Vascular Endothelial Barrier Function Research Articles

Exposure to Concentrated Ambient Fine Particulate Matter Induces Vascular Endothelial Dysfunction via miR-21.

Vascular endothelial permeability transition does not cause significant lesions, but enhanced permeability may contribute to the development of vascular and other diseases, including atherosclerosis, hypertension, heart failure and cancer. Therefore, elucidating the effect of Particulate Matter 2.5 (PM2.5) on vascular endothelial permeability could help prevent disease that might be caused by PM2.5. Our previous study and the present one revealed that PM2.5 significantly increased the permeability of vascular endothelial cells and disrupted the barrier function of the vascular endothelium in Sprague Dawley (SD) rats. We found that the effect occurred mainly through induction of signal transducer and activator of transcription 3 (STAT3) phosphorylation, further transcriptional regulation of microRNA21 (miR-21) and promotion of miR-21 expression. These changes post-transcriptionally repress tissue inhibitor of metalloproteinases 3 (TIMP3) and promote matrix metalloproteinases 9 (MMP9) expression. This work provides evidence that PM2.5 exerts direct inhibitory action on vascular endothelial barrier function and might give rise to a number of vascular diseases.

Exposure to concentrated ambient fine particulate matter disrupts vascular endothelial cell barrier function via the IL-6/HIF-1α signaling pathway.

Exposure to concentrated ambient fine particulate matter (PM2.5) has been associated with cardiovascular diseases (CVDs). The barrier function of vascular endothelial cells is critical for the development of CVDs. Here, we employed human umbilical vein endothelial cells to clarify the function of ambient PM2.5 pollution in the regulation of membrane permeability of vascular endothelial cells. The results show that a high concentration of PM2.5, which mainly includes heavy metals and polycyclic aromatic hydrocarbons, induces barrier dysfunction of vascular endothelial cells. This was mediated in part by promoting IL‐6 expression, which then increases the transcriptional activity of HIF‐1α by promoting its translocation to the nucleus. Our findings indicate that concentrated PM2.5 can destroy membrane integrity and promote permeability in vascular endothelial cells, thereby contributing to the development of CVDs.

Platelets regulate vascular endothelial stability: assessing the storage lesion and donor variability of apheresis platelets.

In current blood banking practices, platelets (PLTs) are stored in plasma at 22°C, with gentle agitation for up to 5 days. To date, the effects of storage and donor variability on PLT regulation of vascular integrity are not known. In this study, we examined the donor variability of leukoreduced fresh (Day 1) or stored (Day 5) PLTs on vascular endothelial barrier function in vitro and in vivo. In vitro, PLT effects on endothelial cell (EC) monolayer permeability were assessed by analyzing transendothelial electrical resistances (TEER). PLT aggregation, a measure of hemostatic potential, was analyzed by impedance aggregometry. In vivo, PLTs were investigated in a vascular endothelial growth factor A (VEGF-A)-induced vascular permeability model in NSG mice, and PLT circulation was measured by flow cytometry. Treatment of endothelial monolayers with fresh Day 1 PLTs resulted in an increase in EC barrier resistance and decreased permeability in a dose-dependent manner. Subsequent treatment of EC monolayers with Day 5 PLTs demonstrated diminished vasculoprotective effects. Donor variability was noted in all measures of PLT function. Day 1 PLT donors were more variable in their effects on TEER than Day 5 PLTs. In mice, while all PLTs regardless of storage time demonstrated significant protection against VEGF-A-induced vascular leakage, Day 5 PLTs exhibited reduced protection when compared to Day 1 PLTs. Day 1 PLTs demonstrated significant donor variability against VEGF-A-challenged vascular leakage in vivo. Systemic circulating levels of Day 1 PLTs were higher than those of Day 5 PLTs In vitro and in vivo, Day 1 PLTs are protective in measures of vascular endothelial permeability. Donor variability is most prominent in Day 1 PLTs. A decrease in the protective effects is found with storage of the PLT units between Day 1 and Day 5 at 22°C, thereby suggesting that Day 5 PLTs are diminished in their ability to attenuate vascular endothelial permeability.

