scholarly journals Staphylococcus aureus iron-regulated surface determinant B (IsdB) protein interacts with von Willebrand factor and promotes adherence to endothelial cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Mariangela J. Alfeo ◽  
Anna Pagotto ◽  
Giulia Barbieri ◽  
Timothy J. Foster ◽  
Karen Vanhoorelbeke ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Endothelial Cells ◽  
Von Willebrand Factor ◽  
Iron Transport ◽  
Matrix Proteins ◽  
Potential Therapeutic Target ◽  
A1 Domain ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Micromolar Range

AbstractStaphylococcus aureus is the cause of a spectrum of diseases in humans and animals. The molecular basis of this pathogenicity lies in the expression of a variety of virulence factors, including proteins that mediate adherence to the host plasma and extracellular matrix proteins. In this study, we discovered that the iron-regulated surface determinant B (IsdB) protein, besides being involved in iron transport and vitronectin binding, interacts with von Willebrand Factor (vWF). IsdB-expressing bacteria bound to both soluble and immobilized vWF. The binding of recombinant IsdB to vWF was blocked by heparin and reduced at high ionic strength. Furthermore, treatment with ristocetin, an allosteric agent that promotes the exposure of the A1 domain of vWF, potentiates the binding of IsdB to vWF. Both near-iron transporter motifs NEAT1 and NEAT2 of IsdB individually bound recombinant A1 domain with KD values in the micromolar range. The binding of IsdB and adhesion of S. aureus expressing IsdB to monolayers of activated endothelial cells was significantly inhibited by a monoclonal antibody against the A1 domain and by IsdB reactive IgG from patients with staphylococcal endocarditis. This suggests the importance of IsdB in adherence of S. aureus to the endothelium colonization and as potential therapeutic target.

scholarly journals Staphylococcus Aureus Iron-Regulated Surface Determinant B (IsdB) Protein Interacts with von Willebrand Factor and Promotes Adherence to Endothelial Cells.

2021 ◽  
Author(s):  
Mariangela J Alfeo ◽  
Anna Pagotto ◽  
Giulia Barbieri ◽  
Timothy J Foster ◽  
Karen Vanhoorelbeke ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Endothelial Cells ◽  
Von Willebrand Factor ◽  
Iron Transport ◽  
Matrix Proteins ◽  
Potential Therapeutic Target ◽  
A1 Domain ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Micromolar Range

Abstract Staphylococcus aureus is the cause of a spectrum of diseases in humans and animals. The molecular basis of this pathogenicity lies in the expression of a variety of virulence factors, including proteins that mediate adherence to the host plasma and extracellular matrix proteins. In this study, we discovered that the iron-regulated surface determinant B (IsdB) protein, besides being involved in iron transport and vitronectin binding, interacts with von Willebrand Factor (vWF). IsdB-expressing bacteria bound to both soluble and immobilized vWF. The binding of recombinant IsdB to vWF was blocked by heparin and reduced at high ionic strength. Furthermore, treatment with ristocetin, an allosteric agent that promotes the exposure of the A1 domain of vWF, potentiates the binding of IsdB to vWF. Both near-iron transporter motifs NEAT1 and NEAT2 of IsdB individually bound recombinant A1 domain with KD values in the micromolar range. The binding of IsdB and adhesion of S. aureus expressing IsdB to monolayers of activated endothelial cells was significantly inhibited by a monoclonal antibody against the A1 domain and by IsdB reactive IgG from patients with staphylococcal endocarditis. This suggests the importance of IsdB in adherence of S. aureus to the endothelium colonization and as potential therapeutic target.

