Species Specific and Developmental Expression of Corneal Keratocyte Crystallin Proteins

Abstract

Abstract: : Purpose: Previous rabbit studies suggest an intracellular component to overall corneal transparency involving abundant expression of a few keratocyte specific crystallin proteins. The purpose of this study was to establish the generalized expression of keratocyte crystallin proteins in different species and determine the relationship between neonatal keratocyte differentiation and protein expression in the rabbit. Methods: Water-soluble proteins were isolated from adult keratocytes obtained from human (Hu), mouse (Mo), rabbit (Ra), chicken (Ch) and pig (P), and characterized by SDS-Page, tryptic peptide sequence analysis and western blotting. Developmental expression of rabbit keratocyte crystallins from day 1 to day 42 after birth were correlated with differentiation as determined by keratocyte density, proliferation (anti-Ki67 immunostaining), stromal thickness and light scattering. Results: In the 5 species evaluated keratocyte crystallin proteins transketolase (TKT; Ra,Hu,Mo,Ch), aldehyde dehydrogenase (ALDH 1/3; Ra,Hu,Mo,P), α-enolase (Hu,Ch) and G3PDH (Hu,Ch) were uniquely identified to different species showing both differing patterns and levels of expression. Keratocyte crystallins were also distinct from epithelial crystallins which included, in addition to the above, protein disulfide isomerase (Ch), annexin II (Ch), and LDH (Ra,Ch). Developmentally, increased rabbit keratocyte crystallin protein expression at day 16 after birth was associated with loss of Ki67 staining and markedly decreased keratocyte density. Interestingly, peak corneal haze observed at day 4 after birth was associated with peak Ki67 staining and elevated keratocyte density. Conclusions: Corneal keratocytes express a few abundant crystallin proteins that vary depending on species and are differentially expressed compared to corneal epithelial crystallins. Additionally, developmental expression appears linked to the quiescent keratocyte phenotype with actively proliferating keratocytes showing lowest crystallin protein expression and the greatest backscattering of light.

Keywords: cornea: stroma and keratocytes • cornea: basic science • crystallins