scholarly journals Adhesive properties of the beta 3 integrins: comparison of GP IIb-IIIa and the vitronectin receptor individually expressed in human melanoma cells.

1991 ◽  
Vol 113 (2) ◽  
pp. 451-461 ◽  
Author(s):  
N Kieffer ◽  
L A Fitzgerald ◽  
D Wolf ◽  
D A Cheresh ◽  
D R Phillips
Keyword(s):  
Von Willebrand Factor ◽  
Cell Attachment ◽  
Melanoma Cells ◽  
Human Melanoma ◽  
Adhesive Properties ◽  
Vitronectin Receptor ◽  
Hel Cells ◽  
Alpha V Beta 3 ◽  
Von Willebrand ◽  
Willebrand Factor

Glycoprotein IIb-IIIa (alpha IIb beta 3) and the vitronectin receptor (alpha v beta 3), two integrins that share the common beta 3 subunit, have been reported to function as promiscuous receptors for the RGD-containing adhesive proteins fibrinogen, vitronectin, fibronectin, von Willebrand factor, and thrombospondin. The present study was designed to establish a cell system for the expression of either GP IIb-IIIa or the vitronectin receptor in an otherwise identical cellular environment and to compare the adhesive properties of these two integrins with those of native GP IIb-IIIa and the vitronectin receptor constitutively expressed in HEL cells or platelets. M21 human melanoma cells lack GP IIb-IIIa and use the vitronectin receptor to attach to vitronectin, fibrinogen, fibronectin, and von Willebrand factor. To study the functional properties of GP IIb-IIIa in these cells, we transfected GP IIb into M21-L cells, a variant of M21 cells (Cheresh, D.A., and R.C. Spiro. 1987. J. Biol. Chem. 262:17703-17711), which lack the expression of functional alpha v and are therefore unable to attach to vitronectin, fibrinogen, and von Willebrand factor. Transfectants expressing GP IIb were isolated by immunomagnetic beads and surface expression of the GP IIb-IIIa complex was documented by FACS analysis and immunoprecipitation experiments performed with 125I-labeled M21-L/GP IIb cells. Comparative functional studies demonstrated that GP IIb-IIIa expressed in M21-L/GPIIb cells as well as native GP IIb-IIIa constitutively expressed in HEL-5J20 cells (an HEL variant lacking alpha v beta 3) mediated cell attachment to immobilized fibrinogen, but not to vitronectin or von Willebrand factor, whereas the vitronectin receptor expressed in M21 cells and HEL-AD1 cells (an HEL variant expressing alpha v beta 3) mediated cell attachment to fibrinogen, vitronectin, and von Willebrand factor. Similarly, PGl2-treated resting platelets attached to immobilized fibrinogen but not to vitronectin or von Willebrand factor, and this attachment could be inhibited by mAb A2A9 (directed against a functional site on the GP IIb-IIIa complex). However, in contrast to platelets, which adhered to vitronectin and von Willebrand factor after stimulation by thrombin or PMA, activation of the protein kinase C pathway in M21-L/GP IIb or HEL cells did not induce cell adhesion to vitronectin or von Willebrand factor.(ABSTRACT TRUNCATED AT 400 WORDS)

Glycoprotein Ib Can Mediate Endothelial Cell Attachment to a von Willebrand Factor Substratum