Abstract 434: Formin Dependent Regulation of Adherens Junction Assembly

Adherens junctions are cadherin-dependent structures that mediate intercellular signaling and structural integrity. In endothelial cells, adherens junctions are the primary determiners of vascular permeability and endothelial barrier function. Adherens junctions connect directly to the actin cytoskeleton and this connection is essential for junction formation and maintenance. Formins are a highly conserved family of cytoskeletal remodeling proteins whose activity has been implicated in regulating junction formation in other cell-types. Therefore, we tested the hypothesis that formin activity is essential for adherens junction assembly in endothelial cells. A small-molecule formin inhibitor (smiFH2) was used to determine the effect of formin inhibition on junction formation in Telomerase-Immortalized Microvascular Endothelial (TIME) cells. Using an in vitro vascular permeability assay, we determined that smiFH2 treatment caused a dose-dependent inhibition of junction formation. We used siRNA to knockdown expression of seven of the formins expressed in TIME cells in order to determine which specific formins were required to maintain endothelial barrier function. We determined that individual knockdown of three formins, FHOD1, FHOD3 and Dia1, significantly increased the permeability of the endothelial monolayer. Interestingly, the knockdown of a fourth formin, FMNL2, had the opposite effect and actually potentiated barrier function. Knockdown of the remaining formins had little or no effect on junction formation. Knockdown of FHOD3 had the greatest inhibitory effect on junction assembly and VE-cadherin, β-catenin and α-catenin protein levels were decreased in FHOD3-depleted cells. The FHOD3 knockdown cells were also elongated in comparison to controls and formed thin linear adherens junctions and few focal adherens junctions. In contrast, the morphology of FMNL2-depleted cells did not appear obviously different from controls. In conclusion, our results suggest that multiple formins play diverse roles in endothelial cell adherens junction formation and maintenance.

Heat shock protein A12B decreases lipopolysaccharide-induced permeability of human umbilical vein endothelial cells

Objective To explore the lipopolysaccharide(LPS)-induced changes of heat shock protein A12B(HSPA12B)expression and its effect on the permeability of human umbilical vein endothelial cells(HUVECs).Methods The HUVECs cultured in vitro were divided into four groups: control group(without any treatment); LPS group(1 μg / m L LPS); LPS + p IRES2-EGFPHSPA12B-3Flag group(The HSPA12 B gene overexpression plasmid was transiently transfected into HUVECs and then LPS of 1 μg / m L was added); and LPS + p IRES2-EGFP-3Flag group(The negative control plasmid was transiently transfected into HUVECs and then LPS of 1 μg / m L was added).The transendothelial electrical resistance(TEER) of HUVECs was measured by MERSST × 01 Electrode.The expression of VE-cadherin was studied by flow cytometry.RT-PCR and Western blotting analysis were used to examine the expression changes of HSPA12 B mRNA and protein in HUVECs stimulated with LPS at various time points.Results It was showed that after stimulated with LPS,HSPA12 B mRNA and protein were gradually upregulated,and peaked at 12 h.HSPA12 B significantly increased the TEER value of HUVECs(P 0.001),and it could completely offset the decline of TEER value induced by LPS(P 0.001).Besides,HSPA12 B could cause upregulation of VE-cadherin expression.Conclusion HSPA12 B can reduce the permeability of vascular endothelial cells by up-regulating the expression of VE-cadherin,thus protecting the vascular endothelial barrier function of endothelial cells.

Cytoskeletal mechanisms regulating vascular endothelial barrier function in response to acute lung injury

Endothelial cells (EC) form a semi-permeable barrier between the interior space of blood vessels and the underlying tissues. In acute lung injury (ALI) the EC barrier is weakened leading to increased vascular permeability. It is widely accepted that EC barrier integrity is critically dependent upon intact cytoskeletal structure and cell junctions. Edemagenic agonists, like thrombin or endotoxin lipopolysaccharide (LPS), induced cytoskeletal rearrangement, and EC contractile responses leading to disruption of intercellular contacts and EC permeability increase. The highly clinically-relevant cytoskeletal mechanisms of EC barrier dysfunction are currently under intense investigation and will be described and discussed in the current review.

