Coordinated activation of corneal wound response genes in vivo as observed by in situ hybridization

We used subtractive screening of a cDNA library prepared from corneoscleral rims after cauterizing rat corneas. We identified 76 clones whose corresponding mRNA increased during the wound healing process in an in vivo model of injury which damages the corneal epithelium, stroma, and endothelium. Of these clones, 31 sequences encode known proteins. Another 45 clones are novel sequences based on comparison with the GenBank/EMBL databases. Changes in the level of expression of the novel genes, and a selected number of the known genes, were examined by in situ hybridization 22 and 72 hr after corneal injury. The majority produced a ‘wound pattern’ of expression such that the mRNAs were highly induced in all cell types adjacent to the wound site at 22 hr post injury. This signal decreased in intensity with distance from the wound site. In a subset of corneoscleral rims examined by in situ hybridization, the mRNAs for these genes were also highly induced in the limbal epithelium, where the progenitor corneal epithelial stem cells reside. By 72 hr, when acute tissue damage had been repaired, the induced mRNA was only faintly present in the thickened epithelium. Our results provide a useful framework for further studies defining the pathophysiological roles of the known and novel proteins encoded by the isolated cDNA clones