1995 ◽  
Vol 73 (02) ◽  
pp. 309-317 ◽  
Author(s):  
Dorothy A Beacham ◽  
Miguel A Cruz ◽  
Robert I Handin
Keyword(s):  
Endothelial Cell ◽  
Von Willebrand Factor ◽  
Cell Attachment ◽  
Umbilical Vein ◽  
Single Amino Acid ◽  
Cell Surface Expression ◽  
Platelet Glycoprotein ◽  
Glycoprotein Ib ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryIntroduction of single amino acid substitutions into the C-terminal Arg-Gly-Asp-Ser (RGDS) site of von Willebrand Factor, referred to as RGD mutant vWF, selectively abrogated vWF binding to platelet glycoprotein IIb/IIIa (GpIIb/IIIa, αIIbβ3 and abolished human umbilical vein endothelial cell (HUVEC) spreading, but not attachment, to RGD mutant vWF (Beacham, D. A., Wise, R. J., Turci, S. M. and Handin, R. I. 1992. J. Biol. Chem. 167, 3409-3415). These results suggested that in addition to the vitronectin receptor (VNR, αvβ3), a second endothelial membrane glycoprotein can mediate HUVEC adhesion to vWF. HUVEC attachment to wild-type (WT) and RGD-mutant vWF was reduced by two proteins known to block the vWF-platelet glycoprotein Ib/IX (GpIb/IX) interaction, the monoclonal antibody AS-7 and the recombinant polypeptide, vWF-A1. The addition of cytochalasin B or DNase I to disrupt potential GPIbα-cytoskeletal interactions enhanced the immunoprecipitation of endothelial GPIbα, caused HUVEC to round up, and increased HUVEC adhesion to RGD mutant vWF. These results indicate that while the VNR is the primary adhesion receptor for vWF, endothelial GPIbα can mediate HUVEC attachment to vWF. GpIb-dependent attachment could contribute to HUVEC adhesion under conditions when cell surface expression of the VNR is downregulated, and VNR-dependent adhesion is reduced.

scholarly journals Adhesive properties of the carbohydrate-modified von Willebrand factor (CHO-vWF)

Blood ◽  
1988 ◽  
Vol 71 (4) ◽  
pp. 947-952 ◽  
Author(s):  
AB Federici ◽  
C De Romeuf ◽  
PG De Groot ◽  
B Samor ◽  
R Lombardi ◽  
...  
Keyword(s):  
Sialic Acid ◽  
Von Willebrand Factor ◽  
Platelet Adhesion ◽  
Platelet Rich Plasma ◽  
Significant Loss ◽  
Total Carbohydrate ◽  
Adhesive Properties ◽  
Normal Platelet ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract In this cooperative study, we explored the role of the carbohydrate moiety (CHO) of von Willebrand factor (vWF) in supporting platelet adhesion. Because of previous discrepant results, all purification steps and CHO modifications by various enzymes were critically evaluated. Under our conditions, CHO-modified vWF preparations contained less than 5% of the initial sialic acid ([Neu]-ase-vWF) and less than 45% ([Neu-Gal]-ase-vWF) or 21% ([Neu-Gal-eF]-ase-vWF) of the D-galactose. These preparations usually showed increased electrophoretic mobility but no significant loss of high-mol-wt multimers when proteolysis had been prevented. Some degree of proteolysis was noted in some carbohydrate-modified vWFs, but the degree of degradation observed did not correlate with the removal of D- galactose. Platelet adhesion to various matrices increased after removal of the terminal sialic acid ([Neu]-ase-vWF) and approximately 45% of the D-galactose ([Neu-Gal]-ase-vWF), but returned to normal values when greater than 70% of the total carbohydrate had been removed by endoglycosidase F [Neu-Gal-ef]-ase-vWF). These changes in reactivity were also reflected in the spontaneous aggregation in normal platelet- rich plasma (PRP) after CHO removal.

THE EFFECT OF PHORBOL DIESTERS AND INTERLEUKINS ON THE SECRETION OF VON WILLEBRAND FACTOR BY HUMAN ENDOTHELIAL CELLS IN CULTURE

1987 ◽  
Author(s):  
J C Giddings ◽  
L Shall
Keyword(s):  
Endothelial Cells ◽  
Von Willebrand Factor ◽  
Umbilical Vein ◽  
Interleukin 1 ◽  
Interleukin 2 ◽  
Morphological Evidence ◽  
Adhesive Properties ◽  
Umbilical Vein Endothelial Cells ◽  
Von Willebrand ◽  
Willebrand Factor

