High proportion of patients with bleeding of unknown cause in persons with a mild-to-moderate bleeding tendency: Results from the Vienna Bleeding Biobank (VIBB).

INTRODUCTION: Data on clinical characteristics and the prevalence of underlying coagulopathies in patients with mild-to-moderate bleeding disorders (MBDs) are scarce. AIM: We established the Vienna Bleeding Biobank (VIBB) to characterize and thoroughly investigate Austrian patients with MBDs. RESULTS: Four hundred eighteen patients (female = 345, 82.5%) were included. A platelet function defect (PFD) was diagnosed in 26 (6.2%) and a possible PFD in 30 (7.2%) patients. Eight patients (1.9%) were diagnosed with von Willebrand disease (VWD) (type 1 n = 6; type 2 n = 2), and 29 patients had low VWF (30-50 IU/dL). Deficiencies in factor VIII, IX, XI or XIII were found in 11 (2.6%), 3 (0.7%), 3 (0.7%) and 1 patient(s), 2 patients had dysfibrinogenaemia, and further 2 had possible PFD and FXI deficiency. Probable causal mutations were detected in 8 of 11 patients with FVIII deficiency, 2 of 3 patients with FIX deficiency and 2 of 8 patients with VWD. Three hundred three patients (72.5%) had normal results in the coagulation assays and were categorized as patients with bleeding of unknown cause (BUC). The bleeding score did not differ between patients with and without established diagnosis. A diagnosis of a bleeding disorder was more frequently made in men than in women (49.3% vs 22.9%). Male sex (OR 3.55, 95% CI: 2.02-6.22; P < .001) and blood group 0 (OR 1.86, 95% CI: 1.17-2.94; P = .008) were independently associated with diagnosis of a bleeding disorder. CONCLUSION: The high rate of patients with BUC despite in-depth haemostatic assessment underlines the incompleteness of available routine laboratory tests. Males with MBDs were more likely to be diagnosed with an established bleeding disorder than females.

Von Willebrand factor indicates bacterial translocation, inflammation, and procoagulant imbalance and predicts complications independently of portal hypertension severity.

BACKGROUND: Elevated plasma von Willebrand factor antigen (vWF) has been shown to indicate the presence of clinically significant portal hypertension, and thus, predicts the development of clinical events in patients with cirrhosis. AIM: To investigate the impact of bacterial translocation and inflammation on vWF, as well as the association between vWF and procoagulant imbalance. Moreover, we assessed whether vWF predicts complications of cirrhosis, independent of the severity of portal hypertension. METHODS: Our study population comprised 225 patients with hepatic venous pressure gradient (HVPG) ≥ 10 mm Hg without active bacterial infections or hepatocellular carcinoma. RESULTS: vWF correlated with markers of bacterial translocation (lipopolysaccharide-binding protein [LBP; ρ = 0.201; P = 0.021]), inflammation (interleukin 6 [IL-6; ρ = 0.426; P < 0.001] and C-reactive protein [CRP; ρ = 0.249; P < 0.001]), and procoagulant imbalance (factor VIII/protein C ratio; ρ = 0.507; P < 0.001). Importantly, the associations between vWF and these parameters were independent of HVPG. Moreover, vWF (per 10%) independently predicted variceal bleeding (hazard ratio [HR]: 1.08 [95% confidence interval (95% CI): 1.01-1.16]; P = 0.023), requirement of paracentesis (HR: 1.05 [95% CI: 1.01-1.1]; P = 0.023) and bacterial infections (HR: 1.04 [95% CI: 1-1.09]; P = 0.04) including spontaneous bacterial peritonitis (HR: 1.09 [95% CI: 0.999-1.18]; P = 0.053) on a trend-wise level. After backward elimination, vWF (HR: 1.05 [95% CI: 1.02-1.08]; P = 0.003) and CRP (per 10 mg/L; HR: 1.53 [95% CI: 1.14-2.05]; P = 0.005) remained in the final model for transplant-free mortality. Finally, the independent prognostic value of vWF/CRP groups for mortality was confirmed by competing risk analysis. CONCLUSION: Our results demonstrate that vWF is not only a marker of portal hypertension but also independently linked to bacterial translocation, inflammation and procoagulant imbalance, which might explain its HVPG-independent association with most clinical events. Prognostic groups based on vWF/CRP efficiently discriminate between patients with a poor 5-year survival and patients with a favourable prognosis.

