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Byung Joo Lee, Hyoung Oh Jun, Jin Hyoung Kim, Jeong Hun Kim; Hyperglycemia induced transcriptional regulation of aquaporins in ARPE-19 cell. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):1535.
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Aquaporin (AQP) is a plasma membrane water channel, the expression of which has been documented in retina, including the retinal pigment epithelium (RPE). Recently, the significance of outer blood-retinal barrier disruption in diabetic retinopathy has been noted and AQP expressed in RPE is suggested to play an important role in the pathogenesis of diabetic macular edema. In the present study, we identified AQP subclasses which are expressed in human RPE and determined the effect of hyperglycemia on the transcription level of AQPs. The effect of glucose concentration on the level of gap junction protein connexin 43 expression, which is related to the proliferation and differentiation of RPE, was also evaluated.
In ARPE-19, a human retinal pigment epithelial cell line, the basal expression of AQP subclasses (AQP1~12) was determined by quantitative real-time PCR (RT-PCR). Transcription levels of each AQPs in ARPE-19 cells grown under normoglycemic (5 mM glucose), hyperglycemic (30 mM glucose), or normoglycemic-hyperosmolar (5 mM glucose and 25 mM mannitol) conditions were measured with quantitative RT-PCR. The expression levels of connexin 43 protein in normoglycemic, hyperglycemic, and normoglycemic-hyperosmolar condition were evaluated with immunocytochemistry.
The mRNA of AQP 1, 2, 3, 4, 5, 6, 7, and 11 were expressed in ARPE-19 cells. When compared with normoglycemic control, the expression levels of AQP1, 3, 5, and 11 were increased by 2.0, 1.8, 1.4, and 2.5 folds respectively, after hyperglycemic treatment. Among those 4 subclasses, AQP1, 3, 11 showed increased transcription level in hyperglycemic condition, even when compared with that in normoglycemic-hyperosmolar condition: 1.2, 4.5, and 1.9 folds, respectively. The level of plasma membrane connexin 43 expression was lower in hyperglycemic group than that in normoglycemic-hyperosmolar condition.
Several subclasses of AQPs are expressed by human RPE cell line and the transcription levels of some AQP subclasses are changed according to glucose concentration. Our results show that, under hyperglycemic condition, the AQPs are up-regulated only by osmotic stimuli, but also by high glucose concentration itself. In contrast, gap junction protein connexin 43 expression is suppressed in a hyperglycemic condition.
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