scholarly journals von Willebrand factor bound to glycoprotein Ib is cleared from the platelet surface after platelet activation by thrombin

Blood ◽  
1992 ◽  
Vol 79 (8) ◽  
pp. 2011-2021
Author(s):  
P Hourdille ◽  
HR Gralnick ◽  
E Heilmann ◽  
A Derlon ◽  
AM Ferrer ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Platelet Activation ◽  
Von Willebrand Factor ◽  
Platelet Rich Plasma ◽  
Surface Expression ◽  
Von Willebrand Disease ◽  
Platelet Surface ◽  
Immunogold Staining ◽  
Von Willebrand ◽  
Willebrand Factor

We recently reported that after activation of human platelets by thrombin, glycoprotein (GP) Ib-IX complexes are translocated to the surface-connected canalicular system (SCCS) (Blood 76:1503, 1990). As GPIb is a major receptor for von Willebrand factor (vWF) in platelet adhesion, we have now examined the consequences of thrombin activation on the organization of vWF bound to GPIb on the platelet surface. Studies were performed using monoclonal or polyclonal antibodies in either immunogold staining and electron microscopy (Au-EM) or in flow cytometry. When unstirred platelet-rich plasma was incubated with ristocetin, bound vWF was located by Au-EM as discrete masses regularly distributed over the cell surface. Platelets from a patient with Glanzmann's thrombasthenia, lacking GPIIb-IIIa complexes, gave a similar pattern, confirming that this represented binding to GPIb. That ristocetin was not precipitating vWF before their binding to the platelets was shown by the detection of similar masses on the surface of platelets of a patient with type IIB von Willebrand disease. Experiments were continued using washed normal platelets incubated in Tyrode-EDTA, the purpose of the EDTA being to limit the surface expression of endogenous vWF after platelet stimulation. Under these conditions, platelets were treated with ristocetin for 5 minutes at 37 degrees C in the presence of increasing amounts of purified vWF. This was followed by incubation with thrombin (0.5 U/mL) for periods of up to 10 minutes. Flow cytometry showed a time-dependent loss in the surface expression of vWF bound to GPIb and these changes were confirmed by Au-EM. In particular, immunogold staining performed on ultrathin sections showed that the bulk of the vWF was being cleared to internal membrane systems. Surface clearance of vWF during thrombin- induced platelet activation is a potential mechanism for regulating platelet adhesivity.

scholarly journals von Willebrand factor bound to glycoprotein Ib is cleared from the platelet surface after platelet activation by thrombin

Blood ◽  
1992 ◽  
Vol 79 (8) ◽  
pp. 2011-2021 ◽  
Author(s):  
P Hourdille ◽  
HR Gralnick ◽  
E Heilmann ◽  
A Derlon ◽  
AM Ferrer ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Platelet Activation ◽  
Von Willebrand Factor ◽  
Platelet Rich Plasma ◽  
Surface Expression ◽  
Von Willebrand Disease ◽  
Platelet Surface ◽  
Immunogold Staining ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract We recently reported that after activation of human platelets by thrombin, glycoprotein (GP) Ib-IX complexes are translocated to the surface-connected canalicular system (SCCS) (Blood 76:1503, 1990). As GPIb is a major receptor for von Willebrand factor (vWF) in platelet adhesion, we have now examined the consequences of thrombin activation on the organization of vWF bound to GPIb on the platelet surface. Studies were performed using monoclonal or polyclonal antibodies in either immunogold staining and electron microscopy (Au-EM) or in flow cytometry. When unstirred platelet-rich plasma was incubated with ristocetin, bound vWF was located by Au-EM as discrete masses regularly distributed over the cell surface. Platelets from a patient with Glanzmann's thrombasthenia, lacking GPIIb-IIIa complexes, gave a similar pattern, confirming that this represented binding to GPIb. That ristocetin was not precipitating vWF before their binding to the platelets was shown by the detection of similar masses on the surface of platelets of a patient with type IIB von Willebrand disease. Experiments were continued using washed normal platelets incubated in Tyrode-EDTA, the purpose of the EDTA being to limit the surface expression of endogenous vWF after platelet stimulation. Under these conditions, platelets were treated with ristocetin for 5 minutes at 37 degrees C in the presence of increasing amounts of purified vWF. This was followed by incubation with thrombin (0.5 U/mL) for periods of up to 10 minutes. Flow cytometry showed a time-dependent loss in the surface expression of vWF bound to GPIb and these changes were confirmed by Au-EM. In particular, immunogold staining performed on ultrathin sections showed that the bulk of the vWF was being cleared to internal membrane systems. Surface clearance of vWF during thrombin- induced platelet activation is a potential mechanism for regulating platelet adhesivity.

