The Role of Regulatory T cells in Primary Infection with Epstein-Barr Virus

Abstract

Infection with Epstein-Barr virus (EBV) during adolescence results in an immunopathological disease, Infectious Mononucleosis (IM), in around 25% of cases. A role for Regulatory T cells (Treg) in IM has yet to be established. These suppressive cells may affect the well-characterised cytotoxic T cell (CTL) response to EBV and thus the level of viral persistence and reactivation, potentially creating an environment conducive to the outgrowth of EBV-infected cells and tumour development. The work in this thesis examines the frequency and functional capacity of Treg in primary EBV infection. The results show that the frequency of Treg within the CD4+ T cell population of IM patients was reduced with borderline significance (p=0.05) compared with healthy controls as revealed by fluorescence activated cell sorting. Treg function was confirmed using suppression assays on peripheral blood mononuclear cells (PBMC) from healthy controls but could not be assessed in IM patients due to low cell numbers. EBV-specific Treg function was analysed using Interferon (IFN)-γ ELISPOT assays in which PBMC from IM patients and healthy controls were stimulated with phytohaemagglutinin (PHA) and EBV peptides in the presence or absence of Treg. The IFN-gamma response of PBMC to PHA stimulation was significantly reduced in IM patients compared to healthy controls (p=0.009) but the IFN-gamma response to EBV peptides did not alter, irrespective of the presence or absence of Treg. Investigation of FOXP3 expression by immunohistochemistry provided evidence of Treg presence and preliminary data indicated an increased expression in IM tonsil sections compared with healthy tonsil sections. The proliferative responses and cytokine profiles of healthy controls, as measured by proliferation assays and ELISAs, in response to stimulation with the recall antigen PPD did not significantly alter upon the addition of latent membrane protein (LMP)-1 peptide. In IM patients, the same treatment resulted in a significant reduction in IFN-gamma (p=0.026) but no significant differences in IL-10 production or cell proliferation. The significantly reduced frequency of Treg in peripheral blood of IM patients and abundant FOXP3 expression in IM tonsils provides evidence for a Treg role in primary EBV infection. One plausible explanation is the recruitment of Treg to the site of primary infection by an as yet unidentified EBV-specific mechanism. Clarification of Treg activity in IM may expose opportunities for immunomanipulation during early stages of infection.