Presence Of Receptor For Activated Factor VIII On The Surface Of Released Platelets

1981 ◽  
Author(s):  
K Brodén ◽  
L-O Andersson
Keyword(s):  
Factor Viii ◽  
Von Willebrand Factor ◽  
Factor X ◽  
Factor Viii Activity ◽  
Platelet Receptor ◽  
Plasma Factor ◽  
Centrifuge Tube ◽  
Dilute Suspension ◽  
Von Willebrand ◽  
Willebrand Factor

In normal plasma Factor VIII activity is associated with a series of high molecular weight glycoprotein complexes also containing von Willebrand Factor related activities. To study the possible binding of various forms of Factor VIII to released platelets, a solution containing Factor VIII was mixed with a dilute suspension of platelets, which were released by addition of collagen. After 10 minutes of incubation the mixture was layered over 1.5 ml of 30% human serum albumin solution in a centrifuge tube and subjected to centrifugation at 7,000xg. Fractions were collected and analyzed for Factor VIII activity and phospholipid-related procoagulant activity. When purified Factor VUI/von Willebrand Factor complex was studied no significant association between the Factor VIII activity and the platelets were found. When purified Factor VUI/von Willebrand Factor complex was activated with 10-3 units/ml of thrombin and then tested, the main part of the Factor VIII activity became associated with the platelets. Even at very low platelet counts this binding was clearly detectable. The binding occurred both in the presence and in the absence of Ca2+. Thus released platelets bind thrombin-activated Factor VIII but not the Factor VUI/von Willebrand Factor complex. It is known that activation of Factor VIII by thrombin causes dissociation of the Factor VIII from the von Willebrand Factor part of the complex. The data obtained indicate that this dissociation is necessary in order to get the Factor VIII to bind to the platelet receptor. It may work as an amplification mechanism where the first traces of Thrombin formed upon initiation of coagulation dissociates Factor VIII from von Willebrand Factor, followed by binding to receptor on released platelets and formation of Factor X activator complex on the surface of the platelets.

scholarly journals Interaction of factor VIII-von Willebrand Factor with phospholipid vesicles

1981 ◽  
Vol 200 (1) ◽  
pp. 161-167 ◽  
Author(s):  
L O Andersson ◽  
J E Brown
Keyword(s):  
Factor Viii ◽  
Density Gradient ◽  
Von Willebrand Factor ◽  
Proteinase Inhibitors ◽  
Phospholipid Vesicles ◽  
Factor X ◽  
Factor Viii Activity ◽  
Von Willebrand ◽  
Willebrand Factor

The interaction of Factor VIII-von Willebrand Factor with phospholipid vesicles has been studied by using sucrose-density-gradient ultracentrifugation. When purified Factor VIII-von Willebrand Factor was run alone. Factor VIII activity and Factor VIIIR-Ag sedimented together to the lower half of the tube. Addition of phosphatidylserine/phosphatidylethanolamine vesicles at concentrations above 250 microgram/ml resulted in complete separation of Factor VIII activity and Factor VIIIR-Ag, the former appearing with the phospholipid on the top of the tube and the latter sedimenting as before. This separation was obtained even in the presence of proteinase inhibitors. Activation of Factor VIII-von Willebrand Factor by thrombin resulted in formation of a slow sedimenting component containing essentially all the Factor VIII activity, whereas the Factor VIIIR-Ag sedimented towards the bottom of the tube as before. The thrombin-induced Factor VIII activity was strongly bound to phospholipid vesicles as determined by density-gradient centrifugations at various Factor VIII concentrations and low concentrations of phospholipid. Based on certain assumptions a dissociation constant of 2.5 nM was calculated, a mechanism for the formation in vivo of the Factor X-activator complex is suggested.

scholarly journals Factor VIII/von Willebrand factor in subendothelium mediates platelet adhesion

