[Immune response of HBsAg gene-modified dendritic cell-based vaccine in HepG2. 2. 15 hepatocellular carcinoma cells].
OBJECTIVE
To study the cytotoxic T lymphocyte (CTL) response induced by dendritic cells (DC) transduced with recombinant adenovirus vector bearing hepatitis B virus surface antigen (HBsAg) gene in hepatocellular carcinoma HepG2. 2. 15 cells in vitro.


METHODS
Full length HBsAg cDNAs were subcloned into pIND vector, followed by being cloned into pShuttle vector. The HBsAg gene fragments resulted from the pShuttle-S digested with PI-Sce and I-Ceu were linked to the linear adeno-X virus DNA. After packaged with HEK293 cells, the adenovirus expression vector was obtained. Then the recombinant adenovirus expression plasmid AdVHBsAg was transfected into human monocyte-derived dendritic cells, to construct AdVHBsAg hepatocarcinoma tumor vaccine. The effectiveness of transfection was detected by Western blot. Surface molecules of AdVHBsAg-DC were detected by FACS. Autologous T cell proliferation stimulated by AdVHBsAg-DC was detected by 3H-TdR assay. Cytotoxic CTL activity induced by AdVHBsAg-DC in vitro was detected by LDH assay.


RESULTS
HBsAg gene in the inserted DNA of AdVHBsAg was confirmed by PCR, and predictive fragments proved by restriction enzyme digestion analysis were exhibited. Cell pathological changes appear after 10 days HEK293 cells transfected AdVHBsAg. Western blot analysis showed that HBV surface antigen gene was expressed in transfected DC, indicating that the transfection was effective. AdVHBsAg-DC was able to upregulate CD1a, CD11c, CD80, CD86 and HLA-DR. Autologus T cell proliferation induced by AdVHBsAg-DCs was significantly higher than that in DC control group and LacZ-DC group (P < 0.05). AdVHBsAg-DC activated CTL presented the specific killer ability to the hepatocellular carcinoma cells expressing HBsAg.


CONCLUSIONS
DC transduced with recombinant adenovirus HBsAg can express HBV-related hepatocellular carcinoma antigen (HBsAg), and AdVHBsAg-DC can induce potent immune response against HBsAg-positive hepatocellular carcinoma cells in vitro.
Yang
Jing-Yue
JY
Department of Oncology, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China.
Liu
Wen-Chao
WC
Cao
Da-Yong
DY
Si
Xiao-Ming
XM
Teng
Zeng-Hui
ZH
D000256:Adenoviridae / Q000235:genetics; D018949:Antigens, CD1 / Q000378:metabolism; D039521:CD11c Antigen / Q000378:metabolism; D019496:Cancer Vaccines / Q000276:immunology; D006528:Carcinoma, Hepatocellular / Q000276:immunology / Q000473:pathology / Q000821:virology; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D003602:Cytotoxicity, Immunologic / Q000276:immunology; D003713:Dendritic Cells / Q000166:cytology / Q000276:immunology* / Q000378:metabolism; D005822:Genetic Vectors; D006514:Hepatitis B Surface Antigens / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008113:Liver Neoplasms / Q000276:immunology / Q000473:pathology / Q000821:virology; D010957:Plasmids; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013602:T-Lymphocytes, Cytotoxic / Q000166:cytology / Q000276:immunology; D014162:Transfection
D018949:Antigens, CD1; D039521:CD11c Antigen; C419393:CD1a antigen; D019496:Cancer Vaccines; D006514:Hepatitis B Surface Antigens; D011994:Recombinant Proteins