A2E, a Pigment of the Lipofuscin of Retinal Pigment Epithelial Cells, Is an Endogenous Ligand for Retinoic Acid Receptor

Abstract

Lipofuscin contains fluorophores, which represent a biomarker for cellular aging. Although it remains unsubstantiated clinically, experimental results support that the accumulation of lipofuscin is related to an increased risk of choroidal neovascularization due to age-related macular degeneration, a leading cause of legal blindness. Here, we report that a major lipofuscin component, A2E, activates the retinoic acid receptor (RAR). In vitro experiments using luciferase reporter assay, competitional binding assay, analysis of target genes, and chromatin immunoprecipitation (ChIP) assay strongly suggest that A2E is a bona fide ligand for RAR and induces sustained activation of RAR target genes. A2E-induced vascular endothelial growth factor (VEGF) expression in a human retinal pigment epithelial cell line (ARPE-19) and RAR antagonist blocked the up-regulation of VEGF. The conditioned medium of A2E-treated ARPE-19 cells induced tube formation in human umbilical vascular endothelial cells, which was blocked by the RAR antagonist and anti-VEGF antibody. These results suggest that A2E accumulation results in the phenotypic alteration of retinal pigment epithelial cells, predisposing the environment to choroidal neovascularization development. This is mediated through the agonistic function of A2E, at least in part. The results of this study provide a novel potential therapeutic target for this incurable condition.Retinal age pigments, or lipofuscin granula, contain the fluorophores that accumulate with age and that are thought to represent a biomarker for cellular aging (1). Lipofuscin results from an incomplete degradation of altered material trapped in lysosomes (2) and the accumulation of lipofuscin is related to an increased risk of choroidal neovascularization (CNV) 2 due to age-related macular degeneration (AMD) (1, 2). AMD is a leading cause of legal blindness in developed countries (3), and even with the recent advent of several treatment options (4), treatment of AMD remains difficult (5). Thus, a better understanding of the pathogenesis is needed to pursue a novel potential pharmaceutical target. Visual loss in AMD is caused by CNV, i.e. the neovascular vessels extending from the choroid below the sensory retina, and the subsequent atrophy of the RPE. The process preceding AMD, early age-related maculopathy (4), is pathologically characterized by age-related changes in the RPE, such as the accumulation of a deposit called drusen in the basement membrane of the RPE, i.e. Bruch's membrane, and fluorescent lipofuscin granules in the RPE cells (6). It is generally considered that the age-related accumulation of these potentially toxic deposits affects normal RPE functions (4). Although it remains unsubstantiated whether the accumulation of lipofuscin is related to the development of exudative AMD, the fact that most abundant accretion is in the RPE cells under the central retina suggests that there may be a causal relationship between lipofuscin accumulation and exudative AMD (2). Laboratory studies also support...

Di-retinoid-pyridinium-ethanolamine (A2E) Accumulation and the Maintenance of the Visual Cycle Are Independent of Atg7-mediated Autophagy in the Retinal Pigmented Epithelium

“…Cell Culture Treatments-A2E, prepared as previously described (39), was conjugated to BSA (bovine serum albumin, Sigma-Aldrich) by adding 20 M A2E to 0.2% BSA solution. The A2E-BSA mixture was incubated at 4°C for 2 days and then centrifuged to remove insoluble debris.…”

PLGA Microparticles Entrapping Chitosan-Based Nanoparticles for the Ocular Delivery of Ranibizumab

“…Wells were viewed using a Leitz Diavert phase contrast microscope and images captured using a Nikon digital camera (Nikon, Kingston upon Thames, UK). The images were then processed using Infinity Capture application (v. 6.2, Lumenera Corp., Canada) and tubules quantified using AngioSys 2.0 software (v. 1.0.0.2, TCS Cellworks, Buckingham, UK) for their length as previously described (46,47) . Data was presented as the total tubule length relative to the VEGF control, for each sample (mean ± S.D, n=3).…”

Lipofuscin Accumulation into and Clearance from Retinal Pigment Epithelium Lysosomes: Physiopathology and Emerging Therapeutics

“…Toxicity of lipofuscin could also involve other less-studied mechanisms, including mitochondrial poisoning, as it has been shown that lysosomal A2E progressively leaks into the mitochondrial compartment [100], where it destabilizes the membrane [101] and inhibits oxidative phosphorylation [102], derail of cholesterol traicking [103], activation of Retinoid Acid Receptor (RAR)-dependent VEGF secretion in RPE [104,105], and inhibition of RPE-65 isomerohydrolase activity, which limits the RPE supply of 11-cis retinal [106]. In summary, although numerous mechanisms of toxicity elicited by pathologic accumulation of LBs in RPE lysosomes have been proposed, no viable therapeutic options have resulted yet from targeting them.…”