scholarly journals Human platelet surface binding of endogenous secreted factor VIII-von Willebrand factor and platelet factor 4

Blood ◽  
1982 ◽  
Vol 59 (1) ◽  
pp. 194-197 ◽  
Author(s):  
JN George ◽  
AR Onofre
Keyword(s):  
Factor Viii ◽  
Von Willebrand Factor ◽  
Membrane Surface ◽  
Platelet Factor 4 ◽  
Secreted Proteins ◽  
Surface Binding ◽  
Platelet Factor ◽  
Platelet Surface ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract Washed human platelets in buffers containing either 2 mM Ca++ or 4 mM EDTA were stimulated by human alpha-thrombin to induce secretion. The binding of two endogenous secreted proteins, factor-VIII-related protein (VIII-R) (von Willebrand factor) and platelet factor 4, was measured by reacting thrombin-treated and control platelets with specific antibodies to these proteins, then quantifying antibody binding with 125I-staphylococcal protein A. Both of these granule proteins were associated with the platelet membrane surface by a calcium-dependent mechanism after thrombin-induced secretion. This ability to bind endogenous secreted proteins to the plasma membrane surface may provide a mechanism by which the platelet can concentrate and organize its secreted proteins for subsequent physiologic reactions.

scholarly journals Human platelet surface binding of endogenous secreted factor VIII-von Willebrand factor and platelet factor 4

Blood ◽  
1982 ◽  
Vol 59 (1) ◽  
pp. 194-197 ◽  
Author(s):  
JN George ◽  
AR Onofre
Keyword(s):  
Factor Viii ◽  
Von Willebrand Factor ◽  
Membrane Surface ◽  
Platelet Factor 4 ◽  
Secreted Proteins ◽  
Surface Binding ◽  
Platelet Factor ◽  
Platelet Surface ◽  
Von Willebrand ◽  
Willebrand Factor

Washed human platelets in buffers containing either 2 mM Ca++ or 4 mM EDTA were stimulated by human alpha-thrombin to induce secretion. The binding of two endogenous secreted proteins, factor-VIII-related protein (VIII-R) (von Willebrand factor) and platelet factor 4, was measured by reacting thrombin-treated and control platelets with specific antibodies to these proteins, then quantifying antibody binding with 125I-staphylococcal protein A. Both of these granule proteins were associated with the platelet membrane surface by a calcium-dependent mechanism after thrombin-induced secretion. This ability to bind endogenous secreted proteins to the plasma membrane surface may provide a mechanism by which the platelet can concentrate and organize its secreted proteins for subsequent physiologic reactions.

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 Identification of platelet glycoprotein IIb/IIIa as the major binding site for released platelet-von Willebrand factor [published erratum appears in Blood 1987 Jun;69(6):1789]

Blood ◽  
1986 ◽  
Vol 68 (3) ◽  
pp. 732-736 ◽  
Author(s):  
RI Parker ◽  
HR Gralnick
Keyword(s):  
Monoclonal Antibodies ◽  
Binding Site ◽  
Von Willebrand Factor ◽  
Plasma Fibrinogen ◽  
Platelet Glycoprotein ◽  
Surface Binding ◽  
Human Fibrinogen ◽  
Platelet Surface ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract We studied the effects(s) of two monoclonal antibodies, 6D1 and 10E5 (directed against platelet glycoprotein Ib [GPIb] and the GPIIb/IIIa complex, respectively), and purified human plasma fibrinogen on the binding of released platelet-von Willebrand factor (vWf) to the platelet surface. Neither of the monoclonal antibodies nor fibrinogen had any effect on the amount of platelet-vWf expressed on unstimulated platelets or on the amount expressed on platelets stimulated in the absence of extracellular Ca++. However, the antibody directed against GPIIb/IIIa inhibited 72% of the thrombin-induced increase in the platelet-vWf bound to the platelet surface when platelets were stimulated in the presence of 5 mmol/L Ca++. The antibody against GPIb did not inhibit the surface expression of platelet-vWf on stimulated platelets in the presence of Ca++. Purified normal human fibrinogen inhibited the surface binding of platelet-vWf to thrombin-stimulated platelets to a degree similar to that observed with the monoclonal antibody directed against the GPIIb/IIIa complex. These data indicate that platelet-vWf released from platelets binds primarily to the GPIIb/IIIa complex at or near the plasma fibrinogen binding site.

