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ABSTRACT: The thrombin generation test (TGT) has demonstrated utility in evaluating overall hemostatic capacity both in bleeding and thrombotic disorders. Although the test is currently well accepted as a research tool, its role in clinical practice has not yet been defined through large prospective multicenter clinical studies. Such prospective studies have been limited by the lack of official standardization of the assay and its large inter-laboratory variability. This international study assessed the intra- and inter-assay imprecision of TGT as well as the inter-centre variability of results in one US and four European centres. Contact-inhibited plasmas from six healthy volunteers, one mild haemophilia A patient, and five patients with heterozygous prothrombin G20210 mutation were assayed. We demonstrated that, using identical equipement, standardized reagents, a carefully selected reference plasma for normalization of results and the same test procedure as described in our DVD, the assay variability was highly reduced compared to previously published data. Our results emphasize the importance of preheating on TGT results and the variability of the assay. In conclusion, our data demonstrated that the standardized TGT methodology evaluated in this study effectively reduces the variability of the assay to acceptable limits and may be used in clinical trials.
Thrombosis Research 08/2012; · 2.44 Impact Factor
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ABSTRACT: Methylene blue (MB) treatment of plasma is known to reduce the activity of clotting factors, but its effect on thrombin generation and clot formation is not well documented.
Individual clotting factors and inhibitors and global tests of thrombin generation and clot formation using rotational thrombelastometry (ROTEM) were assessed in a paired study of standard or MB plasma and cryoprecipitate (n = 20 each).
MB treatment resulted in a 10 percent reduction in endogenous thrombin potential and 30 percent decrease in peak thrombin as well as the expected 20 to 35 percent loss of Factor (F)VIII, fibrinogen, and FXI activity. MB treatment had no effect on the rate of clot formation and increased the clot firmness by 20 percent as assessed by ROTEM. There were minimal further changes in either coagulation factor levels or thrombin generation when thawed plasma was stored for an additional 24 hours. FVIII and fibrinogen content of MB cryoprecipitate was reduced by 30 and 40 percent, respectively, but this was not associated with altered clot time or rate of clot formation by ROTEM and only an 8 percent decrease in clot firmness.
It is concluded that MB treatment is associated with a reduction in the thrombin-generating capacity of plasma, but has very little effect on the strength of clot formation as assessed by thrombelastometry. The thrombin-generating capacity of standard and MB plasma is relatively unaltered by subsequent storage of thawed plasma at 4 degrees C for 24 hours.
Transfusion 02/2009; 49(4):696-703. · 3.22 Impact Factor
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ABSTRACT: Transmittance waveform (TW) analysis has been proposed as a method of both prediction and monitoring of non-overt and overt disseminated intravascular coagulation. This study assessed the use of the rapidTW of the activated partial thromboplastin time in the detection of sepsis in 49 consecutive neutropenic haemato-oncology patients. A slope 1 cut-off value of -0.050 was found to be optimum giving 85% sensitivity with 92% specificity and positive and negative predictive values of 62% and 98%, respectively. Furthermore a worsening slope 1 value at 24 hours was indicative of a 60% increase in mortality risk. Haemato-oncology patients have a significantly increased risk of developing sepsis during intensive chemotherapy, exacerbated by the resultant neutopenia. This sepsis may progress extremely rapidly and is associated with a high mortality. Early diagnosis is therefore critical and is currently made on a predominantly clinical basis with supporting microbiological evidence 2-3 days later. This study showed that TW offers an early marker, predictive of sepsis in neutropenic patients. It correlates with subsequent microbiological results and may identify patients at greater risk of clinical deterioration who may require more intensive early therapy or observation. It may also provide a useful marker to monitor the effects of treatment.
Thrombosis and Haemostasis 08/2008; 100(1):146-8. · 5.04 Impact Factor
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ABSTRACT: Haemostatic changes in septic patients are complex, with both procoagulant and anticoagulant changes. Thirty-eight patients with severe sepsis and 32 controls were investigated by coagulation screens, individual factor assays, calibrated automated thrombography (CAT), whole blood low-dose-tissue factor activated (LD-TFA) Rotem and LD-TFA waveform analysis. Thirty-six of 38 patients had an abnormal coagulation screen. The mean levels of factors II, V (P < 0.05), VII, X, XI and XII, antithrombin and protein C (P < 0.01) was decreased in sepsis compared with controls. The mean factor VIII and fibrinogen level (P < 0.001) was increased. CAT in platelet rich and poor plasma showed a prolonged lag time (P < 0.02), decreased peak thrombin (P < 0.02) and delayed time to peak thrombin (P < 0.001) in sepsis patients, however, the endogenous thrombin potential was equivalent in sepsis and controls. In LD-TFA Rotem, septic patients had delayed clot times (P = 0.04) but an increased maximum velocity of clot formation (P < 0.01) and area under the clot elasticity curve (P < 0.01). LD-TFA waveform analysis showed a delayed onset time but an increased rate of clot formation (P < 0.005). In conclusion, global tests of haemostasis suggest that in this patient group, activation of haemostasis is delayed but once initiated thrombin generation and clot formation are normal or enhanced.
