Objective We aimed to determine if carbohydrate response element binding protein (ChREBP) plays a role in the transcriptional up-regulation of hypoxia-inducible factor-1α (HIF-1α) and downstream vascular endothelial growth factor (VEGF) expression in retinal pigment epithelial (RPE) cells exposed to normoxia and high glucose.Methods ARPE19 cells were exposed to 5.6-30 mM glucose for 48h in serum-free culture media under normoxic (21%O2) conditions.Protein and mRNA expressions of indicated genes were determined by immunoblot analyses and real-time RT-PCR, respectively.Enzyme-linked immunosorbent assay (ELISA) was used to detect the concentrations of VEGF in the media.Immunofluorescent (IF) study and chromatin immunoprecipitation (CHIP) for ChREBP were used to demonstrate nuclear translocation and HIF-1α gene promoter association, respectively.Results Immunoblot analyses showed that HIF-1 o levels were positively related to glucose concentrations between 5.6-25 mM in the RPE cells, indicating the induction and stabilization of HIF-lα by 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-1α and VEGF and ELISA for VEGF indicated that high glucose is associated with elevated production of HIF-1α-induced VEGF, an established inducer of neovascularization, in the RPE cells.IF and ChIP analyses suggested that RPE cells use cytosolic ChREBP as a glucose sensor to up-regulate HIF-1α expression.Conclusion These results suggest a high glucose-induced, ChREBP-mediated, and normoxic HIF activation that may be partially responsible for the neovascularization in both diabetic and age-related retinopathy.