The objective of this study was to determine if treatment of non-insulin-dependent diabetes mellitus (NIDDM) patients with the "insulin sensitizer" troglitazone, both as monotherapy and in combination with insulin, corrects the impaired fibrinolysis and activated coagulation associated with NIDDM. Patients participating in two clinical trials comparing troglitazone and placebo in patients with NIDDM were studied at the time of randomization and after <em>2</em>6 weeks of treatment. Eighteen patients were treated with troglitazone (ten in combination with insulin and eight as monotherapy) and eight were treated with placebo (four in each trial). Plasma concentrations of plasminogen activator inhibitor (PAI-<em>1</em>), <em>prothrombin</em> <em>fragment</em> F<em>1</em>+<em>2</em>, fibrinogen, and von Willebrand Factor (vWF) activity were measured. Plasma PAI-<em>1</em> concentrations fell significantly from a mean of 68.8 +/- 3<em>2</em>.3 ng/mL to 40.4 +/- <em>2</em>0.4 in the troglitazone treated group, but did not change significantly in the placebo treated group. Plasma PAI-<em>1</em> concentrations were elevated in <em>1</em>5 patients treated with troglitazone and fell to normal in eight of them. There was no significant change in plasma F<em>1</em>+<em>2</em>, vWF, and fibrinogen, but plasma C-peptide and triglyceride concentrations fell significantly with troglitazone. This study demonstrates that troglitazone treatment is associated with a significant fall in plasma PAI-<em>1</em> antigen concentrations in patients with NIDDM and, therefore, may have a beneficial effect on fibrinolysis.

ESSENTIALS: The lectin pathway's MASP-<em>1</em>/<em>2</em> activates coagulation factors but the trigger of the activation is unknown. MASP-<em>1</em>/<em>2</em> activation was assessed by quantifying complexes between MASPs and antithrombin/C<em>1</em>-inhibitor. Activated platelets and fibrin were demonstrated to activate MASP-<em>1</em> and MASP-<em>2</em> both in vitro and in vivo. These findings may represent a crossroad between the complement and the coagulation systems.

CONCLUSIONS

BACKGROUND

The activated forms of the complement lectin pathway (LP) proteases MASP-<em>1</em> and MASP-<em>2</em> are able to cleave the coagulation factors prothrombin, fibrinogen, factor XIII and thrombin-activatable fibrinolysis inhibitor in vitro. In vivo studies also show that MASP-<em>1</em> is involved in thrombogenesis.

OBJECTIVE

To clarify the not yet identified mechanisms involved in triggering activation of the LP during thrombotic reactions.

METHODS

Novel sandwich-ELISAs for detection of complexes between MASP-<em>1</em> or MASP-<em>2</em> and the serpins C<em>1</em> inhibitor (C<em>1</em>-INH) or antithrombin (AT), were used to specifically detect and quantify the activated forms of MASP-<em>1</em> and MASP-<em>2</em>.

RESULTS

Activated platelets were shown by flow cytometry to bind Ficolin-<em>1</em>, -<em>2</em> and -3 but not MBL, which was associated with activation of MASP-<em>1</em> and MASP-<em>2</em>. We also demonstrated that fibrin and the plasmin-generated fibrin fragment DD in plasma, bind and activate MASP-<em>1</em> and MASP-<em>2</em>. As demonstrated by the ELISA and SDS-PAGE/Western blotting, the fibrin-associated activation was reflected in a specific inactivation by AT during clotting without the assistance of heparin. In all other cases the MASPs were, as previously reported, inactivated by C<em>1</em>-INH. In systemic lupus erythematosus patients with thrombotic disease and in polytrauma patients, the levels of activated MASP-<em>1</em> and MASP-<em>2</em> in complex with both AT and C<em>1</em>-INH were associated with markers of thrombotic disease and contact/coagulation system activation.

CONCLUSIONS

MASP-<em>1</em> and MASP-<em>2</em> are activated during blood clotting. This activation is triggered by activated platelets and by the generation of fibrin during thrombotic reactions in vitro and in vivo, and may represent a novel activation/amplification mechanism in thromboinflammation.

OBJECTIVE

The degree of oxidative stress and its association with a thrombophilic condition, if any, were investigated in alcoholics before the onset of severe liver disease.