Ristocetin-induced platelet aggregation for monitoring of bleeding tendency in CLL treated with ibrutinib.

Bleeding because of impaired platelet function is a major side effect of the Bruton's tyrosine kinase (BTK) inhibitor ibrutinib. We quantitatively assessed ristocetin-induced platelet aggregation (RIPA) in 64 patients with chronic lymphocytic leukemia (CLL) under ibrutinib at 287 time points. Eighty-seven bleeding episodes in 39 patients were registered (85 Common Toxicity Criteria (CTC) grade 1 or 2, 2 CTC grade 3) during a median observation period of 10.9 months. At times of bleeding, RIPA values were significantly lower (14 vs 28 U; P<0.0001). RIPA was impaired in patients receiving concomitant antiplatelet therapy or anticoagulation (14 vs 25 U, P=0.005). A gradual decline of median RIPA values was observed with increasing bleeding severity. Importantly, no CTC grade 2 or 3 bleeding were observed with RIPA values of >36 U. Sequential monitoring indicated a decrease of RIPA values from a median of 17 to 9 U within 2 weeks after initiation of treatment as well as an increase above the critical threshold of 36 U within 7 days when ibrutinib was paused. Low RIPA values were similar during treatment with another BTK inhibitor, CC292. Quantitative assessment of platelet function is a practical tool to monitor bleeding tendency under BTK-inhibitor therapy.

Comparison between the impact of morning and evening doses of rivaroxaban on the circadian endogenous coagulation rhythm in healthy subjects.

UNLABELLED: ESSENTIALS: It is unknown whether single rivaroxaban doses should best be administered in the morning or evening. Circadian rhythm of coagulation/fibrinolysis was measured after morning or evening intake of rivaroxaban. Evening intake of rivaroxaban leads to prolonged exposure to rivaroxaban concentrations. Evening intake of rivaroxaban better matches the morning hypofibrinolysis. BACKGROUND: A circadian variation of the endogenous coagulation system exists with hypercoagulability and hypofibrinolysis and a corresponding peak of cardiovascular thromboembolic events in the morning. So far, no information is given as to whether single daily doses of the new oral anticoagulant drug rivaroxaban should best be administered in the morning or the evening. MATERIALS AND METHODS: Sixteen healthy male or female volunteers with a mean age of 26 ± 7 years were included in this randomized, controlled, analyst-blinded cross-over clinical trial. All subjects were given three morning and three evening single doses of 10 mg rivaroxaban. Circadian rhythms of prothrombin fragment 1 + 2, plasminogen activator inhibitor, and plasmin-antiplasmin complex were measured before any medication intake, as well as after morning or evening medication intake. Rivaroxaban concentrations were determined by an anti-activated factor X assay and liquid chromatography-mass spectrometry. MAIN RESULTS: Concentrations of rivaroxaban were higher 12 h after evening intake of rivaroxaban than 12 h after morning intake (53.3 ng mL(-1) [95% confidence interval 46.0-67.8] vs. 23.3 ng mL(-1) [19.4-29.1, respectively]). Rivaroxaban intake in the evening reduced morning F1+2 concentrations better at 8:00 AM than did administration on awakening (85 ± 25 nmol L(-1) vs. 106 ± 34 nmol L(-1) , CI: 9.4-32.1). In addition, this suppression effect was longer lasting after evening intake. CONCLUSIONS: Evening intake of rivaroxaban leads to prolonged exposure to rivaroxaban concentrations and better matches the morning hypofibrinolysis. These results might help to further improve the efficacy and safety of rivaroxaban treatment.

Longitudinal analysis of hemostasis biomarkers in cancer patients during antitumor treatment.

