Synthesis of Glycosaminoglycans in Rabbit Corneal Wounds after Extracapsular Lens Extraction

Objective: To determine the production site of corneal wound hyaluronan. To study glycosaminoglycan synthesis during corneal wound healing. Study Design: Extracapsular lens extractions were performed in pigmented rabbits. The cornea was labelled with [³H]-D-gluocosamine and [³⁵S]-sulphate with in-vitro incubation in enriched medium or with intracameral in vivo injections, 7, 14 and 28 days after surgery. Setting: Ophthalmic Research Department and Department of Medical and Physiological Chemistry, University of Lund, Sweden. Main Outcome Measures: Non-incorporated isotope was eliminated with ionexchange chromatography. The samples were treated with Streptomyces hyaluronate lyase, chondroitinase-ABC or keratanase II and again chromatographed to measure the incorporation of label into individual glycosaminoglycans. Synthesis of individual glycosaminoglycans is shown as the amount of [³H] and [³⁵S] incorporated dpm:s after treatment with the different enzymes. The relative synthesis of glycosaminoglycans was calculated as percentage of the total amount of labelled glycosaminoglycans. Results: There was good correlation between in-vivo and in-vitro labelling. Incorporation of D-[³H] glucosamine into hyaluronan in the operated corneas was found seven, 14 and 28 days after surgery, but could not be demonstrated in control cornea. Seven days after surgery the net synthesis of hyaluronan was about 9.5% in vitro (5% in vivo). Fourteen days after surgery the net synthesis had increased to around 19.5% in vitro (9% in vivo) and after 28 days it was around 3.5% in vitro (not detectable in vivo). The net synthesis and absolute synthesis of keratan sulphate was thoroughly decreased during the healing period in in-vitro labelled corneas. In both keratan sulphate and chondroitin sulphate/dermatan sulphate the [³⁵S]/[³H]-ratio was lowered in the operated corneas. Conclusions: Hyaluronan is synthesized in corneal wounds after extracapsular lens extraction. The synthesis of keratan sulphate is decreased in corneal wounds.