METHODS

Reactive oxygen species and total antioxidant capacity were evaluated using two new kinetic spectrophotometric methods in a selected group of 45 consecutive chronic alcohol abusers and 4<em>2</em> apparently healthy moderate drinkers, used as controls. The hemostatic system was explored by detecting the plasma levels of <em>prothrombin</em> <em>fragment</em> <em>1</em>+<em>2</em> (F<em>1</em>+<em>2</em>) and thrombin-antithrombin complexes (TAT) with enzyme-linked immunosorbent assays, while D-dimer plasma levels were measured with a turbidimetric immunoassay.

RESULTS

Reactive oxygen species were significantly higher (p<0.00<em>1</em>) in heavy drinkers than in controls: 3<em>2</em>8.<em>1</em> (<em>1</em>43.4-847.<em>2</em>) U.CARR vs <em>2</em>50 (<em>2</em>00.7-366.8) U.CARR, respectively. The total antioxidant capacity was similar in chronic alcohol abusers and in moderate drinkers: 360.<em>2</em> (336.8-374.4) microMol HClO/mL vs 369 (36<em>2</em>-378.4) microMol HClO/mL, respectively. All molecular markers of hemostatic system activation were significantly increased in chronic alcohol abusers in comparison with those in moderate drinkers, as follows: TAT: <em>2</em>.5 (<em>1</em>.4-<em>1</em>3) microg/L vs <em>1</em>.5 (<em>1</em>-4.<em>1</em>) mocrog/L, respectively (p<0.00<em>1</em>); F<em>1</em>+<em>2</em>: <em>1</em>.7 (0.5-5.<em>2</em>) nMol/L vs 0.9 (0.4-<em>1</em>.<em>1</em>) nMol/L, respectively (p<0.0<em>1</em>); D-dimer: <em>2</em>35.5 (<em>2</em>08-46<em>2</em>) ng/mL vs <em>1</em>63.5 (7<em>1</em>-<em>2</em>33) ng/mL, respectively (p<0.00<em>1</em>).

CONCLUSIONS

Our results suggest that oxidative stress and a thrombophilic condition can be observed in heavy drinkers without severe liver disease. The new test available for measuring reactive oxygen species in serum proved to be reliable and useful as an early marker of tissue damage.

Systemic sclerosis (SSc) is a disease characterized by progressive microvascular occlusion and fibrosis resulting in irreversible organ damage, the pathogenesis of which is felt to be of vascular origin. To gain a comprehensive view of the coagulation/fibrinolytic balance in SSc, a number of haemostatic and fibrinolytic variables were measured in <em>2</em>6 SSc patients (<em>1</em><em>1</em> limited, <em>1</em>5 diffuse) and in <em>2</em><em>2</em> control subjects. Of the coagulation activation markers, the mean plasma level of <em>prothrombin</em> <em>fragment</em> <em>1</em> + <em>2</em> (F<em>1</em> + <em>2</em>), but not of thrombin-antithrombin complexes (TAT), was higher in SSc patients than in controls (P < 0.00<em>1</em>). Plasma levels of fibrin split product D-dimer (DD), fibrinogen (FNG) and von Willebrand factor (vWF) were higher amongst patients than controls (P < 0.00<em>1</em>). vWF and FNG levels were positively correlated (P < 0.00<em>1</em>). Mean levels of DD and vWF were more elevated in patients with diffuse than limited disease (P = 0.00<em>1</em> and P = 0.04, respectively). On the fibrinolytic side, defective tissue plasminogen activator (tPA) release (venous occlusion test, stimulated level < basal level) was noted in 46% (<em>1</em><em>2</em>/<em>2</em>6) of SSc patients, but only in 4% (<em>1</em>/<em>2</em><em>2</em>) of controls. Patients had higher mean levels of tPA inhibitor (PAI) than controls (P < 0.00<em>1</em>), levels being more elevated amongst patients with diffuse than limited disease (P = 0.0<em>1</em>). An abnormally high lipoprotein (a) [Lp(a)] level was found in 9% (<em>2</em>/<em>2</em>0) of control subjects, but in 30% (8/<em>2</em>6) of SSc patients (P = 0.04) where it clustered with fibrinolytic defects. Altogether, these data suggest that patients with SSc are in a hypercoagulable state characterized by elevated plasma levels of FNG and vWF, by a dual hypofibrinolytic pattern (defective tPA release and elevated PAI), and by increased thrombin generation with enhanced fibrin formation. Higher levels of vWF, DD and PAI in patients with diffuse disease are consistent with more extensive (micro)vascular involvement, although no causal relationship can be inferred. The lack of a parallel increase of TAT with F<em>1</em> + <em>2</em>, in the presence of normal levels of antithrombin III (ATIII), indirectly suggests an impairment of the heparan sulphate-ATIII system which would favour thrombin generation. Since thrombin may act as a mitogen for fibroblasts, may upregulate vWF, PAI and endothelin production by endothelial cells, and may promote fibrin deposition on the vessel wall leading to worsening of microvascular occlusions, limitation of thrombin generation, besides fibrinolytic enhancement, could represent a possible coadjuvant interventional strategy.

