ROR2 involvement in endothelial cells responses to flow

Abstract

Endothelial cells (ECs) are sensitive to physical forces created by blood flow, especially to shear stress (SS). ECs transduce this mechanical forces into intracellular signals leading to gene expression, cytoskeleton reorganization, cell elongation and polarization against blood flow direction. Defects in ECs adaption to blood flow are highly correlated with cardiovascular diseases including atherosclerosis. Recently, it has been shown that non-canonical Wnt signaling modulates the endothelial shear stress flow sensor in vascular remodeling. ROR2, a receptor tyrosine kinase, is known to regulate non canonical Wnt/PCP signaling. This study investigates the role of ROR2 in ECs response to flow. Different types of ECs were exposed to low orbital shear stress (OSS) (3,6 dynes/cm 2 ). After 24 h or 72 h of OSS, ROR2 and Klf2, a flow-induced gene, expression were assessed by qRT-PCR and/or Western Blot. ROR2 gain- (lentivirus) and loss-of-function (siRNA) were used to decipher its role in ECs flow response. ECs polarization under laminar shear stress (LSS) were studied using laminar flow chamber slide. After 18 h of LSS (5 dynes/cm 2 ), ECs were immunostained for VE-cadherin, WGA, ROR2 and pericentrin. As expected, OSS induced Klf2 expression as compared to static condition. Interestingly, ROR2 expression was induced at the protein and gene level by OSS. To analyse whether ROR2 could be upstream or downstream Klf2 induction, ECs were treated with ROR2 or control siRNA and subjected to OSS for 24 h. siRNA depletion of ROR2 decreased flow-induced Klf2 expression. In LSS conditions, ECs was mostly polarized against flow direction (pericentrin, Hoechst localization) and we observed ROR2 relocalization at cell-cell junctions, as compared to static conditions. This study shows that ROR2 is involved in ECs responses to flow. Study of ROR2 signaling is under investigation to understand how ROR2 could regulate ECs properties.