EBNA1-specific CD4+ T cells in healthy carriers of Epstein-Barr virus are primarily Th1 in function
Kara Bickham, Christian Münz, Ming Li Tsang, Marie Larsson, Jean-Francois Fonteneau, Nina Bhardwaj, Ralph Steinman
Kara Bickham, Christian Münz, Ming Li Tsang, Marie Larsson, Jean-Francois Fonteneau, Nina Bhardwaj, Ralph Steinman
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EBNA1-specific CD4+ T cells in healthy carriers of Epstein-Barr virus are primarily Th1 in function

Abstract

The Epstein-Barr virus (EBV) nuclear antigen-1 (EBNA1) maintains the viral episome in all host cells infected with EBV. Recently, EBNA1 was found to be the main EBV latency antigen for CD4+ T cells and could be recognized in cultures from all donors tested. We now identify a polarized Th1 phenotype and obtain evidence for its presence in vivo. When T cells were stimulated with dendritic cells infected with vaccinia vectors expressing EBNA1, 18 of 19 donors secreted IFN-γ, whereas only two of 19 secreted IL-4. Magnetic selection was then used to isolate cells from fresh blood based on EBNA1-induced cytokine production. Specific IFN-γ CD4+ cell lines were established from six of six donors and IL-4 lines from three of six. Only the Th1 lines specifically lysed targets expressing three different sources of EBNA1 protein. When the IgG isotype of EBNA1 plasma Ab’s was tested, most specific Ab’s were IgG1 and of a high titer, confirming a Th1 response to EBNA1 in vivo. Ab’s to other microbial antigens generally were not skewed toward IgG1. Given emerging evidence that Th1 CD4+ T cells have several critical roles in host defense to viral infection and tumors, we propose that EBNA1-specific CD4+ Th1 cells contribute to resistance to EBV and EBV-associated malignancies.

Authors

Kara Bickham, Christian Münz, Ming Li Tsang, Marie Larsson, Jean-Francois Fonteneau, Nina Bhardwaj, Ralph Steinman

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EBNA1-specific responses can be detected at very low doses of antigen. A...
EBNA1-specific responses can be detected at very low doses of antigen. A recombinant EBNA1 protein or control PCNA protein was eluted from E. coli–expressing vectors. Proteins were dialyzed overnight and tested for purity with SDS PAGE (left) (a). The recovered rEBNA1 protein was tested for specificity by Western blot analysis using an anti-EBNA1 Ab MAB8173 (right). The Ab AD1.1.10. recognizes a histidine tag contained in the rEBNA1 protein. The vvEBNA1ΔGA-infected DCs were used to expand CD4+ T cells in a 1-week culture. The expanded T cells were restimulated using DCs pulsed with the indicated concentration of rEBNA1 protein or rPCNA control protein and read out using ELISPOT (b). The rEBNA1 protein was added to the DCs during the maturation phase (days 6–8) of the DC culture. The inset graph shows the ELISPOT results of CD4+ T cells expanded with vvEBNA1ΔGA-infected DCs for 1 week and restimulated with either vvEBNA1ΔGA-infected DCs or vvTK– control. LMW, low molecular weight marker.