MicroRNA-147b regulates vascular endothelial barrier function by targeting ADAM15 expression.

A disintegrin and metalloproteinase15 (ADAM15) has been shown to be upregulated and mediate endothelial hyperpermeability during inflammation and sepsis. This molecule contains multiple functional domains with the ability to modulate diverse cellular processes including cell adhesion, extracellular matrix degradation, and ectodomain shedding of transmembrane proteins. These characteristics make ADAM15 an attractive therapeutic target in various diseases. The lack of pharmacological inhibitors specific to ADAM15 prompted our efforts to identify biological or molecular tools to alter its expression for further studying its function and therapeutic implications. The goal of this study was to determine if ADAM15-targeting microRNAs altered ADAM15-induced endothelial barrier dysfunction during septic challenge by bacterial lipopolysaccharide (LPS). An in silico analysis followed by luciferase reporter assay in human vascular endothelial cells identified miR-147b with the ability to target the 3′ UTR of ADAM15. Transfection with a miR-147b mimic led to decreased total, as well as cell surface expression of ADAM15 in endothelial cells, while miR-147b antagomir produced an opposite effect. Functionally, LPS-induced endothelial barrier dysfunction, evidenced by a reduction in transendothelial electric resistance and increase in albumin flux across endothelial monolayers, was attenuated in cells treated with miR-147b mimics. In contrast, miR-147b antagomir exerted a permeability-increasing effect in vascular endothelial cells similar to that caused by LPS. Taken together, these data suggest the potential role of miR147b in regulating endothelial barrier function by targeting ADAM15 expression.

Vascular Endothelial Tight Junctions and Barrier Function Are Disrupted by 15(S)-Hydroxyeicosatetraenoic Acid Partly via Protein Kinase Cϵ-mediated Zona Occludens-1 Phosphorylation at Threonine 770/772

Disruption of tight junctions (TJs) perturbs endothelial barrier function and promotes inflammation. Previously, we have shown that 15(S)-hydroxyeicosatetraenoic acid (15(S)-HETE), the major 15-lipoxygenase 1 (15-LO1) metabolite of arachidonic acid, by stimulating zona occludens (ZO)-2 tyrosine phosphorylation and its dissociation from claudins 1/5, induces endothelial TJ disruption and its barrier dysfunction. Here, we have studied the role of serine/threonine phosphorylation of TJ proteins in 15(S)-HETE-induced endothelial TJ disruption and its barrier dysfunction. We found that 15(S)-HETE enhances ZO-1 phosphorylation at Thr-770/772 residues via PKCε-mediated MEK1-ERK1/2 activation, causing ZO-1 dissociation from occludin, disrupting endothelial TJs and its barrier function, and promoting monocyte transmigration; these effects were reversed by T770A/T772A mutations. In the arteries of WT mice ex vivo, 15(S)-HETE also induced ZO-1 phosphorylation and endothelial TJ disruption in a PKCε and MEK1-ERK1/2-dependent manner. In line with these observations, in WT mice high fat diet feeding induced 12/15-lipoxygenase (12/15-LO) expression in the endothelium and caused disruption of its TJs and barrier function. However, in 12/15-LO(-/-) mice, high fat diet feeding did not cause disruption of endothelial TJs and barrier function. These observations suggest that the 12/15-LO-12/15(S)-HETE axis, in addition to tyrosine phosphorylation of ZO-2, also stimulates threonine phosphorylation of ZO-1 in the mediation of endothelial TJ disruption and its barrier dysfunction.