Post-DDAVP Thrombocytopenia in Type 2B von Willebrand Disease Is not Associated with Platelet Consumption: Failure to Demonstrate Glycocalicin Increase or Platelet Activation

1999 ◽  
Vol 81 (02) ◽  
pp. 224-228 ◽  
Author(s):  
A. Steffan ◽  
E. Pontara ◽  
A. Zucchetto ◽  
C. Rossi ◽  
L. De Marco ◽  
...  
Keyword(s):  
Platelet Count ◽  
Platelet Activation ◽  
Surface Expression ◽  
Von Willebrand Disease ◽  
Platelet Surface ◽  
Platelet Counts ◽  
Normal Platelet ◽  
Thrombotic Complications ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryThrombocytopenia is frequently reported in type 2B von Willebrand disease (vWD), and thought to be related to the abnormally high affinity of 2B von Willebrand factor (vWF) for platelet GPIb-IX. To gain an insight into the nature of this thrombocytopenia, we measured plasma glycocalicin (GC) levels (as a marker of platelet turnover), and platelet surface expression of the alpha granule protein P-selectin (as a marker of platelet activation) in 9 patients with type 2B vWD before, and in 4 patients also following the infusion of 1-desamino-8-d-arginine vasopressin (DDAVP). Three patients presented a persistent decrease of platelet counts in the resting condition. GC levels were within the normal range, regardless of the platelet counts, in all but one patient who presented, on the other hand, a normal platelet count. Moreover, platelets expressed normal amounts of P-selectin on their surface, regardless of platelet counts. These findings suggest that the thrombocytopenia observed in type 2B vWD is not due to platelet activation and subsequent consumption in circulation.Despite a significant, albeit transient, decrease in platelet count, DDAVP did not induce an increase in plasma GC levels, nor enhance P-selectin expression. These observations indicate that the acute post-DDAVP thrombocytopenia in type 2B vWD is not related to platelet activation and consumption. We advance that the post-DDAVP 2B vWF is hemostatically more active, and able to induce agglutination but not aggregation of circulating platelets. This would explain both the prompt recovery of basal platelet counts after the post-DDAVP decrease, and the lack of reported thrombotic complications in this disorder.Therefore, even though 2B vWF is characterized by an enhanced affinity for the platelet surface, its binding to platelet GPIb-IX in the soluble phase is not able to induce true platelet aggregation; vWF thus appears to be mainly an adhesive protein, rather than an aggregating agent.

Factor VIII’s C Domains Bind to Platelets.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1715-1715
Author(s):  
Ting-Chang Hsu ◽  
Kathleen P. Pratt ◽  
Arthur R. Thompson
Keyword(s):  
Monoclonal Antibody ◽  
Flow Cytometry ◽  
Factor Viii ◽  
Platelet Activation ◽  
Von Willebrand Factor ◽  
C2 Domain ◽  
Platelet Surface ◽  
Activated Platelets ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract The C domains of factor VIII contain the primary binding site for the cofactor, activated factor VIII, to interact with the phospholipid membranes, including those on the platelet surface. Isolated C2 domain has been shown to bind to phosphotidyl-L-serine-rich lipids and platelets; under flow cytometry, binding to activated platelets was confirmed. For comparison, C1C2, expressed in E.coli, was prepared with up to mg quantities isolated. Fresh, gel-filtered platelets were then studied in a flow cytometer either with or without activation by the thrombin receptor peptide, SFLLRN-amide. Depending upon the conditions, up to 80% of the platelets could be stained with a monoclonal antibody to C2 (ESH8) that is known not to compete with lipid or von Willebrand factor binding. The results were confirmed using a S2296C mutant C1C2 where the free suflhydryl group was either biotinylated and detected by fluorescein labeled streptavidin or directly labeled with fluorescein. As shown in the figure, essentially all platelets bound directly fluorescein labeled C1C2. Using standardized, labeled microbeads, it was estimated that there are 7000–10,000 binding sites per platelet. After platelet activation, the number of platelets binding C1C2 increased with all three detecting systems but only by 15–30%. In contrast, binding of isolated C2, as determined either by ESH8 or as a C2296 biotinylated species, was much lower when the same molar amounts were added, and was primarily detectable following platelet activation. C1C2 binding appeared independent of von Willebrand factor as platelets from two unrelated subjects with severe, type 3 von Willebrand disease gave the same patterns on flow cytometry as seen in platelets from normal subjects. ESH4, a monoclonal antibody known to inhibit binding of C2 to lipid membranes effectively competed C1C2 binding to platelets. Although an indirect alteration the C2 domain conformation cannot be excluded, results support a direct role of C1 in enhancing platelet binding. Binding of direct florescein-labeled C1C2 to SFLLRN-amide-activated platelets Binding of direct florescein-labeled C1C2 to SFLLRN-amide-activated platelets

