Sorsby Fundus Dystrophy (SFD)-associated mutations in Tissue Inhibitor of Metalloproteinase 3 (Timp3) leads to metabolic dysfunction in the RPE and Retina.

Abstract

Purpose : Although it is well established that photoreceptor death underlies retinal degeneration (RD), the pathological mechanism(s) remain unclear. We hypothesize that aberrant energy metabolism in the retinal pigment epithelial (RPE), results in nutrient deprivation in the retina and contributes to pathology in SFD.

Methods : Proteomic and metabolic analysis of RPE and retinas from mice expressing S179C TIMP3 and age-matched littermate controls was performed. To directly explore metabolic alterations, targeted metabolomics was performed via gas chromatography-mass spectroscopy (GC-MS) with ¹³C₆ glucose isotopic tracing on murine SFD RPE and retina. Retina and RPE were isolated and separately incubated in media containing ¹³C₆ glucose for 24 hours following which intra and extra cellular metabolites were extracted with 70% methanol. Metabolite abundance and isotopic distribution of metabolites were quantified using GC/MS and Agilent MassHunter. Finally, IsoCor software was used to correct for natural isotopic abundance.

Results : Proteomic analysis identified changes in several metabolic pathways suggesting metabolic dysfunction in TIMP3 S179C RPE.Targeted metabolomics demonstrated an increased abundance of extracellular M3 labeled lactate along with intracellular TCA cycle intermediates, indicative of increased RPE glycolytic flux, suggesting metabolic dysfunction. Mass isotopic distributions reveal minimal changes which suggests that TCA flux (reflective of rate of metabolite production and breakdown) is unperturbed. In the retina, we observed decreased M3 lactate enrichment intracellularly, characteristic of decreased glycolytic flux. A decrease in abundance of retinal intracellular TCA cycle intermediates was also observed with increased enrichment of M4 glutamate, M3 fumarate and M3 malate. This suggests major perturbation of TCA cycle operations.

Conclusions : In mice expressing SFD-associated Timp3 mutations, the RPE and retina both display aberrant energy metabolism. RPE exhibits an increased glycolytic flux while the retina shows a decreased glycolytic flux along with increased TCA cycle flux. This adds to the growing body of evidence implicating metabolic dysfunction in retinal degeneration and opens the door to unique hypotheses of metabolic changes following extracellular matrix disruptions.

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.