Corneal wound-associated glycoconjugates analyzed by lectin histochemistry

Purpose. To examine changes in corneal glycoconjugates during wound healing in a pig model of refractive surgery. Methods. Pig corneas were wounded using a mechanized microkeratome. Lamellar keratectomy, without replacement of a corneal cap, or automated lamellar keratoplasty, with replacement of a corneal cap, was performed. Corneas were prepared for microscopy 1, 7, or 28 days after surgery. Cryosections of corneas were probed with lectins recognizing a variety of carbohydrate moieties. Glycoconjugates were characterized by glycosidase digestion and chemical extraction. Results. Of the lectins used, only that from Dolichus biflorus (DBA), specific for terminal N-acetylgalactosamine, exhibited a dramatic change in staining pattern after wounding. In unwounded corneas, DBA labeled the superficial epithelium, Bowman's layer, and occasional keratocytes. After wounding, DBA intensely labeled exposed stromal surfaces and keratocytes near wound sites. Results of glycosidase treatments indicated that the DBA-binding material contains alpha-linked N-acetylgalactosamine. After extraction with lipid solvents, stromal DBA labeling was unchanged, while that of keratocytes was greatly diminished. Conclusions. Corneal wounding results in the accumulation of specific glycoconjugates at surfaces and in keratocytes. After keratectomy, the exposed stromal surface is infiltrated with ocular surface components, including DBA-binding glycoproteins. The presence of these components may influence the course of wound healing. In addition, changes in keratocyte glycoconjugates may accompany their mobilization to the wound-repair phenotype.