Von Willebrand Factor Binds to the Endothelial Growth Factor Angiopoietin-2 Both within Endothelial Cells and Upon Release From Weibel Palade Bodies

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 698-698 ◽  
Author(s):  
Thomas A J Mckinnon ◽  
Richard D Starke ◽  
Kushani Ediriwickrema ◽  
Anna Maria Randi ◽  
Michael Laffan
Keyword(s):  
Extracellular Matrix ◽  
Endothelial Cells ◽  
Von Willebrand Factor ◽  
Extracellular Matrix Proteins ◽  
Matrix Proteins ◽  
Dependent Manner ◽  
Platelet Receptor ◽  
Angiopoietin 2 ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract Abstract 698 Von Willebrand Factor (VWF) is a large multimeric plasma glycoprotein essential for homeostasis, also involved in inflammation and angiogenesis. The majority of VWF is synthesised by endothelial cells (EC) and is either constitutively secreted or stored in Weibel-Palade bodies (WPB), ready to be released in response to endothelial stimulation. Several studies have shown that formation of WPB is dependent on the presence of VWF, and deletion of VWF in human umbilical vein EC (HUVEC) results in loss of WPB. Amongst the other proteins shown to co-localise to WPB is angiopoietin-2 (Ang2), a ligand of the receptor tyrosine kinase Tie-2. Ang2 regulates endothelial cell survival, vascular stability and maturation, by destabilizing quiescent endothelium and facilitating the response to inflammatory and angiogenic stimuli. VWF is required for storage of Ang2, and release of Ang-2 from EC is increased in VWF-deficient HUVEC. Recently, we have shown that VWF itself regulates angiogenesis, raising the hypothesis that some of the angiogenic activity of VWF may be mediated by Ang-2. In the present study we investigated the interaction between Ang2 and VWF. Binding analysis demonstrated that recombinant human Ang2 bound to purified plasma-derived VWF in a pH and calcium dependent manner, with optimal binding occurring at pH 6.5 and 10mM calcium, indicative of binding within the Golgi body. Generation of binding isotherms established that Ang2 bound to VWF with high affinity (KD∼3nM); furthermore binding affinity was not dependent on VWF conformation. Using an array of VWF constructs we determined that Ang2 bound predominantly to the VWF A1 domain, which also contains binding sites to the platelet receptor GPIb and extracellular matrix proteins. Co-immunoprecipitation experiments performed on TNFα- and ionomycin-stimulated HUVECs, to induce WPB exocytosis, confirmed that a portion of Ang2 remained bound to secreted VWF. Moreover, immunofluorescence staining of histamine-stimulated HUVECs to induce VWF release demonstrated the presence of Ang2 on VWF strings secreted from ECs. Finally we demonstrated that Ang2 bound to VWF was still able to interact with Tie-2. These data demonstrate that binding of Ang2 to VWF occurs within the cell; we propose that this is the mechanism mediating storage of Ang2 in WPB. Moreover, the finding that the Ang2-VWF interaction is preserved following secretion raises the intriguing possibility VWF may affect Ang2 function, possibly by localising Ang2 to the Tie 2 receptor under the shear forces experienced in flowing blood. Similarly, Ang-2 binding to VWF may modulate its interaction with receptors and extracellular matrix proteins, and ultimately influence the role of VWF in the angiogenic processes. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Adhesion of Staphylococcus aureus to the vessel wall under flow is mediated by von Willebrand factor–binding protein

Blood ◽  
2014 ◽  
Vol 124 (10) ◽  
pp. 1669-1676 ◽  
Author(s):  
Jorien Claes ◽  
Thomas Vanassche ◽  
Marijke Peetermans ◽  
Laurens Liesenborghs ◽  
Christophe Vandenbriele ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Endothelial Cells ◽  
Von Willebrand Factor ◽  
Vessel Wall ◽  
Binding Protein ◽  
Factor Binding ◽  
Key Points ◽  
Platelet Aggregates ◽  
Von Willebrand ◽  
Willebrand Factor

Key PointsvWbp mediates adhesion of S aureus under flow to activated endothelial cells and the subendothelium via VWF. vWbp activates prothrombin and triggers the formation of bacteria–fibrin–platelet aggregates, which enhance adhesion to vessels under flow.