British Journal of Haematology 10/2006; 135(2):220-7. · 4.94 Impact Factor
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ABSTRACT: The thrombin-platelet feedback loop was examined at low concentration tissue factor using calibrated automated thrombography in combination with the elimination of contact factor activation by corn trypsin inhibitor. The results indicated that, when contact factor activation was eliminated, the thrombin-platelet feedback loop was a major determinant of thrombin generating capacity and that platelets had a greater role in regulating the propagation of thrombin generation than its initiation. This method has potential application to the measurement of platelet-dependent thrombin generation in clinical diagnostic laboratories and hence the investigation of patients with apparent hypo- or hypercoagulable phenotypes.
British Journal of Haematology 09/2006; 134(3):323-5. · 4.94 Impact Factor
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ABSTRACT: In order to develop decentralized anticoagulant care by off-site blood sampling and transport of samples to a centralized laboratory for International Normalized Ratio (INR) determination we have performed a direct comparative study of INR stability. Analysis was performed daily for 5 d using nine thromboplastins. The overall mean difference of INR after 3 d was only 0.05 INR units for samples with a therapeutic INR. After 5 d there was a mean difference of 0.11 INR units with ‘non-Manchester’ reagents and 0.44 INR units with ‘Manchester’ reagents. With over-anticoagulated samples mean differences of 0.55–0.72 INR units were observed after 3 d and 1.16–2.46 INR units after 5 d. Although there was some variation in stability of results with different thromboplastins, the difference over time with each thromboplastin was much less than the difference between thromboplastins.In conclusion, there is no clinically significant change in INR when analysis is delayed for up to 3 d. Off-site blood sampling can accommodate a large increase in patient workload without a major revenue increase in primary care and with continued total quality management and central expert advice.
British Journal of Haematology 10/2003; 96(3):431 - 434. · 4.94 Impact Factor
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ABSTRACT: Stratification for risk of recurrence after a first episode of venous thromboembolism (VTE) would affect the duration of anticoagulant therapy. We aimed to determine the incidence of recurrence of VTE in relation to clinical risk factors and standard laboratory testing for heritable thrombophilic defects.
We established a database to prospectively follow-up a cohort of unselected patients who had had a first episode of objectively proven VTE. We excluded patients with malignant disease and antiphospholipid syndrome. All patients were offered testing for heritable thrombophilia.
At 2 years, the cumulative recurrence rate in 570 patients was 11%. Incidence was lowest after surgery-related VTE (0%) and highest after unprecipitated VTE (19.4%) (p<0.001). 85% of patients were tested for heritable thrombophilic defects. Recurrence rates were not related to presence or absence of laboratory evidence of heritable thrombophilia (hazard ratio 1.50 [95% CI 0.82-2.77]; p=0.187). In patients with a first event that was unprecipitated or was associated with a non-surgical trigger, recurrence rates did not differ in patients with or without thrombophilia (1.34 [0.73-2.46]; p=0.351).
In unselected patients who have had a first episode of VTE, testing for heritable thrombophilia does not allow prediction of recurrent VTE in the first 2 years after anticoagulant therapy is stopped. However, assessment of clinical risk factors associated with the first episode of VTE does predict risk of recurrence. Patients with postoperative VTE have a very low rate of recurrence.
The Lancet 08/2003; 362(9383):523-6. · 38.28 Impact Factor
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ABSTRACT: Three cohorts of patients with the factor V Leiden mutation were recruited independently (heterozygotes, homozygotes and combined thrombophilia). The antithrombotic efficacy of oral anticoagulation and the predictive value for recurrence of an idiopathic as opposed to a precipitated first event were determined. Idiopathic first events occurred at an older age than precipitated events (43 v 26 years, LR = 23.31, P < 0.001). None of the patients had a recurrent event while on warfarin but the median time to recurrence after stopping warfarin was 9 years (95%CI 0.7–17.3 years). The time to recurrence was shorter when the first event was idiopathic as opposed to precipitated (3.5 v 13 years, LR = 4.76, P = 0.029). A calculation of benefit to risk of oral anticoagulation with a target INR of 2.5 does not support the use of long-term therapy in all patients with the factor V Leiden mutation following a first thrombotic event.
British Journal of Haematology 12/2001; 100(4):764 - 768. · 4.94 Impact Factor
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ABSTRACT: A high prevalence of a common mutation in the Hfe gene (C282Y) has recently been reported in patients with the factor V Leiden mutation and a history of thrombosis. The aim of this study was to estimate the relative risk of venous thromboembolism in a large case–control study. 56/481 patients (11.6%) and 57/497 controls (11.5%) were heterozygous for the C282Y allele giving an odds ratio of 1.02 (95%CI 0.69–1.51). 12/81 patients with the factor V Leiden mutation were heterozygous for the C282Y allele compared to 1/13 controls, odds ratio 2.09 (95%CI 0.25–17.6). An analysis of a further group of patients and controls selected for the factor V Leiden mutation did not indicate a higher prevalence of the C282Y allele in symptomatic patients, odds ratio 0.17 (95%CI 0.34–0.81). This study does not support the hypothesis that the C282Y allele is an additional risk factor for venous thrombosis in patients with the factor V Leiden mutation.
British Journal of Haematology 03/1999; 105(1):95 - 97. · 4.94 Impact Factor
