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Journal ArticleDOI

A critical role for phosphatidylinositol (3,4,5)-trisphosphate-dependent Rac exchanger 1 in endothelial junction disruption and vascular hyperpermeability.

07 Dec 2012-Circulation Research (NIH Public Access)-Vol. 111, Iss: 12, pp 1517-1527
TL;DR: These results demonstrate for the first time that P-Rex1 expressed in endothelial cells is activated downstream of TNF-&agr;, which is not a G protein–coupled receptor agonist.
Abstract: Rationale: The small GTPase Rac is critical to vascular endothelial functions, yet its regulation in endothelial cells remains unclear. Understanding the upstream pathway may delineate Rac activation mechanisms and its role in maintaining vascular endothelial barrier integrity. Objective: By investigating P-Rex1, one of the Rac-specific guanine nucleotide exchange factors (GEFs) previously known for G protein-coupled receptor (GPCR) signaling, we sought to determine whether Rac-GEF is a nodal for signal integration and potential target for drug intervention. Methods and Results: Using gene deletion and siRNA silencing approach, we investigated the role of P-Rex1 in lung microvascular endothelial cells (HLMVECs). TNF-α exposure led to disruption of endothelial junctions, and silencing P-Rex1 protected junction integrity. TNF-α stimulated Rac activation and ROS production in a P-Rex1-dependent manner. Removal of P-Rex1 significantly reduced ICAM-1 expression, PMN transendothelial migration and leukocyte sequestration in TNF-α challenged mouse lungs. The P-Rex1 knockout mice were also refractory to lung vascular hyper-permeability and edema in a LPS-induced sepsis model. Conclusions: These results demonstrate for the first time that P-Rex1 expressed in endothelial cells is activated downstream of TNF-α, which is not a GPCR agonist. Our data identify P-Rex1 as a critical mediator of vascular barrier disruption. Targeting P-Rex1 may effectively protect against TNF-α; and LPS-induced endothelial junction disruption and vascular hyper-permeability.
Citations
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Journal ArticleDOI
TL;DR: Recent insights into the post-translational modifications of junctional proteins and signaling pathways regulating plasticity of adherens junctions and endothelial permeability are discussed.
Abstract: The monolayer of endothelial cells lining the vessel wall forms a semipermeable barrier (in all tissue except the relatively impermeable blood–brain and inner retinal barriers) that regulates tissue–fluid homeostasis, transport of nutrients, and migration of blood cells across the barrier. Permeability of the endothelial barrier is primarily regulated by a protein complex called adherens junctions. Adherens junctions are not static structures; they are continuously remodeled in response to mechanical and chemical cues in both physiological and pathological settings. Here, we discuss recent insights into the post-translational modifications of junctional proteins and signaling pathways regulating plasticity of adherens junctions and endothelial permeability. We also discuss in the context of what is already known and newly defined signaling pathways that mediate endothelial barrier leakiness (hyperpermeability) that are important in the pathogenesis of cardiovascular and lung diseases and vascular inflammation.

313 citations


Cites background from "A critical role for phosphatidylino..."

  • ...lymerization of branched actin networks within lamellipodia protrusions,(187,189,270,294) Cdc42 facilitates polymerization of linear F-actin filaments into filopodia.(295,296) On activation,...

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  • ...through phosphatidylinositol (3,4,5)-trisphosphate–dependent Rac exchanger 1 (P-Rex1).(296) In this case, Rac1 signals through...

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Journal ArticleDOI
11 Mar 2016-Thorax
TL;DR: The role of the pulmonary endothelium is considered as a key modulator and orchestrator of ARDS, highlighting advances in understanding of endothelial pathobiology and their implications for the development of endotheric-targeted therapeutics including cell-based therapies.
Abstract: The pulmonary endothelium is a dynamic, metabolically active layer of squamous endothelial cells ideally placed to mediate key processes involved in lung homoeostasis. Many of these are disrupted in acute respiratory distress syndrome (ARDS), a syndrome with appreciable mortality and no effective pharmacotherapy. In this review, we consider the role of the pulmonary endothelium as a key modulator and orchestrator of ARDS, highlighting advances in our understanding of endothelial pathobiology and their implications for the development of endothelial-targeted therapeutics including cell-based therapies. We also discuss mechanisms to facilitate the translation of preclinical data into effective therapies including the application of biomarkers to phenotype patients with ARDS with a predominance of endothelial injury and emerging biotechnologies that could enhance delivery, discovery and testing of lung endothelial-specific therapeutics.

