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VWF
Kinds of VWF Terms modified by VWF Selected AbstractsProcoagulant factors and the risk of myocardial infarction in young womenEUROPEAN JOURNAL OF HAEMATOLOGY, Issue 1 2006Bea Tanis Abstract:,Objectives:,We investigated whether elevated levels of factor VIII, IX and XI is associated with myocardial infarction (MI) in young women. In addition, we studied ABO blood group, von Willebrand factor (VWF) and C-reactive protein (CRP). Methods and results:,We compared 200 women with MI before age 49 years with 626 controls from a population-based case,control study. Mean levels of factor VIII activity (VIII), von Willebrand factor antigen (VWF), factor IX activity (IX) were higher in patients (133, 134 and 132 IU/dL) than in controls (111, 107 and 120 IU/dL, respectively). Mean levels of factor XI (XI) were equal in patients (114 IU/dL) and controls (113 IU/dL). The odds ratio (OR) for MI for blood group non-O vs. O was 1.6 [95% confidence interval (CI) 1.1,2.3]. The OR adjusted for age, index year and area of residence for the highest quartile >150 IU/dL of factor VIII was 2.7 (95% CI 1.6,4.6), of VWF 4.7 (95% CI 2.3,9.7), of factor IX 2.6 (95% CI 1.3,5.4) and of factor XI 0.9 (95% CI 0.5,1.4), all compared with the lowest quartile <100 IU/dL. Conclusions:,Non-O blood group, high VWF, factor VIII and factor IX levels are associated with an increased risk of MI in young women, while high factor XI levels are not. [source] Factor VIII and von Willebrand factor interaction: biological, clinical and therapeutic importanceHAEMOPHILIA, Issue 1 2010V. TERRAUBE Summary., The interaction of factor VIII (FVIII) with von Willebrand Factor (VWF) is of direct clinical significance in the diagnosis and treatment of patients with haemophilia A and von Willebrand disease (VWD). A normal haemostatic response to vascular injury requires both FVIII and VWF. It is well-established that in addition to its role in mediating platelet to platelet and platelet to matrix binding, VWF has a direct role in thrombin and fibrin generation by acting as a carrier molecule for the cofactor FVIII. Recent studies show that the interaction affects not only the biology of both FVIII and VWF, and the pathology of haemophilia and VWD, but also presents opportunities in the treatment of haemophilia. This review details the mechanisms and the molecular determinants of FVIII interaction with VWF, and the role of FVIII,VWF interaction in modulating FVIII interactions with other proteases, cell types and cellular receptors. The effect of defective interaction of FVIII with VWF as a result of mutations in either protein is discussed. [source] Properties of a concentrated minipool solvent-detergent treated cryoprecipitate processed in single-use bag systemsHAEMOPHILIA, Issue 5 2008T. BURNOUF Summary., Cryoprecipitate is still used to treat factor VIII (FVIII), von Willebrand factor (VWF) and/or fibrinogen deficiency. Recently a solvent-detergent (S/D) process of minipools of cryoprecipitate performed in a closed bag system has been designed to improve its viral safety. Still, cryoprecipitate has other drawbacks, including low concentration in active proteins, and presence of haemolytic isoagglutinins. We report here the biochemical evaluation of S/D-treated minipools of cryoprecipitates depleted of cryo-poor plasma. Cryoprecipitates were solubilized by 8 mL of a sterile glucose/saline solution, pooled in batches of 40 donations and subjected to S/D treatment in a plastic bag system using either 2% TnBP or 1% TnBP-1%Triton X-45, followed by oil extractions (n = 10). Mean (±SD) FVIII and fibrinogen content was 8.86 (±1.29) IU mL,1 and 16.02 (±1.98) mg mL,1, and 8.92 (±1.05) IU mL,1 in cryoprecipitate minipools treated with 2% TnBP, and 17.26 (±1.71) mg mL,1, in those treated by TnBP-Triton X-45, respectively. The WWF antigen, ristocetin cofactor and collagen binding activities were close to 10, 7 and 8 IU mL,1, respectively, and were not affected by either SD treatment. VWF multimeric pattern of SD-treated cryoprecipitates were similar to that of normal plasma, and the >15 mers and >10 mers content was identical to that of the starting cryoprecipitates. The anti-A and anti-B titre was 0,1 and 0,1/8, respectively. Therefore, it is possible to prepare virally inactivated cryoprecipitate minipools depleted of isoagglutinins and enriched in functional FVIII, VWF and clottable fibrinogen. [source] The molecular analysis of von Willebrand disease: a guideline from the UK Haemophilia Centre Doctors' Organisation Haemophilia Genetics Laboratory NetworkHAEMOPHILIA, Issue 5 2008S. KEENEY Summary., von Willebrand disease (VWD) is a common autosomally inherited bleeding disorder associated with mucosal or trauma-related bleeding in affected individuals. VWD results from a quantitative or qualitative deficiency of von Willebrand factor (VWF), a glycoprotein that is essential for primary haemostasis and that carries and protects coagulation factor VIII (FVIII) in the circulation. Through characterization of the phenotype and identification of mutations in the VWF gene in patients with VWD, understanding of the genetics and biochemistry of VWF and VWD has advanced considerably. The importance of specific regions of VWF for its interaction with other components of the vasculature has been revealed, and this has facilitated the formal classification of VWD into three subtypes based upon quantitative (types 1 and 3) and qualitative (type 2) deficiency of VWF. The underlying genetic lesions and associated molecular pathology have been identified in many cases of the qualitative type 2 VWD variants (2A, 2B, 2M, 2N) and in the severe quantitative deficiency, type 3 VWD. However in the partial quantitative deficiency, type 1 VWD, the picture is less clear: there is a variable relationship between plasma levels of VWF and bleeding, there is incomplete penetrance and variable expressivity within affected families, the causative molecular defect is unknown in a substantial number of cases, and even in those cases where the causative mutation is known, the associated molecular pathology is not necessarily understood. This guideline aims to provide a framework for best laboratory practice for