Thrombin generation test in children and adolescents with chronic liver disease.

INTRODUCTION: The aim of this study was to evaluate if real-time thrombin generation assay provides additional information to assess the hemostatic balance in children with liver disease as compared to routine coagulation tests. PATIENTS AND METHODS: Sixty-three children with chronic liver disease were enrolled at our tertiary referral center for pediatric hepatology, eight whose routine coagulation tests gave abnormal results (Group A) and 55 whose test results were normal (Group B). Abnormal routine coagulation test was defined as at least one of the following: international normalized ratio≥1.4, activated partial thromboplastin time>44 sec., fibrinogen<1.5 g/L. Platelet-poor plasma was analyzed with the fluorogenic Calibrated Automated Thrombogram to test for thrombin generation, including endogenous thrombin potential. Further, routine coagulation tests and plasma levels of pro- and anticoagulant factors were measured. Twenty age-matched children without liver disease served as controls. RESULTS: The endogenous thrombin potential in the 55 patients with normal routine coagulation tests was not significantly different from that in controls. Group A had significantly lower levels not only of procoagulant factors (II, V, VII, X) but simultaneously also of the anticoagulant factors antithrombin, protein S free, and protein C. These patients had a reduced endogenous thrombin potential compared to Group B, in agreement with their routine coagulation test results. CONCLUSION: Thrombin generation analysis seems to give information on the hemostatic balance consistent with routine coagulation test results in children with liver disease. Further development and clinical evaluation of the method are warranted.

The international normalized ratio according to Owren in liver disease: interlaboratory assessment and determination of international sensitivity index.

INTRODUCTION: The international normalized ratio (INR) is used to prioritize liver disease patients for transplantation. Previous studies have shown high interlaboratory variability in Quick-based INR determinations in samples of patients with liver disease. We assessed Owren-based INR reagents for analyzing INR in patients with liver disease. Further, we determined the difference between international sensitivity index (ISI) for patients on vitamin K antagonists (ISIVKA) and ISI for patients with liver disease (ISIliver). PATIENTS AND METHODS: Twenty patients with liver disease were included, 10 with INR 1.8-3.6 (group A1) and 10 with INR 1.2-1.5 (group C1). Plasma from these patients was analyzed for Owren-based INR in eight Swedish laboratories using either of following reagents: SPA+, Owrens PT or Nycotest PT. To determine ISI liver, the reference thromboplastin RBT/05 and additional 41 patients with liver disease and 20 normal controls were included. ISIVKA was determined according to the WHO procedure. The difference between the ISIVKA and ISIliver was calculated. RESULTS: The coefficients of variance for the Owren based INR methods were 6.2% in group A1, 3.9 % in group C1 and 5.3% for all patients. The difference between ISIVKA and ISIliver were -0.4%, -0.7% and -0.2% for SPA+, Owrens PT and Nycotest PT respectively. CONCLUSIONS: Interlaboratory variation in INR analyses according to Owren in patients with liver disease is low and the difference between ISIVKA and ISIliver is below 10% with this method. ISIVKA can therefore be used in the INR calibration, for the Owren reagents studied, when analyzing plasma from patients with liver disease.

Mild primary hyperparathyroidism: vitamin D deficiency and cardiovascular risk markers.

CONTEXT: The extent and clinical significance of cardiovascular (CV) abnormalities associated with mild primary hyperparathyroidism (PHPT) are still matters for discussion. OBJECTIVE: The main objective of the present study was to evaluate biochemical CV risk markers in PHPT patients before and after parathyroidectomy (PTX) in comparison with controls. DESIGN AND SUBJECTS: In a prospective case-control design, 49 patients with PHPT and 49 healthy matched controls were included. METHODS: Blood pressure (BP), 25-OH-D, plasminogen activator inhibitor-1 activity, von Willebrand factor antigen, homocysteine, high-sensitivity C-reactive protein, IGF-I, and lipid profile were evaluated at baseline and 15 ± 4 months after PTX. RESULTS: At baseline, the level of 25-OH-D was significantly lower in patients compared with controls (40.1 ± 16.5 vs. 64.6 ± 20.8 nmol/liter, P < 0.001) and increased after PTX (58.9 ± 19.5, P < 0.001). Postoperatively, 25-OH-D was inversely correlated to the PTH level (r = -0.34; P < 0.05). Systolic BP (127.2 ± 17.4 vs. 119.3 ± 12.5 mm Hg, P < 0.05) and triglyceride (TG; 1.04 ± 0.60 vs. 0.86 ± 0.43 mmol/liter, P < 0.05) were higher in patients compared with controls and decreased slightly in patients after PTX (BP, 124.4 ± 16.8 mm Hg, and TG, 0.94 ± 0.50 mmol/liter, P < 0.05). Otherwise, there were no intergroup differences in coagulation, inflammatory, metabolic, and lipid status. CONCLUSIONS: Except for a lower 25-OH-D level and slightly higher systolic BP and TG levels, patients with mild PHPT without other CV risk factors did not differ from healthy controls as regards biomarkers predicting CV diseases. PTX had an overall positive effect on TG level, BP, and vitamin D status.

Effects of the oral, direct thrombin inhibitor dabigatran on five common coagulation assays.

