Impaired integrin‐mediated adhesion contributes to reduced barrier properties in VASP‐deficient microvascular endothelium

Schlegel, Nicolas; Waschke, Jens

doi:10.1002/jcp.21772

Cited by 34 publications

(31 citation statements)

References 44 publications

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“…For example, VASP plays a regulatory role on FA in endothelial cells. 6,8 However, in mouse cardiac fibroblasts, the effects of VASP on FA were not as evident. 7,25 Ena/VASP proteins are required for neuritogenesis, 53 and, interestingly, the HSC used in this study may share a common origin with neuronal cells as suggested by expression of the neuronal makers, glial fibrillary acid protein (GFAP) and NG2 chondroitin sulfate proteoglycan.…”

Section: Discussionmentioning

confidence: 96%

Focal Adhesion Assembly in Myofibroblasts Fosters a Microenvironment that Promotes Tumor Growth

Kang

Yaqoob

Geng

et al. 2010

The American Journal of Pathology

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Tumor stroma importantly influences tumor growth and progression with pericytes representing an important stromal cell owing to their importance in angiogenesis and their transformation into highly motile tumor myofibroblasts.1 Focal adhesions (FAs) are specialized structures that connect extracellular environment to actin cytoskeleton thus facilitating the signal transduction and actin remodeling requisite for the process of pericyte migration during angiogenesis.2,3 Thus, mechanisms that govern FA assembly and pericyte motility may present potential targets for anti-cancer therapy but are not yet fully defined.vasodilator-stimulated phosphoprotein (VASP) is a member of the Ena/VASP family of proteins that regulate actin cytoskeleton and cell migration. 4 It contains distinct subdomains that facilitate specific protein interactions and actin binding characteristics that culminate in an anticapping/branching function within the cytoskeleton.4,5 Although VASP resides within FAs, its precise role in FA dynamics has not been completely defined, probably due in part to cell-type specific VASP functions. 6 -8 Because the role of VASP in pericyte function and ensuing effects within the tumor microenvironment remain unexplored, we investigated the role of VASP in FA development in human hepatic stellate cells (HSC), liver pericytes that express a high level of VASP and which on transformation into myofibroblasts, develop a highly motile state that has been postulated to influence tumor growth. Our data reveal a requirement of VASP in FA dynamics and pericyte motility, which requires cooperation of each distinct VASP subdomain. Indeed, overexpression of the EVH2 domain alone results in a dominant negative function that inhibits FA assembly, cell motility, and angiogenesis. We also identify a role of Rac1 as a key binding partner of VASP that promotes its ability to regulate FA and cell motility. Perturbation of Ena/VASP function in tumor myofibroblast precursor cells signifi-

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Section: Discussionmentioning

confidence: 96%

Focal Adhesion Assembly in Myofibroblasts Fosters a Microenvironment that Promotes Tumor Growth

Kang

Yaqoob

Geng

et al. 2010

The American Journal of Pathology

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“…There is precedence for this sort of interaction. VASP bundles actin with Rac1 in endothelium and in so doing enhances Rac activation by PKA (23,26).…”

Section: Tablementioning

confidence: 98%

“…Transient VASP phosphorylation by PKG in fMLP-activated neutrophils alters actin polymerization that augments ␤ 2 integrin adherence (24,25). In endothelial cells, VASP is required for ␤ 1 integrin function in a process that involves establishing a protein complex between actin and PKA, followed by PKA-mediated activation of Rac1 (23,26). In fibroblasts, VASP appears to constrain Rac activity (27).…”

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confidence: 99%

Neutrophil β2 Integrin Inhibition by Enhanced Interactions of Vasodilator-stimulated Phosphoprotein with S-Nitrosylated Actin

Thom

Bhopale

Yang

et al. 2011

Journal of Biological Chemistry

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Production of reactive species in neutrophils exposed to hyperoxia causes S-nitrosylation of ␤-actin, which increases formation of short actin filaments, leading to alterations in the cytoskeletal network that inhibit ␤ 2 integrin-dependent adherence (Thom, S. R., Bhopale, V. M., Mancini, D. J., and Milovanova, T. N. (2008) J. Biol. Chem. 283, 10822-10834). In this study, we found that vasodilator-stimulated protein (VASP) exhibits high affinity for S-nitrosylated short filamentous actin, which increases actin polymerization. VASP bundles Rac1, Rac2, cyclic AMP-dependent, and cyclic GMP-dependent protein kinases in close proximity to short actin filaments, and subsequent Rac activation increases actin free barbed end formation. Using specific chemical inhibitors or reducing cell concentrations of any of these proteins with small inhibitory RNA abrogates enhanced free barbed end formation, increased actin polymerization, and ␤ 2 integrin inhibition by hyperoxia. Alternatively, incubating neutrophils with formylmethionylleucylphenylalanine or 8-bromo-cyclic GMP activates either cyclic AMP-dependent or cyclic GMP-dependent protein kinase, respectively, outside of the short F-actin pool and phosphorylates VASP on serine 153. Phosphorylated VASP abrogates the augmented polymerization normally observed with S-nitrosylated actin, VASP binding to actin, elevated Rac activity, and elevated formation of actin free barbed ends, thus restoring normal ␤ 2 integrin function.Reactive species generated when neutrophils are exposed to high oxygen partial pressures (hyperbaric oxygen (HBO 2 )2 ) in vivo or in suspensions ex vivo inhibit ␤ 2 integrin-dependent adherence in animals and humans (1-4). This effect ameliorates a variety of ischemia-reperfusion disorders in animal studies, yet HBO 2 does not impair neutrophil immune surveillance processes (5). The unexplained basis for these observations was the impetus for this project.The reversible nature of HBO 2 -mediated neutrophil ␤ 2 integrin inhibition can be shown by incubating cells with chemoattractants, such as fMLP, or with membrane-permeable cyclic GMP (cGMP) analogs such as 8-bromo-cyclic GMP (8-Br-cGMP) (1-4). The goal of this investigation was to evaluate the mechanism for reversal of HBO 2 effects by these agents and improve understanding of how hyperoxia disturbs the neutrophil cytoskeleton.Neutrophils migrate by coordinating ␤ 2 integrin adhesion with turnover of filamentous actin (F-actin). Integrin adherence is controlled by conformational alterations in the extracellular structure to increase affinity and by clustering in the plane of the cell membrane to improve avidity. HBO 2 impedes avidity but not affinity changes by increasing production of reactive species derived from nitric-oxide synthase and myeloperoxidase (MPO), which cause S-nitrosylation of the four cysteine moieties closest to the carboxyl-terminal end of ␤-actin (2). This increases intracellular formation of short actin filaments that appears as a dense mass of intracellular F-actin by confocal mic...

