Subretinal expression of Galectin-3 after light-induced retinal damage

Abstract

Purpose : Galectin-3 (Gal-3), is a β-galactoside-binding protein is purported to be a prognostic biomarker in patients with severe disease (e.g., heart failure or depression). We hypothesized that manipulation of the Vitamin A cycle during light-induced retinal damage (LIRD) could reveal whether increased Gal-3 expression is a predictor of microglia/macrophage deposition within the RPE during damage resolution.

Methods : High activity Rpe65 (L450 (^L/L)) and the low activity variant (M450 (^M/M)) were bred to Cx3CR1-GFP-mice on C57Bl/6J background. Light damage was conducted at 50,000 lux for 5 h during the dark phase of the circadian cycle. Mice were imaged and sacrificed at days 1, 3, & 7 after damage (age= P60-P365, n=3-4/group). Retinal thickness and morphology were measured via SD-OCT, cSLO, and histopathology. Tissue was collected for RPE flat mounts. GFP+ (microglia/macrophages) and Gal-3+ cells were counted and classified. ZO-1 spatial patterns were measured with CellProfiler and Imaris.

Results : Differences in damage patterns and kinetics between RPE 65 ^M/M and RPE65^L/L mice were evident in both functional and morphological outcomes. On days 1 & 3 post LIRD, GFP⁺ cells were not significantly different in number between genotypes. Later, on day 7, more GFP⁺ cells were observed in the RPE 65 ^L/L mice compared to RPE65 ^M/M and no damage controls [BJ1] (p<0.0001). Notably, the RPE65^L/L mice lost most of their ERG a-, b-, and c-wave amplitudes within the first 24-72 hours post LIRD induction compared to RPE65^M/M mice (p<0.005). This phenotype worsened by day 7 post LIRD induction (p<0.001). Retinal detachments occurred earlier in RPE65^L/L animals than in the RPE65^M/M mice. Expression of Gal-3 in RPE65^M/M mice was comparable to RPE65 ^L/L during early stages of damage (up to day 3 post LIRD induction); however, at day 7, RPE65^M/M mice exhibited low levels of Gal-3 while RPE65^L/L animals continued to express high levels of the protein heterogeneously in both RPE cells and CX3CR1-GFP⁺ cells. The distribution of GFP⁺ cells was correlated with Gal-3 expression patterns in RPE cells in a time-dependent manner after LIRD induction

Conclusions : Changes in subretinal expression of Gal-3 correlate to CX3CR1-GFP⁺ cell deposition and correspond to functional and morphological damage. Significance: Galectin inhibitory drugs may therapeutically modulate sub-retinal damage via microglia/macrophage-RPE interactions

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.