scholarly journals Effect of aspirin on platelet-von Willebrand factor surface expression on thrombin and ADP-stimulated platelets

Blood ◽  
1989 ◽  
Vol 74 (6) ◽  
pp. 2016-2021 ◽  
Author(s):  
RI Parker ◽  
HR Gralnick
Keyword(s):  
Von Willebrand Factor ◽  
Shape Change ◽  
Adenosine Diphosphate ◽  
Surface Expression ◽  
Platelet Surface ◽  
Granule Secretion ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Platelet Shape ◽  
Stimulation Of

Abstract Platelets contain a pool of endogenous platelet-von Willebrand factor (vWF) that becomes expressed on the platelet surface when platelets are stimulated by a variety of agonists. Maximal platelet-vWF expression occurs in concert with platelet alpha-granule secretion. Aspirin (ASA) is known to impair platelet activation and alpha-granule secretion by irreversible inhibition of platelet cyclo-oxygenase. We studied native and ASA-treated platelets for their ability to mobilize and to express platelet-vWF in response to adenosine diphosphate (ADP) or thrombin. We found that each agonist was effective in promoting increased platelet- vWF surface expression on native and ASA-treated platelets. ASA-treated platelets responded identically to native platelets to low (0.01 U/mL) and high (1.0 U/mL) concentrations of thrombin, while the ADP-induced increase in ASA-treated platelets was only 50% to 60% of that for control platelets. Measurement of secreted platelet-vWF and beta- thromboglobulin indicated that the increase seen with ADP was largely independent of alpha-granule secretion. Using monoclonal antibodies (MoAbs) against the platelet glycoproteins (GP) IIb/IIIa and Ib (MoAbs 10E5 and 6D1, respectively), we demonstrated that the ADP-induced increase in platelet-vWF expression on control platelets primarily involved the binding of secreted platelet-vWF to the platelet GPIIb/IIIa. In contrast, the increase in platelet-vWF that occurred following ADP stimulation of ASA-treated platelets was largely insensitive to GPIIb/IIIa blockade. No effect of GPIb blockade in platelet-vWf expression was noted for either control or ASA-treated platelets. When platelet shape change was prevented by the addition of cytochalasin D, ADP-induced platelet-vWf surface expression on ASA- treated platelets was reduced by more than 80%. Our data indicate that platelets in which the cyclooxygenase pathway is blocked by the action of aspirin can increase surface expression of platelet-vWf as a consequence of platelet shape change. We speculate that this process exposes platelet-vWf bound to GPIIb/IIIa, or possibly GPIb, within the surface connected canalicular system.

scholarly journals Unique interactions of asialo von Willebrand factor with platelets in platelet-type von Willebrand disease

Blood ◽  
1987 ◽  
Vol 70 (6) ◽  
pp. 1804-1809 ◽  
Author(s):  
JL Miller ◽  
ZM Ruggeri ◽  
VA Lyle
Keyword(s):  
Monoclonal Antibody ◽  
Von Willebrand Factor ◽  
Specific Binding ◽  
Platelet Rich Plasma ◽  
Von Willebrand Disease ◽  
Normal Platelet ◽  
High Affinity Binding Site ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Formalin Fixed