the genetic diagnosis of VWD, based upon current knowledge and understanding. [source] Type 1 von Willebrand disease: application of emerging data to clinical practiceHAEMOPHILIA, Issue 4 2008P. W. COLLINS Summary., There has been much recent data published on type 1 von Willebrand disease (VWD) predominantly from three multi-centre cohort studies. These data have influenced a revision of the classification of type 1 VWD and have important implications for the management of this disorder. Patients with low von Willebrand factor (VWF) levels tend to have VWF mutations and VWD is transmitted predictably within families. In patients with VWF levels close to the lower end of the normal range, candidate mutations are found less often, ABO blood group is a more important factor and the disease has variable heritability within families. The importance of bleeding symptoms, in addition to VWF levels, in the diagnosis of type 1 VWD has been highlighted. [source] Hypothyroidism and acquired von Willebrand's syndrome: a systematic reviewHAEMOPHILIA, Issue 3 2008E. MANFREDI Summary., Acquired von Willebrand's syndrome type I is the supposed main underlying cause of bleeding tendency in hypothyroid patients. The purpose of this systematic review was to summarize the published evidence on the association between hypothyroidism and acquired von Willebrand's syndrome. All published clinical epidemiological and interventional studies, case reports and in vitro studies that investigated the association between hypothyroidism and acquired von Willebrand's syndrome were identified by a computer-assisted search of the MEDLINE and EMBASE electronic databases. A quality assessment was performed for clinical epidemiological studies. A total of 41 papers were included. A total of 22 epidemiological in vivo studies, two in vitro studies and 47 case reports were finally analyzed. No high quality in vivo study was identified. Almost all bleeding episodes described in the case reports were mucocutaneous. von Willebrand factor (VWF) antigen value was available for 23 patients: median value 28 U/dL (range: 4,45); VWF activity was available for 24 patients: median value 28.5 U/dL (range: <3,55); factor VIII activity was available for 16 patients: median value 47 U/dL (range: 9,74). Acquired von Willebrand's syndrome may be the main factor responsible for bleeding diathesis in overt hypothyroid patients. Even if bleeding episodes are mainly mild and mucocutaneous, blood transfusion, drug administration or surgical procedure may be required. [source] C1584 in von Willebrand factor is necessary for enhanced proteolysis by ADAMTS13 in vitroHAEMOPHILIA, Issue 4 2007S. KEENEY Summary., The cysteine variant of the amino acid change tyrosine/cysteine 1584 (Y/C1584) in von Willebrand factor (VWF) has previously been shown to cosegregate with increased susceptibility of VWF to proteolysis by ADAMTS13. It is not known whether C1584 itself confers increased proteolysis or is linked to a causative change elsewhere in VWF. To address whether C1584 underlies enhanced susceptibility of VWF to ADAMTS13-mediated proteolysis, a single family comprising two heterozygous Y/C1584 individuals and four homozygous Y/Y1584 individuals was investigated. The essential regions of the VWF gene were sequenced in all six individuals and ADAMTS13-mediated proteolysis of plasma VWF was assessed for each individual. Comparison of the VWF coding sequences for the Y/C1584 individuals with those for the Y/Y1584 individuals revealed that two amino acid variants were unique to the heterozygotes: R484 and C1584. The plasma VWF of the two heterozygotes showed increased susceptibility to proteolysis in vitro compared with that of the four homozygotes. In the present study we demonstrate that R484, in the absence of C1584, does not influence VWF proteolysis. Enhanced proteolysis occurred only in the presence of Cys1584. Thus, Cys1584 is necessary for increased susceptibility of VWF to proteolysis by ADAMTS13. [source] Evaluation of pharmacokinetics, efficacy and safety of Immunate® solvent detergent in previously treated patients with severe haemophilia AHAEMOPHILIA, Issue 1 2007L. NEMES Summary., Immunate® Solvent Detergent (S/D) is a plasma derived, purified, human factor VIII (FVIII) , von Willebrand factor (VWF) complex subjected to two virus inactivation/removal processes: S/D and vapor heat treatment. This prospective, multicentre, three-part clinical study evaluated the pharmacokinetics (in comparison to the predecessor product Immunate®), efficacy and safety of Immunate® S/D in 56 previously treated patients with severe haemophilia A. Subjects received Immunate® S/D on-demand, as a prophylactic regimen or both. The results of the pharmacokinetic population demonstrate that Immunate® and Immunate® S/D were equivalent with respect to the FVIII , and to the retrospectively VWF , parameters assessed. A total of 623 bleeding episodes were reported in 47/56 subjects. The duration of prophylaxis ranged from 0.1--5.2 months with a total of 175.6 months. The median number of bleeds per month in subjects on prophylaxis was 0 (range 0--10). Ninety-six percent of bleeding episodes were rated as having an excellent or good response. For most bleeding episodes (89%), subjects required only one infusion with a mean dose of 29.6 IU kg,1. No FVIII inhibitory antibodies were observed in any subject. No related serious adverse events were reported. Thus, the introduction of S/D treatment did not alter the PK characteristics and function of VWF and FVIII molecules of Immunate® S/D which is effective and safe for treatment of bleeding episodes, management of surgical procedures, and prophylaxis. [source] The 80th anniversary of von Willebrand's disease: history, management and researchHAEMOPHILIA, Issue 6 2006A. B. FEDERICI Summary., The history of von Willebrand's disease (VWD) is fascinating because it demonstrates how good clinical observations, genetic studies and biochemical skills can improve basic understanding of a disease and its management. The continuous efforts of scientists and clinicians during the last 80 years have significantly improved the knowledge of von Willebrand factor (VWF) structure and function and the management of VWD. Diagnosis