UNLABELLED: ESSENTIALS: Hemostasis biomarkers impact thrombosis occurrence and survival in cancer patients. We performed a longitudinal analysis of hemostatic parameters in 112 cancer patients. Hemostatic parameters are associated with disease state, patients' prognosis, and the risk of VTE. The procoagulant state exists not only at diagnosis, but also during the course of disease. BACKGROUND: Hemostasis biomarkers are known to have an impact on venous thromboembolism (VTE) occurrence and survival in cancer patients. OBJECTIVES: As there are almost no data on longitudinal changes, we aimed to evaluate those in the present prospective observational study during chemotherapy and the course of disease. PATIENTS/METHODS: Patients with cancer of the brain (n = 39), lung (n = 41), colon (n = 15) or pancreas (n = 17) were included before initiation of antitumor therapy. Blood samples for determination of factor VIII, thrombin peak height, D-dimer, F1 + 2 , fibrinogen and soluble P-selectin (sP-selectin) were drawn on a monthly basis. The study endpoints were death, VTE occurrence, or completion of the study period. RESULTS: Overall, 546 blood samples of 112 patients were analyzed. D-dimer and sP-selectin levels were significantly higher in patients with distant metastasis than in those without. Patients with complete remission had significantly lower levels of F1 + 2 , D-dimer and fibrinogen. Peak height thrombin levels showed a decrease over time in all tumor types. Levels of biomarkers behaved differently in the various tumor types. Patients who developed VTE (n = 14) showed increasing levels of FVIII, sP-selectin, and D-dimer. At the last blood sampling time-point before VTE occurrence, in 13 patients the D-dimer level was above the median, and in seven of these patients it was even above the 75th percentile; however, the individual course was highly variable. Regarding survival, steadily increased FVIII, sP-selectin and D-dimer levels were associated with higher mortality. CONCLUSIONS: Hemostatic parameters show an association with disease state, prognosis, and the risk of VTE, not only at diagnosis, but also during the course of antineoplastic treatment.

Parameters influencing FVIII pharmacokinetics in patients with severe and moderate haemophilia A.

In haemophilia A patients factor VIII (FVIII) recovery and half-life can vary substantially. There are parameters known to modulate FVIII pharmacokinetics (PK), but they explain only about 34% of the variability. The aim of this study was to identify new parameters that influence FVIII PK and thus to expand the current knowledge. FVIII PK were determined in 42 haemophilia A patients (37 severe, 5 moderate) without inhibitor. Patients' characteristics and laboratory parameters were evaluated for an association with FVIII PK. We analysed plasma levels of low-density lipoprotein receptor-related protein 1 (LRP1) and protein C (PC) activity, which had been hypothesized to influence FVIII activity. Furthermore, four variations in intron 6 of the LRP1 gene, which had been shown to influence LRP1, were investigated. FVIII half-life differed widely from 6.2 to 20.7 h, with a median of 10.0 h. Patients with blood group O had shorter FVIII half-life compared to patients with non-O blood group (median FVIII half-life 9.0 h vs. 10.4 h, P = 0.018). Age was significantly associated with FVIII half-life (r = 0.32, P = 0.035). Besides age, also VWF antigen (r = 0.52, P < 0.001) and blood group (r = -0.37, P = 0.015) was associated with FVIII half-life. No correlation was found with FVIII- or LRP1-genotype, LRP1 or PC concentrations. Our data showed large differences in FVIII PK between individual patients and revealed age, blood group and VWF levels as important determining factors for FVIII half-life. FVIII genotype or levels of LRP1 or PC had no influence on FVIII PK.

Lupus anticoagulant and thrombosis in splenic marginal zone lymphoma.

INTRODUCTION: Splenic marginal zone lymphoma (SMZL) is a rare low-malignant Non-Hodgkin lymphoma (NHL), in which immune mediated paraneoplastic phenomena such as autoimmune hemolytic anemia (AIHA), autoimmune thrombocytopenia (ITP) and C1 esterase inhibitor deficiency are relatively common. MATERIALS AND METHODS: We performed a multicenter retrospective study in 70 patients on the prevalence and clinical features of antiphospholipid antibodies (aPLA) in SMZL. RESULTS AND CONCLUSIONS: Nine patients (13%) had the diagnosis of a lupus anticoagulant (LA). The occurrence of venous thromboembolic events was significantly higher in LA positive patients compared to LA negative patients (4/9 [44%] vs 5/61 [8%], p = 0.002), especially within 12 months after splenectomy (3/6 [50%] vs 2/28 [7%], p = 0.007). None of the patients with LA had a persistent complete remission of LA after splenectomy, but complete remission of LA was achieved in 2/2 patients after rituximab-bendamustine immuno-chemotherapy. In conclusion, our data show a relatively high prevalence of aPLA in SMZL and an increased risk of postsplenectomy thrombosis in these patients. The fact that rituximab-bendamustine was effective for eradicating LA may be considered as an argument for using immuno-chemotherapy as first line therapy in SMZL patients with LA.

No impact of endogenous prothrombotic conditions on the risk of central venous line-related thrombotic events in children: results of the KIDCAT study (KIDs with Catheter Associated Thrombosis).

