Thrombin generation during a regular menstrual cycle in women with von Willebrand disease

Introduction Von Willebrand disease is the most common bleeding disorder with a prevalence of 1-2% of the population. Nevertheless diagnosis of a von Willebrand Syndrom Typ 1 is still challenging. In a newer publication (1), 30 studies about the haemostatic variables during the menstrual cycle were compared. 11 studies were focused on the von Willebrand parameters but only in one study these parameters in patients with von Willebrand disease were observed. We investigated possible cyclic variations in women with menorrhagia, which can lead to a diagnosis of a coagulation disorder. Samples and Methods We conducted a laboratory workup in 122 women sent to our lab for menorrhagia. The following tests were conducted: Blood count, VWF:RCo, VWF:Ag, VWF:CB, Fibrinogen (Clauss), activities of FII, FV, FVII, FVIII (clotting and chromogenic), FIX, FX, FXI, FXII, FXIII during the menstrual cycle on predefined time points (day 1-6, day 7-11, day 12-18, day 19-23, day 24-28). Results In 51 (40%) patients a von Willebrand disease could be detected, 37% hat other coagulation disorders like p. e. factor-VII-deficieny and factor-XIII-deficiency. 11.5% had an iron deficiency. In 11.5% no coagulation disorder could be found. In patients with von Willebrand disease we found cyclic variations especially in the VWF:Ag (p = 0.02). They showed the lowest level during the ovulation. For other coagulation parameters no significance for variations during the menstrual cycle were found. Conclusion There are cyclic variations in von Willebrand antigen. To investigate women on predefined time points during the menstrual cycle can be useful to diagnose a von Willebrand disease particularly in mild cases in which no other suspicious bleeding symptoms exists. Literature Knol H.M., Kemperman R.F.J., Kluin-Nelemans C., Mulder A.B., Meijer K., Haemostatic variables during normal menstrual cycle, Thrombosis and Haemostasis 107.1/2012 Disclosures: Halimeh: Octapharma AG: Investigator Other, Research Funding.

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The Effect of DDAVP Infusion on Thrombin Generation in Platelet-rich Plasma of von Willebrand Type 1 and in Mild Haemophilia A Patients

Thrombosis and Haemostasis ◽

10.1055/s-0037-1614080 ◽

2000 ◽

Vol 84 (10) ◽

pp. 638-642 ◽

Cited By ~ 32

Author(s):

Irene Keularts ◽

K. Hamulyak ◽

H.C. Hemker ◽

Suzette Béguin

Keyword(s):

Factor Viii ◽

Thrombin Generation ◽

Platelet Rich Plasma ◽

Von Willebrand Disease ◽

Clotting Factor ◽

Haemophilia A ◽

The Individual ◽

Von Willebrand ◽

Mild Haemophilia

SummaryIn von Willebrand disease (vWD) type 1 and mild haemophilia A patients we studied the effect of an infusion of DDAVP (0.3 µg/kg body weight) on thrombin generation in platelet-rich plasma (PRP) and platelet-poor plasma (PPP). Baseline thrombin generation in PRP was diminished both in the haemophilia A and vWD patients. It was normal in vWD plasma when sufficient procoagulant phospholipids were present, either via adding phospholipid vesicles to PPP or via scrambling of the platelet membrane with ionomycin in PRP. In haemophilia A plasma, thrombin generation did not normalize by providing procoagulant phospholipids. Treatment with DDAVP temporarily restored thrombin generation in PRP to normal in both diseases.To investigate the individual roles of von Willebrand factor (vWF) and factor VIII, we also studied the effect of factor VIII infusion on thrombin generation in a severe haemophilia patient. It appears that at a fixed normal vWF concentration, <25% factor VIII is sufficient for normal thrombin generation in PRP. At a sufficient factor VIII concentration, however, thrombin generation is still lower than normal in vWD patients; ∼40% of vWF is required for half-normal thrombin generation in PRP.It thus appears that vWF is also a clotting factor, in the sense that it is required for normal thrombin generation. This underlines the importance of the interaction between coagulation and the platelets in normal haemostasis. Thrombin generation in PRP appears to be a suitable test to reflect the combined function.

