Abstract
Purpose :
Aerobic glycolysis is a major mechanism for photoreceptor metabolism, but implications for retinal health are still being elucidated. The byproduct of aerobic glycolysis is lactate, which is transported to the retinal pigment epithelium (RPE). Diseased photoreceptors may decrease the supply of lactate to the RPE. Our study examines how this may affect RPE health in both a mouse model and iPSC-RPE.
Methods :
rhoCre:ldha fl/fl mice were sacrificed at 2 months, 6 months, and 1 year. Ldha fl/fl mice were used as controls for transmission electron microscopy (TEM). Eyes were fixed in 2.5% glutaraldehyde and 0.1 M cacodylate buffer, then postfixed in 1% osmium tetroxide/1.5% potassium ferrocyanide, taken through an ethanol gradient, equilibrated in prooplyene oxide, and infiltrated and embedded In epoxy resin for sectioning on ultramicrotome and then viewed on a JEOL 1200EX microscope. Real-time bioenergetic profiles of iPSC-RPE cells were assessed using the Seahorse XFe96. iPSC-RPE were grown in RPE maintenance media for 6 weeks, with lactate instead of glucose for the last week. Then, they were placed in Seahorse XF base medium and a Mito Stress Test was performed, with injections of oligomycin, BAM15, and a combination of rotenone and antimycin.
Results :
At two months, rhoCre:ldha fl/fl mice showed accumulation of membranous electron-lucent vesicles in Bruchs membrane. At 6 months, vesicles increased, disrupting Bruchs. At one year, there was significant accumulation of basal laminar deposits. Basal infoldings were shortened and lost. In parallel, mitochondrial cross sections became smaller and more numerous with abnormal cristae, consistent with stress. Incomplete digestion of photoreceptor segments was also seen. iPSC-RPE cells in lactate had significantly higher (p<0.05) oxygen consumption rates (OCR) on Mito Stress Test than those switched back to conventional RPE maintenance media with glucose, or with RPE maintenance media containing glucose and lactate.
Conclusions :
In a mouse model deficient in photoreceptor-derived lactate, the RPE mitochondria show signs of increased stress with age, accompanied by degeneration of Bruchs membrane and accumulation of basal laminar deposits (BlamD). iPSC-RPE cells grown in lactate show significantly higher OCR on Mito Stress Test than those in glucose. This suggests that photoreceptor-derived lactate may have a previously unrecognized benefit to RPE mitochondria and metabolism.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.