BACKGROUND AND OBJECTIVE: Central venous lines (CVLs) are the major exogenous risk factor for deep venous thrombosis (DVT) in children. The study objective was to assess whether endogenous prothrombotic conditions contribute to the risk of CVL-related DVT in children. METHODS: This was a cohort study of consecutive children with heart disease requiring CVLs for perioperative care. CVLs were inserted percutaneously in the upper venous system and patients received prophylaxis with continuous unfractionated heparin (50 u kg(-1)  d(-1) ). Blood samples to test for prothrombotic conditions were collected prospectively and assayed in a blinded fashion. Outcome assessment was by screening for DVT by venography, venous ultrasound and echocardiography. RESULTS: The study population consisted of 90 children, median age 2.7 years (0 months-18 years). Prevalence rates of antithrombin deficiency, protein C deficiency, protein S deficiency, heterozygous factor V Leiden, prothrombin G20210A mutation, methylentetrahydrofolate C677TT genotype, hyperhomocysteinemia, lupus anticoagulant, anticardiolipin antibodies and increased levels of lipoprotein (a) were within the range reported for the general population. At least one prothrombotic condition was present in 38% of children and combined abnormalities in 8%. The incidence of DVT was 28% (25/90), and most DVTs were asymptomatic. None of the prothrombotic conditions showed a significant association with DVT. The population attributable risk (i.e. the risk of DVT in the overall population attributable to a specific condition) did not exceed 2.2%. CONCLUSION: Prothrombotic conditions did not have an important impact on the risk of DVT in children with short-term CVLs. The results of the study suggest that screening for prothrombotic conditions is not justified in this setting.

The role of fibrinogen plasma levels, the -455G>A fibrinogen and the factor XIII A subunit (FXIII-A) Val34Leu polymorphism in cancer-associated venous thrombosis.

Venous thromboembolism (VTE) is a life-threatening complication in cancer patients. Identification of risk factors has been in focus in the past years. Functional single nucleotide polymorphisms (SNP) of coagulation factors known to influence the concentration or function may be considered to influence the risk of VTE in cancer patients. We evaluated the influence of fibrinogen plasma levels, the -455G>A SNP in the fibrinogen beta gene and the Val34Leu (163G>T) SNP in the factor XIII A-subunit (FXIII-A) gene on the risk of VTE. In 1,079 tumour patients recruited for the prospective Vienna Cancer and Thrombosis Study (CATS) fibrinogen levels were determined by the Clauss method. The FXIII-A Val34Leu and the fibrinogen -455G>A SNPs were tested by allele-specific PCR. The median follow-up time was 604 days, 83 thrombotic events occurred. The median fibrinogen level was 381 mg/dl (25th-75th percentile: 312 to 467). In a multivariable Cox model adjusted to chemotherapy, surgery, radiotherapy, age and sex, neither the fibrinogen concentration (hazard ratio [HR] =1.05, confidence interval [CI] 0.839-1.310 p=0.68), nor the -455G>A SNP (HR=0.77, 95%CI 0.491-1.197; p=0.24), nor the Val34Leu SNP (HR=0.99, 95%CI 0.646-1.542 p=0.99) were associated with occurrence of VTE. The fibrinogen concentration was not significantly different among the fibrinogen -455G or A genotype carriers (p = 0.33). Disseminated intravascular coagulation was observed in only five patients, none of these developed VTE. In conclusion, fibrinogen plasma levels, the fibrinogen -455G>A and the FXIII-A Val34Leu polymorphisms were not associated with VTE in our study.

Thrombin generation in morbid obesity: significant reduction after weight loss.