of phenotype and genotype is now available in many countries and treatment is becoming more specific according to the VWD type. Any therapeutic agents must correct the dual defect of haemostasis, i.e. the abnormal platelet adhesion due to reduced and/or dysfunctional and low levels of factor VIII (FVIII) associated with VWF defects. Desmopressin (DDAVP) is the treatment of choice for type 1 VWD because it induces release of VWF from cellular compartments. Plasma virally inactivated VWF concentrates containing FVIII are effective and safe in patients unresponsive to DDAVP. There are advanced plans to develop a recombinant VWF but this product will require the concomitant administration of FVIII for the control of acute bleeds. Basic research studies on cellular biology, biochemistry and immunology have confirmed the role of VWF as a crucial participant in both haemostasis and thrombosis as its main biological activity is to support platelet adhesion,aggregation in the circulation. Retrospective and prospective clinical research studies, including bleeding history and laboratory markers for diagnosis as well as the use of DDAVP and VWF concentrates to manage or prevent bleeds in patients with VWD have been essential to provide general guidelines for VWD management. The large number of publications quoting VWD and VWF emphasizes the important role of VWF in medicine. [source] Management of acquired von Willebrand's sryndrome in a patient requiring major surgeryHAEMOPHILIA, Issue 6 2005J. M. Maddox Summary., We present the case of a patient with acquired von Willebrand's syndrome and a monoclonal gammopathy of undetermined significance who required cystectomy for relapsed transitional cell carcinoma (TCC) of the bladder. We demonstrated that infused von Willebrand factor (VWF) containing factor VIII concentrates had an unacceptably short half-life, but that this was significantly prolonged following combined therapy with plasma exchange and intravenous immunoglobulin (IVIgG). This approach was successfully utilized peri-operatively, with the total surgical blood loss less than would be expected even for a haemostatically normal patient. Trough VWF antigen and Ristocetin co-factor activity levels fell on the second postoperative day and we therefore administered further IVIgG. Levels again fell on the fifth postoperative day with the development of a Staphylococcus aureus septicaemia. At this point bleeding occurred from a surgical drain site requiring ,factor VIII inhibitor bypass activity' to secure haemostasis while further plasma exchange and IVIgG were administered. Now 5 years later, there is no evidence of recurrence of the TCC or progression of the monoclonal gammopathy. [source] The role of the platelet function analyser (PFA-100TM) in the characterization of patients with von Willebrand's disease and its relationships with von Willebrand factor and the ABO blood groupHAEMOPHILIA, Issue 3 2003I. C. Nitu-Whalley Summary. Determination of the closure time (CT) with the platelet function analyser (PFA-100TM) is a useful screening test for von Willebrand's disease (VWD) but its role in the characterization of VWD is not well established. We studied the relationship between the prolongation of the CT with adenosine diphosphate (ADP) (CT-ADP) and epinephrine (CT-EPI) cartridges and the von Willebrand factor (VWF) in 53 patients with VWD. We found that a relatively small percentage of the prolongation of the CT-ADR and CT-ADP (16 and 29%, respectively) was determined by a reduction in VWF levels. The CT-ADP was significantly more prolonged in the presence of qualitative defects in VWF but could not discriminate between the VWD subtypes. The ABO blood group had no effect on the prolongation of the CT or the bleeding time. In conclusion, the PFA-100TM appears of little use in the characterization of severity and subtype of VWD. [source] A new ELISA assay for diagnosis of acquired von Willebrand SyndromeHAEMOPHILIA, Issue 3 2003C. Siaka Summary. The pathophysiology of acquired von Willebrand syndrome (AVWS), a rare bleeding disorder, is not fully understood. Circulating antibodies to Von Willebrand factor (VWF) are found in patients with AVWS associated with lymphoproliferative disorders but these autoantibodies are difficult to detect with routine laboratory tests and neutralisation assays. We have developed a simple enzyme-linked immunosorbent assay (ELISA) to detect serum antibody binding to VWF protein immobilized on polystyrene plates. Ten patients with AVWS were studied, eight of whom also had lymphoproliferative disorders. We found antibodies in eight patients; all of them were positive for IgG and five were also positive for IgM. This simple method appears to be more sensitive than functional assays, which failed to identify two of the patients who were positive with the ELISA. In conjunction with other tests, this ELISA method may be useful for demonstrating the immunological mechanism underlying some cases of AVWS. Such patients would qualify for intravenous immunoglobulin therapy, which can correct the clotting disorder. [source] Evaluation of an automated screening assay for von Willebrand Disease Type 2NINTERNATIONAL JOURNAL OF LABORATORY HEMATOLOGY, Issue 6 2002S. L. Taylor Summary Evaluating the factor VIII (FVIII) binding activity of von Willebrand factor (VWF) is an important step in the diagnostic work-up of families affected by apparent mild haemophilia A. In von Willebrand's disease (VWD) type 2N (Normandy), mutations at the N-terminal end of the mature VWF subunit gene prevent the binding of FVIII. Individuals heterozygous for type 2N VWD are generally asymptomatic. Homozygotes and compound heterozygotes present with a clinical picture which mimics haemophilia A, with a markedly reduced FVIII : C activity and VWF within the normal range, but instead of exhibiting X-linked inheritance they show an autosomal recessive inheritance pattern. The distinction between haemophilia A and VWD type 2N has important implications for therapy and genetic counselling. We present a highly specific enzyme-linked immunosorbent assay screening method for the Normandy variant, which measures VWF : FVIII binding activity in parallel with VWF antigen, using monoclonal