(from abstract). There is a significant need to discover the basic molecular mechanisms responsible for corneal inflammation and ulceration in the alkali-injured eye. This project attacks the afferent limb of the inflammatory reaction by attempting to inhibit the chemoattractants which lure polymorphonuclear leukocytes (PMNs) into the damaged corneal tissue. Previous studies have discovered that the alkali-injured eye releases two chemoattractants, N-acetyl-profine-glycine-proline (N-acetyl-PGP) and N-methyl- proline-glycine-proline (N-methyl-PGP), which result directly from hydrolysis of protein. The acetylated tripeptide is the more potent chemoattractant. In the current grant proposal, the PI efforts will center on enhancing the activity of inhibitors developed from sense-antisense and phage display technology that bind to N-acetyl-PGP; many of which would probably bind to both chemoattractants. Inhibitors with the highest binding affinities to the chemoattractants will be rapidly screened for toxicity and inhibition of PMNs in the in vitro polarization assay. They will then be subjected to the ultimate in vitro test for a specific inhibitor of chemotaxis, the collagen gel-visual chemotactic assay. Successful inhibitors will then be used in a battery of in vivo tests for safety and effectivity. The effect of the inhibitors on N-acetyl-PGP will be tested in vivo by intracorneal injections using PMN invasion as a histological yardstick. The effect of each inhibitor on corneal ulceration occurring after chemoattractant and respiratory burst stimulant injection into the normal cornea is an efficient time and effort approach to an otherwise lengthy animal study. Nevertheless the ultimate test of effectiveness for the proven inhibitor will be in the alkali-injured eye. In the alkali-injury experiment, the absence of corneal ulceration and perforation is the final measure of success. There is considerable expectation that this basic pathophysiologic approach will lead to the use of novel treatments, which are based on the molecular pathology found in the alkali-injured eye.
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