BACKGROUND: Patients with morbid obesity (MO; body mass index > 40 kg m(-2)) suffer from an increased risk of cardiovascular disease, stroke, venous thromboembolism and all-cause mortality. OBJECTIVES: Because weight loss by bariatric surgery reduces cardiovascular and all-cause mortality, we hypothesized that the plasmatic clotting system might be involved in cardiovascular risk. PATIENTS/METHODS: Thirty-six MO patients [mean age 42 (+/-13) years; 29 female) were investigated before and 2 years after bariatric surgery. Thrombin generation was measured with a commercially available assay (Technothrombin-TGA,Technoclone). Metabolic parameters and parameters of the hemostatic system, such as tissue factor (TF), TF pathway inhibitor (TFPI), plasminogen activator inhibitor-1 (PAI-1) and prothrombinfragment 1.2 (F1.2), were determined. To investigate associations of changing parameters, deltas were calculated. RESULTS: Metabolic parameters improved with a mean weight loss of 41 (+/-19) kg. Postoperatively, the lag phase was significantly extended compared with preoperative values [median (25th-75th percentile), 7 (4-12) vs. 12 (7-19) min, P = 0.005]. Peak thrombin decreased after weight loss from 345 (232-455) to 282 (111-388) nm (P = 0.015) and the area under the curve from 3962 (3432-5023) to 3227 (2202-4030) nm thrombin (P < 0.001). TF, PAI-1 and F1.2 significantly decreased after weight loss. Analyses of the deltas showed a significant correlation between peak thrombin and total cholesterol (r = 0.50), triglycerides (r = 0.46) and HbA1c (r = 0.55). Moreover, an inverse correlation was found between insulin resistance and the lag phase (r = -0.46). CONCLUSION: Thrombin generation, a marker of the overall coagulation potential, decreased significantly with weight reduction. This might, at least in part, explain the decreased risk of cardiovascular disease after bariatric surgery.

Determinants of factor VIII plasma levels in carriers of haemophilia A and in control women.

Factor VIII (FVIII) levels show a considerable variability in female carriers of haemophilia A. Presently, the reasons for this are poorly understood. The aim of the study was to elucidate the influence of genetic and non-genetic parameters on FVIII plasma levels in carriers (n = 42). Results were compared with age-matched healthy women without carriership of haemophilia A (n = 42). Each carrier was tested for the family-specific mutation, ABO blood group, FVIII level, von Willebrand factor (VWF) antigen and activity and C-reactive protein (CRP). FVIII levels were lower in carriers compared to non-carriers [74% (51-103) vs. 142% (109-169), P < 0.001]. No statistically significant differences were observed between the two groups with respect to VWF activity, prothrombin-time, hs-CRP, fibrinogen, body mass index (BMI), age and smoking status as well as the distribution of ABO blood groups. In non-carriers, FVIII was statistically significantly correlated with BMI, activated partial thromboplastin time (APTT), VWF antigen, hs-CRP and fibrinogen. In carriers, significant correlations between FVIII and APTT, VWF antigen and activity were found, whereas BMI, hs-CRP or fibrinogen did not correlate with FVIII. In non-carriers, the association of FVIII with ABO blood groups was statistically significant (P = 0.006), but not in carriers of haemophilia A (P = 0.234). The type of FVIII gene mutation did not influence FVIII levels. Carrier status is the major determinant of a carrier;s FVIII plasma level. Factors known to influence FVIII levels in the general population do not significantly affect FVIII activity in carriers, neither does the type of mutation influence FVIII levels.

Persistent arterial stiffness and endothelial dysfunction following successful pancreas-kidney transplantation in Type 1 diabetes.

OBJECTIVE: Successful simultaneous pancreas-kidney transplantation (SPK) in Type 1 diabetic (T1DM) patients results in improved cardiovascular outcome and survival. However, it is doubtful whether the impairment of cardiovascular and endothelial function in T1DM can be completely reversed. METHODS: Pulse-wave velocity, stroke volume, heart rate, serological markers of endothelial dysfunction (soluble intercellular, vascular cell-adhesion molecules, E-selectin, and plasminogen-activator-inhibitor-1) were measured in 10 T1DM patients after SPK with non-diabetic glucose levels, 10 T1DM patients with poor [T1DM>8; glycated haemoglobin (HbA1c)>8%], and 10 with good glucose control (T1DM<7, HbA1c<7%), in 6 non-diabetic patients after kidney transplantation (KT) and 9 non-diabetic control subjects (CON), matching for major anthropometric characteristics. RESULTS: Pulse-wave velocity was increased in SPK (P < 0.02 vs. CON, KT, T1DM<7) and in T1DM>8 (P < 0.02 vs. T1DM<7). Systolic blood pressure was increased in SPK (P < 0.05 vs. CON). Stroke volume was reduced in SPK, T1DM>8 and T1DM<7 and KT (P < 0.01 vs. CON). Heart rate was elevated in SPK and in T1DM>8 (P < 0.0003 vs. CON and T1DM<7). In SPK, soluble intercellular and vascular cell-adhesion molecules were 100% and 44% higher (P < 0.03 vs. CON), respectively, while plasminogen-activator-inhibitor-1 was decreased in SPK (P < 0.02 vs. CON). CONCLUSION: T1DM patients after SPK experience arterial stiffness, a higher heart-rate and blood pressure, reduced stroke volume and serological signs of endothelial dysfunction. Thus, functional and structural cardiovascular alterations as a result of glucotoxicity, uraemia and hypertension in T1DM might not be completely resolved by SPK.