Stimulation of G Protein–Coupled Bile Acid Receptor Enhances Vascular Endothelial Barrier Function via Activation of Protein Kinase A and Rac1

Bile acids are end products of cholesterol metabolism, and they constantly exist at high concentrations in the blood. Since vascular endothelial cells express G protein-coupled bile acid receptor (GPBAR), bile acids potentially modulate endothelial function. Here, we investigated whether and how GPBAR agonism affects endothelial barrier function. In bovine aortic endothelial cells (BAECs), treatment with a GPBAR agonist, taurolithocholic acid (TLCA) increased the transendothelial electrical resistance. In addition, TLCA suppressed the thrombin-induced dextran infiltration through the endothelial monolayer. Knockdown of GPBAR abolished the inhibitory effect of TLCA on hyperpermeability. These results indicate that stimulation of GPBAR enhances endothelial barrier function. TLCA increased intracellular cAMP production in BAECs. Inhibition of protein kinase A (PKA) or Rac1 significantly attenuated the TLCA-induced endothelial barrier protection. TLCA induced cortical actin polymerization, which was attenuated by a Rac1 inhibitor. In vivo, local administration of TLCA into the mouse ear significantly inhibited vascular leakage and edema formation induced by croton oil or vascular endothelial growth factor. These results indicate that stimulation of GPBAR enhances endothelial barrier function by cAMP/PKA/Rac1-dependent cytoskeletal rearrangement.

Sirolimus-FKBP12.6 Impairs Endothelial Barrier Function Through Protein Kinase C-α Activation and Disruption of the p120–Vascular Endothelial Cadherin Interaction

Sirolimus (SRL) is an immunosuppressant drug used to prevent rejection in organ transplantation and neointimal hyperplasia when delivered from drug-eluting stents. Major side effects of SRL include edema and local collection of intimal lipid deposits at drug-eluting stent sites, suggesting that SRL impairs endothelial barrier function (EBF). The aim of this study was to address the role of SRL on impaired EBF and the potential mechanisms involved. Cultured human aortic endothelial cells (HAECs) and intact human and mouse endothelium was examined to determine the effect of SRL, which binds FKBP12.6 to inhibit the mammalian target of rapamycin, on EBF. EBF, measured by transendothelial electrical resistance, was impaired in HAECs when treated with SRL or small interfering RNA for FKBP12.6 and reversed when pretreated with ryanodine, a stabilizer of ryanodine receptor 2 intracellular calcium release channels. Intracellular calcium increased in HAECs treated with SRL and normalized with ryanodine pretreatment. SRL-treated HAECs demonstrated increases in protein kinase C-α phosphorylation, a calcium sensitive serine/threonine kinase important in vascular endothelial (VE) cadherin barrier function through its interaction with p120-catenin (p120). Immunostaining of HAECs, human coronary and mouse aortic endothelium treated with SRL showed disruption of p120-VE cadherin interaction treated with SRL. SRL impairment of HAEC EBF was reduced with protein kinase C-α small interfering RNA. Mice treated with SRL demonstrated increased vascular permeability by Evans blue albumin extravasation in the lungs, heart, and aorta. SRL-FKBP12.6 impairs EBF by activation of protein kinase C-α and downstream disruption of the p120-VE cadherin interaction in vascular endothelium. These data suggest this mechanism may be an important contributor of SRL side effects related to impaired EBF.

Neutrophil Proteinase 3 Acts on Protease-Activated Receptor-2 to Enhance Vascular Endothelial Cell Barrier Function

The principle role of the vascular endothelium is to present a semi-impermeable barrier to soluble factors and circulating cells, while still permitting the passage of leukocytes from the bloodstream into the tissue. The process of diapedesis involves the selective disruption of endothelial cell junctions, which could compromise vascular integrity. It is therefore somewhat surprising that neutrophil transmigration does not significantly impair endothelial barrier function. We examined whether neutrophils might secrete factors that promote vascular integrity during the latter stages of neutrophil transmigration, in particular, the role of neutrophil serine proteinase 3 (PR3). Endothelial cells were treated with PR3 either in its soluble form or in a complex form with cell surface NB1. We observed that PR3 mediated the enhancement of endothelial cell junctional integrity and that this required its proteolytic activity, as well as endothelial cell expression of the protease-activated receptor-2. Importantly, PR3 suppressed the vascular permeability changes and disruption of junctional proteins induced by the action of protease-activated receptor-1 agonists. These findings establish the potential for neutrophil-derived PR3 to play a role in reestablishing vascular integrity after leukocyte transmigration and in protecting endothelial cells from protease-activated receptor-1-induced permeability changes that occur during thrombotic and inflammatory events.

