[Functional effects of dendritic cells transfected with Epstein-Barr virus latent membrane protein 2A recombinant adenovirus].
BACKGROUND
To study the effects of dendritic cells (DC) transfected with recombinant adenovirus encoding Epstein-Barr virus(EBV)latent membrane protein 2A(LMP2A)gene, and to provide evidence for further investigation on the therapeutic vaccine against EBV-associated malignancies.


METHODS
DCs were transfected with EBV-LMP2A recombinant adenovirus (Ad5-LMP2A), which was generated by homologous recombination. The expression of LMP2A protein on mature DC transfected with Ad5-LMP2A at different multiplicity of infection(MOI)was analyzed by fluorescence activated cell sorting(FACS), and the dead cells were counted by trypan blue staining. The alteration of surface markers on mature DC including CD1a, CD83, CD40, CD80, HLA?DR was detected by means of FACS before and after transfection. Meanwhile, the functions including stimulating allogenetic T cells reaction and expressing IL12 P40 mRNA on transfected DC were measured by methods of 3H-thymidine uptake and fluorescent semi?quantitative polymerase chain reaction (PCR), respectively.


RESULTS
About 80% mature DC expressed LMP2A protein and >92% cells were viable after transfection at the MOI of 200 No. significant changes in the surface markers and the cytomorphology of mature DCs were detected during the transfection. Transfected DC still have strong potential to stimulate the proliferation of allogenetic T cells and to express IL-12 P40 mRNA.


CONCLUSIONS
EBV-LMP2A gene,which was carried by adenovirus vectors, could be transferred into DC with high efficiency. The function of mature DC was not affected significantly by the transfection of Ad5-LMP2A.
Peng
Guanyong
G
Department of Microbiology and Immunology, Nanjing Medical University, Nanjing 210029, China.
Yau
Kun
K
Xie
Fangyi
F
Xu
Jijun
J
Ding
Chuanlin
C
D000256:Adenoviridae / Q000235:genetics*; D002478:Cells, Cultured; D003713:Dendritic Cells / Q000502:physiology*; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D011995:Recombination, Genetic; D014162:Transfection; D014763:Viral Matrix Proteins / Q000235:genetics*
C037662:EBV-associated membrane antigen, Epstein-Barr virus; D014763:Viral Matrix Proteins