capture and detection antibodies. The assay is fully automated using a robotic microtitre plate processor, requiring minimal user intervention and providing the capacity to screen large numbers of patients. [source] Development of anti-VWF antibody in a patient with severe haemophilia A following the development of high-grade non-Hodgkin's lymphomaINTERNATIONAL JOURNAL OF LABORATORY HEMATOLOGY, Issue 3 2002K. Ghosh A 9-year-old-boy with severe haemophilia A (factor VIII < 1%) developed colicky abdominal pain with swelling in the left iliac fossa for 4 weeks. His LDH level was 1423 IU/l (normal range < 220 IU/l) and his uric acid, 6.8 mg/dl. A computerised tomography (CT) scan of the abdomen demonstrated a tumour of the terminal ileum and mild hepatosplenomegaly. Pre-operative screening for factor VIII inhibitor was negative. Post-operatively, the patient needed high doses of factor VIII to maintain haemostasis. The tumour was found to be a high-grade lymphoma of Burkitt's type. He recovered from his operation and chemotherapy was commenced. Investigations demonstrated an anti-von Willebrand factor (VWF) antibody. He subsequently relapsed and died of progressive disease. Development of anti-VWF antibody in lymphoma is well known, but development of this antibody in a haemophilia A patient developing lymphoma has not been reported. The present case shows that antibody to VWF should be considered as a possible reason for an increased factor VIII requirement in such patients. [source] Effect of blood group on idiopathic thrombotic thrombocytopenic purpuraJOURNAL OF CLINICAL APHERESIS, Issue 4 2009Lara Zuberi Abstract Thrombotic thrombocytopenic purpura (TTP) is a condition caused by deficiency of ADAMTS13 resulting in accumulation of ultra large Von Willebrand factor multimers (ULVWF), leading to micro thrombi in multiple organs. The varying susceptibilities of blood group antigens to ADAMTS13 have been demonstrated. A and B antigens are protective of VWF; and VWF purified from blood group O individuals has been shown to be cleaved faster by ADAMTS13 compared to VWF from blood group AB individuals. We proposed that there may be a difference in the incidence of blood groups in TTP patients compared with the general population. We felt this to be important for a life-threatening disease with poorly understood epidemiology. We report a retrospective analysis of 74 patients presenting from 1993 to 2008 with idiopathic TTP. We studied the incidence across various blood groups and also estimated the recurrence and mortality in each group. The incidence of various blood groups were as follows: O 36%, A 36%, B 25%, and AB 2%, compared with expected frequencies in the Detroit area: O 44%, A 33% B 20%, and AB 3%. There was a trend of lower than expected frequency of blood group O. There were 24 recurrences and 14 deaths, uniform across blood groups. We hypothesized that there may be an association between blood groups and the risk of TTP; however the differences in our study were not statistically significant. Recurrence and disease specific mortality did not appear to be impacted by blood group. J. Clin. Apheresis 2009. © 2009 Wiley-Liss, Inc. [source] Potential Role of Enhanced Cytokinemia and Plasma Inhibitor on the Decreased Activity of Plasma ADAMTS13 in Patients With Alcoholic Hepatitis: Relationship to EndotoxemiaALCOHOLISM, Issue 2010Masatoshi Ishikawa Background:, Deficiency of ADAMTS13 (adisintegrin-like and metalloproteinase with thrombospondin type-1 motifs 13) results in an increase in unusually large von Willebrand factor multimer (UL-VWFM) of the plasma and finally causes microcirculatory disturbance. Our previous study demonstrated that the imbalance of increased UL-VWFM over decreased ADAMTS13 activity may contribute to the development of multiorgan failure in patients with alcoholic hepatitis (AH). The aim of this study was to explore the potential mechanism to reduce the activity of plasma ADAMTS13. Methods:, Plasma cytokine levels including interleukin (IL)-6, IL-8, and tumor necrosis factor-, (TNF-,), plasma endotoxin concentration, and the plasma inhibitor against ADAMTS13 were determined together with ADAMTS13 activity, VWF antigen (VWF:Ag), and UL-VWFM in 24 patients with AH and 5 patients with severe alcoholic hepatitis (SAH). Results:, The concentrations of IL-6, IL-8, and TNF-, on admission were significantly higher in patients with SAH than in those with AH and controls. The ADAMTS13 activity concomitantly decreased, and the VWF:Ag progressively elevated with increasing concentrations of these cytokines from normal range to over 100 pg/ml. Plasma endotoxin concentration was markedly higher in patients with SAH (mean 52.3 pg/ml) and AH (21.7 pg/ml) than in controls (7.9 pg/ml). The endotoxin concentration inversely correlated with ADAMTS13 activity and was higher in patients with UL-VWFM than those without. The inhibitor was detected in 4 patients with SAH (0.9 to 2.1 BU/ml) and 6 patients with AH (0.5 to 1.6 BU/ml). Patients with the inhibitor showed lower functional liver capacity, higher endotoxin concentration, and marked inflammatory signs than those without. At the recovery stage, the ADAMTS13 activity increased to normal range, the VWF:Ag decreased, and the UL-VWFM disappeared with the decrease in the concentrations of cytokines and endotoxin, and the disappearance of the inhibitor. Conclusion:, Decreased ADAMTS13 activity and increased VWF:Ag could be induced not only by pro-inflammatory cytokinemia, but also by its inhibitor, both of which may be closely related to enhanced endotoxemia in patients with AH and SAH. [source] Improved performance characteristics of the von Willebrand factor ristocetin cofactor activity assay using a novel automated assay protocolJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 10 2010A. HILLARP Summary.,Background, objectives and methods:,An accurate, sensitive and precise assay for reliable determination of the ristocetin cofactor activity of von Willebrand factor (VWF:RCo) in plasma and von Willebrand Factor (VWF)-containing concentrates has been evaluated. The assay is based on a commercially available automated protocol with modifications including a combination of adding additional ristocetin and the use