scholarly journals Phospholipase abolishes the effect of stimulated platelets on the thrombin activation of factor VIII

Blood ◽  
1988 ◽  
Vol 71 (1) ◽  
pp. 173-177 ◽  
Author(s):  
ME Rick ◽  
DM Krizek
Keyword(s):  
Factor Viii ◽  
Von Willebrand Factor ◽  
Calcium Ionophore ◽  
Platelet Membrane ◽  
Activate Factor ◽  
Membrane Glycoproteins ◽  
Membrane Phospholipids ◽  
Thrombin Activation ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract Factor VIII functions as a cofactor in the intrinsic coagulation pathway and must first be activated to function optimally in this capacity. Low concentrations of thrombin activate factor VIII, and the presence of stimulated platelets is known to enhance the activation of factor VIII complexed to von Willebrand factor. The current studies show that platelets stimulated by thrombin, collagen, or calcium ionophore will increase the activation of isolated factor VIII by thrombin. Ongoing platelet release is not necessary for the enhanced factor VIII activation, nor is platelet von Willebrand factor or platelet membrane glycoproteins Ib or IIb/IIIa. Platelet membrane phospholipids, on the other hand, are important for the enhanced activation of factor VIII by thrombin because the effect of stimulated platelets is abolished by incubation of the stimulated platelets with phospholipases. These results suggest that the enhanced activation of factor VIII by thrombin in the presence of stimulated platelets may be mediated by factor VIII binding to platelet phospholipid or to a receptor whose functional integrity is dependent on surrounding membrane phospholipid.

scholarly journals Phospholipase abolishes the effect of stimulated platelets on the thrombin activation of factor VIII

Blood ◽  
1988 ◽  
Vol 71 (1) ◽  
pp. 173-177
Author(s):  
ME Rick ◽  
DM Krizek
Keyword(s):  
Factor Viii ◽  
Von Willebrand Factor ◽  
Calcium Ionophore ◽  
Platelet Membrane ◽  
Activate Factor ◽  
Membrane Glycoproteins ◽  
Membrane Phospholipids ◽  
Thrombin Activation ◽  
Von Willebrand ◽  
Willebrand Factor

Factor VIII functions as a cofactor in the intrinsic coagulation pathway and must first be activated to function optimally in this capacity. Low concentrations of thrombin activate factor VIII, and the presence of stimulated platelets is known to enhance the activation of factor VIII complexed to von Willebrand factor. The current studies show that platelets stimulated by thrombin, collagen, or calcium ionophore will increase the activation of isolated factor VIII by thrombin. Ongoing platelet release is not necessary for the enhanced factor VIII activation, nor is platelet von Willebrand factor or platelet membrane glycoproteins Ib or IIb/IIIa. Platelet membrane phospholipids, on the other hand, are important for the enhanced activation of factor VIII by thrombin because the effect of stimulated platelets is abolished by incubation of the stimulated platelets with phospholipases. These results suggest that the enhanced activation of factor VIII by thrombin in the presence of stimulated platelets may be mediated by factor VIII binding to platelet phospholipid or to a receptor whose functional integrity is dependent on surrounding membrane phospholipid.

Role of Platelet Membrane Glycoproteins and Von Willebrand Factor in Adhesion of Platelets to Subendothelium and Collagen

1987 ◽  
Vol 516 (1 Blood in Cont) ◽  
pp. 52-65 ◽  
Author(s):  
KJELL S. SAKARIASSEN ◽  
EDITH FRESSINAUD ◽  
JEAN-PIERRE GIRMA ◽  
DOMINIQUE MEYER ◽  
HANS R. BAUMGARTNER
Keyword(s):  
Von Willebrand Factor ◽  
Platelet Membrane ◽  
Membrane Glycoproteins ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Adhesion Of Platelets

scholarly journals Platelet-collagen interaction: inhibition by ristocetin and enhancement by von Willebrand factor-platelet binding

