Interleukin-1 (IL-1) is a cytokine that has been widely implicated in inflammatory states in multiple organ systems including the eye. The potential role(s) of IL-1 in human cornea are essentially unproven. WE HYPOTHESIZE THAT IL-1 PLAYS A PIVOTAL ROLE IN CORNEAL BIOLOGY AND DISEASE. Our studies provide substantial evidence that human corneal stromal cells synthesize significant amounts of IL-1alpha and IL-1beta and express IL-1 receptor mRNA. In the past year there have been a number of reports supporting our hypothesis regarding the importance of IL-1 in corneal disease. The long term objective of our program is to define in a comprehensive manner the source and role of IL-1 in corneal biology.
The Specific Aims of this project are: (1) To assess the source(s) and modulation of IL-1 in human cornea; (2) To determine if IL- 1 is able to modulate human corneal cell: (i) proliferation, (ii) collagen and collagenase production, (iii) cellular adhesion molecule expression, and (iv) cytokine production; (3) To measure the expression of (i) IL-1 receptor and (ii) IL-1 receptor antagonist (IL-1ra) by normal corneal cells; (4) To assess the in vivo role of IL-1 in mediating corneal inflammation, neovascularization, allograft rejection, and wound healing using an animal model and to determine the feasibility of blocking corneal IL-1 activity by recombinant IL-1ra and/or the IL-1ra gene in an HSV-1 (herpes) vector system. The experimental design/methods of this project are: (1) IL-1 production and modulation by freshly isolated or early passaged epithelial, stromal, or endothelial cells will be determined by bioassay, Northern blot, and immunoblot; (2) IL-1 modulation of corneal cells; (i) proliferation will be determined by direct cell counts and 3H-thymidine uptake, (ii) collagen and collagenase production will be measured by Northern blot and immunoblot, (iii) cellular adhesion molecule expression will be measured by FACS analysis and immunohistochemistry and (iv) cytokine production will be determined by Northern blot, bioassay and ELISA; (3) The expression of corneal cell (i) IL-1 receptor expression will be measured by Northern blot, Scatchard analysis, and in situ hybridization and (ii) IL-1ra will be measured by Northern blot and ELISA; (4) In vivo expression of IL-1 in corneal inflammation, neovascularization, allograft rejection, and wound healing in an animal model will be assessed by immunohistochemistry, in situ hybridization, and corneal IL-1 production will be inhibited by local delivery of recombinant IL-1ra peptide and/or the IL-1ra gene in an HSV-1 deletion mutant vector system. In selected experiments the human corneal cell culture studies will be compared to cultured human corneal explants by immunohistochemistry and in situ hybridization. These studies will lead to new and important information regarding the role of IL-1 in human corneal biology and disease. In addition, the use of recombinant IL-1ra protein or the IL-1ra gene in an HSV-1 vector in the cornea could be a paradigm for the use of specific cytokine antagonists or agonists in corneal disease.