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Fibrin-Dependent Platelet Procoagulant Activity Requires GPIb Receptors and von Willebrand Factor

Blood ◽

10.1182/blood.v93.2.564 ◽

1999 ◽

Vol 93 (2) ◽

pp. 564-570 ◽

Cited By ~ 67

Author(s):

S. Béguin ◽

R. Kumar ◽

I. Keularts ◽

U. Seligsohn ◽

B.S. Coller ◽

...

Keyword(s):

Von Willebrand Factor ◽

Thrombin Generation ◽

Platelet Rich Plasma ◽

Von Willebrand Disease ◽

Procoagulant Activity ◽

Increased Risk ◽

Coagulation Proteins ◽

A Minor ◽

Von Willebrand ◽

Willebrand Factor

Abstract Thrombin generation in platelet-rich plasma (PRP) involves complex interactions between platelets and coagulation proteins. We previously reported that the addition of fibrin to PRP enhances tissue-factor initiated thrombin generation by ≈ 40%, and the current studies were designed to assess the mechanism(s) underlying thrombin generation in the absence and presence of fibrin. Blocking platelet GPIIb/IIIa + vβ3 receptors with a monoclonal antibody (MoAb) inhibited basal thrombin generation, but did not affect the enhancement produced by fibrin. In contrast, blocking GPIb with any of three different MoAbs had no effect on basal thrombin generation, but essentially eliminated fibrin enhancement of thrombin generation. When thrombin generation was tested in PRP deficient in von Willebrand factor (vWF), both basal and fibrin-enhanced thrombin generation were markedly reduced, and the addition of factor VIII did not normalize thrombin generation. Botrocetin, which induces the binding of vWF to GPIb, enhanced thrombin generation. In all studies, the ability of PRP to support thrombin generation correlated with the production of platelet-derived microparticles and serum platelet-derived procoagulant activity. Thus, two separate mechanisms, both of which depend on vWF, appear to contribute to platelet-derived procoagulant activity: one is independent of fibrin and relies primarily on GPIIb/IIIa, but with a minor contribution from vβ3; and the other is fibrin-dependent and relies on GPIb. These data may have implications for understanding the mechanisms of the abnormalities in serum prothrombin times reported in Bernard-Soulier syndrome, hemorrhage in von Willebrand disease (vWD), and the increased risk of thrombosis associated with elevated vWF levels.

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Clinical significance of diagnostic algorithm in detection of mild hemostasis disorders in women with menorrhagia

Vojnosanitetski pregled ◽

10.2298/vsp180330123d ◽

2020 ◽

Vol 77 (6) ◽

pp. 601-606

Author(s):

Svetlana Djukic ◽

Nebojsa Andjelkovic ◽

Vladimir Vukomanovic ◽

Ivana Simic-Vukomanovic ◽

Aleksandar Djukic ◽

...

Keyword(s):

