The proposed studies are a continuation of a project to investigate the characteristics of corneal ulceration and the development of new therapeutic modalities. In the present phase of the project we will focus upon inflammatory aspects of the corneal response to trauma and the ultimate role of the polymorphonuclear leukocyte (PMN) and its mediators in the ulceration process. We will attempt to verify citrate inhibition of the PMN degranulation and phagocytosis in vivo by histological ultrastructural and clinical studies in established corneal ulcers and alkali burned eyes (rabbit). Further in vivo studies will focus on combined citrate and ascorbate therapy in alkali burned eyes and the effect of citrate on the prevention of ulceration in the cornea of vitamin A deficient eyes. The effect of citrate on mediation of the first and second waves of PMN entering the cornea after alkali burning will be ascertained. The pharmacological basis of citrate effects on PMN will be investigated by studies of leukocyte adherence in rat mesentery and in vitro by nylon fiber columns and vascular endothelial cell cultures. The effect of citrate on PMN chemotaxis, by mediators likely to be present in alkali burned corneas, will be studied in Boyden chambers and by exacting visual assay techniques. We will attempt to mimic the inhibitory effect of citrate on the PMN response in the alkali burned cornea by applying these chemoattractants to normal rabbit corneas. The pyroantimonate technique will be used to determine if citrate can affect the intracellular Ca2+ stores of PMN. Investigations on alkali burned collagen breakdown products will complete work which shows the release of a less than 1,000 M.W. substance causing PMN to undergo chemotaxis and a respiratory burst but without B-glucuronidase release. The ultrastructural characteristics, including granule counts, of PMN exposed to this factor(s) will be studied and correlated with these previous results. Subfraction of the crude preparation by high performance liquid chromatography will separate individual peptides for confirmation of stimulatory activity. Gas liquid chromatography will then be employed to do a preliminary characterization of the stimulant.
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