Ocular Epigenetic Memory and Blindness

Abstract

Purpose : Epigenetic memory is characterized by methylation/demethylation including genomic imprinting/off-printing. Hyperhomocysteinemia (HHcy) causes metabolic dysfunction epigenetically via glutamate-mediated neurovascular cytotoxicity in eye that leads to retinal artery/vein occlusion but also increases probability of developing AMD, glaucoma and sphincter muscle atrophy. However; underlying mechanism(s) how HHcy derails ocular homeostasis remains unknown. Recent developments recognize circular RNA (circRNA) molecules as an enigmatic gene-transcripts since they have been shown to exert profound regulatory functions across animal kingdom. Our research has led to the discovery of a novel target in eye that reflects an unbiased increase in the absolute levels of circGrm4 followed by a concomitant decrease in miRNAs specific to mGluR4; hence underscoring the existence of an unknown regulatory system (circGrm4 – miRNA - mGluR4 axis). These findings highlight the importance of a ‘competitive endogenous RNA (ceRNA) network’ that operates in an environment which favors homeostatic imbalance

Methods : To determine role of circGrm4 in glutamate cytotoxicity, we treated WT and CBS^+/- (HHcy) mice with/without anti-mGluR4 antibody. Ocular levels of circGrm4, mGluR4, glutamate, DNMT1, EMMPRIN/CD147, β-Catenin, Wnt-1, MFN2, DRP1, peripherin, REDD1 and EAAT-2 were measured. Retinal architecture and intraocular pressure were recorded by optical coherence tomography (OCT) and tonometry. Vision guided behavior was also tested in a light-dark chamber and with novel object recognition test (NORT) along with measurement of retinal functions employing electroretinography (ERG). In addition, microvascular permeability was determined by FITC-BSA administration followed by intravital fluorescence confocal microscopy

Results : When results were compared with WT, the CBS^+/- strain revealed differential expression profile of key proteins such as DNMT1, mGluR4, EAAT-2/GLT-1, Wnt-1, claudin-5, etc. Structural changes indicated that retinal layers were also affected in CBS^+/− mice compared to controls. Intervention with anti-mGluR4 antibody was able to reverse many of the pathological alterations as revealed by ERG and vision guided behavior analyses

Conclusions : Our findings reveal that circGrm4 regulates the expression dynamics of mGluR4 receptor in the eye most likely by serving as a molecular sponge for the miRNAs that specific to this receptor

This is a 2020 ARVO Annual Meeting abstract.