Intermedin (adrenomedullin2) stabilizes the endothelial barrier and antagonizes thrombin‐induced barrier failure in endothelial cell monolayers

Abstract

BACKGROUND AND PURPOSEIntermedin is a member of the calcitonin gene-related-peptide (CGRP) family expressed in endothelial cells and acts via calcitonin receptor-like receptors (CLRs). Here we have analysed the receptors for intermedin and its effect on the endothelial barrier in monolayers of human umbilical vein endothelial cells (HUVECs). EXPERIMENTAL APPROACHWe analysed the effect of intermedin on albumin permeability, contractile machinery, actin cytoskeleton and VE-cadherin in cultured HUVECs. KEY RESULTSIntermedin concentration-dependently reduced basal endothelial permeability to albumin and antagonized thrombin-induced hyperpermeability. Intermedin was less potent (EC50 1.29 Ϯ 0.12 nM) than adrenomedullin (EC50 0.24 Ϯ 0.07 nM) in reducing endothelial permeability. These intermedin effects were inhibited by AM22-52 and higher concentrations of aCGRP8-37, with pA2 values of aCGRP8-37 of 6.4 for both intermedin and adrenomedullin. PCR data showed that HUVEC expressed only the CLR/RAMP2 receptor complex. Intermedin activated cAMP/PKA and cAMP/Epac signalling pathways. Intermedin's effect on permeability was blocked by inhibition of PKA but not of eNOS. Intermedin antagonized thrombin-induced contractile activation, RhoA activation and stress fibre formation. It also induced Rac1 activation, enhanced cell-cell adhesion and antagonized thrombin-induced loss of cell-cell adhesion. Treatment with a specific inhibitor of Rac1 prevented intermedin-mediated barrier stabilization. CONCLUSION AND IMPLICATIONSIntermedin stabilized endothelial barriers in HUVEC monolayers via CLR/RAMP2 receptors. These effects were mediated via cAMP-mediated inactivation of contractility and strengthening of cell-cell adhesion. These findings identify intermedin as a barrier stabilizing agent and suggest intermedin as a potential treatment for vascular leakage in inflammatory conditions.

cAMP-dependent protein kinase A (PKA) regulates angiogenesis by modulating tip cell behavior in a Notch-independent manner

“…Usually, PKA is activated in response to stimulation of GPCRs and consecutive elevation of cAMP. Several GPCRs and their ligands have been shown to regulate cAMP level or PKA activity in endothelial cells (Aslam et al, 2012;Bakre et al, 2002;Dormond et al, 2002), some of which have been proposed to activate PKA during vascular development (Jin et al, 2010). However, it remains unclear as to which GPCRs indeed regulate endothelial PKA during vascular development.…”

Intermedin

“…24 Furthermore, IMD was able to stabilize the endothelial barrier function via the maintenance of VE-cadherin on the cell surface. 26 Thus, IMD may serve as an integrator in the regulation of VE-cadherin and VEGF activities so as to affect the angiogenic process.…”

Intermedin/adrenomedullin2: an autocrine/paracrine factor in vascular homeostasis and disease

“…IMD stabilized endothelial barriers in human umbilical vein endothelial cell (HUVEC) monolayers via CRLR/RAMP2 receptors [23]. These effects were mediated via cAMPmediated inactivation of contractility and strengthening of cell-cell adhesion.…”