Potential Mechanisms of NUR77‐Mediated Signaling in the Aging Retinal Pigment Epithelium: Implications for Age‐Related Macular Degeneration

Abstract

PurposeRetinal pigmented epithelium (RPE) dysfunction is associated with age‐related macular degeneration (AMD), the leading cause of vision loss in the elderly in the western world. A better understanding of RPE‐related signaling pathways is the key to developing therapies and potential drug treatments for the early stages of AMD. Nuclear receptors are master regulators of cellular processes including lipid metabolic dysregulation, apoptosis, and inflammation, pathogenic pathways associated with AMD development and progression. Previously we found that orphan nuclear receptor member 1(NR4A1) or NUR77 is expressed by human RPE. The role of NUR77 in the eye or RPE‐mediated signaling pathways, however is not known. Given NUR77’s purported role in regulating inflammation, angiogenesis, and apoptosis in a number of body tissues, herein we investigated the role of NUR77 in RPE as a function of age.MethodsTotal RNA was extracted from RPE/choroid fractions of young and old mouse eyes, RPE/choroid from human donors, and human primary RPE cultures harvested from young and old eye tissue (ages 18‐93). Expression of NUR77/Nurr77 and its target genes were measured by qRT‐PCR. Cellular bioenergetics of young versus old RPE and NUR77 overexpressing RPE were measured using the Seahorse XF Real‐Time ATP Rate Assay. RPE cells overexpressing NUR77 were analyzed for expression of senescence‐related and epithelial‐mesenchymal transition (EMT) markers. To investigate the function of Nur77, in vivo, ocular imaging and retinal function assessments of 4‐8 month old Nur77‐null mice was done. Ex vivo, ocular morphology was examined. Statistical methods for data analysis included two‐tailed Student’s t‐test and two‐way ANOVA. Values were considered statistically significant at p < 0.05.ResultsNur77/NUR77 expression in mouse and human RPE decreases with age. Overexpression of NUR77 (induction of the ‘young phenotype’) in RPE cells isolated from old donors results in a decrease in expression of the senescence associated genes p21 and p16, while low levels of NUR77 were associated with elevated senescence. Expression of the metabolic markers COX III, pyruvate kinase M1 and 2were differentially regulated in the RPE/choroid fractions from old versus young mice. Seahorse assays revealed that overexpression of NUR77 in RPE cells from young but not old donors resulted in a metabolic shift from glycolysis towards mitochondrial respiration. In the absence of Nur77, enhanced expression of markers of inflammation (CCL2, TNFa), oxidative stress (HMOX1, SOD1), metabolism (COXIII, PKM2) and EMT (N‐cadherin, Snail 2, ZEB1, FN1, Vimentin) was observed in the mouse RPE/choroid. Initial assessments of ocular imaging revealed the presence of hypopigmented regions within the posterior pole of Nur77‐null mice at 4 months of age.ConclusionsOur preliminary results indicate an age‐dependent role of NUR77 in balancing a metabolic switch towards mitochondrial respiration in the RPE. Studies are ongoing to further dissect the NUR77‐mediated signaling pathways in aging and ocular disease, and in determining if NUR77 overexpression may serve as potential therapy in diseases wherein RPE is compromised.