of two calibration curves for the high and low measuring ranges. Results:,Addition of extra ristocetin resulted in improved measurement of VWF recoveries from various VWF-containing concentrates that were underestimated using the standard automated protocol. The modifications resulted in improved assay performance over an extended measuring range (2.00,0.03 IU mL,1). Accuracy was tested using VWF deficiency plasma spiked with the 1st international standard (IS) for VWF concentrate. Seven dilutions, ranging from 1.80 to 0.05 IU mL,1, were analyzed and resulted in measured concentrations between 80% and 100% of the assigned potency of the standard. Linearity was determined from the regression plot of the same concentrate dilutions and resulted in a correlation coefficient of 0.998. The repeatability, expressed as coefficient of variation, was 2% in the normal range (0.90 IU mL,1) and 8% at the level of 0.05 IU mL,1. The corresponding reproducibility results were 2% and 15% at the normal and low measuring ranges, respectively. Conclusions:,Analysis of patients with von Willebrand disease (VWD) indicates that the modified automated BCS® protocol has a superior discrimination power compared with the standard protocol. This is especially true in samples with low VWF, as in patients with type 3 VWD. [source] Homozygous type 2N R854W von Willebrand factor is poorly secreted and causes a severe von Willebrand disease phenotypeJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 9 2010G. CASTAMAN Summary.,Background:,von Willebrand disease (VWD) type Normandy (VWD 2N) is caused by mutations at the factor (F)VIII-binding site of von Willebrand factor (VWF), located in the D,and D3 domains on the N-terminus of mature VWF. The R854Q mutation is the most frequent cause of this phenotype. Objectives:,We report the characterization of a homozygous VWD 2N mutation, R854W, detected in a patient with a severe VWD phenotype. Methods:,The plasma VWF phenotype was studied, transient expression of recombinant mutant full-length VWF in 293 EBNA cells was performed, and the results were compared with those obtained with wild-type (WT) VWF. Furthermore, expression was also examined in HEK293 cells, which form Weibel,Palade body-like granules when transfected with WT VWF. Results:,The multimer analysis of plasma VWF showed the lack of the typical triplet structure, with the presence of the central band only, and a relative decrease in the high molecular mass multimers. Homozygous expression of recombinant R854W VWF resulted in normal amounts of cellular VWF, but with a severe reduction in secretion into the medium. Severe reductions in FVIII binding to R854W VWF, glycoprotein Ib binding activity and collagen binding of secreted W854 VWF was observed, and reproduced the phenotypic parameters of plasma VWF. In HEK293 cells, homozygous R854W VWF failed to form Weibel,Palade body-like granules. Conclusions:,Our results demonstrate that a homozygous R854W mutation in the D, domain of VWF induces impaired secretion and activity of the protein, thereby explaining the severe phenotype of the patient. [source] Polymicrobial sepsis and endotoxemia promote microvascular thrombosis via distinct mechanismsJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 6 2010K. N. PATEL Summary.,Background:,We reported recently that endotoxemia promotes microvascular thrombosis in cremaster venules of wild-type mice, but not in mice deficient in toll-like receptor 4 (TLR4) or von Willebrand factor (VWF). Objective:,To determine whether the clinically relevant model of polymicrobial sepsis induced by cecal ligation/perforation (CLP) induces similar responses via the same mechanisms as endotoxemia. Methods:,We used a light/dye-injury model of thrombosis in the cremaster microcirculation of wild-type mice and mice deficient in toll-like receptor-4 (C57BL/10ScNJ), toll-like receptor 2 (TLR2), or VWF. Mice underwent CLP or sham surgery, or an intraperitoneal injection of endotoxin (LPS) or saline. In the CLP model, we assessed the influence of fluid replacement on thrombotic responses. Results:,Both CLP and LPS enhanced thrombotic occlusion in wild-type mice. In contrast to LPS, CLP enhanced thrombosis in TLR4- and VWF-deficient strains. While TLR2-deficient mice did not demonstrate enhanced thrombosis following CLP, LPS enhanced thrombosis in these mice. LPS, but not CLP, increased plasma VWF antigen relative to controls. Septic mice, particularly those undergoing CLP, developed significant hemoconcentration. Intravenous fluid replacement with isotonic saline prevented the hemoconcentration and prothrombotic responses to CLP, though fluids did not prevent the prothrombotic response to LPS. Conclusions:,Polymicrobial sepsis induced by CLP and endotoxemia promote microvascular thrombosis via distinct mechanisms; enhanced thrombosis induced by CLP requires TLR2 but not TLR4 or VWF. The salutary effects of intravenous fluid replacement on microvascular thrombosis in polymicrobial sepsis remain to be characterized. [source] von Willebrand factor activation, granzyme-B and thrombocytopenia in meningococcal diseaseJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 5 2010M. J. HOLLESTELLE Summary.,Background:,During invasive meningococcal disease, severe thrombocytopenia is strongly associated with a poor outcome. Objectives:,In order to elucidate the pathophysiological mechanism behind the development of thrombocytopenia, we studied the role of von Willebrand factor (VWF) in meningococcal disease. Patients/methods:,Thirty-two children with severe meningococcal disease admitted to our university hospital were included in this study. VWF and related parameters were measured and results were correlated with the development of shock and thrombocytopenia. Results:,At admission, all patients had increased levels of (active) VWF and VWF propeptide. The highest VWF propeptide levels were observed in patients with shock, indicating acute endothelial activation. Although VWF propeptide levels in patients with shock, with or without thrombocytopenia, were similar, increased active VWF was significantly lower in patients with thrombocytopenia as compared with patients without thrombocytopenia. ADAMTS13 was moderately decreased. However, the