[Adeno-associated virus-mediated Her2/neu gene transfection enhances the immunostimulatory capacity of human dendritic cells in vitro].
OBJECTIVE
To evaluate the immunostimulatory capacity of human peripheral blood dendritic cells (DCs) with Her2/neu gene transfection mediated by adeno-associated virus.


METHODS
The HLA genotypes of the breast cancer cells SK-BR-3 and MCF7 were determined, and the mononuclear cells from healthy donors with matching HLA genotype were isolated by Ficoll-Hypaque density gradient separation. The isolated cells were divided into two groups with or without transfection with the recombinant virus harboring Her2/neu gene. The cells were cultured for 7 days in RPMI 1640 medium supplemented with 10% AB human serum, GM-CSF, interleukin-4 (IL-4) and tumor necrosis factor-alpha (TNF-alpha). The mature DCs were then harvested from the cell culture and their phenotypes were identified using flow cytometry. MTT assay was employed to examine the specific killing activity of the T cells induced by the DCs.


RESULTS
The DCs transfected with the recombinant adeno-associated virus expressed CD1a, CD86 and CD83 at the rate of 98.10%, 99.42%, and 84.59%, and those without the viral transfection expressed the markers at the rate 92.69%, 98.07%, and 82.72%, respectively, showing no obvious differences in the phenotypes of the two DCs. The transfected DCs, however, showed markedly higher expression rates of CD40 and CD80 than the non-transfected DCs (61.02% vs 36.19%, and 97.61% vs 55.5%, respectively). The DCs, irrespective of the transfection, showed comparable capacities in stimulating T cell proliferation. The transfected DCs exhibited the capacity of inducing the T cells to specifically kill the target tumor cells, with the highest killing rate of (39.7-/+7.2)%.


CONCLUSIONS
The immunostimulatory capacity of human peripheral blood DCs are enhanced by Her2/neu gene transfection mediated by adeno-associated virus.
Cencer of Oncology, Nangfang Hospital, Southern Medical University, Guangzhou, China. tracyxulei@126.com
Lei
L
Xu
Rong-cheng
RC
Luo
Zhong-hong
ZH
Bai
Ruan-cheng
RC
Xue
Fang-ju
FJ
Wang
D001943:Breast Neoplasms / Q000473:pathology; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D003713:Dendritic Cells / Q000166:cytology / Q000276:immunology* / Q000378:metabolism; D000229:Dependovirus / Q000235:genetics* / Q000378:metabolism; D018734:Genes, erbB-2 / Q000235:genetics*; D005822:Genetic Vectors; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000166:cytology; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000096:biosynthesis* / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D014162:Transfection
D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D018719:Erb-b2 Receptor Tyrosine Kinases; D011994:Recombinant Proteins