Transition from Non-Platelet-Binding to Platelet-Binding Conformation of Von Willebrand Factor Occurs upon Exocytosis

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3917-3917
Author(s):  
Evelyn Groot ◽  
Rob Fynheer ◽  
Silvie AE Sebastian ◽  
Peter J Lenting ◽  
Philip G De Groot
Keyword(s):  
Monoclonal Antibody ◽  
Endothelial Cells ◽  
Vascular Injury ◽  
Von Willebrand Factor ◽  
Binding Conformation ◽  
Platelet Binding ◽  
A1 Domain ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Adhesion Of Platelets

Abstract Introduction Von Willebrand factor (VWF) is a large multimeric glycoprotein that contributes to platelet recruitment at sites of vascular injury. VWF is mainly produced in endothelial cells from where it is secreted directly into the circulation or stored in the rod-shaped organelles called Weibel-Palade bodies. VWF present in the circulation does not bind to platelets. Stimulated endothelial cells secrete VWF that has the capacity to spontaneously interact with platelets. Conversion of the platelet-binding conformation of secreted VWF into the non-binding conformation of plasma VWF involves proteolytic processing by the metalloprotease ADAMTS13. At sites of vascular injury, binding of VWF to the exposed subendothelial collagen induces a conformational change in VWF allowing a strong interaction with the platelet receptor glycoprotein (Gp)Ibα. Undesired secretion of active VWF may also occur in several pathological conditions. One example is von Willebrand disease type 2B (VWD2B), where a gain of function mutation in the VWF/A1 domain induces a permanent platelet-binding state in the VWF molecule. As a consequence, VWF can spontaneously interact with platelets in the circulation, leading to thrombocytopenia, a hallmark of VWD2B. Objective The aim of this study was to investigate whether VWF present in the Weibel-Palade bodies of endothelial cells is stored in a platelet-binding conformation. Methods Immunofluorescence experiments were performed on wildtype and VWD2B endothelial cells. Monoclonal antibody AU/VWF-a11 is directed against the VWF/A1 domain and recognizes VWF only when it is in its GpIb-binding conformation. Monoclonal antibody AU/VWF-C37H is directed against the VWF/A3 domain and recognizes both the platelet-binding and the non-platelet- binding conformation. Experiments were performed on cultured endothelial cells to study the conformation of VWF in the Weibel-Palade bodies. To study the conformation of secreted VWF, stimulated endothelial cells were perfused with washed platelets. Results AU/VWF-C37H fluorescence was observed in the Weibel-Palade bodies of both wildtype and VWD2B endothelial cells, whereas AU/VWF-a11 fluorescence was only detected in the Weibel-Palade bodies of the VWD2B cells. Perfusion of washed platelets over wildtype and VWD2B endothelial cells resulted in adhesion of platelets to thin strings of secreted VWF. These strings stained positive for both AU/VWF-C37H and AU/VWF-a11. Of note, significantly more platelets adhered to VWF secreted from VWD2B than from wildtype endothelial cells. This hyperactive VWD2B-like platelet adhesion pattern could be mimicked by wildtype endothelial cells upon perfusion with platelets that were mixed with ristocetin. Conclusions VWF stored within the Weibel-Palade bodies of endothelial cells does not possess platelet-binding capacities. Upon secretion, VWF undergoes a conformational change that allows the adhesion of platelets. The presence of ADAMTS13 is necessary to prevent the release of platelet-binding VWF in the circulation. In the absence of ADAMTS13, secreted VWF does not expose all its GpIb-binding sites as more platelets adhere to ristocetin activated VWF-strings or strings released from VWD2B endothelial cells.

Requirement for Both D Domains of the Propolypeptide in von Willebrand Factor Muitimerization and Storage

1993 ◽  
Vol 70 (06) ◽  
pp. 1053-1057 ◽  
Author(s):  
Agnès M Journet ◽  
Simin Saffaripour ◽  
Denisa D Wagner
Keyword(s):  
Endothelial Cells ◽  
Endoplasmic Reticulum ◽  
Von Willebrand Factor ◽  
Deletion Mutants ◽  
Relative Importance ◽  
D2 Domain ◽  
Constitutive Secretion ◽  
Von Willebrand ◽  
And Storage ◽  
Willebrand Factor

