Abstract
Purpose :
Chronic hypoxia accompanied by decreased glycolysis in the retina and ON has been observed in retinal cells following long-term ocular hypertension. Declines in glycolysis despite its hypoxia-associated promotion suggest the retina may transition into a pseudohypoxic state, with HIF-1αstabilization despite sufficient oxygen. This project investigates the role of chronic HIF-1α stabilization in metabolic dysfunction in mouse eyes.
Methods :
To induce chronic HIF-1α stabilization, the prolyl hydroxylase inhibitor Roxadustat (Roxa) was injected into CAG-creERT2-ODD HIF-1α reporter mice for 2 or 4 weeks. These mice express tdTomato protein when HIF-1α is stabilized. Protein and mRNA isolated from retina and optic nerve was used to analyze glucose transporters, metabolic enzymes, and mitochondrial proteins. We also used the Seahorse XFe24 Analyzer to measure dependency and flexibility of Roxa retina on glycolysis.
Results :
Roxa injection resulted in tdTomato expression in the retina, confirming the stabilization of HIF-1α. Chronic HIF-1α stabilization was associated with significant changes in the levels of key metabolic proteins. There was a progressive decrease in the mitochondrial protein TOM20 and metabolic enzymes pyruvate dehydrogenase and succinate dehydrogenase. The antioxidant protein SOD2 levels also decreased over time. Glucose transporter 1 (GLUT-1), specific for glial cells, significantly decreased after 4 weeks; however, the levels of glucose transporter 3 (GLUT-3), specific for neurons, did not change. Seahorse results indicated Roxa-injected mouse retinas were less dependent on glycolysis than control retinas. More relevant for function and perhaps indicative of pseudohypoxia, chronic HIF-1αstabilization reduced the flexibility of retinas to use other metabolic substrates.
Conclusions :
Our results show that chronic hypoxia leads to significant decline in mitochondrial function in mice retinas. This is expected because hypoxia promotes glycolysis. However, the limit on glycolysis dependence along with limited metabolic flexibility indicates HIF-1αstabilization may contribute to metabolic dysfunction in glaucoma. These indicate that chronic hypoxia, as we observe in glaucoma, possibly impairs the primary metabolic pathways in mice retina and cellular response to metabolic stress.
This is a 2021 ARVO Annual Meeting abstract.