In many ways the process of corneal wound-healing resembles such processes of embryonic development as cell migration, reabsorption of tissues and synthesis of new tissue components. However, injuries to corneas frequently lead to blindness because the healing processes seldom restore normal tissue function. Our goal is to provide a better understanding of these devastating processes at the molecular and cellular levels. It is hoped that with greater understanding, better treatments can be designed to prevent the loss of vision. At least three possible mechanisms can account for the failure of corneal wound-healing to regenerate normal tissues. They are: 1) unidirectional cell differentiation; 2) altered cell-cell interactions, i.e., direct cell-cell contact and humoral factors; 3) altered cell-matrix interactions. We believe that cell-cell interactions and cell-matrix interactions play the major roles in modulating corneal wound-healing. In order to provide a better understanding of corneal wound healing, it is necessary to identify proteins which are synthesized by injured corneal tissues but not by normal corneas, or vice versa. To obtain this information, cDNA libraries have been prepared from mRNA from normal and alkali-burned rabbit corneas. 32P-Labeled cDNA from mRNA from normal and injured corneas have been used to screen the replicas of cDNA libraries. Using this procedure, we have isolated 18 cDNA clones encoding mRNA abundant in alkali- burned corneas. We are currently characterizing these cDNA clones by restriction endonuclease digestion, DNA sequencing, slot-blot hybridization, in situ hybridization and Northern-blot hybridization. To further characterize the proteins encoded by the cDNA clones, antisera will be prepared against the synthetic oligopeptides using information available from cDNA sequences. This series of experiments will yield useful information regarding the syntheses of various proteins during corneal wound-healing. In addition the specific proteins synthesized by normal or alkali- burned corneas will be analyzed by 2-dimensional gel electrophoresis. Furthermore, synthetic oligonucleotides complimentary to the amino acid sequence of proteins of interest will be used to isolate the cDNA clones from cDNA libraries. In another series of experiments, we will characterize factors which inhibit the epithelial cell proliferation of injured rabbit corneas.
