Objective: To observe the enhancement of cytotoxic T lymphocyte responses in patients with chronic hepatitis B following vaccination with dendritic cell stimulated by Poly (I:C) in vitro.
Methods: Peripheral blood mononuclear cells (PBMC) were isolated from whole blood by density gradient centrifugation on Ficoll-Hypaque, the adherent cells were cultured in the medium AIM-V contained recombinant human IL-4 and recombinant human GM-CSF. On day 7, one part of wells were added with Poly (I:C). On day 9, mature DCs (mDCs) were harvested and used to phenotype analysis. Both of the immature DCs and mature DCs were concultured with the autologous T cells for another two-three days. According to the source of dendritic cells concultured with the T cells, the subjects were divided into three groups: the T cells isolated from CHB group; the T cells stimulated by dendritic cells pulsed with HBcAg18-27 CTL epitope peptide group; the T cells stimulated by dendritic cells concultured with Poly (I:C) and pulsed with HBcAg18-27 CTL epitope peptide group. The function and frequency of HBV specific CTL were detected by Elispot (Enzyme-linked immunospot assay, Elispot) and tetramer staining.
Results: The average of percentage of HBV specific CD8(+) cells of total CD8(+) cells was 0.77% (0.45% approximately 1.74%) in the T cells isolated from 22 chronic hepatitis B patients and the average of the spots of antigen-specific IFN-gamma-releasing effector cells was 16 (9 approximately 28); The percentage of HBV specific CD8(+) cells of the T cells stimulated by dendritic cells pulsed with HBcAg18-27 CTL epitope peptide group rose to 1.92% (1.36% approximately 2.65%) and the average of the spots of antigen-specific IFN-gamma-releasing effector cells was 46 (30 approximately 67); while stimulated by dendritic cells added with Poly (I:C) and pulsed with HBcAg18-27 CTL epitope peptide, the percentage of HBV specific CD8(+) cells of total CD8(+) cells rose to 3.49% (2.02% approximately 4.60%) and the average of the spots was 98 (59 approximately 130). There was statistical difference between the results of Elispot and tetramer of the three groups (P < 0.01).
Conclusion: The T lymphocyte of patients with chronic hepatitis B stimulated by the autologous dendritic cells may result in obtaining high percentage and functional antigen-specific T lymphocyte. The addition of Poly (I:C) which was used as maturation promoting factor in DC culture can enhance the function of DC significantly and can get even higher percentage antigen-specific T lymphocyte with enhanced function.