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

Abstract 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 Antigen and substrate withdrawal in the management of autoimmune thrombotic disorders

Blood ◽  
2012 ◽  
Vol 120 (20) ◽  
pp. 4134-4142 ◽  
Author(s):  
Douglas B. Cines ◽  
Keith R. McCrae ◽  
X. Long Zheng ◽  
Bruce S. Sachais ◽  
Eline T. Luning Prak ◽  
...  
Keyword(s):  
Thrombotic Thrombocytopenic Purpura ◽  
Antiphospholipid Syndrome ◽  
Von Willebrand Factor ◽  
Platelet Factor 4 ◽  
Platelet Factor ◽  
Thrombocytopenic Purpura ◽  
Heparin Induced Thrombocytopenia ◽  
Systemic Anticoagulation ◽  
Von Willebrand ◽  
Willebrand Factor

AbstractPrevailing approaches to manage autoimmune thrombotic disorders, such as heparin-induced thrombocytopenia, antiphospholipid syndrome and thrombotic thrombocytopenic purpura, include immunosuppression and systemic anticoagulation, though neither provides optimal outcome for many patients. A different approach is suggested by the concurrence of autoantibodies and their antigenic targets in the absence of clinical disease, such as platelet factor 4 in heparin-induced thrombocytopenia and β2-glycoprotein-I (β2GPI) in antiphospholipid syndrome. The presence of autoantibodies in the absence of disease suggests that conformational changes or other alterations in endogenous protein autoantigens are required for recognition by pathogenic autoantibodies. In thrombotic thrombocytopenic purpura, the clinical impact of ADAMTS13 deficiency caused by autoantibodies likely depends on the balance between residual antigen, that is, enzyme activity, and demand imposed by local genesis of ultralarge multimers of von Willebrand factor. A corollary of these concepts is that disrupting platelet factor 4 and β2GPI conformation (or ultralarge multimer of von Willebrand factor oligomerization or function) might provide a disease-targeted approach to prevent thrombosis without systemic anticoagulation or immunosuppression. Validation of this approach requires a deeper understanding of how seemingly normal host proteins become antigenic or undergo changes that increase antibody avidity, and how they can be altered to retain adaptive functions while shedding epitopes prone to elicit harmful autoimmunity.

scholarly journals Multimeric structure of platelet factor VIII/von Willebrand factor: the presence of larger multimers and their reassociation with thrombin- stimulated platelets

Blood ◽  
1982 ◽  
Vol 60 (5) ◽  
pp. 1132-1138 ◽  
Author(s):  
MF Fernandez ◽  
MH Ginsberg ◽  
ZM Ruggeri ◽  
FJ Batlle ◽  
TS Zimmerman
Keyword(s):  
Factor Viii ◽  
Von Willebrand Factor ◽  
Divalent Cations ◽  
Polyacrylamide Gel Electrophoresis ◽  
Structural Difference ◽  
Platelet Factor ◽  
Cell Extracts ◽  
Platelet Enzymes ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract The multimeric structure of platelet factor VIII/von Willebrand factor (FVIII/vWF) in cell extracts and in collagen and thrombin releasates has been analyzed by SDS polyacrylamide gel electrophoresis followed by detection with 125I-anti-FVIII/vWF. Platelets contained larger multimers than those normally present in plasma. When secreted FVIII/vWF was analyzed, all platelets. In contrast, in thrombin releasates the larger multimers were lost in a manner dependent on divalent cations, time, and thrombin dose. This loss could not be accounted for by modification of FVIII/vWF by thrombin or platelet enzymes since no effect of thrombin on the multimeric structure of FVIII/vWF in the absence of platelets or in the presence of platelet lysates was observed. Large multimers of 125I-labeled purified FVIII/vWF underwent divalent cation-dependent association with platelets in the presence of thrombin, indicating that the loss of FVIII/vWF from thrombin releasates was due to reassociation with the platelet. These studies show a structural difference between platelet and plasma FVIII/vWF that suggests a specific role for platelet FVIII/vWF in hemostasis.

scholarly journals Multimeric structure of platelet factor VIII/von Willebrand factor: the presence of larger multimers and their reassociation with thrombin- stimulated platelets