VWF multimeric pattern was minimally increased. We assume that these findings are explained by VWF consumption and perhaps by granzyme B (GrB). In vitro experiments showed that GrB is able to cleave VWF multimers in plasma, whereas GrB was high in patients with shock, who developed thrombocytopenia. Conclusions:,Our results demonstrate that consumption of VWF, derived from endothelial cells, could be a key feature of meningococcal disease and primary to the development of thrombocytopenia during shock. [source] The glycoprotein Ib,,von Willebrand factor interaction induces platelet apoptosisJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 2 2010S. LI Summary.,Background: The interaction of glycoprotein (GP) Ib, with von Willebrand factor (VWF) initiates platelet adhesion, and simultaneously triggers intracellular signaling cascades leading to platelet aggregation and thrombus formation. Some of the signaling events are similar to those occurring during apoptosis, however, it is still unclear whether platelet apoptosis is induced by the GPIb,,VWF interaction. Objectives: To investigate whether the GPIb,,VWF interaction induces platelet apoptosis and the role of 14-3-3, in apoptotic signaling. Methods: Apoptotic events were assessed in platelets or Chinese hamster ovary (CHO) cells expressing wild-type (1b9) or mutant GPIb,IX interacting with VWF by flow cytometry or western blotting. Results: Ristocetin-induced GPIb,,VWF interaction elicited apoptotic events in platelets, including phosphatidylserine exposure, elevations of Bax and Bak, gelsolin cleavage, and depolarization of mitochondrial inner transmembrane potential. Apoptotic events were also elicited in platelets exposed to pathologic shear stresses in the presence of VWF; however, the shear-induced apoptosis was eliminated by the anti-GPIb, antibody AK2. Furthermore, apoptotic events occurred in 1b9 cells stimulated with VWF and ristocetin, but were significantly diminished in two CHO cell lines expressing mutant GPIb,IX with GPIb, truncated at residue 551 or a serine-to-alanine mutation at the 14-3-3,-binding site in GPIb,. Conclusions: This study demonstrates that the GPIb,,VWF interaction induces apoptotic events in platelets, and that the association of 14-3-3, with the cytoplasmic domain of GPIb, is essential for apoptotic signaling. This finding may suggest a novel mechanism for platelet clearance or some thrombocytopenic diseases. [source] The plasma von Willebrand factor O -glycome comprises a surprising variety of structures including ABH antigens and disialosyl motifsJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 1 2010K. CANIS Summary.,Background: von Willebrand factor (VWF) is a key component for maintenance of normal hemostasis. Its glycan moieties, accounting for about 20% of its molecular weight, have been shown to affect many of its properties. Previous studies reported correlations between VWF secretion, half-life and the nature or presence of its N -glycans, and more importantly between VWF plasma level and the type of N -linked ABH antigens. Despite the presence of 10 predicted O -glycosylation sites, the O -glycome remains poorly characterized, impairing the complete elucidation of its influence on VWF functions. So far only a single glycan structure, a disialyl core 1 glycan, has been identified. Objectives: To define an exhaustive profile of the VWF O -glycan structures to help the understanding of their role in VWF regulation and properties. Methods: Plasma-derived VWF O -linked sugars were isolated and analyzed using state-of-the-art mass spectrometry methodologies. Results and conclusions: We provide here a detailed analysis of the human plasma-derived VWF O -glycome. Eighteen O -glycan structures including both core 1 and core 2 structures are now demonstrated to be present on VWF. Amongst the newly determined structures are unusual tetra-sialylated core 1 O -glycans and ABH antigen-containing core 2 O -glycans. In conjunction with current models explaining VWF activity, knowledge of the complete O -glycome will facilitate research aimed at providing a better understanding of the influence of glycosylation on VWF functions. [source] von Willebrand factor is a major determinant of ADAMTS-13 decrease during mouse sepsis induced by cecum ligation and punctureJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 5 2009N. LEROLLE Summary.,Background:,During sepsis, von Willebrand factor (VWF) is abundantly secreted; the main mechanism regulating its size involves specific proteolysis by the metalloprotease ADAMTS-13. Objectives:,To determine whether ADAMTS-13 consumption due to its binding to, and/or cleavage, of VWF contributes to its decrease during sepsis and whether abrogating or enhancing ADAMTS-13 activity influences sepsis outcome. Methods:,ADAMTS-13 activity was evaluated in a model of sepsis induced by cecum ligature and puncture (CLP) in wild-type and Vwf,/, mice. Sepsis outcome was studied in those mice and in Adamts-13,/, mice. Finally, survival was studied in wild-type mice injected hydrodynamically with the human ADAMTS-13 gene. Results:,In wild-type mice, CLP-induced sepsis elicited a significant ADAMTS-13 decrease, and a strong negative correlation existed between VWF and ADAMTS-13. In Vwf,/, mice, CLP also induced severe sepsis, but ADAMTS-13 was not significantly diminished. Notably, Vwf,/, mice lived significantly longer than wild-type mice. In contrast, Adamts-13,/, mice and wild-type mice were comparable with regard to thrombocytopenia, VWF concentrations, absence of thrombi, and survival. Hydrodynamic hADAMTS-13 gene transfer with the pLIVE expression vector resulted in high and stable ADAMTS13 activity in CLP mice; however, no impact on survival was observed. Conclusions: VWF secretion is a major determinant of ADAMTS-13 decrease in the CLP model, and plays an important role in sepsis-induced mortality, but the complete absence of its regulating protease, ADAMTS-13, had no detectable impact in this sepsis model. Furthermore, increasing ADAMTS-13 activity had no impact on survival. [source] VH1-69 germline encoded antibodies directed towards ADAMTS13 in patients with acquired thrombotic thrombocytopenic purpuraJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 