Lipid-retinoid Accumulation in the Retinal pigment epithelial cells of Recessive Stargardt Disease is mediated by DGAT1

Abstract

Purpose : Recessive Stargardt disease (STGD1), the most prevalent juvenile maculopathy, is caused by mutations in ATP-binding transporter (ABCA4) gene. STGD1 is characterized by the deposition of lipid-rich lipofuscin in the retinal pigment epithelium (RPE). A pivotal event in the progression of STGD1 is the death of RPE cells by a mechanism yet to be elucidated. Our previous studies indicated increased lipid-retinoid load and lipid metabolism defects in the RPE cells of STGD1 models. In the current study, we aimed to define the role of Diacylglycerol acyltransferase 1 (DGAT1), an enzyme with dual activity, in the lipid and retinoid turnover in the RPE cells of the STGD1 (Abca4^-/-) mouse model.

Methods : Following normalization to GAPDH, the level of DGAT1 protein in RPE cells was determined by immunoblotting. All-trans-retinyl palmitate (at-RP) levels were quantified in RPE extracts by HPLC. DGAT1 enzyme activity was assessed using mouse RPE homogenates with all-trans-retinol substrate in the presence of palmitoyl-Co-enzyme A. Triglycerides and related-lipids levels were quantified by MS lipidomics analysis. Lipid droplets were stained by LipidTOX in RPE flatmounts and visualized by confocal microscopy. All experiments included n= 3-6 biological samples and t-test for statistical analysis.

Results : At 6-mo, DGAT1 protein level was ~6-fold higher in the Abca4^-/- vs wild-type RPE cells. These data correlated with ~3-fold higher DGAT1-mediated at-RP synthesis in the Abca4^-/- RPE homogenates. Remarkably, the Abca4^-/- mice showed an age-dependent at-RP accumulation in the RPE cells, at several-fold higher levels than age-matched wild-type mice. Importantly, lipid droplets accumulation was elevated by ~2.5 fold in 6-mo-old Abca4^-/- RPE versus wild-type, consistent with the observed higher triglycerides levels by lipidomics analysis.

Conclusions : In summary, we established that RPE lipid buildup is a key pathogenic feature in STGD1. We identified DGAT1 as a molecular driver responsible for lipid-retinoid accumulation in STGD1 RPE cells. These results further our understanding of STGD1 pathogenesis and opens the door to therapeutic avenues targeting DGAT1 activity in mitigating RPE dysfunction and cell death.

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