High factor VIII levels independently predict venous thromboembolism in cancer patients: the cancer and thrombosis study.

OBJECTIVE: Patients with cancer are at an increased risk for venous thromboembolism (VTE). Clotting factor VIII activity (FVIII) has been established as risk factor of primary and recurrent VTE. We investigated FVIII as predictive parameter of VTE in cancer patients. METHODS AND RESULTS: The prospective observational Cancer and Thrombosis Study (CATS) includes patients with newly diagnosed cancer or disease progression, study end point is symptomatic VTE. FVIII was measured on a Sysmex CA 7000 analyzer. Data on 840 patients (median age: 62 years, 25th to 75th percentile 53 to 68, 378 women) were available for analyses, of these 111 patients had hematologic malignancies and 729 solid cancer. During a median observation time of 495 days 62 events occurred. Cumulative probability of VTE after 6 months was 14% in patients with elevated FVIII-levels and 4% in those with normal levels (P=0.001). The association was strongest in younger patients: whereas in 40-year-old patients a 2-fold VTE risk per factor VIII increase of 20% was observed (HR=2.0 [95% CI: 1.5 to 2.7], P<0.0001), this association was still present but attenuated in older patients. CONCLUSIONS: FVIII is independently associated with an increased risk of VTE in cancer patients. The association between FVIII and VTE risk declines with increasing age.

Mechanisms underlying acquired von Willebrand syndrome associated with an IgM paraprotein.

Acquired von Willebrand (vW) syndrome is a rare bleeding disorder which is frequently associated with immunological, malignant or cardiovascular disorders. The underlying pathomechanisms, particularly in patients with IgM monoclonal gammopathies, often remain unknown. We report a patient with indolent small B-cell lymphoma (immunocytoma) and plasmacytic differentiation with an IgM kappa paraprotein who was admitted with retroperitoneal haematoma. Medical history and coagulation testing were consistent with acquired vW syndrome. vW immunohistochemistry showed normal cytoplasmic labelling of endothelial cells and megakaryocytes, whereas the lymphomatous infiltrate was negative. Acquired vW syndrome due to adsorption of vW factor on malignant cells was thus excluded. In the multimeric analysis, all multimers were present similar to that in type 1 vW syndrome, but the triplet structures were blurred. The bands on serum immunofixation electrophoresis were also atypically broadened, which suggested complex formation between the IgM and vW factor. Immunoprecipitation studies showed that the 176-kDa proteolytic fragment of vW factor co-precipitated with the IgM paraprotein in the patient but not in the controls, suggesting a specific interaction between vW factor and the paraprotein in the patient. The patient required surgery and was successfully managed by chemotherapy consisting of rituximab and fludarabin as well as plasma exchange.

Thrombin generation in haemophilia A patients with factor VIII inhibitors after infusion of recombinant factor VIIa.

BACKGROUND: Development of factor VIII inhibitors is a serious complication in haemophilia A patients. Recombinant factor VIIa (rVIIa) is clinically effective, but its effects on haemostatic system need still to be fully elucidated. MATERIAL AND METHODS: In an open controlled study, we measured thrombin generation (peak thrombin) in venous blood and prothrombin fragment F1 + 2 (F1 + 2) and D-dimer in venous and in shed blood in five haemophilia A patients with inhibitors before and after rVIIa infusion. A total of five healthy individuals who did not receive rVIIa served as controls. RESULTS: At baseline, patients had lower mean (min-max) peak thrombin levels than controls [0.12 (0.0-0.6) vs. 186.9 (116.0-254.4) nM, P = 0.001]. After infusion, peak thrombin levels increased in average to 40.7 (28.3-51.6) nM, which translates into 80.2% (95% CI 65.4-88.6%) lower levels compared to that of controls. Mean (min-max) F1 + 2 levels in venous blood did not differ significantly between patients and controls [160.7 (89.8-331.3) vs. 160.8 (104.4-242.3) pmol L(-1)], but increased in average (min-max) by 39.4% (14.1-58.5%) after infusion. In blood emerging from incisions made to determine the bleeding (shed blood), F1 + 2 levels were lower in patients than controls [1383.3 (906.4-2044.6) vs. 2981.7 (1610.0-4539.6) pmol L(-1); P = 0.04], but were not affected by rVIIa; D-dimer levels were significantly higher in haemophiliacs than in controls and remained unchanged after infusion. CONCLUSIONS: Haemophilia A patients with factor VIII inhibitors have low thrombin generation. After rVIIa, the extent of coagulation activation as measured by levels of F1 + 2 is increased, but thrombin generation is restored to only 20%. Peak thrombin levels could reflect the effects of rVIIa on coagulation mechanisms, and their relevance with regard to the clinical coagulation defect of haemophilia A patients with factor VIII inhibitors might be evaluated.