Human umbilical vein endothelial cells (EC) were cultured in the presence of 4p-phorbol 12-myristate 13-acetate (PMA, 10ug/l), interleukin 1 (IL-1, 1 unit/ml) and interleukin 2 (IL-2, 1 unit/ml), and secretion of von Willebrand factor activity (vWF, Ristocetin co-factor) and von Willebrand factor antigen (vWFAG, ELISA Technique) measured at intervals. Confluent control EC were treated with PMA, IL-1 and IL-2, and the supernatant medium assayed for release of vWF and vWFAg. Treated cells were also examined for vWFAg by immuno-fluorescence. The levels of both vWF and vWFAg in cultures containing IL-1 were significantly higher than those in control cultures after 5-6 days growth. Moreover, vWF and vWFAg increased significantly in the supernatant of confluent control EC incubated further in the presence of IL-1. Furthermore, the characteristic fluorescence pattern of endothelial vWFAg was markedly reduced in EC treated with IL-1. The levels of vWF and vWFAg in cultures containing PMA were also significantly higher than those of control cultures. In these conditions, however, the growth of cells appeared to be enhanced, and confluence was observed after about 6 days in the presence of PMA compared to 9 - 10 days in control cultures. The mean levels of vWF and vWFAg in the supernatant of EC incubated with PMA were higher than the control values but the differences were not statistically significant. Immunofluorescence of PMA-treated cells suggested that vWFAg might be less granular than in control cells but the differences were not as marked as those seen with IL-1. The results of all assays in the presence of IL-2 were not significantly different from those of control cells. In all instances no morphological evidence of endothelial injury was observed and more than 90% of cells remained viable at the termination of cultures. The results indicated that the synthesis and release of vWF were increased in the presence of PMA, and secretion of vWF was stimulated by IL-1. The data suggest that secreted vWF might contribute to the previously reported enhanced procoagulant and adhesive properties of EC treated with these substances.

Retention Test Homburg To Monitor the Adhesive Properties of von Willebrand Factor after Substitution or Desmopressin Therapy

2005 ◽  
Vol 31 (04) ◽  
pp. 441-448
Author(s):  
J. Ulrich Wieding ◽  
Juliane Matern ◽  
Heinz Köstering ◽  
Cornelia Wermes ◽  
Ernst Wenzel
Keyword(s):  
Von Willebrand Factor ◽  
Retention Test ◽  
Adhesive Properties ◽  
Von Willebrand ◽  
Willebrand Factor

scholarly journals Solid-phase von Willebrand factor contains a conformationally active RGD motif that mediates endothelial cell adhesion through the alpha v beta 3 receptor

Blood ◽  
1993 ◽  
Vol 82 (12) ◽  
pp. 3622-3630 ◽  
Author(s):  
C Denis ◽  
JA Williams ◽  
X Lu ◽  
D Meyer ◽  
D Baruch
Keyword(s):  
Monoclonal Antibody ◽  
Cell Adhesion ◽  
Endothelial Cell ◽  
Von Willebrand Factor ◽  
Solid Phase ◽  
Rgd Sequence ◽  
Alpha V Beta 3 ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Endothelial Cell Adhesion

Abstract The interaction of von Willebrand factor (vWF) with the alpha v beta 3 integrin of human umbilical vein endothelial cells is dependent on the RGD sequence present at residues 1744–1746 of the mature vWF subunit. We compared vWF and its two dimeric fragments, SpIII (residues 1–1365) and SpII (residues 1366–2050), as adhesion substrates. Solid-phase vWF and SpII supported endothelial cell adhesion, whereas SpIII, which contains the glycoprotein (GP) Ib binding domain, did not. Soluble SpII inhibited adhesion to immobilized ligands, whereas soluble vWF did not, suggesting that exposure of the cell attachment domain involves a conformational modification of vWF. Dendroaspin and albolabrin, two RGD- containing peptides of the disintegrin family, were potent inhibitors of cell adhesion to vWF (IC50 approximately 15 nmol/L). Complete inhibition of endothelial cell adhesion to vWF was obtained in the presence of F(ab')2 of monoclonal antibody 9 to vWF, which blocks vWF binding to platelet GPIIb/IIIa. In contrast, monoclonal antibody 713 to vWF, which blocks its binding to platelet GPIb, did not inhibit cell adhesion to vWF. These results indicate that endothelial cell adhesion to vWF is mediated by an RGD-dependent interaction with alpha v beta 3, but does not seem to involve a GPIb-like receptor, and show the importance of the conformation of the RGD sequence.