Blood ◽  
1985 ◽  
Vol 65 (4) ◽  
pp. 823-831 ◽  
Author(s):  
VT Turitto ◽  
HJ Weiss ◽  
TS Zimmerman ◽  
II Sussman
Keyword(s):  
Factor Viii ◽  
Von Willebrand Factor ◽  
Vessel Wall ◽  
Platelet Adhesion ◽  
Human Factor ◽  
Rabbit Aorta ◽  
Human Factor Viii ◽  
Plasma Factor ◽  
Von Willebrand ◽  
Willebrand Factor

The present studies were undertaken to determine whether factor VIII/von Willebrand factor (vWF) present in the vessel wall (in addition to that in plasma) may mediate the attachment of platelets to subendothelium. Subendothelium from everted rabbit aorta was exposed to human citrated blood flowing through an annular perfusion chamber at 40 mL/min (wall shear rate of 2,600 s-1 for five minutes). The vessel segments were incubated at 37 degrees C for one hour with various dilutions of either goat-anti-rabbit factor VIII/vWF serum or an IgG fraction prepared from the serum. Control segments were incubated with serum or IgG from a nonimmunized goat. Values of platelet contact (C), platelet adhesion (C + S), and thrombus formation (T) on the subendothelium were evaluated by a morphometric technique. Compared with vessels incubated with fractions prepared from a normal goat, a significant decrease in platelet adhesion (C + S), ranging from 45% to 65%, was observed on vessels incubated with various dilutions (1:5 to 1:50) of either serum or IgG fractions of goat-anti-rabbit factor VIII/vWF. A similar decrease in platelet adhesion was observed with vessels incubated with an F(ab')2 fragment against rabbit factor VIII/vWF prepared in the goat. When goat-anti-rabbit factor VIII/vWF IgG was added to rabbit blood (1:75 dilution), platelet adhesion was reduced to the same extent (65%) on normal rabbit vessels and on vessels pre-incubated with goat-anti-rabbit factor VIII/vWF. Immunofluorescence studies revealed the presence of rabbit factor VIII/vWF in the subendothelium of rabbit aorta and the continued binding of the goat-anti-factor VIII/vWF antibodies on subendothelium during the perfusion studies. No uptake of human factor VIII/vWF on the rabbit subendothelium was observed by this immunologic technique; human factor VIII/vWF was found to be entirely associated with the attached human platelets. Thus, factor VIII/vWF in the vessel wall may mediate platelet attachment to subendothelium in a manner similar to that of plasma factor VIII/vWF.

scholarly journals Characterization of the defect of the factor VIII/von Willebrand factor protein in von Willebrand's disease

Blood ◽  
1982 ◽  
Vol 59 (3) ◽  
pp. 542-548 ◽  
Author(s):  
HR Gralnick ◽  
MC Cregger ◽  
SB Williams
Keyword(s):  
Sialic Acid ◽  
Factor Viii ◽  
Von Willebrand Factor ◽  
Platelet Rich Plasma ◽  
Molecular Forms ◽  
Von Willebrand's Disease ◽  
Von Willebrand’S Disease ◽  
Platelet Receptor ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract The factor VIII/von Willebrand factor (f.VIII/vWf) protein was purified from the plasma of a patient with von Willebrand's disease (vWd). The patient had all of the classic laboratory findings of vWd except for the ristocetin-induced platelet aggregation of his own platelet-rich plasma. The disease has been documented in three generations. Comparison of the purified normal and vWd f.VIIi/vWf protein revealed several abnormalities, including decreased concentration of f.VIII/vWf antigen; decreased specific vWf activity; absence of the larger molecular forms of the f.VIII/vWf protein; carbohydrate deficiencies affecting the sialic acid, penultimate galactose and N- acetylglucosamine moieties; and decreased binding of the f.VIII/vWf protein to its platelet receptor. These studies indicate the multiplicity of biochemical and functional abnormalities associated with the f.VIII/vWf protein in vWd. f.VIII/vWf protein to normal f.VIII/vWf protein that had been treated with 2-mercaptoethanol (2-ME) to reduce the multimer size and then treated with specific exoglycosidases to remove the sialic acid and penultimate galactose residues revealed similar biologic properties.

scholarly journals Effect of von Willebrand factor coexpression on the synthesis and secretion of factor VIII in Chinese hamster ovary cells.

