Abstract
Purpose :
Recently impaired glycolysis was reported in patients with age-related macular degeneration (AMD) with a high lactate/pyruvate ratio suggesting that increased lactate levels may be implicated in AMD pathogenesis. Elevated serum homocysteine (Hcy) (known as Hyperhomocysteinemia, HHcy) was reported in many AMD clinical studies, suggesting an association between HHcy and the risk of AMD. We reported a direct impact of HHcy on retinal pigment epithelium (RPE) structure, barrier function, and choroidal neovasularization(CNV). We hypothesis that HHcy contributes to AMD via inducing metabolic switch in the mitochondria, in which cells predominantly produce energy by the high rate of glycolysis simulating cancer cells “Warburg” effect, rather than by comparatively low rate of glycolysis followed by oxidation of pyruvate in mitochondria as in normal cells. Increased glycolysis results in an increase in the rate of glucose uptake and preferential production of lactate, increasing cellular acidity with subsequent activation of angiogenesis, RPE barrier dysfunction, and CNV induction.
Methods :
Evaluation of cellular energy production under HHcy was evaluated (both in vivo, using mice with HHcy (cbs+/- & cbs-/-) and in vitro, using RPE cells) by Seahorse Analyzer, multitracker-based cytofluorimetry, immunofluorescence, and the western blot (WB) experiments. The Seahorse extracellular flux (XF) analyzer evaluates mitochondrial respiration by measuring oxygen concentration and proton flux in the cell supernatant over time. These measurements are expressed as mitochondrial oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) as indicative of oxidative phosphorylation and glycolysis.
Results :
HHcy showed a significant increase of both OCR and ECAR both in CBS mice (cbs+/- &cbs-/-) and in vitro (Hcy-treated Human ARPE-19) compared to wild-Type mice retina and untreated ARPE-19 cells. Furthermore, HHcy upregulated glycolytic enzyme in RPE cells and inhibition of Glut1(the main glucose transporter in the RPE) was able to reduce glycolysis in Hcy-treated RPE and improve albumin leakage& CNV induction in Hcy-injected mouse eyes.
Conclusions :
The current study suggests that Homocysteine causes a metabolic switch from mitochondrial respiration to glycolysis in RPE during AMD. Resulting in the activation of angiogenesis via activation of VEGF. Therefore, targeting Homocysteine clearance could be a novel therapeutic strategy for AMD.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.