ALDEHYDE DEHYDROGENASE 1A1 IN GUINEA PIG CORNEA: MOLECULAR FEATURES AND TISSUE LOCALIZATION

Abstract

Purpose: Climatic droplet keratopathy (CDK) is a human degenerative corneal disease characterized by protein aggregation under the epithelium and caused by unfavorable environmental conditions (chronic exposure to ultraviolet radiation (UVR), constant erosions of the cornea) which induce oxidative stress. In an attempt to develop an experimental animal model for CDK, we previously studied the effect of ascorbate (antioxidant agent present in the cornea) in the cornea of healthy guinea pigs exposed to ultraviolet radiation (UVR) and erosions. Since aldehyde dehydrogenase 1A1 (ALDH1A1) also contribute to corneal preservation against oxidative stress, we decided to study the molecular aspects and localization of this enzyme in guinea pig cornea.

Methods: Total corneal extract as well as epithelium, stroma and endothelium extracts from male guinea pigs were used. The expression of ALDH1A1 was studied by means of immunoblotting, and immunofluorescence assays. The existence of isoforms for the enzyme was investigated by isoelectric focusing.

Results: ALDH1A1 from guinea pig corneas presents some unique characteristics compared with other mammals. This enzyme has a molecular weight of about 54 kDa, as another crystalline present in the cornea (ALDH3A1), previously characterized by our group in corneal epithelial cells of normal guinea pigs. Isoelectric focusing studies revealed the existence of a single isoenzyme for ALDH1A1 with a pI of approximately 9 as opposed to the two isoforms of the enzyme ALDH3A1. Immunofluorescence studies showed that ALDH1A1 enzyme as well as ALDH3A1 enzyme is expressed in epithelial cells, keratocytes and endothelial cells from the cornea.

Conclusions: In this work we identified and molecularly characterized for the first time, the enzyme ALDH1A1 in the different cellular compartments of normal guinea pig cornea. We also describe their tissue location and compare it to the location of the other crystalline (ALDH3A1).