PKCα Activation of p120-Catenin Serine 879 Phospho-Switch Disassembles VE-Cadherin Junctions and Disrupts Vascular Integrity

Adherens junctions (AJs) are the primary intercellular junctions in microvessels responsible for endothelial barrier function. Homophilic adhesion of vascular endothelial (VE) cadherin forms AJs, which are stabilized by binding of p120-catenin (p120). p120 dissociation from VE-cadherin results in loss of VE-cadherin homotypic interaction and AJ disassembly; however, the signaling mechanisms regulating p120 dissociation from VE-cadherin are not understood. To address the mechanism of protein kinase C (PKC)-α function in increasing endothelial permeability, we determined the role of PKCα phosphorylation of p120 in mediating disruption of AJ integrity. We showed that PKCα phosphorylation of p120 at serine (S)879 in response to thrombin or lipopolysaccharide challenge reduced p120 binding affinity for VE-cadherin and mediated AJ disassembly secondary to VE-cadherin internalization. In studies in mouse lung vessels, expression of the phosphodeficient S879A-p120 mutant prevented the increase in vascular permeability induced by activation of the thrombin receptor PAR-1. PKCα phosphorylation of p120 at S879 is a critical phospho-switch mediating disassociation of p120 from VE-cadherin that results in AJ disassembly. Therefore, blocking PKCα-mediated p120 phosphorylation represents a novel targeted anti-inflammatory strategy to prevent disruption of vascular endothelial barrier function.

Regulation of Lymphocyte Function by Adenosine

Adenosine regulates the interaction between lymphocytes and the vasculature, and is important for controlling lymphocyte trafficking in response to tissue injury or infection. Adenosine can blunt the effects of T cell receptor activation primarily by activating adenosine A(2A) receptors and signaling via cyclic AMP and protein kinase A. Protein kinase A reduces proximal T cell receptor signaling by phosphorylation of C-terminal Src kinase, nuclear factor of activated T cells and cyclic AMP response element-binding protein. Protein kinase A activation can either enhance or inhibit the survival of T cells depending on the strength and duration of signaling. Inducible enzymes such as CD73 and CD39 regulate adenosine formation and degradation in vivo. The extravasation of lymphocytes through blood vessels is influenced by A(2A) receptors-mediated suppression of intercellular adhesion molecule 1 expression on lymphocytes and diminished production of interferon γ and interferon γ-inducible chemokines that are chemotactic to activated lymphocytes. Adenosine also decreases the barrier function of vascular endothelium by activating A(2B)Rs. In sum, adenosine signaling is influenced by tissue inflammation and injury through induction of receptors and enzymes and has generally inhibitory effects on lymphocyte migration into inflamed tissues due to protein kinase A-mediated effects on adhesion molecules, interferon γ production, and endothelial barrier function.

Degradation of the endothelial glycocalyx is associated with chylomicron leakage in mouse cremaster muscle microcirculation