157 citations


Cites background from "A critical role for phosphatidylino..."

  • ...Rac1 can either positively or negatively regulate barrier function in a stimulus dependent manner [48-50] whilst Rap1, enhances barrier function via inhibition of Rho and activation of Cdc42 [51 52] as well as a co-...

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Journal ArticleDOI
TL;DR: This review provides an overview on endothelial barrier regulation and focuses on the endothelial signaling mechanisms controlling the opening and closing of paracellular pathways for solutes and cells such as leukocytes and metastasizing tumor cells.
Abstract: A central function of the vascular endothelium is to serve as a barrier between the blood and the surrounding tissue of the body. At the same time, solutes and cells have to pass the endothelium to...

152 citations

Journal ArticleDOI
TL;DR: The signaling networks that contribute to endothelial permeability and leukocyte transendothelial migration are compared, focusing particularly on signals mediated by small GTPases that regulate cell adhesion and the actin cytoskeleton.

150 citations

Journal ArticleDOI
TL;DR: The pivotal role of Rac-mediated generation of reactive oxygen species (ROS) to regulate the integrity of endothelium cell-cell junctions and the ability of endothelial adhesion receptors, involved in leukocyte transendothelial migration, to control endothelial permeability to small molecules are discussed.
Abstract: The decrease of endothelial barrier function is central to the long-term inflammatory response. A pathological alteration of the ability of endothelial cells to modulate the passage of cells and solutes across the vessel underlies the development of inflammatory diseases such as atherosclerosis and multiple sclerosis. The inflammatory cytokine tumour necrosis factor (TNF) mediates changes in the barrier properties of the endothelium. TNF activates different Rho GTPases, increases filamentous actin and remodels endothelial cell morphology. However, inhibition of actin-mediated remodelling is insufficient to prevent endothelial barrier disruption in response to TNF, suggesting that additional molecular mechanisms are involved. Here we discuss, first, the pivotal role of Rac-mediated generation of reactive oxygen species (ROS) to regulate the integrity of endothelial cell-cell junctions and, second, the ability of endothelial adhesion receptors such as ICAM-1, VCAM-1 and PECAM-1, involved in leukocyte transendothelial migration, to control endothelial permeability to small molecules, often through ROS generation. These adhesion receptors regulate endothelial barrier function in ways both dependent on and independent of their engagement by immune cells, and orchestrate the crosstalk between leukocyte transendothelial migration and endothelial permeability during inflammation.

130 citations


Cites background from "A critical role for phosphatidylino..."

  • ...In human endothelial cells, P-Rex knockdown, expression of dominant-negative Rac and specific Rac inhibitors abrogate ROS production in response to TNF (37)....

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  • ...Rac-induced ROS regulate endothelial permeability by elevating the tyrosine phosphorylation of VE-cadherin (68), which prevents the binding of β-catenin and p120-catenin and results in inhibition of endothelial barrier function (37, 69, 70)....

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  • ...Upstream of Rac, the Rac GEFs Vav, Tiam-1, Trio and P-Rex, are activated in response to TNF (37, 39, 40)....

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  • ...Rac activity increases during the first minutes of TNF activation (21, 37) but decreases during longterm stimulation (32, 38)....

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  • ...However, to date, there has been no comparative study of the relative contribution of each of these GEFs to TNF-induced endothelial barrier disturbance (37, 40)....

