Purpose: Leukocytes such as neutrophils contribute to the pathogenesis of corneal ulcer. The effect of the proinflammatory cytokine tumor necrosis factor (TNF)-alpha on the expression of intercellular adhesion molecule (ICAM)-1 by cultured human keratocytes was investigated because the interaction of leukocytes with ICAM-1 expressed on the surface of structural cells mediates leukocyte infiltration into tissue at sites of inflammation.
Methods: Cultured human keratocytes were incubated with various concentrations of TNF-alpha. The surface expression of ICAM-1 was evaluated by whole-cell enzyme-linked immunosorbent assay, flow cytometry, and immunohistochemistry. The abundance of ICAM-1 mRNA in cell lysate was determined by quantitative reverse transcription and polymerase chain reaction analysis. Adhesion of neutrophils to corneal fibroblasts was assayed by measuring the fluorescence of Calcein-AM-labeled neutrophils.
Results: Incubation of keratocytes with TNF-alpha induced increased expression of ICAM-1 on the surface of keratocytes in a dose- and time-dependent manner. The abundance of ICAM-1 mRNA in keratocytes was increased by the incubation of cells with TNF-alpha. Exposure of keratocytes to TNF-alpha increased the adherence of human neutrophils to these cells.
Conclusions: Stimulation of keratocytes with TNF-alpha resulted in an increase in the abundance of ICAM-1 mRNA, the cell surface expression of ICAM-1 protein, and enhanced adhesion of neutrophils to these cells. The expression of ICAM-1 on human keratocytes may thus contribute to leukocyte infiltration into the corneal stroma of individuals with inflammatory ocular diseases.