SummaryBiosynthesis of the adhesive glycoprotein von Willebrand factor (vWf) by endothelial cells results in constitutive secretion of small multimers and storage of the largest multimers in rodshaped granules called Weibel-Palade bodies. This pattern is reproduced by expression of pro-vWf in heterologous cells with a regulated pathway of secretion, that store the recombinant protein in similar elongated granules. In these cells, deletion of the vWf prosequence prevents vWf storage. The prosequence, composed of two homologous domains (D1 and D2), actively participates in vWf multimer formation as well. We expressed deletion mutants lacking either the D1 domain (D2vWf) or the D2 domain (D1vWf) in various cell lines to analyze the relative importance of each domain in vWf muitimerization and storage. Both proteins were secreted efficiently without being retained in the endoplasmic reticulum. Despite this, neither multimerized past the dimer stage and they were not stored. We conclude that several segments of the prosequence are jointly involved in vWf muitimerization and storage.

Adverse Influence of Cigarette Smoking on the Endothelium

1993 ◽  
Vol 70 (04) ◽  
pp. 707-711 ◽  
Author(s):  
Andrew D Blann ◽  
Charles N McCollum
Keyword(s):  
Endothelial Cells ◽  
Von Willebrand Factor ◽  
Tobacco Smoke ◽  
Lipid Peroxides ◽  
Short Exposure ◽  
Acute Effects ◽  
Adverse Influence ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryThe effect of smoking on the blood vessel intima was examined by comparing indices of endothelial activity in serum from smokers with that from non-smokers. Serum from smokers contained higher levels of von Willebrand factor (p <0.01), the smoking markers cotinine (p <0.02) and thiocyanate (p <0.01), and was more cytotoxic to endothelial cells in vitro (p <0.02) than serum from non-smokers. The acute effects of smoking two unfiltered medium tar cigarettes was to briefly increase von Willebrand factor (p <0.001) and cytotoxicity of serum to endothelial cells in vitro (p <0.005), but lipid peroxides or thiocyanate were not increased by this short exposure to tobacco smoke. Although there were correlations between von Willebrand factor and smokers consumption of cigarettes (r = 0.28, p <0.02), number of years smoking (r = 0.41, p <0.001) and cotinine (r = 0.45, p <0.01), the tissue culture of endothelial cells with physiological levels of thiocyanate or nicotine suggested that these two smoking markers were not cytotoxic. They are therefore unlikely to be directly responsible for increased von Willebrand factor in the serum of smokers. We suggest that smoking exerts a deleterious influence on the endothelium and that the mechanism is complex.

Epinephrine Induces von Willebrand Factor Release from Cultured Endothelial Cells: Involvement of Cyclic AMP-dependent Signalling in Exocytosis

1997 ◽  
Vol 77 (06) ◽  
pp. 1182-1188 ◽  
Author(s):  
Ulrich M Vischer ◽  
Claes B Wollheinn
Keyword(s):  
Endothelial Cells ◽  
Adenylate Cyclase ◽  
Von Willebrand Factor ◽  
Umbilical Vein ◽  
Free Calcium ◽  
Camp Content ◽  
Von Willebrand ◽  
Willebrand Factor

Summaryvon Willebrand factor (vWf) is released from endothelial cell storage granules after stimulation with thrombin, histamine and several other agents that induce an increase in cytosolic free calcium ([Ca2+]i). In vivo, epinephrine and the vasopressin analog DDAVP increase vWf plasma levels, although they are thought not to induce vWf release from endothelial cells in vitro. Since these agents act via a cAMP-dependent pathway in responsive cells, we examined the role of cAMP in vWf secretion from cultured human umbilical vein endothelial cells. vWf release increased by 50% in response to forskolin, which activates adenylate cyclase. The response to forskolin was much stronger when cAMP degradation was blocked with IBMX, an inhibitor of phosphodiesterases (+200%), whereas IBMX alone had no effect. vWf release could also be induced by the cAMP analogs dibutyryl-cAMP (+40%) and 8-bromo-cAMP (+25%); although their effect was weak, they clearly potentiated the response to thrombin. Epinephrine (together with IBMX) caused a small, dose-dependent increase in vWf release, maximal at 10-6 M (+50%), and also potentiated the response to thrombin. This effect is mediated by adenylate cyclase-coupled β-adrenergic receptors, since it is inhibited by propranolol and mimicked by isoproterenol. In contrast to thrombin, neither forskolin nor epinephrine caused an increase in [Ca2+]j as measured by fura-2 fluorescence. In addition, the effects of forskolin and thrombin were additive, suggesting that they act through distinct signaling pathways. We found a close correlation between cellular cAMP content and vWf release after stimulation with epinephrine and forskolin. These results demonstrate that cAMP-dependent signaling events are involved in the control of exocytosis from endothelial cells (an effect not mediated by an increase in [Ca2+]i) and provide an explanation for epinephrine-induced vWf release.

