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Title: High glucose activates ChREBP-mediated HIF-1a and VEGF expression in human RPE cells under normoxia

Author
item CHANG, MIN-LEE - Jean Mayer Human Nutrition Research Center On Aging At Tufts University
item CHIU, CHUNG-JUNG - Jean Mayer Human Nutrition Research Center On Aging At Tufts University
item SHANG, FU - Jean Mayer Human Nutrition Research Center On Aging At Tufts University
item TAYLOR, ALLEN - Jean Mayer Human Nutrition Research Center On Aging At Tufts University

Submitted to: Book Chapter
Publication Type: Book / Chapter
Publication Acceptance Date: 12/1/2013
Publication Date: 1/15/2014
Citation: Chang, M., Chiu, C., Shang, F., Taylor, A. 2014. High glucose activates ChREBP-mediated HIF-1a and VEGF expression in human RPE cells under normoxia. IN: Lambris, J.D., Cohen, I.R., Lajtha, A., Paoletti, R. Advances in Experimental Medicine and Biology. Springer, New York. 801:609-621.

Interpretive Summary:

Technical Abstract: Because retina-damaging angiogenesis is controlled by vascular endothelial growth factor (VEGF) and people with higher glucose intakes are more susceptible to retinal complications that may be due to increased VEGF, it is crucial to elucidate relations between glucose exposure and VEGF expression. We aimed to determine if a carbohydrate response element binding protein (ChREBP) plays a role in the transcriptional up-regulation of hypoxia-inducible factor-1alpha (HIF-1alpha) and the downstream VEGF expression in retinal pigment epithelial (RPE) cells exposed to high glucose under normoxic conditions. ARPE19 cells were exposed to 5.6, 11, 17, 25 and 30 mM glucose for 48 h in serum-free culture media under normoxic (21'% O2) conditions. Protein and mRNA expression of indicated genes were determined by immunoblot analyses and real-time RT-PCR, respectively. An enzyme-linked immunosorbent assay (ELISA) was used to detect the concentrations of VEGF in the media. Immunofluorescence (IF) and chromatin immunoprecipitation (ChIP) for ChREBP were used to demonstrate nuclear translocation and HIF-1alpha gene promoter association, respectively. Immunoblot analyses showed that HIF-1alpha levels were positively related to levels of glucose exposure between 5.6-25 mM in the RPE cells, indicating the induction and stabilization of HIF-1alpha by elevated glucose under normoxic conditions. Human lens epithelial cells and HeLa cells did not respond to high glucose, implying that this phenomenon is cell type-specific. Real-time RT-PCR for HIF-1alpha and VEGF and ELISA for VEGF indicated that high glucose is associated with elevated production of HIF-1alpha-induced VEGF, an established inducer of neovascularization, in the RPE cells. IF analyses showed that, although ChREBP was expressed under both low (5.6 mM) and high (25 mM) glucose conditions, it appeared more in the nuclear region than in the cytosol of the RPE cells after the high glucose treatment. ChIP analyses suggested a HIF-1alpha gene promoter association with ChREBP under the high glucose condition. These results imply that RPE cells use cytosolic ChREBP as a glucose sensor to up-regulate HIF-1alpha expression. These results suggest a high glucose-induced, ChREBP-mediated, and normoxic HIF-1alpha activation that may be partially responsible for neovascularization in both diabetic and age-related retinopathy.