Abstract The present studies demonstrate that platelets from patients with platelet-type von Willebrand disease show specific and saturable binding of asialo von Willebrand factor (AS-vWF) under conditions where such binding is not observed with normal platelets. Although specific binding of 125I-AS-vWF to formalin-fixed normal platelets could not be demonstrated, specific binding to fixed patient platelets was seen with an apparent Kd of 1.3 micrograms/mL and specific maximally bound ligand of 0.40 micrograms/10(8) platelets. Preincubation of patient platelets with the antiglycoprotein Ib (anti-GPIb) monoclonal antibody AS-2 reduced total binding close to the level of computer-estimated nonspecific binding. In contrast, binding was not reduced by preincubation with anti-GPIIb/IIIa monoclonal antibody or with 5 mmol/L EDTA. Under stirring conditions, the binding of AS-vWF to fixed patient platelets was accompanied by a strong agglutination response. AS-vWF- induced agglutination was similarly observed in patient but not normal platelet-rich plasma (PRP) in the presence of 5 mmol/L EDTA. In the absence of EDTA, AS-vWF produced a full aggregation response in patient PRP at concentrations as low as 0.1 microgram/mL in contrast to the 2 to 20 micrograms/mL required by normal PRP. Both thromboxane B2 formation and adenosine triphosphate secretion showed an AS-vWF concentration dependence paralleling the aggregation responses. These studies show that a major difference in the platelets from patients with platelet-type von Willebrand disease is the presence of an exposed, high-affinity binding site associated with GPIb that recognizes AS-vWF.

scholarly journals Neutrophil cathepsin G modulates the platelet surface expression of the glycoprotein (GP) Ib-IX complex by proteolysis of the von Willebrand factor binding site on GPIb alpha and by a cytoskeletal-mediated redistribution of the remainder of the complex

Blood ◽  
1994 ◽  
Vol 84 (1) ◽  
pp. 158-168 ◽  
Author(s):  
CA LaRosa ◽  
MJ Rohrer ◽  
SE Benoit ◽  
MR Barnard ◽  
AD Michelson
Keyword(s):  
Binding Site ◽  
Von Willebrand Factor ◽  
Cytochalasin B ◽  
Surface Expression ◽  
Cathepsin G ◽  
Prostaglandin I2 ◽  
Platelet Surface ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Platelet Content

Abstract The effects of neutrophil cathepsin G on the glycoprotein (GP) Ib-IX complex of washed platelets were examined. Cathepsin G resulted in a concentration- and time-dependent decrease in the platelet surface GPIb- IX complex, as determined by flow cytometry, binding of exogenous von Willebrand factor (vWF) in the presence of ristocetin, and ristocetin- induced platelet agglutination. Cathepsin G resulted in proteolysis of the vWF binding site on GPIb alpha (defined by monoclonal antibody [MoAb] 6D1), as determined by increased supernatant glycocalicin fragment (a proteolytic product of GPIb alpha); decreased total platelet content of GPIb; and lack of effect of either cytochalasin B (an inhibitor of actin polymerization), prostaglandin I2 (an inhibitor of platelet activation), or prior fixation of the platelets. However, cathepsin G resulted in minimal decreases in the binding to fixed platelets of MoAbs TM60 (directed against the thrombin binding site on GPIb alpha) and WM23 (directed against the macroglycopeptide portion of GPIb alpha). In contrast to its proteolytic effect on GPIb alpha, the cathepsin G-induced decrease in platelet surface GPIX and the remnant of the GPIb-IX complex (defined by MoAbs FMC25 and AK1) was via a cytoskeletal-mediated redistribution, as determined by lack of change in the total platelet content of GPIX and the GPIb-IX complex; complete inhibition by cytochalasin B, prostaglandin I2, and prior fixation of platelets. Experiments with Serratia protease-treated and Bernard- Soulier platelets showed that neither platelet surface GPIb nor cathepsin G-induced proteolysis of GPIb were required for the cathepsin G-induced redistribution of the remnant of the GPIb-IX complex or the cathepsin G-induced increase in platelet surface P-selectin. In summary, neutrophil cathepsin G modulates the platelet surface expression of the GPIb-IX complex both by proteolysis of the vWF binding site on GPIb alpha and by a cytoskeletal-mediated redistribution of the remainder of the complex. Prior studies show that, although thrombospondin 1, antiserine proteases, and plasma are all inhibitors of cathepsin G, the effects of cathepsin G on platelets, including an increase in surface GPIIb-IIIa, occur during close contact between neutrophils and platelets in a protective microenvironment (eg, thrombosis and local inflammation).(ABSTRACT TRUNCATED AT 400 WORDS).