3 2009W. POS Summary.,Background: Autoantibodies directed towards ADAMTS13 are present in the majority of patients with acquired thrombotic thrombocytopenic purpura (TTP). Analysis of a set of antibodies derived from two patients with acquired TTP revealed frequent use of the VH1-69 heavy chain gene segment for the assembly of anti-ADAMTS13 antibodies. Objective: We explored the ability of two VH1-69 germline gene-encoded antibodies to inhibit the von Willebrand factor (VWF)-processing activity of ADAMTS13 under different experimental conditions. Furthermore, the presence of VH1-69 encoded anti-ADAMTS13 antibodies in 40 patients with acquired TTP was monitored using monoclonal antibody G8, which specifically reacts with an idiotype expressed on VH1-69 encoded antibodies. Methods and Results: Binding of the two VH1-69 encoded monoclonal antibodies was dependent on the presence of the spacer domain. Both antibodies inhibited ADAMTS13 activity under static conditions, as measured by cleavage of FRETS-VWF73 substrate and cleavage of VWF multimers. The recombinant antibodies were also capable of inhibiting the processing of UL-VWF strings on the surface of endothelial cells. G8-reactive antibodies directed towards ADAMTS13 were present in plasma of all patients containing anti ADAMTS13 antibodies. Conclusions: These results suggest that VH1-69 derived antibodies directed towards ADAMTS13 develop in the majority of patients with acquired TTP. [source] Generation and validation of the Condensed MCMDM-1VWD Bleeding Questionnaire for von Willebrand diseaseJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 12 2008M. BOWMAN Summary.,Background:,Given the challenges involved in obtaining accurate bleeding histories, attempts at standardization have occurred and the value of quantifying hemorrhagic symptoms has been recognized. Patients/methods:,An extensive validated bleeding questionnaire (MCMDM-1VWD) was condensed by eliminating all details that did not directly affect the bleeding score (BS) and the correlation between the two versions was tested. Additionally, the diagnostic utility of the condensed version was prospectively tested. Results:,Data on 259 individuals who were administered the questionnaire are presented here; 217 being prospectively investigated for von Willebrand disease (VWD) (group 1) and 42 previously known to have type 1, 2 or 3 VWD (group 2). Of the 217 prospectively investigated, 35 had positive BS (,4) and 182 had negative scores. Seven individuals (all with positive BS) had laboratory results consistent with type 1 VWD. This results in a sensitivity of 100% and a specificity of 87%. The positive predictive value is 0.20 and the negative predictive value is 1. The correlation between the full MCMDM-1VWD and condensed versions is excellent (Spearman's 0.97, P < 0.001, linear regression r2 = 96.4). Inter-observer reliability for the condensed version is reasonable (Spearman's 0.72, P < 0.001 and intra-class correlation coefficient 0.805, P < 0.001). There was a significant difference in BS between subtypes of VWD, with type 3 >> type 2 >> type 1 VWD (anovaP < 0.001). There is a strong inverse relationship between VWF:Ag level and BS (Spearman's ,0.411, P < 0.001). Conclusions:,The Condensed MCMDM-1VWD Bleeding Questionnaire is an efficient, effective tool in the evaluation of patients for VWD. [source] Protein kinase C-, mediates von Willebrand factor secretion from endothelial cells in response to vascular endothelial growth factor (VEGF) but not histamineJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 11 2008O. LORENZI Summary.,Background:,Vascular endothelial growth factor (VEGF) and histamine induce von Willebrand factor (VWF) release from vascular endothelial cells. Protein kinase C (PKC) is involved in the control of exocytosis in many secretory cell types. Objectives:,We investigated the role of PKC and the interactions between PKC and Ca2+ signaling in both VEGF-induced and histamine-induced VWF secretion from human umbilical vein endothelial cells (HUVECs). Results:,Several PKC inhibitors (staurosporine, Ro31-8220, myristoylated PKC peptide inhibitor and Go6983) block VEGF-induced but not histamine-induced VWF secretion. PKC-, and novel PKCs (PKC-,, PKC-,, and PKC-,), but not PKC-,, are expressed in HUVECs. Both VEGF and histamine activate PKC-,. However, gene inactivation experiments using small interfering RNA indicate that PKC-, (but not PKC-,) is involved in the regulation of VEGF-induced but not histamine-induced secretion. Both VEGF and histamine induce a rise in cytosolic free Ca2+ ([Ca2+]c), but the response to VEGF is weaker and even absent in a significant subset of cells. Furthermore, VEGF-induced secretion is largely preserved when the rise in [Ca2+]c is prevented by BAPTA-AM. Conclusions:,Our study identifies striking agonist specificities in signal,secretion coupling. Histamine-induced secretion is dependent on [Ca2+]c but not PKC, whereas VEGF-induced secretion is largely dependent on PKC-, and significantly less on [Ca2+]c. Our data firmly establish the key role of PKC-, in VEGF-induced VWF release, but suggest that a third, VEGF-specific, signaling intermediate is required as a PKC-, coactivator. [source] ADAMTS-13, von Willebrand factor and related parameters in severe sepsis and septic shockJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 11 2007J. A. KREMER HOVINGA Summary. Background:,Insufficient control of von Willebrand factor (VWF) multimer size as a result of severely deficient ADAMTS-13 activity results in thrombotic thrombocytopenic purpura associated with microvascluar thrombosis and platelet consumption, features not seldom seen in severe sepsis and septic shock. Methods:,ADAMTS-13 activity and VWF parameters of 40 patients with severe sepsis or septic shock were compared with those of 40 healthy controls of the same age and gender and correlated with clinical findings and sepsis outcome. Results:,ADAMTS-13 activity was significantly lower in patients than in healthy controls [median 60% (range 27,160%) vs. 110% (range 63,200%); P < 0.001]. VWF parameters behaved reciprocally and both VWF ristocetin cofactor activity (RCo) and VWF antigen (VWF:Ag) were significantly (P < 0.001) higher