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P.D. Senanayake, A. Calabro, J.G. Hu, V. Bonilha, A. Darr, D. Bok, J.G. Hollyfield; Glucose Utilization by the Retinal Pigment Epithelium: Evidence for Rapid Uptake, Storage in Glycogen and Glycogen Utilization . Invest. Ophthalmol. Vis. Sci. 2005;46(13):3045.
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Purpose: To evaluate the utilization of glucose, synthesis and storage of glycogen by confluent retinal pigment epithelium (RPE) cultures with high resistance junctions. Methods: Human RPE was cultured in Millicell–[PFC] culture plates in medium containing 1 mg/ml glucose with medium changes every three days. Efficiency of separation of the apical and basal compartments was determined by transepithelial resistance. Cell cultures and the respective medium samples were collected from 1 hour to 72 hours following media change. These samples were digested with proteinase K and ethanol precipitated. Supernatant and precipitate fractions were either 2–aminoacridone (AMAC) derivatized directly or digested with glycoamylase. The digestion products were fluorotagged with AMAC and separated by electrophoresis. Glucose transporter expression in the RPE cultures was evaluated with RT–PCR. GLUT 1 localization in the RPE cell cultures was followed with immunocytochemistry. Results: Glucose concentration at the end of three days averaged 0.100 mg/ml in the basal medium and 0.010 mg/ml in the apical medium, indicating more efficient glucose uptake at the RPE apical surface. The de novo synthesis of glycogen by the RPE and the presence of glycogen degradation products followed glucose removal from the culture medium. The highest levels of glycogen and glycogen degradation products occurred at 24 hours, followed by their decline to basal levels at 72 hours. GLUT 1 was the only glucose transporter isoform expressed by the RPE cells. GLUT 1 localizes to the apical and basolateral border of the RPE cells. Conclusions: These studies are the first to demonstrate glycogen synthesis and degradation by the RPE. The rapid depletion of glucose from the culture medium suggests that RPE cell culture studies using longer intervals before medium changes may result in depletion of the energy source.
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