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“…Direct evidence that underscores the physiologic significance of integrin-matrix interactions in vascular integrity comes from the studies showing that disruption of integrin-ECM interaction with RGD peptides increases venular permeability by 2-to 3-fold in vivo 43 and reduces the barrier properties of endothelial monolayers. 44 In mature EC cultures, ␣V␤3 also localizes to and maintains the integrity of the junctions 45 and decreased ␤3 expression with siRNA increases permeability. 46 Mechanistically, it has been suggested that this integrin collaborates with other intercellular molecules to form lateral junctions and specifically participates in formation of adherens junctions.…”

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confidence: 99%

The integrin coactivator Kindlin-2 plays a critical role in angiogenesis in mice and zebrafish

Pluskota

Dowling

Gordon

et al. 2011

Blood

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Kindlin-2, a widely distributed cytoskeletal protein, has been implicated in integrin activation, and its absence is embryonically lethal in mice and causes severe developmental defects in zebrafish. Knockdown of kindlin-2 levels in endothelial cells resulted in defective adhesive and migratory responses, suggesting that angiogenesis might be aberrant even with partial reduction of kindlin-2. This hypothesis has now been tested in the kindlin-2 ؉/؊ mice. RM1 prostate tumors grown in kindlin-2 ؉/؊ mice had fewer blood vessels, which were thinner and shorter and supported less tumor growth compared with wild-type littermates. The vessels that did form in the kindlin-2 ؉/؊ mice lacked smooth muscle cells and pericytes and had thinner basement membranes, indicative of immature vessels. VEGF-induced angiogenesis in matrigel implants was also abnormal in the kindlin-2 ؉/؊ mice. Vessels in the kindlin-2 ؉/؊ mice were leaky, and BM transplantation from kindlin-2 ؉/؊ to WT mice did not correct this defect. Endothelial cells derived from kindlin-2 ؉/؊ mice had integrin expression levels similar to WT mice but reduced ␣V␤3-dependent signaling, migration, adhesion, spreading, and tube formation. Developmental angiogenesis was markedly impaired by kindlin-2 morpholinos in zebrafish. Taken together, kindlin-2 plays an important role in pathologic and developmental angiogenesis, which arises from defective activation of integrin ␣V␤3. IntroductionAlthough endothelial cells (ECs) are the primary cell type that forms blood vessel tubes, other cell types, including BM-derived cells, smooth muscle cells, and pericytes, are all engaged in forming blood-bearing conduits. Growth factor-growth factor receptor interactions are involved in initial recruitment of these cells while other ligand-receptor systems govern migration of the responding cells into angiogenic tissues (reviewed in Folkman 1 ). Among the cellular receptors that specialize in regulating the adhesive and migratory responses of cells during angiogenesis are members of the integrin family. 2 Integrins of the ␤1 and ␤3 subfamilies on ECs have been implicated in angiogenesis in vivo through knockout 3 and inhibitor approaches 4 and integrin ␣V␤3 and ␣5␤1 are targets of antiangiogenic drugs for cancer treatment (reviewed in Avraamides et al 5 ).The regulation of integrin function in angiogenesis as well as many other responses depends on their ability to undergo activation, a rapid transition from a low-to a high-affinity state for recognition of their ligands. Particularly relevant to angiogenesis is the ability of VEGF to activate integrin ␣V␤3 and ␣5␤1 by engaging one of its EC receptors, VEGFR2. 6 Indeed, VEGFR2 and integrin ␣V␤3 can form a physical complex in EC and influence the functions of each other. 6,7 Two sets of cytoskeletal proteins, the talins and the kindlins, bind to the ␤ cytoplasmic tails and are now known to be involved in integrin activation. [8][9][10] Talin has long been known to be critical for integrin activation 11 and more recent studies have sh...

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Impaired integrin‐mediated adhesion contributes to reduced barrier properties in VASP‐deficient microvascular endothelium

Cited by 34 publications

References 44 publications

Focal Adhesion Assembly in Myofibroblasts Fosters a Microenvironment that Promotes Tumor Growth

Focal Adhesion Assembly in Myofibroblasts Fosters a Microenvironment that Promotes Tumor Growth

Neutrophil β2 Integrin Inhibition by Enhanced Interactions of Vasodilator-stimulated Phosphoprotein with S-Nitrosylated Actin

The integrin coactivator Kindlin-2 plays a critical role in angiogenesis in mice and zebrafish

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