The authors assessed hemostasis and fibrinolysis serially: on admission and on the <em>1</em>st and 7th days after surgery for subarachnoid hemorrhage (SAH), examining the complications of aneurysm rupture and its surgical repair. Of 3<em>2</em> patients, <em>2</em>5 with SAH were compared with seven control patients who underwent surgery for an unruptured intracranial aneurysm. On admission, patients with SAH had higher thrombin-antithrombin III complex (TAT) levels (<em>1</em>3.3 +/- 3.8 vs. 3.8 +/- 0.6 ng/ml, p = 0.0<em>1</em>), fibrin degradation product, D-dimer levels (<em>1</em>3<em>1</em>0 +/- <em>2</em><em>2</em>0 vs. 556 +/- 89 ng/ml, p = 0.000<em>1</em>), and leukocyte counts (<em>1</em>4.6 +/- 0.7 vs. <em>1</em>0.6 +/- <em>1</em>.8 x <em>1</em>0(9) cells/L, p < 0.05) than did control patients. Postoperative D-dimer values (p = 0.007) remained higher in the SAH group. Furthermore, admission D-dimer levels were higher in the patients in poor clinical condition than in those in good condition (<em>2</em>0<em>1</em>7 +/- 377 vs. 934 +/- <em>2</em>08 ng/ml, p = 0.007), and D-dimer levels were associated with the outcome at 3 months after admission. Additionally, thrombin generation and fibrinolytic markers measured on admission were related to clinical grade, amount of subarachnoid blood seen on computerized tomography (CT) scanning, and patient fatality. Patients with hypodense lesions verified on follow-up CT scanning or with persistent neurological deficits at 3 months had higher <em>prothrombin</em> <em>fragments</em> <em>1</em> and <em>2</em>, TAT, D-dimer, and plasminogen activator inhibitor-<em>1</em> values on the <em>1</em>st day postoperatively than did patients without such lesions. In short, in patients with SAH, activation of coagulation and fibrinolysis was strongly associated with clinical state, patient fatality, and outcome at 3 months, and postoperatively this activation correlated with the development of brain infarction.

BACKGROUND

Abacavir (ABC) treatment has been associated with an increased incidence of myocardial infarction. The pathophysiological mechanism is unknown. In this study markers of inflammation and coagulation in HIV-infected patients using antiretroviral therapy with or without ABC were examined to pinpoint a pathogenic mechanism. Given the important role of high sensitivity C-reactive protein (hsCRP) levels in predicting cardiovascular risk, patient groups were also analyzed according to hsCRP levels.

METHODS

Patients treated with ABC and a matched control group treated without ABC were selected retrospectively. Vascular endothelial growth factor (VEGF) and markers of endothelial cell activation (von Willebrand factor (vWF), factor VIII), fibrin formation (fibrinogen, D-dimer, <em>prothrombin</em> <em>fragment</em> <em>1</em>+<em>2</em> (F<em>1</em>+<em>2</em>), endogenous thrombin potential (ETP)), anticoagulation markers (protein C and S, activated protein C sensitivity ratio (APCsr)) and inflammation markers (IL-6, hsCRP) were measured in citrated plasma.

RESULTS

A total of 8<em>1</em> patients were included of whom <em>2</em>7 patients used an ABC-containing regimen and 54 used a non-ABC-containing regimen. Patient characteristics were not significantly different between the groups except for longer duration of use of the current antiretroviral regimen in the ABC group (p = 0.0<em>1</em>). The median time on ABC was 68 months (interquartile range 59-80 months). No differences in coagulation and inflammation markers according to ABC use were observed. For the whole patient group elevated vWF and F<em>1</em>+<em>2</em> levels were observed in <em>2</em>3% and 37%, respectively. Compared to the reference ranges for the general population increased APCsr was found in 79% and lower protein C and VEGF levels in 40% and 43%, respectively. Patients in the high-risk category for cardiovascular disease with hsCRP levels>> 3 mg/L had significantly higher fibrinogen, D-dimer, F<em>1</em>+<em>2</em> and ETP levels compared to patients from the low-risk category with hsCRP levels < <em>1</em> mg/L.

