Ag-specific CD4+ T cells are present in peripheral blood in low frequency, where they undergo recruitment and expansion during immune responses and in the pathogenesis of numerous autoimmune diseases. MHC tetramers, which constitute a labeled MHC-peptide ligand suitable for binding to the Ag-specific receptor on T cells, provide a novel approach for the detection and characterization of such rare cells. In this study, we utilized this technology to identify HLA DQ-restricted Ag-specific T cells in the peripheral blood of human subjects and to identify immunodominant epitopes associated with viral infection. Peptides representing potential epitope regions of the VP16 protein from HSV-2 were loaded onto recombinant DQ0602 molecules to generate a panel of Ag-specific DQ0602 tetramers. VP16 Ag-specific DQ-restricted T cells were identified and expanded from the peripheral blood of HSV-2-infected individuals, representing two predominant epitope specificities. Although the VP16 369–380 peptide has a lower binding affinity for DQ0602 molecules than the VP16 33–52 peptide, T cells that recognized the VP16 369–380 peptide occurred at a much higher frequency than those that were specific for the VP16 33–52 peptide.

Herpes stromal keratitis (HSK) is a prevalent and frequently vision-threatening disease associated with herpes simplex virus type 1 (HSV-1) infection. In mice, HSK progression occurs after viral clearance and requires T cells and neutrophils. One model implicates Th1-like CD4 T cells with cross-reactivity between the HSV-1 protein UL6 and a corneal autoantigen. HSK can be prevented by establishing specific immunological tolerance. However, HSK can also occur in T-cell receptor-transgenic X SCID mice lacking HSV-specific T cells. To study the pathogenesis of HSK in the natural host species, we measured local HSV-specific T-cell responses in HSK corneas removed at transplant surgery (n ‫؍‬ 5) or control corneas (n ‫؍‬ 2). HSV-1 DNA was detected by PCR in two specimens. HSV-specific CD4 T cells were enriched in three of the five HSK specimens and were not detectable in the control specimens. Reactivity with peptide epitopes within the tegument proteins UL21 and UL49 was documented. Responses to HSV-1 UL6 were not detected. Diverse HLA DR and DP alleles restricted these local responses. Most clones secreted gamma interferon, but not interleukin-5, in response to antigen. HSV-specific CD8 cells were also recovered. Some clones had cytotoxic-T-lymphocyte activity. The diverse specificities and HLA-restricting alleles of local virus-specific T cells in HSK are consistent with their contribution to HSK by a proinflammatory effect.Herpes simplex keratitis (HSK) is a common and frequently vision-threatening disease. In the natural human host, interstitial inflammation leading to opacification of the light path occurs in 28 to 40% of persons with herpes simplex virus (HSV) infection of the corneal epithelium. Recovery of live virus from the corneal stroma, in the absence of epithelial ulceration, is uncommon, and antiviral therapy does little to prevent the progression from epithelial to stromal disease. In contrast, corticosteroids and cyclosporin A, compounds with antileukocyte effects, are therapeutic. These clinical observations and murine models have recently been reviewed (21, 37). Despite the lack of spontaneous HSV reactivation in mice, these models partially replicate human HSK in that initial productive viral infection is followed by leukocyte infiltration, virus clearance, and an interval of normal histology. A second phase of leukocyte infiltration and chronic inflammation then occurs in the absence of live virus. Hypotheses developed in these models are addressed here using human corneal specimens.Murine models show that both virus and host factors are involved in the pathogenesis of HSK. Live virus scarification into the cornea is required. Inoculation into the anterior chamber leads to antigen-specific tolerance rather than immunopathology (18, 45). The conclusions from various murine models are widely disparate, however. In the congenic mouse strains C.AL-20 and C.B-17, which differ in susceptibility to HSK, an elegant three-way molecular mimicry has been shown to explain both susceptibility and resistance ...

We used CD4 lymphocyte clones from herpes simplex virus type 2 (HSV-2) lesions or the cervix and molecular libraries of HSV-2 DNA to define HSV-2 major capsid protein VP5 and glycoprotein E (gE) as T-cell antigens. Responses to eight HSV-2 glycoprotein, tegument, nonstructural, or capsid antigens were compared in 19 donors. Recognition of VP5 and tegument VP22 were similar to that of gB2 and gD2, currently under study as vaccines. These prevalence data suggest that HSV capsid and tegument proteins may also be candidate vaccine antigens.CD4 responses to herpes simplex virus (HSV) may have several functional roles, including secretion of cytokines with antiviral and immunostimulatory activity, cytotoxic T-lymphocyte (CTL) activity, inhibition of viral replication, and B-cell help (37-39). CD4 reactivity with glycoproteins B (gB) and D is therefore one rationale for their use as HSV vaccines (40, 41). Responses to gB and gD are prevalent in peripheral blood mononuclear cells (PBMC) (31, 41), but few lesion-derived CD4 clones recognize these proteins (13). Recognition of gB or gD occurs in about 40 to 50% of bulk skin-infiltrating lymphocyte cell lines expanded from recurrent genital HSV type 2 (HSV-2) lesions (15).The spectrum of HSV T-cell antigens is expanding. Tegument protein U L 48 (VP16) contains at least eight CD4 T-cell epitopes (14, 16). Tegument proteins U L 21 and U L 49 (VP22), nonstructural protein U L 50 (dUTPase), and gC2 are antigens for HSV-2 lesion-derived CD4 T-cell clones (13,14). Typecommon epitopes in U L 21 and U L 49 are recognized by potentially pathogenic local CD4 T cells in herpes simplex keratitis (15a). CD4 clones from HSV-1 retinitis are stimulated by tegument proteins U L 46 and U L 47 (33), and gH and gL of HSV-1 are also antigenic for PBMC (36). We now add the first virion capsid protein and an additional glycoprotein to the set of known HSV T-cell antigens. We have also begun to compare the prevalence of CD4 responses to specific HSV-2 proteins in PBMC.