Menstrual Cycle ◽

Diagnostic Algorithm ◽

Von Willebrand Disease ◽

Reproductive Age ◽

Cycle Duration ◽

Coagulation Disorders ◽

Estimate Frequency ◽

Von Willebrand ◽

Semiquantitative Method

Background/Aim. Coagulation disorders could be a cause of menorrhagia in women of reproductive age. The aim of the study was to estimate frequency of coagulation disorders and design an appropriate algorithm for detection of coagulation disorders. Methods. We investigated coagulation in 115 women (36.1 ? 9.6 years) with anamnestic data of menorrhagia, verified using semiquantitative method ? Pictorial Bleeding Assessment Chart (PBAC) with score ? 100. Results. Menorrhagia was objectively verified in sixty-four women (55.7%) and in comparison with those with normal menstruation they had higher PBAC score of menstrual cycle [median (Md) = 150.0 vs. Md = 50.0; p < 0.001] but not its duration (7.2 ? 2.1 days vs. 7.3 ? 1.9 days; p > 0.05). Coagulation defects was found in 12 (10.4%) women ? decreased F IX: Ac in 4 (3.5%), decreased F VII: Ac in 1 (0.9%), decreased F X: Ac in 1 (0.9%), decreased F XI: Ac in 1 woman (0.9%), while 5 (4.3%) women matched criteria for mild von Willebrand disease (VWD) type 1. Women with coagulation disorders had prolonged prothrombin time (PT) [Md = 13.1 s, range: 12.2?14.8 s vs. Md = 12.5 s, range 10.6?18.3 s; p = 0.032]. Anemia was diagnosed in 61 (53.0%) women. The strongest predictor of the hemostasis disorder was menorrhagia (Quotient of probability 0.018), then anemia presence (12.43), P? (2.35), menstrual cycle duration (1.16) and the PBAC score (0.98). Conclusion. The results of the study indicate the need to form a diagnostic algorithm for hemostasis disorders, primarily VWD. Sophisticated analysis of hemostasis is required, especially if predictive factors of statistical models are detected: objectively verified menorrhagia, anemia, prolonged menstrual cycle, PBAC score > 100 and extended PT.

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Endothelial Microparticles Improve Primary and Secondary Hemostasis in Patients with Von-Willebrand Disease

Blood ◽

10.1182/blood.v118.21.2287.2287 ◽

2011 ◽

Vol 118 (21) ◽

pp. 2287-2287

Author(s):

Arne Trummer ◽

Ingvild Birschmann ◽

Sonja Werwitzke ◽

Arnold Ganser ◽

Andreas Tiede

Keyword(s):

Platelet Aggregation ◽

Human Plasma ◽

Thrombin Generation ◽

Conflicts Of Interest ◽

Annexin V ◽

Von Willebrand Disease ◽

Endothelial Microparticles ◽

Circulating Microparticles ◽

Von Willebrand

Abstract Abstract 2287 The bleeding phenotype in patients with von Willebrand disease (vWD) type 1 and 3 can usually be correlated to the amount and function of von Willebrand factor (vWF:Ag, vWF:RCo) but also to the impairment of functional coagulation assays: the ristocetin-induced platelet aggregation (RIPA) for primary hemostasis and the parameters of thrombin generation in a calibrated automated thrombogram (CAT, Thrombinoscope®) for secondary hemostasis. We therefore evaluated whether addition and depletion of endothelial microparticles (EMP) might influence these assays for samples of vWD patients. EMP were obtained from stimulated endothelial cell lines, isolated either by ultracentrifugation or magnetic beads selection for Annexin V and detected and quantified by flow cytometry after labelling for Annexin V and vWF. For CAT, EMP concentration was adjusted to 3×103/μl, for RIPA to 2×104/μl. vWD patient samples were diluted 3:1 or 2:1 with control buffer (HEPES/CaCl) or EMP solution. For thrombin generation, there was a significant shortening of lag time (6.8min vs 16.5min, p=.029) and time-to-peak (13.6min vs. 23.0min, p=.049) in vWD type 3 plasma (n=4) after addition of EMP using no reagent or MP reagent while there was no significant effect on these parameters in normal plasma. After addition of a tissue factor blocking antibody, the EMP effect could be largely reversed. For RIPA, we found a median maximum aggregation of 4% in vWD patients (n=6, vWF-Ag <40%). After addition of EMP, aggregation levels increased to 17.3%, which was significantly higher compared to control buffer (9.8%, p=.028), but lower compared to 250 IU/ml of a plasma-derived vWF product (Humate P®, 88.7%). To evaluate if significant amounts of vWF are bound to circulating microparticles in human plasma, we used magnetic bead selection to reduce MP counts in plasma of DDAVP stimulated vWD patients (n=10), because depletion of microparticles by 0.2μm filtration showed significant disruption of large vWF multimers. After MP reduction (1464/μl vs. 1863/μl, p=.036), both vWF:Ag (134.7% vs 161.1%, p=.005) and vWF:RCo (131.8% vs. 167.2%, p=.017) were significantly decreased. In summary, we could show that in vitro generated endothelial microparticles are able to improve thrombin generation and platelet aggregation in vWD patients and that significant amounts of vWF are bound to circulating microparticles in human plasma after DDAVP stimulation. Thus, in vitro generated EMP might have the potential to improve replacement therapy for vWD patients. Disclosures: No relevant conflicts of interest to declare.

