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Jianhai Du, Kaitlen Knight, Christina Lieu, Connor Jankowski, Van Tran, James Hurley, Jennifer R Chao; Nutrient utilization and metabolite transport in retinal pigment epithelium. Invest. Ophthalmol. Vis. Sci. 2016;57(12):1751.
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© ARVO (1962-2015); The Authors (2016-present)
The retinal pigment epithelium (RPE) is essential to maintaining metabolic homeostasis in the outer retina. How RPE utilizes nutrients and transports metabolites is not well understood. We have used 13C tracers to study nutrient consumption and metabolite transport in human fetal RPE cells.
Human fetal RPE cells were grown for 4-6 weeks on transwell filters and then switched into culture medium with 13C tracers including glucose, glutamine and proline on either the apical or basal side. Media from the apical and basal sides were collected at 8h, 24h and 48h for analysis of labeled and unlabeled metabolites by GC-MS and LC-MS.
RPE cells utilize 13C glucose, 13C glutamine and 13C proline from both apical and basal sides. 13C label from glucose appears in intermediates from glycolysis and the mitochondrial TCA cycle including lactate, pyruvate, citrate, oxoglutarate and malate. 13C also appears in amino acids including glutamate, glutamine, serine, glycine, and alanine on both sides. Remarkably, these 13C-glucose-derived metabolites appear predominantly on the apical side at 8h (8-35 times more than basal side). Most of these metabolites are transported gradually to the basal side at 24 h and 48 h (3-9 times and 1.5-5 times more than the basal side, respectively). Both 13C glutamine and 13C proline can provide carbons for synthesis of glutamate and mitochondrial intermediates that are released primarily to the apical side. 13C from 13C-glutamine also labels pyruvate and lactate that accumulate quickly at the apical side and before being transported to basal side. RPE also exports a high level of 3-hydroxybutyrate (3-HB) to the apical side. The 3-HB does not incorporate labeled carbons from glucose, glutamine or proline.
In addition to nutrient transport, RPE also actively utilizes nutrients to synthesize and export metabolic intermediates and amino acids to the apical side to support the outer retina.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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