Infected hematopoietic stem cells and with integrated HBV DNA generate defective T cells in chronic HBV infection patients

A weak T-cell response plays a key role in the persistence of hepatitis B virus (HBV) infection. We aimed to confirm that T-cell defects in patients with chronic HBV infection are associated with HBV DNA infection of bone marrow (BM) hematopoietic stem cells (HSCs). Using reverse transcription polymerase chain reaction (RT-PCR) and fluorescence in situ hybridization (FISH), we observed the transcription of HBsAg coding genes and confirmed the integration of HBV DNA in CD34(+) BM HSCs from chronic HBV infection patients. T cells were generated by coculturing the HSCs with delta-like ligand 1-expressing OP9 (OP9-DL1) cells. The phenotypes of the T cells were then evaluated by flow cytometric (FACS) analysis on days 14 and 25. The results demonstrated that fewer CD3(+)TCRa beta(+) CD3(+)CD4(+) and CD4(+)CD8(+) T cells were generated from the HSCs of the patients than from the healthy controls, (P < 0.01) but the frequency of CD3(+)D8(+) T cells was not significantly different between the two group (P > 0.05). In contrast, CD4(+)CD25(+) T cells were more in the patient group than in healthy controls (P < 0.01) on both days 14 and 25. There were fewer CD3(+)CD4(+)/CD3(+)CD8(+) cells in the patient group than in the healthy control group on day 25 (P < 0.05). Less proliferation and lower levels of IL-2 and IFN-gamma were also observed in the patient group compared with the control group (P < 0.05). These data suggest that HBV DNA infected and integrated into the BM HSCs from patients with chronic HBV infection and that these BM HSCs generated defective T cells.