Evidence of a U-shaped association between factor XII activity and overall survival.

INTRODUCTION: The clinical relevance of decreased coagulation factor XII (FXII) plasma activity as a risk factor for both venous and arterial thrombosis is still discussed controversially. The current study evaluated the predictive value of FXII levels for all-cause mortality in a large Viennese patient cohort. PATIENTS AND METHODS: Individuals, whose FXII activity levels were determined for suspected coagulation disorders or thrombophilia screening between 1991-2003 were included in this study (n = 8936, 51% male, 49% female, median age 43 years). Death/survival was determined by record linkage with the Austrian Death Registry. The median observation period was 5 years covering a total of 46 400 person years; the death rate was 17.1%. For Cox regression analysis, FXII plasma activity was divided into 11 categories of 10% steps with the category of > 100% FXII serving as a reference category. RESULTS: With decreasing FXII plasma activity, hazard ratios for all-cause mortality gradually increased linearly from 1.0 in the > 100% category to 1.5 (95% CI: 1.2-1.9) in the 80-90% category to 4.7 (95% CI: 3.4-6.5) in the 10-20% category. Similar results were obtained, when only vascular mortality or death as a result of ischemic heart disease was considered. No significant increase in all-cause mortality (HR: 1.4, 95%CI 0.7-2.8) was observed in the small group of FXII-deficient subjects [0-10% category (n = 58)]. CONCLUSIONS: This study first demonstrates a strong and almost linear association of FXII plasma activity between 90% and 10% with all-cause mortality in a large Viennese patient cohort. Interestingly, mortality rates are not increased when FXII activity is below 10%, resulting in a U-shaped survival curve.

Low-density lipoprotein receptor-related protein 1 polymorphism 663 C > T affects clotting factor VIII activity and increases the risk of venous thromboembolism.

BACKGROUND: Clotting factor (F) VIII is an independent risk factor for primary and recurrent venous thromboembolism (VTE). The causes for high plasma FVIII levels are not fully understood, but an involvement of genetic factors has been demonstrated. A multifunctional endocytic receptor, low-density lipoprotein receptor-related protein 1 (LRP1), mediates cellular uptake and subsequent degradation of FVIII and may contribute to variations in FVIII levels. OBJECTIVE: We assessed the association of a genetic variation of LRP1 (663C > T) with basal FVIII levels and the risk of venous thrombosis in a group of high-risk patients and in healthy controls. PATIENTS AND METHODS: One hundred and fifty-two patients with a history of recurrent VTE (median age 56 years, 47% women) were compared with 198 age- and sex-matched controls (median age 53 years, 50% women). The LRP1 663C > T genotype was analyzed by mutagenic separated polymerase chain reaction assay and heterozygosity was confirmed by sequence analysis. RESULTS: LRP1 663C > T genotype distribution differed significantly between patients (663CC n = 138, 663CT n = 14) and controls (663CC n = 190, 663CT n = 8; P = 0.048). In multivariable linear regression analysis including LRP1 663C > T, ABO blood group, von Willebrand factor antigen, C-reactive protein and age, LRP1 663CT was independently associated with FVIII activity (P = 0.02). LRP1 663CT was also associated with increased odds for VTE following adjustment for blood group O, FV Leiden and the prothrombin variation 20210G > A in multivariate analysis (odds ratio 3.3, 95% CI 1.3-8.5). CONCLUSIONS: According to our data the LRP1 663C > T polymorphism influences plasma FVIII levels independently of blood group, C-reactive protein and von Willebrand factor and is significantly associated with the risk of VTE.

Prediction of recurrent venous thromboembolism by the activated partial thromboplastin time.