CONCLUSIONS

HIV-infected patients using ABC showed no specific abnormalities in coagulation or inflammation markers that might explain the increased risk of myocardial infarction. For the whole group, regardless of ABC use, evidence of a prothrombotic state was observed. Thirty-three percent of patients with long-term use of antiretroviral treatment had hsCRP levels above 3 mg/L, which is strongly associated with cardiovascular disease in HIV-uninfected individuals.

High-density lipoproteins (HDL) can bind and neutralize lipopolysaccharides (LPS) in vitro and in vivo. HDL can also affect fibrinolytic activity and can directly influence platelet function by reducing platelet aggregation. In this study, the effects of reconstituted HDL (rHDL) on LPS-induced coagulation, fibrinolysis and platelet activation in humans were investigated. In a double-blind, randomized, placebo-controlled, cross-over study, eight healthy male volunteers were injected with LPS (4 ng/kg) on two occasions, once in conjunction with rHDL (40 mg/kg, given as a 4 h infusion starting 3.5 h prior to LPS injection), and once in conjunction with placebo. rHDL significantly reduced LPS-induced activation of coagulation (plasma levels of <em>prothrombin</em> <em>fragment</em> F<em>1</em> + <em>2</em>) and fibrinolysis (plasma levels of tissue type plasminogen activator antigen, t-PA). No effect was observed on LPS-induced inhibition of the fibrinolytic pathway (PAI-<em>1</em>) or on the transient thrombocytopenia elicited by LPS. Furthermore, rHDL treatment significantly enhanced the inhibition of collagen-stimulated inhibition of platelet aggregation during endotoxemia, but had no such effect on arachidonate-stimulated platelet aggregation. rHDL treatment per se also reduced collagen-induced platelet aggregation. These results indicate that rHDL modifies the procoagulant state associated with endotoxemia.

To examine the relationship between exercise intensity and activation of coagulation and fibrinolysis, we measured markers of thrombin, fibrin, and plasmin formation in <em>1</em><em>2</em> male subjects (mean <em>2</em>4+/-4 yr (SD)) before and after running on a treadmill for <em>1</em> h at two different intensities corresponding to moderate (8<em>2</em>% maximal heart rate (HR), 68% VO<em>2</em>max) and very heavy (94% maximal HR, 83% VO<em>2</em>max) exercise. During moderate exercise plasma levels of tissue plasminogen activator (t-PA) antigen rose from 3.7+/-0.5 (mean+/-SE) to <em>1</em>4.6+/-<em>1</em>.8 ng x mL[-<em>1</em>] (P < 0.0<em>1</em>) and of plasmin-alpha-antiplasmin (PAP) complexes from <em>2</em>.<em>1</em>+/-0.3 to 4.<em>2</em>+/-0.7 nmol x L[-<em>1</em>] (P < 0.0<em>1</em>), whereas <em>prothrombin</em> <em>fragment</em> <em>1</em>+<em>2</em> (PTF<em>1</em>+<em>2</em>), thrombin-antithrombin III (TAT) complexes and fibrinopeptide A (FPA) did not change significantly. In response to very heavy exercise, mean plasma levels of t-PA antigen and PAP complexes exceeded the upper limit of normal values <em>2</em>.5- (P < 0.0<em>1</em>) and two-fold (P < 0.0<em>1</em>), respectively, while significant increases of plasma levels of PTF<em>1</em>+<em>2</em> (P < 0.0<em>1</em>), TAT (P < 0.05), and FPA (P < 0.0<em>1</em>) occurred within the range of normal. We conclude that in healthy young individuals, exercise-induced activation of coagulation is well balanced by activation of the fibrinolytic system, since moderate exercise results in increased plasmin formation only, while at very heavy exercise generation of plasmin seems to exceed that of thrombin and fibrin.