scholarly journals Evidence for the Misfolding of the A1 Domain within Multimeric von Willebrand Factor in Type 2 von Willebrand Disease

2020 ◽  
Vol 432 (2) ◽  
pp. 305-323 ◽  
Author(s):  
Alexander Tischer ◽  
Maria A. Brehm ◽  
Venkata R. Machha ◽  
Laurie Moon-Tasson ◽  
Linda M. Benson ◽  
...  
Keyword(s):  
Von Willebrand Factor ◽  
Von Willebrand Disease ◽  
A1 Domain ◽  
Von Willebrand ◽  
Willebrand Factor

scholarly journals Quantification of von Willebrand factor and ADAMTS-13 after traumatic injury: a pilot study

2021 ◽  
Vol 6 (1) ◽  
pp. e000703
Author(s):  
Taleen A MacArthur ◽  
Julie Goswami ◽  
Laurie Moon Tasson ◽  
Alexander Tischer ◽  
Kent R Bailey ◽  
...  
Keyword(s):  
Healthy Volunteers ◽  
Acute Phase ◽  
Von Willebrand Factor ◽  
Thrombin Generation ◽  
Trauma Patients ◽  
Phase Reaction ◽  
Time Points ◽  
A1 Domain ◽  
Von Willebrand ◽  
Willebrand Factor

BackgroundVon Willebrand factor (VWF) is an acute phase reactant synthesized in the megakaryocytes and endothelial cells. VWF forms ultra-large multimers (ULVWF) which are cleaved by the metalloprotease ADAMTS-13, preventing spontaneous VWF–platelet interaction. After trauma, ULVWF is released into circulation as part of the acute phase reaction. We hypothesized that trauma patients would have increased levels of VWF and decreased levels of ADAMTS-13 and that these patients would have accelerated thrombin generation.MethodsWe assessed plasma concentrations of VWF antigen and ADAMTS-13 antigen, the Rapid Enzyme Assays for Autoimmune Diseases (REAADS) activity of VWF, which measure exposure of the platelet-binding A1 domain, and thrombin generation kinetics in 50 samples from 30 trauma patients and an additional 21 samples from volunteers. Samples were analyzed at 0 to 2 hours and at 6 hours from the time of injury. Data are presented as median (IQR) and Kruskal-Wallis test was performed between trauma patients and volunteers at both time points.ResultsREAADS activity was greater in trauma patients than volunteers both at 0 to 2 hours (190.0 (132.0–264.0) vs. 92.0 (71.0–114.0), p<0.002) and at 6 hours (167.5 (108.0–312.5.0) vs. 92.0 (71.0–114.0), p<0.001). ADAMTS-13 antigen levels were also decreased in trauma patients both at 0 to 2 hours (0.84 (0.51–0.94) vs. 1.00 (0.89–1.09), p=0.010) and at 6 hours (0.653 (0.531–0.821) vs. 1.00 (0.89–1.09), p<0.001). Trauma patients had accelerated thrombin generation kinetics, with greater peak height and shorter time to peak than healthy volunteers at both time points.DiscussionTrauma patients have increased exposure of the VWF A1 domain and decreased levels of ADAMTS-13 compared with healthy volunteers. This suggests that the VWF burst after trauma may exceed the proteolytic capacity of ADAMTS-13, allowing circulating ULVWF multimers to bind platelets, potentially contributing to trauma-induced coagulopathy.Level of evidenceProspective case cohort study.

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