Expression of mRNA and protein of cyclooxygenase 2 and activation of nuclear factor kappa B in corneal epithelium following an ethanol exposure.

Abstract

Abstract: : Purpose: To examine the expression pattern of protein and mRNA of cyclooxygenase 2 (COX2) and activation of nuclear factor kappa B (NFkB) following an ethanol (Et) exposure in animal cornea to model a pre–treatment of LASEK. Methods: (1) Central cornea of one eye of adult albino rabbits (n=16) was subjected to an Et exposure (30%, 30 sec) and washed by saline under general anesthesia. After healing intervals up to Day 7, the animals were killed by intravenous overdose of pentobarbital and cornea was processed for immunohistochemistry for proteins of COX2, NFkB, phospholylated inhibitor of nuclear factor kappa B (pIkB). (2) Cornea of one eye of Wistar rats (n=26) was exposed to 20% Et (30 sec) under general anesthesia and allowed to heal up to Day 7. The animals were sacrificed and enucleated eye was subjected to in situ hybridization (ISH) for mRNA of COX2 by using radioisotope–labeled probes and to immunohistochemistry for COX2 protein. Results: Corneal epithelial cells were alive following the Et exposure in both animals. (1) In rabbit cornea COX2 protein was detected in epithelium and keratocytes from Hr 12 to Day 7 follwoing the Et exposure. NFkB protein was detected in epithelium from Hr 12 to Day 2. At Hr 12, pIkB was also detected epithelium. (2) In rat corneas, COX2 mRNA expression was detected at Hr 2 in epithelium post–treatment by ISH. COX2 protein was more markedly expressed in epithelium from Hr 6 to Day 7 and keratocytes from Hr 3 to 12. NFkB was detected in epithelial nuclei from Hr 6 to 24. pIkB was also detected in the affected epithelium at Hr 6. Conclusions: An Et exposure induces expression of COX2 and activation of NFkB in corneal epithelium and/or keratocytes in rats and rabbits. Activation of NFkB and expression of COX2 following ethanol exposure may potentially stimulate inflammatory response in cornea, suggestive of the clinical efficacy of anti–inflammatory drugs.

Keywords: refractive surgery • immunohistochemistry • gene/expression