scholarly journals Structure of Plasmodium falciparum TRAP (thrombospondin-related anonymous protein) A domain highlights distinct features in apicomplexan von Willebrand factor A homologues

2013 ◽  
Vol 450 (3) ◽  
pp. 469-476 ◽  
Author(s):  
Tero Pihlajamaa ◽  
Tommi Kajander ◽  
Juho Knuuti ◽  
Kaisa Horkka ◽  
Amit Sharma ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Von Willebrand Factor ◽  
Metal Ion ◽  
Cell Attachment ◽  
Protein A ◽  
Site Directed Mutagenesis ◽  
Mammalian Host ◽  
Functional Roles ◽  
Von Willebrand ◽  
Willebrand Factor

TRAP (thrombospondin-related anonymous protein), localized in the micronemes and on the surface of sporozoites of the notorious malaria parasite Plasmodium, is a key molecule upon infection of mammalian host hepatocytes and invasion of mosquito salivary glands. TRAP contains two adhesive domains responsible for host cell recognition and invasion, and is known to be essential for infectivity. In the present paper, we report high-resolution crystal structures of the A domain of Plasmodium falciparum TRAP with and without bound Mg2+. The structure reveals a vWA (von Willebrand factor A)-like fold and a functional MIDAS (metal-ion-dependent adhesion site), as well as a potential heparan sulfate-binding site. Site-directed mutagenesis and cell-attachment assays were used to investigate the functional roles of the surface epitopes discovered. The reported structures are the first determined for a complete vWA domain of parasitic origin, highlighting unique features among homologous domains from other proteins characterized hitherto. Some of these are conserved among Plasmodiae exclusively, whereas others may be common to apicomplexan organisms in general.

Catching platelets from the bloodflow: the role of the conformation of von Willebrand factor

2018 ◽  
Vol 13 (5) ◽  
pp. 44 ◽  
Author(s):  
Aleksey V. Belyaev
Keyword(s):  
Von Willebrand Factor ◽  
Chemical Properties ◽  
Growth Dynamics ◽  
Blood Platelet ◽  
Adhesive Properties ◽  
Bulk Fluid ◽  
Initial Adhesion ◽  
Von Willebrand ◽  
Willebrand Factor

The mechanics of platelet initial adhesion due to interactions between GPIb receptor with von Willebrand factor (vWf) multimers is essential for thrombus growth and the regulation of this process. Multimeric structure of vWf is known to make adhesion sensitive to the hydrodynamic conditions, providing intensive platelet aggregation in bulk fluid for high shear rates. But it is still unclear how it affects the dynamics of platelet motion near vessel walls and efficiency of their adhesion to surfaces. Our goal is to resolve the principal issues in the mechanics of platelet initial attachment via GPIb-vWf bonds in near-wall flow conditions: when the platelet tends to roll or slide and how this dynamics depends on the size, conformation and adhesive properties of the vWf multimers. We employ a 3D computer model based on a combination of the Lattice Boltzmann method with mesoscopic particle dynamics for explicit simulation of vWf-mediated blood platelet adhesion in shear flow. Our results reveal the link between the mechanics of platelet initial adhesion and the physico-chemical properties of vWf multimers. This has implications in further theoretical investigation of thrombus growth dynamics, as well as the interpretation of in vitro experimental data.

scholarly journals The Flow Dependent Adhesion of von Willebrand Factor (VWF)-A1 Functionalized Nanoparticles in an in Vitro Coronary Stenosis Model

