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Hannah Whitmore, Dhanesh Amarnani, Santiago Delgado-Tirado, Joseph Arboleda-Velasquez, Leo A. Kim; In vitro analysis of RUNX1 and VEGF pathway interactions. Invest. Ophthalmol. Vis. Sci. 2019;60(9):2687. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Neovascularization is a pathological feature of proliferative diabetic retinopathy (PDR). Current anti-angiogenic therapies typically target vascular endothelial growth factor (VEGF) and are effective treatments for a number of angiogenic diseases of the eye. We have recently identified glucose-dependent up-regulation of Runt-related transcription factor 1 (RUNX1) in vascular endothelial cells and its role in PDR. We have identified RUNX1 as a novel pathway of potential therapeutic interest and implicate RUNX1 as a mediator of aberrant retinal angiogenesis. The aim of this study is to investigate the role of VEGF in regulating RUNX1 expression.
Human Microvascular Endothelial cells (HMRECs) were cultured in endothelial growth media, supplemented with high concentrations of D-glucose (30 mM), h-VEGF (5 or 10 ng/ml) or aflibercept (40 mg/ml) for 12 to 48 hours. The effect of glucose and VEGF on RUNX1 mRNA levels was evaluated by Quantitative RT-PCR (qRT-PCR). Western blot analyses with antibodies against RUNX1 and β-actin were performed to confirm protein expression of RUNX1.
30 mM D-glucose increased RUNX1 RNA (1.22 ± 0.03 fold, p < 0.05) and protein expression (1.28 ± 0.03 fold, p < 0.05) in HMRECs after 48 hours of treatment. Treatment of HMRECs with 5 or 10 ng/ml of VEGF had no effect on RUNX1 RNA levels at 12 hours but caused a decrease in RUNX1 after 48 hours (0.72 ± 0.06, p < 0.05). Treatment with aflibercept caused a non-significant increase in RUNX1 RNA and protein expression.
These findings are consistent with previous findings that RUNX1 is involved in the endothelial cell response to glucose and mediate the behavior of HMRECs in angiogenesis. The results also indicate that increased expression of RUNX1 as found in pathologic ocular angiogenesis is VEGF-independent and thus suggests an alternative VEGF-independent mechanism of endothelial cell behavior in ocular pathology.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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