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Xiaoling Liang; Activation of Prolyl Hydroxylase (PHD) 2 prevents high glucose-induced blood-retina barrier breakdown in diabetic retinopathy. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):5188. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
To investigate the role of PHD2 in high glucose-induced blood-retina barrier (BRB) breakdown in human retinal microvascular endothelial cells (HRMECs) and in retina of streptozotocin(STZ)-induced diabetic retinopathy (DR) rats.
HRMECs were pre-treated with PHD2 activator (R59949) or inhibitor (DMOG) for 1 hour and were subsequently treated with 30mM high glucose for 24 and 48 hours. The 70kD FITC-dextran was used to detect the paracellular permeability and the transendothelial electrical resistance (TER) measurement was used to detect the endothelial barrier dysfunction. Levels of tight junction proteins (claudin-1 and ZO-1), HIF-1α were tested by western blot, Real-time quantitative PCR and immunofluorescence. In parallel, STZ-induced DR rats was used to evaluate the effects of R59949 or DMOG on BRB breakdown. After 1 month of STZ induction, DR rats and age-matched non-diabetic controls were evenly divided into four group: Control group (n = 30), DMSO group (n = 30), R59949 (0.1mg/200g) group (n = 30) and DMOG (2mg/200g) group (n = 30). The retina was investigated for BRB breakdown by Evans Blue Dye. Retinal HIF-1α and VEGF expressions was quantified using western blot analysis.
R59949 (PHD2 activator)ameliorated the endothelial barrier dysfunction and moderated the interrupted distribution of occludin and ZO-1 that induced by high glucose (P<0.01). The expression of HIF-1α and VEGF were reduced by 75% and 35% after being treated with R59949 (P<0.05). In contrast, DMOG (PHD2 inhibitor)substantially aggravated the high glucose-induced endothelial barrier dysfunction (P<0.01). Additionally, treatment with DMOG further elevated the HIF-1α and VEGF expression by 140% and 700% in glucose-induced HRMECs (P<0.01). In parallel, HIF-1α and VEGF expression were increased by 400% and 700% after STZ-induced in retina of rats. R59949 prevented the increase in BRB breakdownand inhibited the expressions of HIF-1α and VEGF induced by STZ. In contrast, DMOG substantially aggravated the STZ-induced BRB breakdown (P<0.01) and further elevated the STZ-induced increased expressions of HIF-1α and VEGF by 15% and 20% (P<0.05).
The results indicated that the activation of PHD2 could prevent high glucose-induced BRB breakdown in HRMECs and in retina of STZ-induced DR rats via regulating HIF-1α-VEGF signaling pathway.
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