in patients compared with controls. Neither ADAMTS-13 activity nor VWF parameters correlated with disease severity, organ dysfunction or outcome. However, a contribution of acute endothelial dysfunction to renal impairment in sepsis is suggested by the significantly higher VWF propeptide and soluble thrombomodulin levels in patients with increased creatinine values as well as by their strong positive correlations (creatinine and VWF propeptide rs = 0.484, P < 0.001; creatinine and soluble thrombomodulin rs = 0.596, P < 0.001). Conclusions:,VWF parameters are reciprocally correlated with ADAMTS-13 activity in severe sepsis and septic shock but have no prognostic value regarding outcome. [source] Two novel monoclonal antibodies to VWFA3 inhibit VWF-collagen and VWF-platelet interactionsJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 9 2007Y. ZHAO Summary.,Background:,The interaction of collagen-von Willebrand factor (VWF)-GPIb is essential for platelet adhesion, especially under high shear conditions. VWF, which acts as a bridge between platelets and exposed subendothelium, interacts with collagen through its A3 domain, which is a new target for the antithrombotic agent. Objective:,To develop functional blockers that specifically inhibit VWF-dependent adhesion of platelets to collagen under high shear stress. Methods:,To develop murine antihuman VWF A3 monoclonal antibodies (mAbs) by standard hybridoma technology, and characterize their abilities to block interactions between VWF A3 and collagen as well as platelet function. Results:,Thirty anti-VWF-A3 mAbs were obtained. Among them, two mAbs, designated as SZ-123 and SZ-125, were found to inhibit VWF-collagen type III interaction. SZ-123 and SZ-125 inhibited the binding of purified human VWF (1.5 or 3 ,g mL,1) to human placenta collagen type III (IC50 = 0.07 ± 0.02 and 0.15 ± 0.03 ,g mL,1, respectively) or to calf skin collagen type III (IC50 = 0.48 ± 0.06 and 0.51 ± 0.07 ,g mL,1, respectively) coated on plates. Under flow shear condition (1000 s,1), SZ-123 and SZ-125 inhibited platelet adhesion on human placenta collagen- or calf skin collagen-coated surfaces. Both mAbs also inhibited platelet aggregation induced by ristocetin, botrocetin or bovine plasma. Conclusions:,SZ-123 and SZ-125 inhibited VWF-collagen and VWF-platelet interactions. [source] Purified A2 domain of von Willebrand factor binds to the active conformation of von Willebrand factor and blocks the interaction with platelet glycoprotein Ib,JOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 7 2007C. MARTIN Summary.,Background:,von Willebrand factor (VWF) does not interact with circulating platelets unless it is induced to expose the binding site for platelet glycoprotein (GP)Ib, in the A1 domain by high shear stress, immobilization, and/or a modulator. Previous studies have implied indirectly that the A2 domain may be involved in regulating A1,GPIb, binding. Objective and methods:,Because the relationship between the A1 and A2 domains has not been defined, we have investigated the effect of the A2 domain on the binding activity of the A1 domain using recombinant A domain polypeptides, multimeric VWF, and monoclonal antibodies (mAb). Results:,The A2 domain polypeptide bound specifically to the immobilized A1 domain polypeptide or full-length VWF, with half-maximal binding being obtained at 60 or 168 nm, respectively. This A1,A2 interaction was inhibited by mAb against the A2 or A1 domain and by the A1 domain polypeptide. The A2 domain polypeptide effectively blocked GPIb,-mediated platelet adhesion under high flow conditions. The A2 domain polypeptide specifically recognizes the GPIb,-binding conformation in the A1 domain, as it only interacted with VWF activated by the modulator ristocetin or immobilized VWF. Furthermore, in contrast to plasma VWF, the ultra-large (UL)VWF multimers or a recombinant VWF,A1A2A3 polypeptide containing a gain-of-function mutation (R1308 L) of type 2B von Willebrand disease bound to the A2 domain polypeptide without the need for ristocetin. Conclusions:,The recombinant A2 domain polypeptide specifically binds to the active conformation of the A1 domain in VWF and effectively blocks the interaction with platelet GPIb, under high-flow conditions. [source] Formation of platelet strings and microthrombi in the presence of ADAMTS-13 inhibitor does not require P-selectin or ,3 integrinJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 3 2007A. K. CHAUHAN Summary. Background:,Ultra-large von Willebrand factor (ULVWF) and the receptor P-selectin are released from endothelial Weibel,Palade bodies during injury or inflammation. VWF mediates platelet adhesion and P-selectin promotes leukocyte rolling. ADAMTS-13 limits the duration of platelet adhesion by cleaving the ULVWF. In the absence of ADAMTS-13, long VWF filaments decorated with platelets form. Recent in vitro studies suggested that P-selectin might anchor these platelet strings to endothelium, but whether the same mechanism exists in vivo remains to be elucidated. Methods:,We address the role of P-selectin and ,3 integrin in platelet string formation in vivo using intravital microscopy by infusing inhibitory ADAMTS-13 antibody in P-selectin-/- and ,3 -deficient mice and activating the endothelium by injecting histamine. Results:,We show that inhibition of ADAMTS-13 combined with endothelial activation leads to similar extents of platelet string formation in wild-type, P-selectin- and integrin ,3 -deficient mice. Further, in venules the platelet strings can coalesce into VWF-platelet aggregates. This process utilizes neither the platelet ,3 integrin nor P-selectin. We also show in vitro that platelets can act as a bridge between the VWF fibers and that VWF can self-associate even in areas devoid of platelets. Conclusions:,The formation or retention of the platelet strings does not require P-selectin or the endothelial VWF receptor ,v,3. Furthermore, in the presence of low ADAMTS-13 activity, VWF-dependent and ,IIb,3 -independent platelet clustering occurs in veins, as has been shown at high arterial shear rates. Our study further supports the importance of regulation of VWF multimer size upon secretion from Weibel,Palade bodies. [source] | ||||||||||||||||