Contribution of the clock gene DEC2 to VEGF mRNA upregulation by modulation of HIF1 protein levels in hypoxic MIO-M1 cells, a human cell line of retinal glial (Muller) cells

PurposeClock genes are components of the molecular clock. Their malfunction is thought to increase the risk of numerous diseases, including cancer. Vascular endothelial growth factor (VEGF) has a pivotal role in angiogenesis, and its expression levels are controlled by clock genes in tumor cells. Ophthalmic diseases such as age-related macular degeneration, proliferative diabetic retinopathy, and neovascular glaucoma are also associated with abnormal angiogenesis followed by upregulation of VEGF in the eye. In the present study, we aimed to uncover the relationship between clock genes and VEGF in the eye.Study designLaboratory investigationMethodsOxygen-induced retinopathy (OIR) mice were prepared to mimic hypoxic conditions in the eye. Deferoxamine (DFO) was used to mimic hypoxic conditions in human Muller cell line MIO-M1 cells. Expression levels of mRNA and protein were quantified by quantitative reverse transcription polymerase chain reaction and Western blot analysis, respectively.ResultsIn the retinas of OIR mice, the expression levels of Vegf and the clock gene Dec2 increased transiently, and their temporal profiles were correlated. Knockdown of DEC2 resulted in a significant (26.7%) reduction of VEGF expression in MIO-M1 cells under hypoxia-mimicking conditions induced by DFO (P < .05). Levels of HIF1 protein were also reduced significantly, by 60.2%, in MIO-M1 cells treated with siRNA against the DEC2 gene (P < .05). Moreover, HIF1 levels showed a significant (2.5-fold) increase in MIO-M1 cells overexpressing DEC2 (P < .05).ConclusionDEC2 could upregulate retinal VEGF gene expression through modulation of HIF1 levels under hypoxic conditions.