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Sandeep Vellanki, Ana Ferrigno, Yuliana Alanis, Brandi S Betts-Obregon, Andrew T C Tsin; Glucose effect on Rat and Human Müller Cells Viability and VEGF Secretion. Invest. Ophthalmol. Vis. Sci. 2016;57(12):4206. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Early stage Diabetic Retinopathy is manifested by excessive angiogenesis of which VEGF is a strong promoter. We hypothesize that cultured Rat Müller Cells (RMC) and Human Müller Cells (HMC) respond to high glucose by an increase in cell number and VEGF secretion.
RMC were plated at 23,000 cells/well and HMC were plated 20,000 cells/well in a 24- well plate and treated with 0mM, 5.5mM or 30mM glucose for 24 hours. The number of viable cells were then counted using Trypan Blue Exclusion Method. ELISA was used to determine VEGF levels in cell media. The amount of VEGF secreted per cell was determined by dividing VEGF level in cell media by cell number.
A concentration dependent change in cell viability was observed with glucose treatment. After 24 hours, RMC and HMC in 30mM increased in cell number by 23.6% and 46.6% respectively, compared to those in 5.5mM. In contrast, number of cells in 0mM glucose decreased by 50% and 28.3% respectively, and this glucose treatment effect is significant (One-way ANOVA; P≤ 0.05). Compared to 5.5mM the level of VEGF in cell media (pg/ml) increased by 9% in HMC and 19.6% in RMC in 30mM and by 47% in HMC and 9% in RMC in 0mM glucose (One-way ANOVA; P> 0.05). In both RMC and HMC the amount of VEGF secreted per cell decreased by 50% when glucose was increased from 0 to 5.5mM. This decrease is significant in RMC. (One-way ANOVA; P≤ 0.05). However, only a slight decrease (17%) was observed when glucose level increased from 5.5 to 30mM.
Our results show that RMC and HMC are highly responsive to change in glucose concentrations. High glucose (30mM compared to 5.5mM) significantly increased HMC cell viability but did not induced a significant change in VEGF secretion (pg/cell). At 5.5mM glucose, HMC secreted a seven-fold higher level of VEGF (pg/cell) than RMC. The mechanism of glucose-induced changes in RMC and HMC cell viability and VEGF secretion remains to be elucidated.
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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