Blue light-induced mitochondrial oxidative damage underlay retinal pigment epithelial cell apoptosis

Abstract

Purpose : Reactive oxygen species (ROS) and the resulting oxidative stress play a pivotal role in apoptosis. We reported that Blue Light (BL) induced an oxidative stress in human retinal pigment epithelial (RPE) cells in vitro, and increased drusen deposition that triggered oxidative stress and RPE cell apoptosis in human eye specimens. We aim to determine the mechanisms underlying BL-induced damage to primary human RPE cells.

Methods : We used A2E-loaded ARPE-19 cells and 6 human donors-derived primary RPE cells. Cells were exposed or not to BL (400 – 500 nm) under a Solar Simulator (TSS-156R, OAI) set at 100 mW/cm² (equivalent to 1 sun intensity), in the presence or absence of the antioxidant N-acetyl cysteine (NAC; 1mM). Cells were analyzed for their (i) oxidative status by assessing the levels of the mitochondrial ROS, (ii) proliferation, (iii) viability, and (iv) mitochondria membrane potential (ΔΨ_M) fluctuation. We analyzed the activation of the caspases 9/3 cascade. Proteomic analyses to search for putative differentially expressed proteins were performed. Data were compared using an ANOVA and Dunnett post-hoc test for multiple comparisons with one control group. A P value < 0.05 was statistically significant.

Results : The levels of the mitochondrial superoxide anion increased significantly following RPE cells exposure to BL (P < 0.001). While increased ROS production did not affect RPE cell proliferation (P ≥ 0.88), it was accompanied by a significant decrease ΔΨ_M (P < 0.018) and increase in RPE cell apoptosis (P < 0.01). Notably, BL-induced RPE cell apoptosis resulted from the activation of the caspase cascade in a ROS-dependent manner (P < 0.01). Proteomic analyses revealed that BL decreased the expression levels of several ROS detoxifying enzymes in exposed RPE cells (P < 0.02) that will prolong the oxidative stress in these cells with a maintenance of the BL cytotoxic effects.

Conclusions : We report that BL-induced oxidative stress is cytotoxic to RPE cells. BL-mediated oxidative stress triggered mitochondria damage, and concomitant caspase cascade activation and antioxidant enzymes expression inhibition. Together, our findings bring new insights on the involvement of BL on RPE cell damage and its putative role in the progression of age-related macular degeneration. The use of antioxidants is an avenue to block or delay BL-mediated RPE cell apoptosis to counteract the disease progression.

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