Soluble VE‐cadherin disrupts endothelial barrier function via VE‐PTP/RhoA signalling

Abstract

Increased levels of soluble VE‐cadherin fragments (sVE‐cadherin) have previously been linked to inflammation‐induced loss of endothelial barrier function. We hypothesized that sVE‐cadherin independent of pro‐inflammatory stimuli may be critically involved in the context of endothelial barrier dysfunction.Recombinant human sVE‐cadherin (extracellular domains EC1‐5) was generated and verified by sequencing and immunoblotting. Application of sVE‐cadherin on human dermal microvascular endothelial cells (HDMECs) dose‐dependently induced loss of endothelial barrier function as revealed by measurements of transendothelial electrical resistance (TER) and flux of 70 kDa FITC‐dextran across HDMEC monolayers. In an in vivo rat model i.v. application of sVE‐cadherin resulted in loss of endothelial barrier function and reduced microcirculatory flow, which was comparable to the conditions when bacterial Lipopolysaccharide (LPS) was applied in vivo. Immunofluorescence staining and western blot analysis revealed that the functional effects of sVE‐cadherin were paralleled by decreased expression of VE‐cadherin and reduction of α‐/ β‐/ γ‐/ δ‐catenin and tight‐junction associated protein ZO‐1 at the cell borders. VE‐protein tyrosine phosphatase (VE‐PTP) which is associated with VE‐cadherin under resting conditions was removed from the cell borders following application of sVE‐cadherin to HDMEC. These changes were accompanied by intercellular gap formation and increased stress fibers in the actin cytoskeleton. On a mechanistic level, sVE‐cadherin did not induce cell death but increased RhoA activity and reduced extracellular VE‐cadherin interaction as revealed by atomic force measurements. Rho‐kinase inhibitor Y27632 blocked all sVE‐cadherin‐induced effects on endothelial barrier function. Furthermore, VE‐PTP inhibitor AKB9778 attenuated both RhoA activation and all sVE‐cadherin‐induced effects.In summary, sVE‐cadherin disrupts endothelial barrier function by dismantling the VE‐cadherin complex at the cell border via VE‐PTP‐dependent RhoA activation. This points to a novel pathophysiological role of sVE‐cadherin in the context of endothelial barrier dysfunction in inflammation.