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PATIENTS REFERRED FOR BLEEDING SYMPTOMS OF UNKNOWN CAUSE: DOES EVALUATION OF THROMBIN GENERATION CONTRIBUTE TO DIAGNOSIS?

Mediterranean Journal of Hematology and Infectious Diseases ◽

10.4084/mjhid.2016.014 ◽

2016 ◽

Vol 8 ◽

pp. 2016014 ◽

Cited By ~ 3

Author(s):

Elena Holm ◽

Eva Zetterberg ◽

Susanna Lövdahl ◽

Erik Berntorp

Keyword(s):

Thrombin Generation ◽

Assessment Tool ◽

Correlation Coefficients ◽

Von Willebrand Disease ◽

Assessment Tools ◽

Clotting Factors ◽

Viii And Ix ◽

Bleeding Score ◽

Von Willebrand ◽

Willebrand Factor

IntroductionPatients with mild to moderate bleeding symptoms referred for coagulation investigation sometimes never receive a definitive diagnosis. Bleed assessment tools have been developed and validated to assess the severity of symptoms. Global coagulation assays, e.g., the thrombin generation test (thrombogram) have a potential to identify hemostatic defects that are not detected in specific assays.Material and MethodsOne hundred and eighty-five patients referred to our centre because of bleeding symptoms were evaluated using the bleeding assessment tool (BAT) described by Tosetto and colleagues in 2006. Blood samples were investigated for thrombin generation (TG) capacity (Technoclone) , in platelet poor (PPP) plasma , and specific clotting factors, i.e, von Willebrand factor, factor VIII and IX, as well as INR, APTT, platelet count, and platelet adhesion.ResultsOf the 185 patients, five women were diagnosed with mild von Willebrand disease and one male with mild hemophilia A. The remaining 179 subjects (76% females and 24% males with average ages of 33 and 28 years, respectively) were evaluated further. In the total cohort and among women, peak TG, and lag time correlated with bleeding score (p=0.01 and p=0.04, respectively with correlation coefficients). No such correlations were found among males. Discussion and conclusion Although our study showed some correlation between TG and bleeding score, results are generally consistent with a previous report which failed to demonstrate the value of TG measurement in a similar setting. In conclusion, the complexity of the mechanisms underlying clinical bleeding complicates the ability to use TG tests as reliable predictors of bleeding. Mild congenital bleeding disorders, especially VWD, should be specifically screened for in patients with mild/moderate symtoms.

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Platelets compensate for poor thrombin generation in type 3 von Willebrand disease

Platelets ◽

10.1080/09537104.2019.1581922 ◽

2019 ◽

Vol 31 (1) ◽

pp. 103-111 ◽

Cited By ~ 4

Author(s):

Timea Szanto ◽

Vuokko Nummi ◽

Annukka Jouppila ◽

Herm Jan M. Brinkman ◽

Riitta Lassila

Keyword(s):

Thrombin Generation ◽

Von Willebrand Disease ◽

Type 3 ◽

Von Willebrand

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Fibrin-Dependent Platelet Procoagulant Activity Requires GPIb Receptors and von Willebrand Factor

Blood ◽

10.1182/blood.v93.2.564.402k04_564_570 ◽

1999 ◽

Vol 93 (2) ◽

pp. 564-570 ◽

Cited By ~ 2

Author(s):

S. Béguin ◽

R. Kumar ◽

I. Keularts ◽

U. Seligsohn ◽

B.S. Coller ◽

...

