Ocular infection by subgroup D adenovirus (Ad) serotypes 8, 19, and 37 causes epidemic kerato-conjunctivitis (EKC), manifested by acute pseudomembranous conjunctivitis, punctate and macro-epithelial erosions, and delayed onset stromal keratitis. In EKC, the delayed infiltration of leukocytes into the subepithelial corneal stroma causes light sensitivity, foreign body sensation, and reduced vision, sometimes lasting for months to years after infection. Our goal is to dissect the molecular mechanisms for the development of stromal keratitis in EKC, which at this time remain obscure. We hypothesize that adenoviral subepithelial infiltrates result when infection of superficial keratocytes induces a specific intracellular signal transduction cascade, leading to expression of proinflammatory mediators into the corneal stroma, that in turn cause leukocyte migration into the corneal extracellular matrix. We propose three specific aims: 1) to test the hypothesis that c-Src acts as a global regulator of intracellular signaling and chemokine gene expression in adenovirus-infected corneal fibroblasts, 2) to test the hypothesis that mitogen-activated protein kinases determine the specificity of chemokine gene expression after adenovirus infection of corneal fibroblasts, and 3) to test the hypothesis that inhibitors of intracellular signaling will reduce inflammation in the mouse adenovirus keratitis model. Our experimental design will utilize both cultured human corneal fibroblasts and a new mouse model of adenovirus keratitis, in which characteristic and reproducible leukocyte infiltration follows Ad inoculation. Interventions will include use of chemical inhibitors of cell signaling, dominant negative and constitutively active kinases, and short interfering RNA against chemokine transcription factors. Our long-term objective is to understand the interplay between adenoviruses and immune responses in the human cornea, so that information-based therapies against adenovirus keratitis can be developed. Adenoviruses represent perhaps the most common ocular pathogen in the industrialized world. The overall health relevance of these studies derives from our discovery and elucidation of novel mechanisms of innate immune responses in the cornea that cause keratitis in EKC.
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