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References
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Journal ArticleDOI
17 Dec 1987-Nature
TL;DR: Protection against shock, vital organ dysfunction, persistent stress hormone release and death was conferred by administration of antibodies 2 h before bacterial infusion, indicating that cachectin is a mediator of fatal bacteraemic shock and suggesting that antibodies against Cachectin offer a potential therapy of life-threatening infection.
Abstract: Bacterial infection of the mammalian bloodstream can lead to overwhelming sepsis, a potentially fatal syndrome of irreversible cardiovascular collapse (shock) and critical organ failure. Cachectin, also known as tumour necrosis factor, is a macrophage-derived peptide hormone released in response to bacterial lipopolysaccharide, and it has been implicated as a principal mediator of endotoxic shock, although its function in bacterial sepsis is not known. Anaesthetized baboons were passively immunized against endogenous cachectin and subsequently infused with an LD100 dose of live Escherichia coli. Control animals (not immunized against cachectin) developed hypotension followed by lethal renal and pulmonary failure. Neutralizing monoclonal anti-cachectin antibody fragments (F(ab′)2) administered to baboons only one hour before bacterial challenge protected against shock, but did not prevent critical organ failure. Complete protection against shock, vital organ dysfunction, persistent stress hormone release and death was conferred by administration of antibodies 2 h before bacterial infusion. These results indicate that cachectin is a mediator of fatal bacteraemic shock, and suggest that antibodies against cachectin offer a potential therapy of life-threatening infection.

2,568 citations

Journal ArticleDOI
TL;DR: With 69 distinct homologues, Dbl-related GEFs represent the largest family of direct activators of Rho GTPases in humans, and they activate RhoGTPases within particular spatio-temporal contexts.
Abstract: Guanine nucleotide-exchange factors (GEFs) are directly responsible for the activation of Rho-family GTPases in response to diverse extracellular stimuli, and ultimately regulate numerous cellular responses such as proliferation, differentiation and movement. With 69 distinct homologues, Dbl-related GEFs represent the largest family of direct activators of Rho GTPases in humans, and they activate Rho GTPases within particular spatio-temporal contexts. The failure to do so can have significant consequences and is reflected in the aberrant function of Dbl-family GEFs in some human diseases.

1,710 citations

Journal ArticleDOI
17 Oct 1991-Nature
TL;DR: The active components of purified ω1 are two proteins that associate as heterodimers17, and here it is reported that these are the small GTP-binding protein p21racl and the GDP-dissociation inhibitor rhoGDI.
Abstract: Professional phagocytes, such as neutrophils and monocytes, have an NADPH oxidase that generates superoxide and other reduced oxygen species important in killing microorganisms. Several components of the oxidase complex have been identified as targets of genetic defects causing chronic granulomatous disease. The complex consists of an electron transport chain that has as its substrate cytosolic NADPH and which discharges superoxide into the cavity of the intracellular phagocytic vacuole. The only electron transport component identified so far is a low-potential cytochrome b, apparently the only membrane component required. At least three cytosolic factors are also necessary, two of which, p67phOx and p47phOx, have been identified by their absence in patients with chronic granulomatous disease. A third component, sigma 1, is required for stimulation of oxidase activity in a cell-free system. The active components of purified sigma 1 are two proteins that associate as heterodimers, and here we report that these are the small GTP-binding protein p21rac1 and the GDP-dissociation inhibitor rhoGDI.

939 citations

Journal ArticleDOI
TL;DR: The characterize of an antibody directed to mouse vascular endothelial (VE)-cadherin, a major adhesive protein of interendothelial adherens junctions, and its results indicate that VE-caderin is required for vascular integrity and normal organ functions.
Abstract: In the present paper, we characterize an antibody, mAb BV13, directed to mouse vascular endothelial (VE)-cadherin, a major adhesive protein of interendothelial adherens junctions. When added to cultured endothelial cells, BV13 induces a redistribution of VE-cadherin from intercellular junctions. VE-cadherin redistribution did not change the localization of platelet endothelial cell adhesion molecule or tight junction markers such as zonula occludens 1, cingulin, and junctional adhesion molecule. Intravenous administration of mAb BV13 induced a concentration- and time-dependent increase in vascular permeability in heart and lungs. By electron microscopy, interstitial edema and accumulation of mixed types of inflammatory cells in heart and lungs were observed. Injection of (rhodamine-labeled) Ricinus communis I lectin showed focal spots of exposed basement membrane in the alveolar capillaries and in some larger pulmonary vessels. These data indicate that VE-cadherin is required for vascular integrity and normal organ functions.