Blood ◽  
1986 ◽  
Vol 68 (4) ◽  
pp. 927-937
Author(s):  
FM LaDuca ◽  
RE Bettigole ◽  
WR Bell ◽  
EB Robson
Keyword(s):  
Platelet Aggregation ◽  
Von Willebrand Factor ◽  
Platelet Rich Plasma ◽  
Platelet Membrane ◽  
Type I ◽  
Membrane Glycoproteins ◽  
Platelet Binding ◽  
Von Willebrand ◽  
Willebrand Factor

The contribution of von Willebrand factor (vWF)-platelet binding to platelet-collagen interaction was examined in vitro. The binding of vWF to platelets was mediated and regulated by ristocetin. Subthreshold concentrations of ristocetin (less than or equal to 1 mg/mL), insufficient to cause ristocetin-induced platelet aggregation (RIPA), were added to platelet-rich plasma (PRP) prior to the addition of collagen. The collagen-induced platelet aggregation (CIPA) was modified by ristocetin and the degree of alteration was dependent on the ristocetin concentration. Response as a function of ristocetin concentration was designated the Collagen-Platelet Aggregation Response (CoI-PAR). In normal PRP the CoI-PAR was a progressive inhibition followed by decreasing inhibition and then an enhanced response. The enhanced response occurred over a narrow range of ristocetin concentrations (0.8 to 1.0 mg/mL). In the absence of vWF (severe von Willebrand's disease, Type I, vWF less than 1%) the CoI-PAR was a progressive, eventually complete inhibition with no enhanced response (with ristocetin concentrations up to 3.0 mg/mL). With addition of vWF to this PRP an enhanced response was observed at a ristocetin concentration inversely proportional to the vWF level. PRP from a patient with severe Hemophilia A showed a response within the normal range. Subthreshold ristocetin did not cause plasma protein precipitation or platelet release of 3H-serotonin, nor induce micro platelet aggregate formation. Digestion of platelet membrane glycoproteins (GP(s] with chymotrypsin demonstrated that upon removal of GPI, RIPA was absent, CIPA retained and the CoI-PAR was progressive inhibition, with no enhancement. With removal of GPs I, II, and III, RIPA, CIPA, and the CoI-PAR were absent. A dose-response 125I-vWF- platelet binding occurred with increasing ristocetin concentrations which was unchanged by the addition of collagen. These results demonstrated that ristocetin-platelet association inhibited CIPA, and vWF-platelet binding enhanced platelet-collagen adhesion and platelet aggregation. The in vitro-enhanced CIPA represents a vWF-dependent aggregation of sufficient magnitude to overcome the inhibitory effect of ristocetin. These studies demonstrate an influential interaction of ristocetin, vWF, and collagen with the platelet membrane and imply an important hemostatic contribution of vWF-platelet binding in platelet- collagen interaction.

scholarly journals Platelet-collagen interaction: inhibition by ristocetin and enhancement by von Willebrand factor-platelet binding