BACKGROUND: Venous thromboembolism (VTE) is a multi-factorial disease. Extensive thrombophilia screening is costly and often inconclusive. Simple laboratory methods are required to predict the risk of recurrence. OBJECTIVE: To assess if measurement of activated partial thromboplastin time (APTT) allows stratification of patients with VTE into high- and low-risk categories with regard to recurrence. PATIENTS AND METHODS: We prospectively followed 918 patients with a first unprovoked VTE and studied the relationship between recurrence and an APTT after discontinuation of anticoagulation. APTT was expressed as a ratio of test to reference coagulation times. Study endpoint was symptomatic recurrent VTE. RESULTS: Venous thromboembolism recurred in 101 (11%) patients. Patients without recurrence had a greater APTT ratio than those with recurrence (0.97 +/- 0.09 vs. 0.93 +/- 0.09, P = 0.001). After 4 years, probability of recurrent VTE was 8.5% (95% CI: 5.5-11.5%) among patients with a ratio equal to or > 0.95 and 15.6% (95% CI: 11.4-19.9%) among patients with a lower ratio (P = 0.005). Compared with patients with an APTT ratio < 0.95, the relative risk (RR) of recurrence among patients with a ratio equal to or > 0.95 was 0.56 (95% CI: 0.38-0.84, P = 0.005) before and 0.58 (95% CI: 0.39-0.87, P = 0.009) after adjustment for sex, age, factor V Leiden, and factor II G20210A. CONCLUSIONS: Measurement of APTT allows stratification of patients with VTE into high- and low-risk categories with regard to recurrence.

Fibrinolytic dysfunction in insulin-resistant women with previous gestational diabetes.

BACKGROUND: Women with a history of gestational diabetes (p-GDM) are at increased risk of developing type 2 diabetes mellitus (DM2) later in life, and therefore at increased risk for future cardiovascular disease. MATERIALS AND METHODS: Three months after delivery we investigated the plasma levels of plasminogen activator inhibitor type 1 (PAI-1), tissue plasminogen activator (t-PA), fibrinogen and von Willebrand factor (vWF) in 74 women with p-GDM and 20 healthy females with normal glucose tolerance during and after pregnancy, as well as the relation of fibrinolytic parameters to insulin resistance and glycaemic control. All women underwent an oral (OGTT) as well as an intravenous glucose tolerance test (FSIGT). Mathematical model analysis disclosed that 50% (n=37 each) of the p-GDM subjects had normal (NIS) or impaired (IIS) insulin sensitivity. Parameters of interest were determined using commercially available test systems. RESULTS: Women with p-GDM and IIS had significantly increased body fat mass (BFM) (P

Validation of rotation thrombelastography in a model of systemic activation of fibrinolysis and coagulation in humans.

BACKGROUND: Thrombelastography (TEG) is a whole blood assay to evaluate the viscoelastic properties during blood clot formation and clot lysis. Rotation thrombelastography (e.g. ROTEM) has overcome some of the limitations of classical TEG and is used as a point-of-care device in several clinical settings of coagulation disorders. Endotoxemia leads to systemic activation of the coagulation system and fibrinolysis in humans. OBJECTIVES: We validated whether ROTEM is sensitive to endotoxin induced, tissue factor-triggered coagulation and fibrinolysis and if its measures correlate with biohumoral markers of coagulation and fibrinolysis. PATIENTS AND METHODS: Twenty healthy male volunteers participated in this randomized placebo-controlled trial. Volunteers received either 2 ng kg(-1) National Reference Endotoxin or saline. RESULTS: Endotoxemia significantly shortened ROTEM clotting time (CT) by 36% (CI 0.26-0.46; P < 0.05) with a strong inverse correlation with the peak plasma levels of prothrombin fragments (F(1 + 2)) (r = -0.83, P < 0.05). Additionally, endotoxin infusion enhanced maximal lysis (ML) 3.9-fold (CI: 2.5-5.2) compared with placebo or baseline after 2 h (P < 0.05). Peak ML and peak tissue plasminogen activator (t-PA) values correlated excellently (r = 0.82, P < 0.05). ROTEM parameters clot formation time and maximal clot firmness were not affected by LPS infusion, whereas platelet function analyzer (PFA-100) closure times decreased. CONCLUSIONS: Rotation thrombelastography (ROTEM) detects systemic changes of in vivo coagulation activation, and importantly it is a point of care device, which is sensitive to changes in fibrinolysis in humans. The ex vivo measures CT and ML correlate very well with established in vivo markers of coagulation activation (F(1 + 2)) and fibrinolysis (t-PA), respectively.