Keyword(s):

Von Willebrand Factor ◽

Thrombin Generation ◽

Platelet Rich Plasma ◽

Von Willebrand Disease ◽

Procoagulant Activity ◽

Increased Risk ◽

Coagulation Proteins ◽

A Minor ◽

Von Willebrand ◽

Willebrand Factor

Thrombin generation in platelet-rich plasma (PRP) involves complex interactions between platelets and coagulation proteins. We previously reported that the addition of fibrin to PRP enhances tissue-factor initiated thrombin generation by ≈ 40%, and the current studies were designed to assess the mechanism(s) underlying thrombin generation in the absence and presence of fibrin. Blocking platelet GPIIb/IIIa + vβ3 receptors with a monoclonal antibody (MoAb) inhibited basal thrombin generation, but did not affect the enhancement produced by fibrin. In contrast, blocking GPIb with any of three different MoAbs had no effect on basal thrombin generation, but essentially eliminated fibrin enhancement of thrombin generation. When thrombin generation was tested in PRP deficient in von Willebrand factor (vWF), both basal and fibrin-enhanced thrombin generation were markedly reduced, and the addition of factor VIII did not normalize thrombin generation. Botrocetin, which induces the binding of vWF to GPIb, enhanced thrombin generation. In all studies, the ability of PRP to support thrombin generation correlated with the production of platelet-derived microparticles and serum platelet-derived procoagulant activity. Thus, two separate mechanisms, both of which depend on vWF, appear to contribute to platelet-derived procoagulant activity: one is independent of fibrin and relies primarily on GPIIb/IIIa, but with a minor contribution from vβ3; and the other is fibrin-dependent and relies on GPIb. These data may have implications for understanding the mechanisms of the abnormalities in serum prothrombin times reported in Bernard-Soulier syndrome, hemorrhage in von Willebrand disease (vWD), and the increased risk of thrombosis associated with elevated vWF levels.

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Clinical Applications, Pitfalls, and Uncertainties of Thrombin Generation in the Presence of Platelets

Journal of Clinical Medicine ◽

10.3390/jcm9010092 ◽

2019 ◽

Vol 9 (1) ◽

pp. 92 ◽

Cited By ~ 2

Author(s):

Marina Panova-Noeva ◽

Paola E.J. van der Meijden ◽

Hugo ten Cate

Keyword(s):

Thrombin Generation ◽

Ex Vivo ◽

Platelet Rich Plasma ◽

Coagulation Factor ◽

Bleeding Risk ◽

Coagulation Factors ◽

Clinical Applications ◽

Von Willebrand Disease ◽

Von Willebrand

Platelet-dependent thrombin generation is a helpful tool to assess ex vivo the interaction between platelets and plasma coagulation factors in the initiation, amplification, and inhibition of thrombin generation (TG). This review article discusses the most relevant available data on the clinical applications of fluorogenic TG, the most widely used TG assay, performed in the presence of platelets, i.e., in platelet-rich plasma. With respect to prothrombotic states, arterial hypertension and obesity were the most prominent cardiovascular conditions linked to increased platelet-dependent TG. In addition, platelet-associated hypercoagulability, assessed by the TG assay, has been shown in individuals with active cancer. In terms of bleeding, platelet-dependent TG has been applied to assess bleeding risk in individuals with hemophilia, von Willebrand disease, and Glanzmann thrombasthenia as well as in subjects with other congenital or acquired coagulation factor deficiencies. In addition to risk prediction, a role of the TG assay has been suggested in monitoring antiplatelet therapy in prothrombotic conditions and replacement therapy in bleeding diathesis. Finally, for the routine clinical use and as a biomarker of disease development and progression, better standardization and clinical validation of platelet-dependent TG are still needed.

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