IL‐33/ST2L Signaling Regulate Blood‐Retinal Barrier Integrity via Affecting Cadherin‐Catenin Adheren Junction Complex

Abstract

PurposeAngiogenesis, neovascularization and vascular remodeling are highly dynamic processes, where endothelial cell‐cell adhesion within the vessel wall controls a range of physiological processes, such as growth, integrity and barrier function. The cadherin–catenin adhesion complex is a key contributor to blood‐retinal barrier (BRB) integrity and dynamic cell movements. However, the pre‐eminent role of cadherins and their associated catenins in adheren junction structure and function is not fully understood. Also, the role of IL33/ST2L signaling in regulating BRB integrity is not known.MethodsUsing a murine model of oxygen‐induced retinopathy (OIR) and human retinal microvascular endothelial cells (HRMVECs), we are trying to understand the significance of IL‐33/ST2L‐signaling on endothelial barrier disruption, leading to abnormal angiogenesis and enhanced vascular permeability.ResultsOur in vivo genetic deletion studies revealed that deletion of IL‐33 resulted in reduced vascular tufts and vascular leakage. Therefore, in order to understand the molecular mechanism by which IL‐33 regulates the BRB integrity, we performed some in vitro studies using HRMVECs. The electric cell‐substrate impedance sensing (ECIS) analysis showed that IL‐33 (20 ng/ml) induce endothelial barrier disruption in HRMVECs. Likewise, FITC‐dextran permeability assay also showed that IL‐33 stimulate the endothelial dysfunction in HRMVECs. As we know, tight junctions (TJs) and adheren junctions (AJs) form the adhesion structure and maintain the BRB integrity, therefore we look for the junctional proteins involved in the IL‐33/ST2L mediated endothelial dysfunction. Furthermore, we found that IL‐33 mediates the phosphorylation of VE‐cadherin and a‐catenin in HRMVECs. Furthermore, mass‐spectroscopy (MS) analysis revealed the phosphorylation of specific Ser439 residue of a‐catenin upon IL‐33 inducement in HRMVECs. Consequently, mutation of a‐catenin at Ser439 residue restores the BRB integrity disrupted by IL‐33. In addition, we have observed that IL‐33 induce the generation of reactive oxygen species (ROS), thereby destabilize adheren junctions in HRMVECs.ConclusionsWe conclude that IL‐33/ST2L signaling plays a significant role in angiogenesis and neovascularization by regulating endothelial permeability and BBB integrity.