Blood ◽  
1982 ◽  
Vol 60 (5) ◽  
pp. 1132-1138 ◽  
Author(s):  
MF Fernandez ◽  
MH Ginsberg ◽  
ZM Ruggeri ◽  
FJ Batlle ◽  
TS Zimmerman
Keyword(s):  
Factor Viii ◽  
Von Willebrand Factor ◽  
Divalent Cations ◽  
Polyacrylamide Gel Electrophoresis ◽  
Structural Difference ◽  
Platelet Factor ◽  
Cell Extracts ◽  
Platelet Enzymes ◽  
Von Willebrand ◽  
Willebrand Factor

The multimeric structure of platelet factor VIII/von Willebrand factor (FVIII/vWF) in cell extracts and in collagen and thrombin releasates has been analyzed by SDS polyacrylamide gel electrophoresis followed by detection with 125I-anti-FVIII/vWF. Platelets contained larger multimers than those normally present in plasma. When secreted FVIII/vWF was analyzed, all platelets. In contrast, in thrombin releasates the larger multimers were lost in a manner dependent on divalent cations, time, and thrombin dose. This loss could not be accounted for by modification of FVIII/vWF by thrombin or platelet enzymes since no effect of thrombin on the multimeric structure of FVIII/vWF in the absence of platelets or in the presence of platelet lysates was observed. Large multimers of 125I-labeled purified FVIII/vWF underwent divalent cation-dependent association with platelets in the presence of thrombin, indicating that the loss of FVIII/vWF from thrombin releasates was due to reassociation with the platelet. These studies show a structural difference between platelet and plasma FVIII/vWF that suggests a specific role for platelet FVIII/vWF in hemostasis.

scholarly journals Identification of platelet glycoprotein IIb/IIIa as the major binding site for released platelet-von Willebrand factor [published erratum appears in Blood 1987 Jun;69(6):1789]

Blood ◽  
1986 ◽  
Vol 68 (3) ◽  
pp. 732-736
Author(s):  
RI Parker ◽  
HR Gralnick
Keyword(s):  
Monoclonal Antibodies ◽  
Binding Site ◽  
Von Willebrand Factor ◽  
Plasma Fibrinogen ◽  
Platelet Glycoprotein ◽  
Surface Binding ◽  
Human Fibrinogen ◽  
Platelet Surface ◽  
Von Willebrand ◽  
Willebrand Factor

We studied the effects(s) of two monoclonal antibodies, 6D1 and 10E5 (directed against platelet glycoprotein Ib [GPIb] and the GPIIb/IIIa complex, respectively), and purified human plasma fibrinogen on the binding of released platelet-von Willebrand factor (vWf) to the platelet surface. Neither of the monoclonal antibodies nor fibrinogen had any effect on the amount of platelet-vWf expressed on unstimulated platelets or on the amount expressed on platelets stimulated in the absence of extracellular Ca++. However, the antibody directed against GPIIb/IIIa inhibited 72% of the thrombin-induced increase in the platelet-vWf bound to the platelet surface when platelets were stimulated in the presence of 5 mmol/L Ca++. The antibody against GPIb did not inhibit the surface expression of platelet-vWf on stimulated platelets in the presence of Ca++. Purified normal human fibrinogen inhibited the surface binding of platelet-vWf to thrombin-stimulated platelets to a degree similar to that observed with the monoclonal antibody directed against the GPIIb/IIIa complex. These data indicate that platelet-vWf released from platelets binds primarily to the GPIIb/IIIa complex at or near the plasma fibrinogen binding site.

scholarly journals Subcellular platelet factor VIII antigen and von Willebrand factor.

1975 ◽  
Vol 141 (5) ◽  
pp. 1101-1113 ◽  
Author(s):  
R L Nachman ◽  
E A Jaffe
Keyword(s):  
Factor Viii ◽  
Von Willebrand Factor ◽  
Human Platelets ◽  
Factor Vii ◽  
Platelet Factor ◽  
Plasma Factor ◽  
Factor Viii Antigen ◽  
Von Willebrand ◽  
Willebrand Factor

Subcellular membrane and granule fractions derived from human platelets contain factor VIIII antigen and von Willebrand factor activity but not factor VII procoagulant activity. Circulating platelets constitute a significant reservoir of plasma factor VIII antigen, containing approximately 15% of the amount of factor VIII antigen present in platelet-poor plasma. The antibiotic ristocetin, which aggregates human platelets in the presence of von Willebrand factor, nonspecifically precipitates platelet membrane factor VIII antigen. Thus normal platelets contain surface-bound as well as internally stored von Willebrand factor, a protein synthesized by endothelial cells which is necessary for normal platelet function in vivo.

Sign in / Sign up

Export Citation Format

Share Document