We have previously demonstrated that CD8+ T cells, including HSV-2 specific CD8+ T cells, persisted in genital skin for months after the herpes lesion completely healed, and preferentially accumulated at the dermal-epidermal junction where sensory nerve endings were enriched. These persisting CD8+ T cells have been postulated in the containment of the reactivating virus before clinical symptomatic lesion occurs. To understand functions of persisting CD8+ T cells in human, we have combined immunofluorescent (IF) staining, laser capture microdissection (LCM) and RT-PCR to analyze the gene expression of CD8+ T cells at the dermal-epidermal junction in post-healed lesion biopsy. CD8+ T cells were captured at a single cell level. The purity was validated by the lack of CD4 and a 100-fold more enriched CD8 expression in LCM captured cells than in whole tissue sections. In captured CD8+ T cells, we detected up-regulation of genes encoding cytolytic granules, such as perforin and granzyme B, as compared to keratinocytes in the epidermis. The expression of cytolytic granules was further confirmed by IF staining of persistent CD8+ T cells, even in post-healed biopsy of acyclovir treated individual. Our data strongly indicated the cytolytic potentials of CD8+ T cells persisting at the dermal-epidermal junction and implicated their role in immune surveillance against HSV-2 reactivation in skin and mucosa.

This work was supported by NIH grants R37 AI042528 and PO1 AI030731.