Purchase this article with an account.
Keith D. Rochfort, Lara S. Carroll, Peter Barabas, Timothy M. Curtis, Balamurali K. Ambati, Niall Barron, Philip M. Cummins; COMP-Ang1 Stabilizes Hyperglycemic Disruption of Blood-Retinal Barrier Phenotype in Human Retinal Microvascular Endothelial Cells. Invest. Ophthalmol. Vis. Sci. 2019;60(10):3547-3555. doi: https://doi.org/10.1167/iovs.19-27644.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Current treatments for diabetic retinopathy (DR) have considerable limitations, underpinning the need for new therapeutic options. In this article, the ability of an engineered angiopoietin-1 variant (COMP-Ang1) to ameliorate the injurious effects of hyperglycemia on barrier integrity in a human retinal microvascular endothelial cell (HRMvEC) model is comprehensively investigated.
Confluent HRMvECs were treated (0–72 hours) with d-glucose (5 or 30 mM) in the absence and presence of COMP-Ang1 (10–200 ng/mL). l-glucose (30 mM) was used as osmotic control. Posttreatment, intact cell monolayers were monitored for permeability to FITC-dextran 40 kDa. Cells were also harvested for analysis of interendothelial junction targets by RT-qPCR and Western blotting. The impact of receptor tyrosine kinase Tie2 gene silencing on COMP-Ang1 efficacy was also evaluated.
Treatment with 30 mM d-glucose (but not l-glucose) demonstrated a time-dependent elevation in the mean rate of FITC-dextran diffusion across intact HRMvEC monolayers, in parallel with significant reductions in mRNA/protein levels of occludin, claudin-5, ZO-1, and VE-Cadherin. These effects were all attenuated by COMP-Ang1 in a concentration-dependent fashion, with 200 ng/mL recovering barrier function by ∼88%, and recovering reduced interendothelial junction protein levels by more than 50%. Finally, Tie2 knockdown by small interfering RNA silencing blocked the ability of COMP-Ang1 to mitigate against hyperglycemia-induced permeabilization of HRMvECs and depletion of junctional expression levels.
In summary, this article presents a reproducible in vitro cell study that quantifies the concentration-dependent efficacy of COMP-Ang1 to mitigate the injurious effects of hyperglycemic challenge on HRMvEC barrier properties via Tie2-mediated signaling.
This PDF is available to Subscribers Only