RESUMEN
PURPOSE: Adenovirus type 19 (Ad19) infection of the human cornea results in a chronic, multifocal, subepithelial keratitis. Existing evidence suggests that early subepithelial corneal infiltrates are composed of polymorphonuclear neutrophils. In this study, the capacity of Ad19-infected human corneal stromal fibroblasts (HCFs) to produce neutrophil chemotactants (chemokines) was tested. METHODS: HCFs grown from human donor corneas and passaged thrice were infected with a corneal isolate of Ad19 or mock-infected with virus-free media. Bioactivity of the cell supernatants was tested by a neutrophil chemotaxis assay. Supernatants were assayed by enzyme-linked immunosorbent assay for the neutrophil chemotactants interleukin-8 (IL-8) and GRO-alpha. Corneal facsimiles were generated with HCFs and collagen type I, infected with Ad19, and assayed by immunohistochemistry. RESULTS: Ad19 infection of HCFs increased neutrophil chemotaxis from a baseline of 0.4+/-0.7 cells/high-powered field (hpf; mock-infected) to 21.8+/-2.3 cells/hpf (Ad19-infected). Chemotaxis was reduced by the addition of neutralizing antibodies against IL-8 and GRO-alpha. Infection of HCFs induced quantities of IL-8 protein 300- and 1000-fold over mock-infected controls at 4 and 24 hours, respectively (33 versus 11,813 pg/mL at 4 hours, and 57 versus 76,376 pg/mL at 24 hours, P< or = 0.001 for both). In contrast, GRO-alpha protein levels were only sevenfold higher at 24 hours postinfection (118 pg/mL in mock-infected controls versus 880 pg/mL in Ad19-infected cell supernatants). Neither chemokine was induced by infection of an immortalized human corneal epithelial cell line. Immunohistochemistry of infected corneal facsimiles demonstrated IL-8 in the extracellular matrix within 3 days after infection. CONCLUSIONS: Production of chemokines in infected tissues facilitates an early innate immune response to infection, and in the infected corneal stroma represents an elementary defense mechanism. Interleukin-8 may play a role in the development of subepithelial infiltrates in adenovirus keratitis.