A thick endothelial glycocalyx contributes to the barrier function of vascular endothelium in macro- and microcirculation. We hypothesised in the current study that diet-induced hyperlipidaemia perturbs the glycocalyx, resulting in decreased dimensions of this layer and increased transendothelial lipoprotein leakage in capillaries. Glycocalyx thickness was measured in mouse cremaster muscle capillaries by intravital microscopy from the distance between flowing red blood cells and the endothelial surface. In control C57BL/6 mice on standard chow, glycocalyx thickness measured 0.58 ± 0.01 (mean ± SEM) μm, and no lipoproteins were observed in the tissue. After three months administration of an either mild or severe high-fat / high-cholesterol diet (HFC) to C57BL/6 and ApoE3-Leiden mice, circulating large lipoproteins appeared into the subendothelial space in an increasing proportion of cremaster capillaries, and these capillaries displayed reduced glycocalyx dimensions of 0.40 ± 0.02 and 0.30 ± 0.01 μm (C57BL/6 mice), and 0.37 ± 0.01 and 0.28 ± 0.01 μm (ApoE3-Leiden mice), after the mild and severe HFC diet, respectively. The chylomicron nature of the accumulated lipoproteins was confirmed by observations of subendothelial deposition of DiI-labeled chylomicrons in capillaries after inducing acute glycocalyx degradation by heparitinase in normolipidaemic C57BL/6 mice. It is concluded that while under control conditions the endothelial glycocalyx contributes to the vascular barrier against transvascular lipoprotein leakage in the microcirculation, diet-induced hyperlipidaemia reduces the thickness of the glycocalyx, thereby facilitating leakage of chylomicrons across the capillary wall.

Role of transglutaminases in cuff-induced atherosclerotic lesion formation in femoral arteries of ApoE3 Leiden mice

Transglutaminases play an important role in vascular remodeling, calcification, cell adhesion and endothelial barrier function. In this study we investigate the influence of combined inhibition of both tissue-type transglutaminase (TG2) and the plasma transglutaminase FXIIIA on early lesion development. Methods A cuff was placed around the femoral arteries of ApoE3 Leiden mice while fed a Western type diet to induce atherosclerotic lesion development. An osmotic minipump was placed in the intraperitoneal cavity containing an irreversible inhibitor of TG2 and FXIIIA activity ((1,3,4,5-tetramethyl-2-[(2-oxopropyl)thio]imidazolium chloride, Zedira). Atherosclerotic lesion composition was analyzed using immunohistochemistry and RT-PCR. Results Inhibition of transglutaminases did not influence lesion size or geometric remodeling of the vessels. However, systemic transglutaminase inhibition resulted in 41% less macrophage infiltrate in the media of the vessels. Additional in vitro experiments on HL60 cells confirmed a decreased migratory response during transglutaminase inhibition. Conclusion Inhibition of TG2 and FXIIIA during early development of lesions reduced the macrophage content in the media of atherosclerotic vessels, while not affecting lesion size or geometric remodeling.

Claudin-5 as a Novel Estrogen Target in Vascular Endothelium

Estrogens have multiple effects on vascular physiology and function. In the present study, we look for direct estrogen target genes within junctional proteins. We use murine endothelial cell lines of brain and heart origin, which express both subtypes of estrogen receptor, ERalpha and ERbeta. Treatment of these cells with 17beta-estradiol (E2) led to an increase in transendothelial electric resistance and a most prominent upregulation of the tight junction protein claudin-5 expression. A significant increase of claudin-5 promoter activity, mRNA, and protein levels was detected in cells from both vascular beds. In protein lysates and in immunoreactions on brain sections from ovariectomized E2-treated mice, we noticed an increase in claudin-5 protein and mRNA content. Treatment of cells with a specific ERbeta agonist, diarylpropionitrile, revealed the same effect as E2 stimulation. Moreover, we detected significantly lower claudin-5 mRNA and protein content in ERbeta knockout mice. We describe claudin-5 as a novel estrogen target in vascular endothelium and show in vivo (brain endothelium) and in vitro (brain and heart endothelium) effects of estrogen on claudin-5 levels. The estrogen-induced increase in junctional protein levels may lead to an improvement in vascular structural integrity and barrier function of vascular endothelium.

