[Retroviral-mediated transfection of hepatitis B virus core gene into bone marrow-derived dendritic cells].
OBJECTIVE
To evaluate the transfection efficiency of recombinant retrovirus vector bearing hepatitis B virus(HBV) core gene to bone marrow-derived dendritic cells(DCs) and the capability of these DCs to induce cytotoxic T lymphocyte(CTL) response.


METHODS
C57BL/6 mice bone marrow cells were stimulated with recombinant mouse granulocyte-macrophage colony-stimulating factor(rmGM-CSF) and interleukin-4(rmIL-4) for 6 days. Expanding DC progenitors were transfected by retrovirus vector containing HBV core gene. Integration and transcription of HBV core gene were determined by PCR and RT-PCR, respectively. Expression of HBcAg was analyzed by fluorescence activated cell sorter (FACS) and Western blot. Cytokines were quantified by enzyme immunoassay. The expressions of CD80 and MHC class II molecules on DCs were measured by FACS. Generation of CTLs in mixed leukocyte reaction were determined by LDH release assays.


RESULTS
Transfected bone marrow cells were capable of differentiating into DCs in-vitro at the presence of rmGM-CSF and rmIL-4. The result of PCR and RT-PCR showed that the HBV core gene was integrated into the genome of infected DCs. Western blot analysis showed that HBV core gene was expressed in DCs. Transfection rate was 28% determined by FACS. Retroviral transfection had no influence on expressions of CD80 and MHC class II molecules, as well as IL-12 production. HBcAg-specific CTLs could be generated by using transfected DCs as antigen presenting cell (APC).


CONCLUSIONS
Retroviral transfected myeloid DC progenitors could efficiently express HBcAg, without significant change on development and function of DCs, which lays a solid foundation for immunotherapy of chronic hepatitis B.
Ding
Chuan-Lin
CL
Department of Immunology and Microbiology, Nanjing Medical University, Nanjing 210029, China.
Yao
Kun
K
Zhang
Tian-Tai
TT
Peng
Guang-Yong
GY
D000818:Animals; D001853:Bone Marrow; D003713:Dendritic Cells / Q000276:immunology; D006515:Hepatitis B virus / Q000235:genetics; D008810:Mice, Inbred C57BL; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology; D014162:Transfection