Changes of hepatitis B core antigen-specific cytotoxic T lymphocytes in chronic hepatitis B patients during antiviral treatment and relapse after withdrawal of treatment

Abstract

Objective To explore the potential mechanism of severe liver injury shortly after withdrawal of antiviral therapy in chronic hepatitis B (CHB) patients. Methods Forty-nine patients with chronic hepatitis B virus (HBV) infection from the Department of Infectious Diseases of the First Affiliated Hospital of Nanchang University and 8 healthy volunteers from August 2014 to March 2015 were included in this study. All of them were human leukocyte antigen (HLA)-A2-positive. CHB patients were classified into three groups, including 15 cases in immune-tolerance group, 20 cases in sustained antiviral treatment group, and 14 cases in recurrence of drug withdrawal group. The frequency of peripheral HLA-A0201-restricted hepatitis B core antigen (HBcAg)18-27 pentamer complex specific CD8+ T cells in CHB patients was analyzed by flow cytometry. Enzyme linked immunospot assay(ELISPOT) was used to detect interferon-gamma (IFN-γ) and tumor necrosis factor-α (TNF-α) secretions of HBcAg18-27-specific CD8+ T cells. The experimental data were analyzed using non-parametric U tests. Results In healthy control group, immune-tolerance group, sustained antiviral treatment group and recurrence of drug withdrawal group, the frequencies of HBcAg-specific CD8+ T cells were (0.17±0.16)%, (1.46±0.72)%, (3.24±1.60)% and (4.67±2.43)%, respectively. Compared with healthy control group, the difference were all statistically significant in the three groups (Z=-3.583, -4.018 and -3.823, respectively; all P<0.01). The frequencies of HBcAg-specific CD8+ T cells in immune-tolerance group or recurrence of drug withdrawal group were both significantly different from that in sustained antiviral therapy group (Z=-3.400 and -2.030, respectively; both P<0.05). The difference between immune-tolerance group and recurrence of drug withdrawal group was also significant (Z=-3.230, P<0.01). The secretion levels of IFN-γ of HBcAg-specific CD8+ T cells in healthy control group, immune-tolerance group, sustained antiviral treatment group and recurrence of drug withdrawal group were 2 (0-6), 16 (2-53), 106 (14-254) and 156 (28-395) spot forming cell (SFC)/106 peripheral blood mononuclear cell (PBMC), respectively. The differences between healthy control group and immune-tolerance group, sustained antiviral treatment group or recurrence of drug withdrawal group were all statistically significant (Z=-3.585, -4.069 and -3.824, respectively; all P<0.01). The IFN-γ level of HBcAg-specific CD8+ T cells in recurrence of drug withdrawal group was significantly higher than that in sustained antiviral therapy group (Z=-2.205, P=0.027), and that in sustained antiviral therapy group was significantly higher than that in immune-tolerance group (Z=-4.700, P<0.01). The TNF-α levels secreted by HBcAg-specific CD8+ T cells in each group were 2 (0—5), 16 (2—32), 112 (15—283), and 195 (55—537) SFC/106 PBMC, respectively. The differences between healthy control group and immune-tolerance group, sustained antiviral treament group or recurrence of drug withdrawal group were all statistically significant (Z=-3.619, -4.069 and -3.824, respectively; all P<0.01). The TNF-α level secreted by HBcAg-specific CD8+ T cells in recurrence of drug withdrawal group was significantly higher than that in sustained antiviral therapy group (Z=-2.449, P=0.014), and that in sustained antiviral therapy group was significantly higher than that in immune-tolerance group (Z=-4.350, P<0.01). Conclusions The changes of frequency and immune function of HBcAg-specific CD8+ T cells in CHB patients may be one of the reasons causing severe liver damage after irregular withdrawal of nucleoside analogues. Key words: Hepatitis B; T-lymphocytes, cytotoxic; Antiviral treatment; Recurrence