1989 ◽  
Vol 9 (3) ◽  
pp. 1233-1242 ◽  
Author(s):  
R J Kaufman ◽  
L C Wasley ◽  
M V Davies ◽  
R J Wise ◽  
D I Israel ◽  
...  
Keyword(s):  
Factor Viii ◽  
Von Willebrand Factor ◽  
Chinese Hamster Ovary ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Translation Product ◽  
Factor Viii Activity ◽  
Ovary Cells ◽  
Von Willebrand ◽  
Willebrand Factor

In plasma, antihemophilic factor (factor VIII) exists as a 200-kilodalton heavy-chain polypeptide in a metal ion association with an 80-kilodalton light-chain polypeptide. This complex is bound by hydrophobic and hydrophilic interactions to a large multimeric glycoprotein, von Willebrand factor (vWF). Accumulation of secreted human factor VIII activity expressed in Chinese hamster ovary cells requires the addition of serum in the growth medium, which provides vWF. Here we report that coexpression of vWF with factor VIII in Chinese hamster ovary cells resulted in increased stable accumulation of factor VIII activity in the absence of serum in the growth medium. In the coexpressing cells, the vWF cDNA transcription unit was transcribed to yield mRNA which was efficiently translated. vWF was properly processed and secreted to yield disulfide-bonded high-molecular-weight multimers similar to those observed in vWF secreted from human endothelial cells. Nuclear run-on assays showed that the factor VIII gene was transcribed at a level similar to that of the vWF gene, but the mRNA did not accumulate to high levels in the cytoplasm. In addition, although the translation efficiency of the factor VIII mRNA was similar to that of vWF, the processing and secretion of the factor VIII primary translation product was dramatically reduced compared with vWF. These results demonstrate that in Chinese hamster ovary cells both factor VIII mRNA accumulation and the processing and secretion of the primary factor VIII translation product are inefficient processes.

scholarly journals Multimeric composition of factor VIII/von Willebrand factor following administration of DDAVP: implications for pathophysiology and therapy of von Willebrand's disease subtypes

Blood ◽  
1982 ◽  
Vol 59 (6) ◽  
pp. 1272-1278 ◽  
Author(s):  
ZM Ruggeri ◽  
PM Mannucci ◽  
R Lombardi ◽  
AB Federici ◽  
TS Zimmerman
Keyword(s):  
Factor Viii ◽  
Von Willebrand Factor ◽  
Resting State ◽  
Bleeding Time ◽  
Type I ◽  
Von Willebrand's Disease ◽  
Von Willebrand’S Disease ◽  
Plasma Factor ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract We have studied the modifications in the multimeric composition of plasma factor VIII/von Willebrand factor and the bleeding time response following administration of 1-Deamino-[8-D-arginine]-Vasopressin (DDAVP) to patients with different subtypes of von Willebrand's disease. In type I, all multimers were present in plasma in the resting state, though they were decreased in concentration. Administration of DDAVP resulted in an increased concentration of these forms as well as the appearance of larger forms than were previously present. There was concomitant correction of the bleeding time. In type IIA, large multimers were absent in the resting state, and although DDAVP induced an average threefold increase in the plasma concentration of factor VIII/von Willebrand factor, the larger multimers did not appear and the bleeding time, although shortened, was not corrected. In contrast, the larger multimers that were also absent from type IIB plasma in the resting state rapidly appeared following DDAVP administration. However, their appearance was transitory and the bleeding time, as in IIA patients, was shortened but not corrected. The characteristic multimeric composition of platelet factor VIII/von Willebrand factor in given subtypes predicted the alteration in plasma factor VIII/von Willebrand factor induced by DDAVP. These studies provide evidence that the different subtypes of von Willebrand's disease represent distinct abnormalities of factor VIII/von Willebrand factor. They also suggest that complete hemostatic correction following DDAVP can be routinely expected only in type I von Willebrand's disease, and only if factor VIII/von Willebrand factor can be raised to normal levels.

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