Complete Removal of N-Linked Glycans from Platelet GPIbα Impairs Platelet Agglutination and von Willebrand Factor Binding In Vitro.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1535-1535
Author(s):  
Suzana M. Zorca ◽  
Emma C. Josefsson ◽  
Viktoria Rumjantseva ◽  
John H. Hartwig ◽  
Karin M. Hoffmeister
Keyword(s):  
Flow Cytometry ◽  
Von Willebrand Factor ◽  
Cho Cells ◽  
Chinese Hamster ◽  
Surface Expression ◽  
Lectin Domain ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract We previously reported that the lectin domain of the αMβ2 receptor on macrophages mediates the rapid clearance of transfused washed murine platelets which have been refrigerated for 2 hrs in the absence of plasma. The clearance is mediated by the recognition of exposed βN-acetylglucosamine (β-GlcNAc) residues on N-linked glycans of clustered platelet GPIbα molecules. Covering the exposed β-GlcNAc residues on GPIbα N-linked glycans via galactosylation prevents the clearance of chilled murine platelets from the circulation. The role of N-linked glycans in platelet function and survival is unclear. To dissect the role of N-linked glycosylation of GPIbα on the binding of von Willebrand factor (vWf), we use human platelets and Chinese Hamster Ovary (CHO) cells, stably expressing human GPIbα/βand GPIX. Deglycosylation of platelet GPIbα N-liked glycans was achieved using the enzyme peptide-N-glycosidase F (PGNaseF), specific for complex N-linked glycans. In agglutination assays using platelets incubated with and without PNGaseF for 16hrs at 37°C, we observed 30-40 % less agglutination in response to ristocetin for platelets depleted of N-linked glycans with PNGaseF. Additionally, a 30 % reduction in vWf binding to PNGaseF-treated platelets compared with control platelets was measured by flow cytometry, using a FITC-conjugated mAb that detects surface-bound vWf. In CHO cells, GPIbα N-linked oligosaccharides were manipulated by adding swainsonine or tunicamycin, two inhibitors of N-linked oligosaccharide synthesis in the Golgi. vWf binding to platelets or to CHO cells was studied by aggregometry or by light microscopy to establish the fraction of CHO-cell aggregates. As was the case with platelets, vWf-dependent aggregation of CHO cells expressing GPIb-IX decreased three fold in response to botrocetin, but only following complete N-linked glycans depletion with tunicamycin. In contrast, partial N-linked carbohydrate modification with swainsonine did not significantly alter aggregate formation in CHO- cells expressing GPIb-IX. Complete inhibition of N-linked glycosylation decreased botrocetin-induced vWf binding to CHO- cells expressing GPIb-IX by ~50%, as determined by flow cytometry. No change was observed following swainsonine treatment. Surface expression of GP1bα remained unchanged after both tunicamycin and swainsonine treatment, and with PGNaseF treatment of platelets. These results confirm that 1) N-linked glycans are not required for GPIbα surface expression, and 2) indicate that N-linked glycans likely play a role in vWf binding to platelet GPIbα.

scholarly journals A new congenital platelet abnormality characterized by spontaneous platelet aggregation, enhanced von Willebrand factor platelet interaction, and the presence of all von Willebrand factor multimers in plasma

Blood ◽  
1989 ◽  
Vol 74 (6) ◽  
pp. 2028-2033
Author(s):  
A Casonato ◽  
L De Marco ◽  
M Mazzucato ◽  
V De Angelis ◽  
D De Roia ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Platelet Aggregation ◽  
Von Willebrand Factor ◽  
Platelet Rich Plasma ◽  
Von Willebrand Disease ◽  
Bleeding Tendency ◽  
Binding Studies ◽  
Spontaneous Platelet Aggregation ◽  
Von Willebrand ◽  
Willebrand Factor

A case is reported of a 49-year-old woman with a mild bleeding tendency. Her bleeding time, platelet count and size, plasma ristocetin cofactor activity, von Willebrand factor (vWF) antigen, and vWF multimeric pattern are all within normal limits. Spontaneous platelet aggregation is observed when citrated platelet-rich plasma (PRP) is stirred in an aggregometer cuvette. This aggregation is completely is only slightly diminished by an antiglycoprotein (GP) IIb/IIIa or by an anti GPIb monoclonal antibody. The patient's PRP shows increased sensitivity to ristocetin. The distinct feature of this patient, also present in two family members studied, is that platelet aggregation is initiated by purified vWF in the absence of any other agonist. The vWF- induced platelet aggregation is abolished by anti-GPIb and anti- GPIIb/IIIa monoclonal antibodies and by EDTA (5 mmol/L). Apyrase inhibits the second wave of aggregation. Patient's platelets in PRP are four to six times more reactive to asialo vWF-induced platelet aggregation than normal platelets. The amount of radiolabeled vWF bound to platelets in the presence of either low concentration of ristocetin or asialo vWF was increased 30% compared with normal. The patient's platelet GPIb was analyzed by SDS page and immunoblotting and by binding studies with anti-GPIb monoclonal antibodies showed one band with slightly increased migration pattern and a normal number of GPIb molecules. Unlike the previously reported patients with pseudo or platelet-type von Willebrand disease, this patient has normal vWF parameters.

