Elucidating the pathomechanisms of aniridia related keratopathy in Pax6^Sey-Neu mice

Abstract

Purpose : Aniridia-associated keratopathy (AAK) is a progressive disorder found in individuals with aniridia, marked by the transition from subclinical abnormalities in infancy to clinical keratopathy in adulthood. Clinical features include inflammation, subepithelial fibrosis, and opacification. Despite several proposed pathophysiological mechanisms, the cellular and molecular changes driving AAK remain poorly understood. This study aims to investigate the hypothesis that Pax6 deficiency in ocular surface cells plays a crucial role in shaping and altering the corneal microenvironment, significantly influencing the progression of AAK.

Methods : Localized changes in vascular structure or nerve fibers can provide essential insights into changes in a tissue’s microenvironment. Neovascularization and decreased corneal sensitivity are characteristic of AAK. As a first test of our hypothesis, we evaluated changes in blood vessel and nerve fiber growth between Pax6^+/+ and Pax6^Sey-Neu mice corneas. Corneas were collected at P0, P7, P15 and adult time points. Blood vessels and nerve fibers were visualized by indirect immunofluorescent labeling using antibodies directed against CD31 and b-III tubulin, respectively, and confocal microscopy. Changes in gene expression were assessed by quantitative RNA-based assays.

Results : Blood vessel growth is evident in the corneas of Pax6^Sey-Neu mice by P15 shortly after eye-opening. Notably, qPCR results indicate alterations in the expression of notch signaling components, with a significant increase in the levels of Delta-like 4 transcripts in Pax6^Sey-Neu corneas. While wild-type and Pax6^Sey-Neu mice exhibit thick corneal nerve fiber bundles at birth, a noteworthy shift occurs post-eye opening. In Pax6^Sey-Neu mice, thick nerve fiber bundles persist in the cornea, contrasting with numerous smaller fibers in wild-type corneas. This pattern is seen in adulthood, with thick nerve fiber bundles persisting in the corneas of Pax6^Sey-Neu mice, particularly in the region affected by keratopathy. Differential transcript analyses of Pax6-normal and Pax6-deficient corneal cells indicate changes in additional signaling pathways.

Conclusions : Our current data suggest that at birth, the corneal microenvironment in Pax6^Sey-Neu mice is more similar to that of wild-type mice but changes as the mouse ages. Notable changes include altered expression of factors associated with angiogenesis and nerve growth.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.