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Audrey Phone, James Lo, Alvina H. Han, Ricardo Lamy, Jay M Stewart; Glucose and Pyruvate Toxicity for Human Retinal Pigment Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2018;59(9):4025.
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Retinal consumption of glucose is several-fold higher than other tissues in human body. Human retinal pigment epithelial cells (ARPE-19) are highly metabolically active and rapidly remove glucose from the culture medium. Many authors suggest using high levels of glucose (25 mmol/L) in the media, and it has been reported that cell culture media should be replenished more frequently than every 72 hours to maintain physiologically relevant glucose concentrations. Sodium pyruvate serves as an intermediate metabolite in the glycolytic pathway, and it is commonly added to cell culture media as an additional source of energy. The purpose of this study is to determine glucose and sodium pyruvate acute toxicity levels on ARPE-19 cells.
ARPE-19 cells were obtained from the American Type Culture Collection (ATCC) and cultured on a 96-well plate for 48 hours at various concentrations of glucose (5.5, 25, 40, 60, 80, 100, 150, 200 mmol/L) and sodium pyruvate (2, 3, 4, 5, 6, 8, 10, 20 mmol/L). To measure the cell viability at these varying glucose concentrations, 10µL of the Cell Counting Kit-8 (Dojindo Molecular Technologies, Inc) was administered after the incubation period. Fluorescence was then measured spectrophotometrically at the wavelength of 450 nm. Statistical analysis was performed using ANOVA followed by Tukey’s test to compare the number of viable cells among the groups.
We observed a 26.4% decrease in cell viability at 80 mmol/L of glucose (p<0.05) and a 19.9% decrease at 10 mmol/L of sodium pyruvate (p<0.05). Higher levels of glucose and sodium pyruvate led to an even higher decrease in cell viability.
ARPE-19 cells can tolerate up to 60 mmol/L (1080 mg/dL) of glucose and up to 8 mmol/L of sodium pyruvate over a period of 48 hours in cell media culture. Glucose concentrations equal to or higher than 80 mmol/L (1440 mg/dL) lead to significant decrease in cell viability. These levels are much higher than physiological levels of glucose in blood (5.5 mmol/L; 100 mg/dL). The result shows that ARPE-19 cells are very resilient and can tolerate high levels of glucose and sodium pyruvate in cell culture, and suggests that these cells can withstand the high blood glucose levels observed in acute hyperglycemic states and diabetic ketoacidosis.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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