Molecules ◽  
2019 ◽  
Vol 24 (15) ◽  
pp. 2679 ◽  
Author(s):  
Yathreb Asaad ◽  
Mark Epshtein ◽  
Andrew Yee ◽  
Netanel Korin
Keyword(s):  
Von Willebrand Factor ◽  
Coronary Stenosis ◽  
Nano Particles ◽  
Adhesive Properties ◽  
Polystyrene Nanoparticles ◽  
A1 Domain ◽  
Von Willebrand ◽  
Willebrand Factor

In arterial thrombosis, von Willebrand factor (VWF) bridges platelets to sites of vascular injury. The adhesive properties of VWF are controlled by its different domains, which may be engineered into ligands for targeting nanoparticles to vascular injuries. Here, we functionalized 200 nm polystyrene nanoparticles with the VWF-A1 domain and studied their spatial adhesion to collagen or collagen-VWF coated, real-sized coronary stenosis models under physiological flow. When VWF-A1 nano-particles (A1-NPs) were perfused through a 75% stenosis model coated with collagen-VWF, the particles preferentially adhered at the post stenotic region relative to the pre-stenosis region while much less adhesion was detected at the stenosis neck (~ 65-fold less). When infused through collagen-coated models or when the A1 coating density of nanoparticles was reduced by 100-fold, the enhanced adhesion at the post-stenotic site was abolished. In a 60% stenosis model, the adhesion of A1-NPs to collagen-VWF-coated models depended on the location examined within the stenosis. Altogether, our results indicate that VWF-A1 NPs exhibit a flow-structure dependent adhesion to VWF and illustrate the important role of studying cardiovascular nano-medicines in settings that closely model the size, geometry, and hemodynamics of pathological environments.

INCREASED ADHESIVE PROPERTIES OF THE CARBOHYDRATE-MODIFIED VON WILLEBRAND FACTOR (CHO-VWF): CRITICAL ROLE OF THE MULTIMERIC STRUCTURE AND CORRELATION WITH PLATELET AGGREGATION

1987 ◽  
Author(s):  
A B Federici ◽  
C De Romeuf ◽  
P G De Groot ◽  
P M Mannucci ◽  
B Samor ◽  
...  
Keyword(s):  
Von Willebrand Factor ◽  
Platelet Adhesion ◽  
Platelet Rich Plasma ◽  
Critical Role ◽  
Freezing And Thawing ◽  
Adhesive Properties ◽  
Spontaneous Aggregation ◽  
Von Willebrand ◽  
Willebrand Factor

We have reexplored the role of the carbohydrate moiety (CHO) on the von Willebrand Factor (vWF) structure and function by critically evaluating its different purification steps and modifications in CHO content by specific enzymes. Structural and functional assays have been evaluated separately in each laboratory (Milano and Lille) and jointly in Utrecht during several organized experiments. Under our conditions, the CHOVWFs obtained were characterized by less than 5% of sialic acid "(Neu)asevWF" and about 45% of D-Galactose "(Neu-Gal)ase-vWF" remaining, by increased electrophoretic mobility without any significant losses of the high molecular weight multimers and by their capacity to induce spontaneous aggregation in normal platelet rich plasma (PRP). Platelet adhesion to these different CHO-vWFs was tested in the flat chamber devised by Sakariassen in the presence of different subendothelial matrices and data expressed as the percentage of the surface covered by platelets. The blood reconstituted with different plasma samples showed the following percentual values of surface coverage (mean ± SD):- Normal plasma = 15 ± 3.8- Severe vWd plasma = 4 ± 1.9- SvWd pl + Native vWF = 14 ± 2.8- SvWd pl + (Neu) ase-vWF = 23 ± 3.5- SvWd pl + (Neu-Gal) ase-vWF = 19 ± 2.9This significantly increased adhesion to the subendothelium of the CHO-vWFs corresponded to the spontaneous aggregation present in normal PRP but it disappeared when the multimeric structure was damaged by in vitro proteolysis and/or by storage conditions (changes in temperature and freezing and thawing). From these results we may conclude that removal of terminal sugars enhances not only platelet-vWF interactions, but also platelet adhesion to the subendothelium.

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