668 citations


"A critical role for phosphatidylino..." refers background or methods in this paper

  • ...Phosphatidylinositol (3,4,5)-trisphosphate–dependent Rac exchanger 1 (P-Rex1) regulates tumor necrosis factor (TNF)-α–induced reactive oxygen species (ROS) production....

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  • ...Role for phosphatidylinositol (3,4,5)-trisphosphate– dependent Rac exchanger 1 (P-Rex1) in tumor necrosis factor (TNF)-α–induced Rac activation....

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  • ...Objective: By investigating phosphatidylinositol (3,4,5)-trisphosphate–dependent Rac exchanger 1 (P-Rex1), one of the Rac-specific guanine nucleotide exchange factors previously known for G protein–coupled receptor signaling, we sought to determine whether Rac-guanine nucleotide exchange factor is nodal for signal integration and potential target for drug intervention....

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  • ...Key Words: acute lung injury ◼ endothelial dysfunction ◼ guanine nucleotide exchange factors ◼ pulmonary edema ◼ reactive oxygen species ◼ small GTPases ◼ vascular permeability A Critical Role for Phosphatidylinositol (3,4,5)-Trisphosphate–Dependent Rac Exchanger 1 in Endothelial Junction Disruption and Vascular Hyperpermeability...

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  • ...Tumor necrosis factor (TNF)-α induces phosphoinositide 3-kinase–dependent membrane translocation of phosphatidylinositol (3,4,5)-trisphosphate–dependent Rac exchanger 1 (P-Rex1)....

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Journal ArticleDOI
TL;DR: Data indicate that 7B4 antigen is an endothelial-specific cadherin that plays a role in the organization of lateral endothelial junctions and in the control of permeability properties of vascular endothelium.
Abstract: mAbs were raised in mice against cultured human endothelial cells (EC) and screened by indirect immunofluorescence for their ability to stain intercellular contacts. One mAb denoted 7B4 was identified which, out of many cultured cell types, specifically decorated cultured human EC. The antigen recognized by mAb 7B4 is bound at the appositional surfaces of cultured EC only as they become confluent and is stably expressed at intercellular boundaries of confluent monolayers. EC recognition specificity was maintained when the antibody was assayed by immuno-histochemistry in tissue sections of many normal and malignant tissues and in blood vessels of different size and type. The antigen recognized by 7B4 was enriched at EC intercellular boundaries similarly in vitro and in situ. In vitro, addition of mAb 7B4 to confluent EC increased permeation of macromolecules across monolayers even without any obvious changes of cell morphology. In addition, when EC permeability was increased by agents such as thrombin, elastase, and TNF/gamma IFN, its distribution pattern at intercellular contact rims was severely altered. mAb 7B4 immunoprecipitated a major protein of 140 kD from metabolically and surface-labeled cultured EC extracts which appeared to be an integral membrane glycoprotein. On the basis of its distribution in cultured cells and in tissues in situ, 7B4 antigen is distinct from other described EC proteins enriched at intercellular contacts. NH2-terminal sequencing of the antigen, immunopurified from human placenta, and sequencing of peptides from tryptic peptide maps revealed identity to the cDNA deduced sequence of a recently identified new member of the cadherin family (Suzuki, S., K. Sano, and H. Tanihara. 1991. Cell Regul. 2:261-270.) These data indicate that 7B4 antigen is an endothelial-specific cadherin that plays a role in the organization of lateral endothelial junctions and in the control of permeability properties of vascular endothelium.

663 citations