[Effect of tenofovir disoproxil fumarate antiviral therapy on virus-specific CD8+T Cells function in patients with chronic hepatitis B].

Objective

To observe the effect of tenofovir disoproxil fumarate (TDF) antiviral therapy on HBV-specific CD8(+)T cell function in peripheral blood of patients with HBeAg-positive chronic hepatitis B, and to assess its correlation with HBeAg sero-negativeness.

Methods

Sixty-three cases with HLA-A02 restricted HBeAg-positive chronic hepatitis B who received TDF (300 mg/d) antiviral therapy were enrolled from October 2016 to July 2018. The peripheral blood CD8(+)T cells were separated at baseline and 48 weeks after treatment. The peripheral blood T cells count were detected by flow cytometry. The frequency of HBV-specific CD8(+)T cells secreting perforin, granzyme B, and interferon-γ (IFN-γ) were detected by enzyme-linked immunoblotting test. Direct and indirect contact co-culture system was established between HBV-specific CD8(+)T cells and HepG2.2.15 cells. HBV DNA was detected in the culture supernatant. Target cell mortality was calculated by lactate dehydrogenase level. Cytokines expression was detected by enzyme-linked immunosorbent assay. Virus-specific CD8(+)T cells cytokilling and non-cytokilling functions were evaluated. Measurement data of the two groups were compared by t-test or paired t-test.

Results

Viral response, biochemical response, and HBeAg seroconversion rate at 48 weeks of TDF treatment were 100%, 90.48% (57/63), and 25.40% (16/63), respectively. There was no statistically significant difference in peripheral blood T cell count when compared with baseline and control group at 48 weeks of TDF treatment (P > 0.05). At 48 weeks of TDF treatment, the frequency of HBV-specific CD8(+)T cells secreting perforin, granzyme B, and IFN-γ in CHB patients was significantly higher than baseline (P < 0.001). Furthermore, the frequency of HBV-specific CD8(+)T cells secreting perforin, granzyme B, and IFN-γ was also significantly higher in CHB patients with HBeAg negative than that of non-negative (P < 0.05). HBV-specific CD8(+)T cells had induced significant down-regulation of HBV DNA in the supernatant of HepG2.2.15 cell culture (P < 0.001) and remarkable IFN-γ and interleukin-2 secretion (P < 0.05) at 48 weeks of TDF therapy in direct and indirect contact co-culture system. However, HepG2.2.15 cells death rate induced by virus-specific CD8(+)T cells was increased only in the direct contact co-culture system (21.7% ± 6.18% vs. 16.1% ± 4.15%, P < 0.001). Compared with HBeAg non-negative patients, HBeAg negative CHB patients with HBV-specific CD8(+)T cells had induced a strong decrease in HBV DNA (P < 0.001) and an increase in IFN-γ secretion level (P < 0.05). However, the target cell death proportion difference between HBeAg negative and non-negative patients was not statistically significant (P > 0.05).

Conclusion

During TDF treatment, with the viral load reduction, virus-specific CD8(+)T cells cytokilling and non-cytokilling functions are significantly enhanced, and are closely related to HBeAg negative.