Nitric Oxide-Mediated Neurotoxicity after Excimer Laser Photorefractive Keratectomy

ABSTRACT

The aim of this study was to evaluate neurotoxicity of Nitric oxide(NO) on cornea after excimer laser photorefractive keratectomy(PRK). PRK was performed on rabbit eyes. According to the time table, tear samples were collected with microcapillary tubes and corneal sensitivity was measured with a Cochet-Bonnet esthesiometer. No generation in the tear fluid was analyzed. To demonstrate NO Synthase(NOS), immunohistochemical localization was performed on frozen sections from rat eyeball tissue. Western blot analysis was used for detection of peroxynitrite, powerful oxidant of NO. NO generation was increased and reached to a maximum value(0.69+/-0.22micrometer/microgram) after 96 hours of PRK, as compared with in normal subjects(Mean 0.30+/-0.08micrometer/microgram) and was not increased in the treated group with topical application of Ng-nitro-L-arginine methyl ester(L-NAME), a competitive inhibitor of constitutive NOS(cNOS) and inducible NOS(iNOS). Corneal sensitivity decreased below pretreatment levers after three postoperative days, but it was not observed in the L-NAME applied group. We have confirmed that a very strong iNOS and BNOS immunoreactivity was present in corneal keratocytes. Western blot analysis identifed the bands of nitrotyro-sine-proteins suggesting in vivo peroxynitrite toxicity. Our results suggested that NO generated from the enzyme after PRK decreased corneal sensitivity by damaging corneal sensory nerve through the NO and iths oxidant peroxitrite. Therefore topical application of a NOS inhibitor may be effective in maintaining corneal sensitivity.