NADPH oxidase 1-PKCδ-dependent ZO-1 phosphorylation mediates IL-33-induced inner blood-retinal barrier disruption in proliferative retinopathies

Proliferative retinopathies are associated with abnormal angiogenesis that can result in visual impairment or vision loss. The tight junction complex regulates blood-retinal barrier integrity; however, its role in proliferative retinopathies is still at an early stage. Here, we used human retinal microvascular endothelial cells (HRMVECs) and a mouse model of oxygen-induced retinopathy (OIR) to investigate the impact of interleukin-33 (IL-33) signaling on tight junction disintegration and pathological angiogenesis. Our experimental findings demonstrate that IL-33 induces zonula occludens-1 (ZO-1) serine/threonine phosphorylation and tight junction disruption in HRMVECs. In addition, mass spectroscopy analysis revealed that treating HRMVECs with IL-33 induces ZO-1 phosphorylation at the Thr861 residue. Furthermore, we observed that NOX1-PKCδ (protein kinase C-δ) signaling modulates IL-33-induced ZO-1 phosphorylation and tight junction integrity in HRMVECs. We also observed that IL-33 depletion significantly reduces OIR-induced NOX1-PKCδ-ZO-1 signaling and vascular leakage in the ischemic retina. We also observed that the NOX1-specific inhibitor, fluoflavine (ML-090), attenuated OIR-induced NADPH oxidase activity and pathological retinal neovascularization in the ischemic retina. Thus, we infer that IL-33-mediated NOX1-PKCδ-ZO-1 signaling regulates ischemia-induced retinal endothelial cell tight junction disruption and retinal neovascularization.<b>NEW & NOTEWORTHY</b> The significance of blood-retinal barrier integrity in proliferative retinopathies, including retinopathy of prematurity and diabetic retinopathy, is still in its nascent phase. Using human retinal microvascular endothelial cells and an oxygen-induced retinopathy animal model, we demonstrate that IL-33-mediated NOX1-PKCδ-ZO-1 signaling regulates tight junction disruption and blood-retinal barrier integrity in ischemic retinal diseases.