Introduction: Varicella Zoster Virus (VZV) causes varicella (chicken pox) upon primary infection; establishes life-long latency, and causes herpes zoster (shingles) upon reactivation. Evidence suggests that T cells are important in the control of varicella zoster virus (VZV) replication. The immediate early proteins, including IE4 & IE63, are thought to be potential targets for the host immune system, as they are expressed during all phases of the viral life cycle and are essential for viral replication. Methods: Using IFNγ ELISpot and intracellular cytokine secretion assays we tested the hypothesis that detectable IE63-specific T cells circulate in the peripheral blood of healthy immune donors. We have subsequently identified and utilized the first VZV-specific MHC class II tetrameric complex to characterize the ex vivo phenotype of IE63-specific CD4+ T cells in adults with a history of varicella infection. Results: Whilst non-immune donors (n = 12) showed no response at any time point tested, responses were observed to a range of the 27 overlapping peptides spanning IE63 in all immune donors (n = 24). We identified several potential epitopes, and determined the HLA restriction using partially HLA-matched targets and blocking antibodies. One epitope, restricted by HLA-DRB1*1501, has been further characterised using an HLA tetrameric complex. The tetramer positive cells were present at a mean of 0.00575% of CD4+ T cells, and are comparable to those seen in other herpes virus infections. To characterize the activation and differentiation status of the tetramer+ CD4+ T cell population we performed detailed phenotypic analysis using a large panel of markers, and showed that the responding cells have a mixed early-intermediate differentiation phenotype. Discussion: Our results show IE63-specific T cells can be identified ex vivo in the peripheral blood of healthy immune donors many years post primary infection, and using HLA-class II tetrameric complexes, we have shown that IE63-specific CD4+ T cells express an early-intermediate differentiation and mixed TEM and TCM phenotype, compatible with the possibility that IE63-specific T cells have a role in the control of viral re-activation. |