Author/Authors :
Yi Xiang-hua نويسنده Department of Pathology, Tongji Hospital of Tongji University, School of Medicine, Shanghai, China , Yong-ge Liu نويسنده Allergy Department, Beijing Key Laboratory for Pediatric Diseases of Otolaryngology, Head and Neck Surgery, Beijing Children’s Hospital, Capital Medical University, Beijing, China Yong-ge Liu , Guicheng Wu نويسنده Chongqing Three Gorges Central Hospital, Chongqing,
404000, China , Xuan An نويسنده Chongqing Three Gorges Central Hospital, Chongqing,
404000, China , Fang Wei نويسنده Chongqing Three Gorges Central Hospital, Chongqing,
404000, China , Lina Xia نويسنده Chongqing Three Gorges Central Hospital, Chongqing,
404000, China , Qing Zhong نويسنده Chongqing Three Gorges Central Hospital, Chongqing,
404000, China , Biao Du نويسنده Chongqing Three Gorges Central Hospital, Chongqing,
404000, China
Abstract :
Background Hepatitis B virus (HBV) infection is a significant
global problem. The main therapies for chronic hepatitis B are
nucleos(t)ide analogues (NAs) and (pegylated) interferon (IFN). However,
the prognosis of chronic hepatitis B is still not so optimistic since
the low serological conversion rate and the high recurrence rate after
drug withdrawal. Objectives This aim was to evaluate the killing
activity of cytotoxic T lymphocytes (CTLs) induced by dendritic cells
transduced recombinant adenovirus associated virus (rAAV) with different
hepatitis B virus gene fragment (HBV-S, C, E, X). Methods Peripheral
blood mononuclear cells (PBMCs) were extracted from chronic hepatitis B
patients , and the adhered cells were harvested to cultivate with
recombinant adeno-associated virus with different hepatitis B virus
(HBV) antigen gene fragment (rAAV-HBV-S, C, E, X) for 7 days by adding
GM-CSF, IL-4 and TNF-α to generate mature dendritic cells (DCs). The
DCs’ state were observed and their differentiation antigen molecules
(CD) were detected by flow cytometry (FACS) to evaluate their maturation
and function. DCs were co-cultured with prepared T lymphocytes to induce
cytotoxic T lymphocytes (CTLs). The activity of CTLs induced by
different DCs were compared by detection of lymphocyte CD molecules and
cytokine levels. HepaG2.2.15 cells were target cells and HepaG2 cells as
control. The specific killing activity of CTLs were compared by cell
killing assay using 3-(4,
5-dimethylthiazol-2-yl)-5(3-carboxymethoxyphenyl)-2-(4-sulfopheny)-2H-tetrazolium
(MTS). Results The expression of phenotype CD14, CD80, CD83, CD86 from
DCs transduced with rAAV-HBV-S, C, E, X and the secretion of cytokine
IL-10, IL-12 were compared, respectively. CD80 in rAAV-HBV-S was the
highest (P < 0.05), and CD83 in rAAV-HBV-X was the highest
(P<0.05), but other results were not significantly different (P
> 0.05). The response of cytotoxic T-lymphocytes induced by
different adeno-associated virus (AAV) vector delivery into dendritic
cells had significant differences. In comparing the four antigens in
terms of CD4+CD25+T cells, it was found that rAAV HBV-E group was the
highest, and rAAV HBV-X group was the lowest (P < 0.05); CD8+
CD69 + T cells in rAAV HBV-X group was the highest, and rAAV HBV-S group
was the lowest (P < 0.05); IFN-γ levels secreted by CTLs in rAAV
HBV-X group was the highest, and rAAV HBV-S group was the lowest (P
< 0.05).The specific cell killing activity of CTLs induced by DCs
transduced by rAAV-HBV-X to the HepaG2.2.15 cells was the highest, but
the four groups were no significant difference, respectively (P >
0.05). Conclusions These data suggest that rAAV-HBV-S, C, E, X delivery
into DC may be all useful for immunotherapeutic strategies against
hepatitis B virus infection and that the HBV X antigen gene may be the
most useful.