Blood ◽  
1986 ◽  
Vol 68 (4) ◽  
pp. 927-937 ◽  
Author(s):  
FM LaDuca ◽  
RE Bettigole ◽  
WR Bell ◽  
EB Robson
Keyword(s):  
Platelet Aggregation ◽  
Von Willebrand Factor ◽  
Platelet Rich Plasma ◽  
Platelet Membrane ◽  
Type I ◽  
Membrane Glycoproteins ◽  
Platelet Binding ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract The contribution of von Willebrand factor (vWF)-platelet binding to platelet-collagen interaction was examined in vitro. The binding of vWF to platelets was mediated and regulated by ristocetin. Subthreshold concentrations of ristocetin (less than or equal to 1 mg/mL), insufficient to cause ristocetin-induced platelet aggregation (RIPA), were added to platelet-rich plasma (PRP) prior to the addition of collagen. The collagen-induced platelet aggregation (CIPA) was modified by ristocetin and the degree of alteration was dependent on the ristocetin concentration. Response as a function of ristocetin concentration was designated the Collagen-Platelet Aggregation Response (CoI-PAR). In normal PRP the CoI-PAR was a progressive inhibition followed by decreasing inhibition and then an enhanced response. The enhanced response occurred over a narrow range of ristocetin concentrations (0.8 to 1.0 mg/mL). In the absence of vWF (severe von Willebrand's disease, Type I, vWF less than 1%) the CoI-PAR was a progressive, eventually complete inhibition with no enhanced response (with ristocetin concentrations up to 3.0 mg/mL). With addition of vWF to this PRP an enhanced response was observed at a ristocetin concentration inversely proportional to the vWF level. PRP from a patient with severe Hemophilia A showed a response within the normal range. Subthreshold ristocetin did not cause plasma protein precipitation or platelet release of 3H-serotonin, nor induce micro platelet aggregate formation. Digestion of platelet membrane glycoproteins (GP(s] with chymotrypsin demonstrated that upon removal of GPI, RIPA was absent, CIPA retained and the CoI-PAR was progressive inhibition, with no enhancement. With removal of GPs I, II, and III, RIPA, CIPA, and the CoI-PAR were absent. A dose-response 125I-vWF- platelet binding occurred with increasing ristocetin concentrations which was unchanged by the addition of collagen. These results demonstrated that ristocetin-platelet association inhibited CIPA, and vWF-platelet binding enhanced platelet-collagen adhesion and platelet aggregation. The in vitro-enhanced CIPA represents a vWF-dependent aggregation of sufficient magnitude to overcome the inhibitory effect of ristocetin. These studies demonstrate an influential interaction of ristocetin, vWF, and collagen with the platelet membrane and imply an important hemostatic contribution of vWF-platelet binding in platelet- collagen interaction.

scholarly journals Effect of Thrombin on Factor VIII/VON Willebrand Factor (FVIII/vWF)

1977 ◽  
Author(s):  
Mary Ellen Switzer ◽  
P.A. McKee
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulfate ◽  
Factor Viii ◽  
Von Willebrand Factor ◽  
Protein Concentration ◽  
Sodium Dodecyl ◽  
Activity Peak ◽  
Thrombin Activation ◽  
Von Willebrand ◽  
Willebrand Factor

The sodium dodecyl sulfate (SDS) gels and immunological properties of FVIII/vWF and thrombin-activated (thr-act) FVIII/vWF are identical. Hence thrombin must either modify FVIII/vWF by very minor proteolysis or cleave only a few FVIII/vWF molecules. The procoagulant (PC) activity of FVIII/vWF and thr-act FVIII/vWF eluted sharply in the void volume (Vo) from 4% agarose in 0.15 M NaCl, with >65% loss of the PC activity of the thr-act FVIII/vWF by 3 hrs. The PC activity of FVIII/vWF and thr-act FVIII/vWF was stabilized by 0.25 M CaCl2. When FVIII/vWF and thr-act FVIII/vWF were filtered on 4% agarose in 0.25 M CaCl2, the protein eluted in the Vo, but most of the PC activity eluted later in a region of little detectable protein; however, the delayed PC peak from thr-act FVIII/vWF was greatly enhanced above control levels. Nonreduced, neither activity peak protein entered a 7.5% SDS gel. After reduction, the gel pattern for the PC activity peak protein from FVIII/vWF showed major bands of 195x, 79x, 61x, 51x and 18x (103) molecular weight (MW) and several minor bands >100,000 MW. The reduced PC protein from thr-act FVIII/vWF lacked all bands >100,000 MW, but the four lower MW bands were present and well-resolved. Thrombin activation did not affect the vWF activity which was proportional to protein concentration throughout any chromatogram; the 195,000-dalton subunit was not necessary for vWF activity. FVIII/vWF, which had the peak PC activity in the Vo and no other PC activity peak on 4% agarose-0.25 M CaCl2, was thr-act before filtration under the same conditions. Then, the greatly enhanced PC activity eluted well after the Vo, with <10% PC activity in the Vo. We conclude that modification of a few FVIII/vWF molecules by thrombin and their stabilization by 0.25 M CaCl2 causes the PC peak that elutes aberrantly from 4% agarose-0.25 M CaCl2.

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