Dabigatran is an oral, reversible thrombin inhibitor that has shown promising results in large clinical trials. Laboratory monitoring is not needed but the effects on common coagulation assays are incompletely known. Dabigatran was added to plasma from healthy subjects in the concentration range 0-1,000 µg/l and analysed using several reagents for activated thromboplastin time (APTT), prothrombin time (PT), fibrinogen, antithrombin, and activated protein C resistance. Typical trough concentrations are about 50 µg/l, peak concentrations 100-300 µg/l. At 100 µg/l all APTT-results were prolonged. The concentration required to double APTT ranged between 227 and 286 µg/l, the responses for all five reagents were similar. PT-reagents were much less affected with almost no samples above INR 1.2 at 100 µg/l. The effect was sample dilution dependent with PT Quick type more sensitive than PT Owren type methods. If a patient on dabigatran has prolonged APTT, >90 seconds, and Quick PT INR>2 or Owren PT INR>1.5 over-dosing or accumulation of dabigatran should be considered. Two of four fibrinogen reagents underestimated the fibrinogen concentration considerably at expected peak concentration. Methods based on inhibition of thrombin over-estimated the antithrombin concentration, but not Xa-based. The APC-resistance methods over-estimated the APC-ratio, which may lead to miss-classification of factor V Leiden patients as being normal. Different coagulation assays, and even different reagents within an assay group, display variable effects at therapeutic concentrations of dabigatran. Some of these assay variations are of clinical importance, thus knowledge is needed for a correct interpretation of results.

Identifying hypocoagulable states with a modified global assay of overall haemostasis potential in plasma.

To test the sensitivity of the global assay of overall haemostasis potential (OHP) in detecting hypocoagulation, the OHP was assayed in plasma containing exogenous thrombin (0.04 IU/ml), tissue-plasminogen activator (330 ng/ml), Ca and a platelet reagent. Commercial plasmas with factor II, V, VIII, IX, X, XI, XII or VII deficiency were mixed with normal plasma in different proportions to imitate different severities. Samples from patients with haemophilia and factor XII deficiency were also examined. No clot was found in the absence of factor II/factor X, indicating that the tiny dose of thrombin worked solely as a trigger for the intrinsic pathway activation. Changed levels of the investigated coagulants, apart from factor XII, influenced the outcome. OHPs were decreased in patients with haemophilia but were unchanged or even increased in those with factor XII deficiency. This modified OHP method may therefore be useful for estimating the bleeding tendency in haemophilic patients and to find suitable doses and intervals for prophylactic treatment. It may also be of use in investigations of the effect of antifibrinolytic drugs as well as for identifying a thrombotic tendency in patients with factor XII deficiency. For detection of other coagulation factor deficiencies, our investigations with the commercial plasmas suggest that the OHP assay is also valuable, especially when the intrinsic pathway of the coagulation cascade is impaired.

The complex between tPA and PAI-1: risk factor for myocardial infarction as studied in the SHEEP project.

INTRODUCTION: The tPA/PAI-1 complex seems to be an important biochemical marker for myocardial reinfarction. Therefore we explored the distribution, correlation and interaction of plasma concentrations of tPA/PAI-1 complex in all available patients and matched controls in the Stockholm Heart Epidemiology Program (SHEEP). METHODS AND PATIENTS: The SHEEP study is a case control study of 2246 patients with a first myocardial infarction (MI), of which 1267 surviving patients were subjected to blood sampling about 3 months after MI and compared with a control group, matched for age, sex and living area within the Stockholm County. The study consists of 886 (591 men and 295 women) patients and 1198 (753 men and 445 women) matched controls, who were all analysed for plasma tPA/PAI-1 complex. RESULTS: The plasma concentration of tPA/PAI-1 complex was significantly associated with the risk for MI, for both genders. Synergistic interactions were observed in men for the co exposure to high plasma tPA/PAI-1 complex concentrations in combination with smoking (OR=4.6) or diabetes mellitus (OR=7.9). Synergism was also seen in combination with exposure to serum hypercholesterolemia or increased levels of apolipoprotein B. An antagonistic effect of the co exposure to high tPA/PAI-1 complex and hypertension was found among men with a similar tendency among women, but an antagonistic effect of increased waist/hip ratio was only observed among the women. CONCLUSIONS: Measuring the plasma concentration of tPA/PAI-1 complex might be of practical value in assessing risk of MI for both genders, especially in those who are smokers or in patients with manifest diabetes mellitus.

PAI-1 level and the PAI-1 4G/5G polymorphism in relation to risk of non-fatal myocardial infarction: results from the Stockholm Heart Epidemiology Program (SHEEP).

This study examines the relationship between plasma Plasminogen Activator Inhibitor-1 (PAI-1) activity and the PAI-1 4G/5G polymorphism, and their association with the risk of myocardial infarction (MI). Furthermore, this study explores whether a high level of PAI-1 or whether the PAI-1 4G/5G polymorphism synergistically interacts with any established environmental risk factor for MI. This case-referent study of subjects aged 45-70 and living in Stockholm includes 851 men and 361 women with first-time MI and 1051 men and 505 women who were randomly chosen as referents from a population register. The adjusted odds ratio (OR) of MI was 1.9 (95% confidence interval [CI] 1.4-2.8) for men with a plasma PAI-1 activity level greater than the 90 th percentile value of referents. The corresponding OR for women was 1.5 (95% CI 0.9-2.5). A strong indication of synergistic interaction was found in men for the co-exposure to high plasma PAI-1 activity and current smoking, an adjusted synergy index score of 3.9 (95% CI 1.2-13.2). In women, the 4G allele was associated slightly with an increased risk of MI, OR 1.4 (95% CI 1.0-1.9). This association was not found in men. There were no clear indications of synergistic interaction effects involving the PAI-1 4G/5G polymorphism and the environmental exposures considered (cigarette smoking, physical inactivity, overweight, diabetes mellitus, hypercholesterolaemia, hypertension, high C-reactive protein and hypertriglyceridaemia).