Delineating Cellular and Molecular Mechanisms of Pre-Retinal Fibrosis

Abstract

Purpose : Retinal fibrosis is a common consequence of ischemic retinal vasculopathies such as Diabetic Retinopathy and Retinopathy of Prematurity, which can progress to tractional retinal detachments and blindness. However, little is known about the cell type(s) and molecular pathways that drive scar tissue formation. We have developed an experimental mouse model of ischemia-driven fibrosis and are using it to identify the cellular and molecular mechanisms of retinal fibrosis, with the hope of developing therapeutics to treat or prevent this process.

Methods : Mice were exposed to hyperoxia from P2 to P16 (called long oxygen-induced retinopathy, L-OIR) and compared with age-matched mice in normoxia two weeks after exiting hyperoxia. Eyes were enucleated and retinas dissected for flatmounts and brightfield imaging. Plaques were assessed by area over total retinal area using two-tailed Mann-Whitney test. Cohorts of mice were exposed to L-OIR and their age-matched controls 0, 1 and 2 weeks post-hyperoxia (WPH) and sectioned for immunohistochemistry. Sections were analyzed by length of vasculature compared to nuclear layer length. Retinal thickness was measured using distance between retinal ganglion cell and photoreceptor nuclei from three areas in two sections per eye. To identify collagen-producing cells, Col1a1-GFP transgenic mice were exposed to L-OIR and evaluated by scRNA-seq.

Results : Fibrotic plaque formation was seen 2 WPH following L-OIR compared to normoxic age-matched controls in approximately 75% of eyes (p<0.05). Compared to normoxic age-matched controls and the standard OIR (P7-P12) mouse model, intermediate and deep vascular plexuses 1 and 2 WPH fail to revascularize in L-OIR, with retinal atrophy and disorganization of retinal layers. Sequencing data from GFP+ cells in mice that underwent L-OIR in comparison to their age-matched normoxic controls demonstrate that retinal pericytes highly express collagen 1.

Conclusions : We have developed a reliable model of ischemia-driven retinal fibrosis. Following L-OIR, there is loss of the intermediate and deep capillary plexus, disorganization of the retinal layers, retinal atrophy, and deposition of scar tissue on the surface of the retina. Collagen 1, a major component of scar tissue, is primarily secreted by pericytes. Future work is aimed at understanding the molecular mechanisms that drive retinal fibrosis.

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