Endothelial Progenitor Cell Adhesion Mediated by α4β1 and α5β1 Integrins is Critical to the Restoration of Lung Vascular Endothelial Barrier Function in a Model of Endotoxin‐induced Acute Lung Injury

Here we tested the hypothesis that integrin‐mediated adhesion event is a critical requirement for the endothelial barrier protection induced by endothelial progenitor cells (EPCs). We used bone‐marrow (BM)‐derived mononuclear (MNCs) as a source for EPCs, and characterized for EPC marker expression including CD34, VE‐cadherin, Flk1, and integrins. To model Acute Lung Injury (ALI), the recipient mice were γ‐irradiated then challenged intraperitonially with bacterial endotoxin Lipopolysaccharide (LPS). Mice were administered with either control endothelial cells (ECs), EPCs, EPCs incubated with adhesion blocking anti‐integrin monoclonal antibodies, or EPCs transduced with specific shRNA integrin construct. The end points assessed were: (a) survival, (b) lung water content, (c) capillary filtration coefficient (Kfc), (d) H&E staining of lung sections, and (e) confocal image analysis of EPC retention. We found that treatment with EPCs afforded 50% survival (P<0.001) and restored lung fluid balance (Kfc, P<0.001) in α4 and α5 integrins‐dependent manner. Confocal image analyses showed increased retention of Red Fluorescent Protein (RFP)‐tagged EPCs until 8 weeks and overall improved lung morphology. Thus, the treatment with EPCs reduced the extent of LPS‐induced lung microvessel injury, restored lung fluid balance, and increased survival in α4β1 and α5β1 integrins dependent manner.

Reprint of “The role of cytoskeleton in the regulation of vascular endothelial barrier function” [Microvascular Research 76 (2008) 202–207

The cytoskeleton is vital to the function of virtually all cell types in the organism as it is required for cell division, cell motility, endo- or exocytosis and the maintenance of cell shape. Endothelial cells, lining the inner surface of the blood vessels, exploit cytoskeletal elements to ensure the integrity of cell monolayer in quiescent endothelium, and to enable the disintegration of the formed barrier in response to various agonists. Vascular permeability is defined by the combination of transcellular and paracellular pathways, with the latter being a major contributor to the inflammation-induced barrier dysfunction. This review will analyze the cytoskeletal elements, which reorganization affects endothelial permeability, and emphasize signaling mechanisms with barrier-protective or barrier-disruptive potential.

FGF Signaling Preserves the Integrity of Endothelial Adherens Junctions

Fibroblast growth factor (FGF)-activated signaling regulates an array of cellular processes ranging from embryonic development to tissue repair. A recent paper by Murakami et al. identifies a potentially important role for FGF signaling in maintenance of endothelial barrier homeostasis through the regulation of adherens junctions.

VE-cadherin and β-catenin binding dynamics during histamine-induced endothelial hyperpermeability

Beta-catenin plays an important role in the regulation of vascular endothelial cell-cell adhesions and barrier function by linking the VE-cadherin junction complex to the cytoskeleton. The purpose of this study was to evaluate the effect of beta-catenin and VE-cadherin interactions on endothelial permeability during inflammatory stimulation by histamine. We first assessed the ability of a beta-catenin binding polypeptide known as inhibitor of beta-catenin and T cell factor (ICAT) to compete beta-catenin binding to VE-cadherin in vitro. We then overexpressed recombinant FLAG-ICAT in human umbilical vein endothelial cells (HUVECs) to study its impact on endothelial barrier function controlled by cell-cell adhesions. The binding of beta-catenin to VE-cadherin was quantified before and after stimulation with histamine along with measurements of transendothelial electrical resistance (TER) and apparent permeability to albumin (P(a)) under the same conditions. The results showed that ICAT bound to beta-catenin and competitively inhibited binding of the VE-cadherin cytoplasmic domain to beta-catenin in a concentration-dependent manner. Overexpression of FLAG-ICAT in endothelial cell monolayers did not affect their basal permeability properties, as indicated by unaltered TER and P(a); however, the magnitude and duration of histamine-induced decreases in TER were significantly augmented. Likewise, the increase in P(a) in the presence of histamine was exacerbated. Overexpression of FLAG-ICAT also significantly decreased the level of beta-catenin-associated VE-cadherin following histamine stimulation. Taken together, these data suggest that inflammatory agents like histamine cause a transient and reversible disruption of binding between beta-catenin and VE-cadherin, during which endothelial permeability is elevated.