scholarly journals von Willebrand factor is present on the surface of platelets stimulated in plasma by ADP

Blood ◽  
1987 ◽  
Vol 70 (5) ◽  
pp. 1362-1366
Author(s):  
B Adelman ◽  
P Carlson ◽  
P Powers
Keyword(s):  
Binding Site ◽  
Von Willebrand Factor ◽  
Platelet Rich Plasma ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Surface Binding ◽  
Platelet Surface ◽  
Von Willebrand ◽  
Willebrand Factor

von Willebrand factor (vWf) can bind to glycoprotein (GP) IIb/IIIa on activated platelets. The significance of this interaction is unclear, however, because it has not been possible to detect vWf binding to GPIIb/IIIa on platelets stimulated in plasma. We have developed an indirect, flow cytometry assay that uses fluorescein-labeled antibodies to detect vWf and fibrinogen on platelets. Using this assay, we found vWf on the surface of platelets stimulated in plasma by ADP. The number of platelets that bound vWf increased in proportion to ADP concentration and incubation time. Washed platelets in a protein-free buffer activated by 1 mumol/L calcium ionophore A23187 or 10 mumol/L ADP also bound vWf, suggesting that we were detecting surface binding of alpha-granule-derived vWf. Monoclonal antibodies against the vWf binding site on GPIb (6D1) and the vWf and fibrinogen binding sites on GPIIb/IIIa (LJP5 and LJ-CP8, respectively) were used to characterize the mechanism of vWf binding to stimulated platelets. Ristocetin- induced binding of vWf was inhibited by 6D1, and ADP-induced binding of fibrinogen was inhibited by LJ-CP8. None of these antibodies inhibited ADP-induced vWf binding. Aspirin and prostaglandin E1 also inhibited ADP-induced binding of vWf in platelet-rich plasma. During platelet activation in plasma, vWf derived from alpha-granules becomes bound to the platelet surface possibly being transferred already associated with a binding site.

scholarly journals Comparative analysis of type 2b von Willebrand disease mutations: implications for the mechanism of von Willebrand factor binding to platelets

Blood ◽  
1996 ◽  
Vol 87 (6) ◽  
pp. 2322-2328 ◽  
Author(s):  
KA Cooney ◽  
D Ginsburg
Keyword(s):  
Von Willebrand Factor ◽  
Binding Domain ◽  
Von Willebrand Disease ◽  
Surface Glycoprotein ◽  
Platelet Surface ◽  
Disease Mutations ◽  
A Domains ◽  
Adhesive Proteins ◽  
Von Willebrand ◽  
Willebrand Factor

von Willebrand factor (vWF) is a multimeric glycoprotein that forms an adhesive link following vascular injury between the vessel wall and its primary ligand on the platelet surface, glycoprotein Ib (GpIb). Type 2b von Willebrand disease (vWD) is a qualitative form of vWD resulting from enhanced binding of vWF to platelets. Molecular characterization of the vWF gene in patients with type 2b vWD has resulted in identification of a panel of mutations associated with this disorder, all clustered within the GpIb binding domain in exon 28 of the vWF gene. We have expressed six of the most common type 2b vWD mutations in recombinant vWF and show that each mutation produces a similar increase in vWF binding to platelets in the absence or presence of ristocetin. Furthermore, expression of more than one type 2b vWD mutation in the same molecule (cis) or in different molecules within the same multimer (trans) failed to produce an increase in vWF platelet binding compared with any of the individually expressed mutations. Taken together, these data support the hypothesis that the vWF GpIb binding domain can adopt either a discrete “on” or “off” conformation, with most type 2b vWD mutations resulting in vWF locked in the on conformation. This model may have relevance to other adhesive proteins containing type A domains.

Sign in / Sign up

Export Citation Format

Share Document