hepatitis B(CHB).2. To investigate the relationship between CD8+ T cell functionality and effect of different antiviral treatment in CHB patients.3. To investigate the relationship between CD4+ T cell functionality and effect of different antiviral treatment in CHB patients.Methods1. 36 CHB patients(HBe Ag positive) were designed to two different antiviral treatment scheme due to clinical facts, 19 cases of them using the interferon(peg-interferon alpha or common interferon alpha) and the other using entecavir(nucleoside analogue). The blood samples were collected in before antiviral treatment(0w), treatment for 4w, 12 w, 24 w, 48 w,respectively, and we isolated peripheral blood mononuclear cells(PBMCs) with Dakota human lymphocyte separation tube.2. Application Real time-PCR was used to detect quantitative of HBV DNA level in CHB patients.Liver function with nine indicators was tested by Abbott automatic biochemical analyzer.Time-resolved fluorescence immunoassay quantitative techniques(TRFIA) was used for quantitative detection of hepatitis B virus markers five indicators of hepatitis B, including surface antigen(HBs Ag) and antibody(HBs Ab); e antigen(HBe Ag) and e antibody(HBe Ab); core antibody(HBc Ab).3. The flow cytometry was applied to reveal the frequency of CD8+T cells and CD4+T cells as well as their cytokine secretion capacity.4. Analysis of clinical data:According to the virological response in 48 th week of treatment,all of the patients receiving antiviral treatment were divided into A, B two groups. A group:ETV—HBV DNA<5*102, IFN—HBV DNA<1*104;B group:ETV—HBV DNA>5*102,IFN—HBV DNA>1*104). The following are the same.5. Graph Pad Prism software was used for mapping, Flowjo Software flow was used to process cytometry graph, and we used SPSS19.0 for statistical analysis of the data,considering p-value <0.05 with significant difference.Results1.ETV and IFN antiviral therapies have different clinical effects.Compared with IFN group,ETV group has lower levels of ALT and HBV DNA at each point of treatment. At48 th week, virus negative rates of ETV group and IFN group are 24% and 84%repectively,and the difference between two groups was statistically significant(p<0.05).HBe Ag level of IFN group showed downward trend along the treatment. Compared with IFN group, ETV group has lower e-antigen negative rate. At 48 th week, HBe Ag serological conversion rates of ETV and IFN group are 0% and 23% respectively, and the difference between the two groups was statistically significant(p<0.05).2.The frequency of CD8+T cell and CD4+T cell and level of their cytokine secretion in CHB patients before antiviral therapy(1) Compared with healthy controls,the frequency of CD3+CD4+T cell in CHB patients was significantly lower(p<0.05),but we find no significant difference of CD3+CD8+T cell frequency between the two groups. CD8+T cell had significantly higher level of TNFαsecretion and lower level of IL-2 secretion(p<0.05) compared with healthy controls. And CD4+T cell had significantly higher level of IFN γ secretion and lower levels of IL-2secretion(p<0.05) compared with healthy contols.(2) In IFN group and ETV group,the frequency of CD 8+T cell showed downward trend,but the difference of CD8+T cell frequency between the two groups was not statistically significant. In IFN group,IFNγ secretion level of CD8+T cell offered upgrade firstly than descending latter tendency,but difference was not statistically significant. IL-2 secretion levels of CD8+T cell showed upward trend,but difference was not statistically significant compared with pre-treatment time.(3) The frequency of CD4+T cell in both of the two groups decreased firstly and increased later along the process of treatment,but difference was not statistically significant between different point of treatment. Secretion of IFNγand TNFα in both two groups had no significant difference. IL-2 secretion of CD4+T cells in both two groups showed upward trend,and in IFN group,secretion of IL-2 was higher at 24 th week compared with pre-treatment(p<0.05). But there are no significant difference between different point of treatment in ETV group.4. Relationship between Antiviral efficacy and CD8+T cell(1)In all CHB patients,IFNγ secretion of CD8+T cell in group A decreased firstly and increased later,and it showed downward trend in group B. But difference between different time of treatment was not statistically significant,and difference between group A and B was statistically significant. In ETV group,secretion of IFNγ in group A offered upgrade firstly than descending latter tendency,but difference was not statistically significant. In IFN group,secretion of IFNγ in group A was higher in 24 th and 48 th week compared with group B.(2)Either in all CHB patients or ETV group,IL-2 secretion of CD8+T cell in both group A and B revealed upward trend,but,compared with pre-treatment, difference was not statistically significant. In IFN group,IL-2 secretion of CD8+T cell in group A showed upward trend,and it was significantly higher than group B in 24 th and 48 th week.5. Relationship between Antiviral efficacy and CD4+T cell(1)Either in all CHB patients or ETV group,IFNγ secretion of CD4+T cell in group A increased firstly and decreased later, but difference between group A and B was not statistically significant. In IFN group, secretion of IFN γ showed upward trend, and compared with group B, it was significantly higher in 24 th and 48 th week in group A(p<0.05).But difference was not statistically significant compared with pre-treatment.(2)In all CHB patients,secretion of IL-2 in both group A and B showed upward trend,and compared with group B,iit was significantly higher in24 th week in group A(p<0.05).But difference was not statistically significant compared with pre-treatment in both group A and B. In IFN group,secretion of IL-2 in group A showed upward trend,and compared with pre-treatment,it was significantly higher in12 th and 24 th week(p<0.05). Compared with group B, IL-2 secretion in group A was significantly higher in 12 th and 24 th week(p<0.05).Conclusion1.Antiviral efficacy of ETV and IFN:in 48 th week of treatment,virus negative rates of ETV group was higher than IFN group. But e-antigen negative rate and HBe Ag serological conversion rates of ETV group was lower compared with IFN group.2.Compared with healthy controls, TNF α secretion of CD8+T cell was significantly higher and IL-2 secretion was lower in CHB patients.3.During antiviral treatment,in ETV group,functional activity of CD4+T and CD8+T cell had changed in some degree,but it has nothing to do with the antiviral effect. IFN treatment can improve functional activity of CD4+T and CD8+T cell,and it has partial relationship with antiviral efficacy. |
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