Currently there are no effective treatments for stimulating healing of stromal endothelial wounds caused by surgery, trauma or disease, yet inadequate healing of corneal wounds can lead to severe complications. Our long term goals are to understand the molecular mechanisms that regulate the natural healing process in the cornea, and to develop ways to stimulate corneal healing. Our hypothesis is that normal healing in the cornea is regulated by peptide growth factors produced by cells in the area of injury. These peptide growth factors operate via autocrine or paracrine mechanisms to stimulate mitosis and migration of corneal cells. This hypothesis implies that inadequate corneal healing results from insufficient production of wound stimulating factors and their receptors or the overproduction of wound inhibiting factors and their receptors. Also, it implies that addition of appropriate stimulating peptide growth factors would stimulate healing of corneal wounds. To test this hypothesis, we propose to analyze bovine and human corneal keratocytes and endothelial cells for production of mRNA for eight different growth factors (EGF, bFGF, aFGF, TGF-alpha, TGF-beta, IGF-I, IGF-II, PDGF) using cDNA hybridization techniques (dot-blot, Northern, in situ hybridization). We will determine if the levels of these growth factors and the EGF receptor change during the course of wound healing, and we will investigate what physiological substances may regulate the level of their expression. Also, we will investigate the molecular mechanism of EGF action on corneal cells by biochemically characterizing the cellular substrates which are phosphorylated by the EGF receptor kinase. To further test this hypothesis in vivo, we will measure the strength of corneal incisions in rabbits treated topically with biosynthetic bFGF, IGF- I, PDGF, or TGF-beta, and we will evaluate the effect of formulations containing growth factors on stimulating mitosis of cat corneal endothelium. Results from these experiments should further our basic understanding of the molecular mechanism regulating corneal wound healing, and indicate the potential clinical usefulness of exogenous growth factor therapy in stimulating corneal healing.
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