Expression pattern of 8–nitroguanosine in rat corneas post–ethanol exposure or post–debridement: possible marker for nucleic acid damage

Abstract

Abstract: : Purpose: To evaluate the expression pattern of 8–nitroguanosine (NGS) in rat corneal epithelium and keratocytes during healing interval following an ethanol exposure or epithelial debridement. NGS is a marker for nucleic acid damage and is reportedly express in response to nitrative cell damage/cell stress. Methods: Wistar rats (n=62) were generally anesthetized by ether inhalation. Central corneal epithelium was exposed to 20% ethanol for 30 sec and washed with phosphate–buffered saline. In another series of experiment, central cornea of one eye was mechanically debrided and allowed to heal for up to 7 days. After specific intervals, the eye in each group was processed for immunofluorescence staining for NGS. Results: No immunofluorescence for NGS was detected in untreated cornea. At 12 hr after ethanol exposure marked immunofluorescence was detected in keratocytes and faint, but positive, staining was seen in corneal epithelium. Keratocyte NGS was decreased thereafter. Expression of NGS in epithelium then became more evident with a peak at 36 hr post–ethanol treatment and turned to be negative at Day 5. In corneas following debridement NGS was expressed at day 1 and 2 and then turned to be negative. NGS was not detected in keratocytes in epithelium–debrided corneas. Conclusion: An ethanol exposure of the corneal epithelium and keratocytes transiently induced nitration of nucleic acid more rapidly and more markedly than in cells of an epithelium–debrided cornea. Nucleic acid damage upon nitrative stress by ethanol exposure implicates its potential toxicity to corneal cells.

Keywords: nitric oxide • stress response • immunohistochemistry