The specificity of thrombin for procoagulant and anticoagulant substrates is regulated allosterically by Na+. Ordered cleavage of <em>prothrombin</em> (ProT) at Arg3<em>2</em>0 by the <em>prothrombin</em>ase complex generates proteolytically active, meizothrombin (MzT), followed by cleavage at Arg<em>2</em>7<em>1</em> to produce thrombin and <em>fragment</em> <em>1</em>.<em>2</em>. The alternative pathway of initial cleavage at Arg<em>2</em>7<em>1</em> produces the inactive zymogen form, the prethrombin <em>2</em> (Pre <em>2</em>).<em>fragment</em> <em>1</em>.<em>2</em> complex, which is cleaved subsequently at Arg3<em>2</em>0. Cleavage at Arg3<em>2</em>0 of ProT or prethrombin <em>1</em> (Pre <em>1</em>) activates the catalytic site and the precursor form of exosite I (proexosite I). To determine the pathway of expression of Na+-(pro)exosite I linkage during ProT activation, the effects of Na+ on the affinity of fluorescein-labeled hirudin-(54-65) ([5F]Hir-(54-65)(SO-3)) for the zymogens, ProT, Pre <em>1</em>, and Pre <em>2</em>, and for the proteinases, MzT and MzT-des<em>fragment</em> <em>1</em> (MzT(-F<em>1</em>)) were quantitated. The zymogens showed no significant linkage between proexosite I and Na+, whereas cleavage at Arg3<em>2</em>0 caused the affinities of MzT and MzT(-F<em>1</em>) for [5F]Hir-(54-65)(SO-3) to be enhanced by Na+ 8- to <em>1</em>0-fold and 5- to 6-fold, respectively. MzT and MzT(-F<em>1</em>) showed kinetically different mechanisms of Na+ enhancement of chromogenic substrate hydrolysis. The results demonstrate for the first time that MzT is regulated allosterically by Na+. The results suggest that the distinctive procoagulant substrate specificity of MzT, in activating factor V and factor VIII on membranes, and the anticoagulant, membrane-modulated activation of protein C by MzT bound to thrombomodulin are regulated by Na+-induced allosteric transition. Further, the Na+ enhancement in MzT activity and exosite I affinity may function in directing the sequential ProT activation pathway by accelerating thrombin formation from the MzT fast form.

OBJECTIVE

We sought to determine whether venous thromboembolism in cancer patients is associated with aberrant plasma levels of hemostatic and angiogenic factors.

METHODS

Peripheral blood was collected before anticoagulant therapy from cancer patients with acute deep venous thrombosis (DVT; DVT + cancer group, n = 3<em>2</em>), those without DVT (cancer control group, n = 36), and patients with acute DVT but no cancer (DVT control group, n = 58). Plasma assays of activation and inhibition of coagulation and fibrinolysis, as well as angiogenesis activation, were then performed.

RESULTS

Median levels of thrombin-antithrombin complex, <em>prothrombin</em> <em>fragments</em> <em>1</em> + <em>2</em>, and von Willebrand factor antigen were significantly greater in the DVT + cancer group than in the cancer control and DVT control groups (<em>1</em>7.8 ng/mL v 4.6 ng/mL and 9.8 ng/mL, P =.000<em>1</em> and P =.003, respectively; 3.65 nmol/L v <em>1</em>.60 nmol/L and <em>2</em>.7<em>1</em> nmol/L, P <.000<em>1</em> and P =.0<em>1</em><em>1</em>, respectively; and 4.04 U/mL v <em>2</em>.<em>2</em>6 U/mL and <em>2</em>.06 U/mL, P <.000<em>1</em>, respectively). Median levels of tissue-type plasminogen activator were also significantly higher, while protein C activity was lower in the DVT + cancer group than in the DVT control group (<em>1</em>4.6 ng/mL v 9.50 ng/mL, respectively, P =.0005; 0.89 U/mL v <em>1</em>.<em>1</em><em>1</em> U/mL, respectively, P =.0008).

CONCLUSIONS

These data not only support prior observations of coagulation activation in patients with malignancy, but also provide new evidence for enhanced coagulation activation in the setting of acute venous thromboembolism in cancer. Future prospective studies are warranted to determine whether these and other potential markers of hypercoagulability may help to identify cancer patients at highest risk for venous thromboembolism.

In order to specifically evaluate the role of Factor Va in the <em>prothrombin</em>ase complex, studies of the activation of <em>prothrombin</em>, <em>Fragment</em> <em>1</em>.<em>2</em>-prethrombin-<em>2</em>, and active-site-blocked meizothrombin were carried out, both in the absence of phospholipid and at concentrations of substrates and Factor Va sufficient to approach saturation in all components. Km values were independent of Factor Va concentrations, whereas kcat (apparent) values approached saturation with respect to Factor Va concentrations. The three respective substrates exhibited the following parameters of kinetics (Km, microM; kcat, s-<em>1</em> at saturating [Factor Va]): <em>prothrombin</em> (9.0 +/- 0.4; 3<em>1</em> +/- <em>1</em>); <em>Fragment</em> <em>1</em>.<em>2</em>-prethrombin-<em>2</em> (5.4 +/- 0.4; <em>1</em>3 +/- <em>2</em>); and meizothrombin (3.6 +/- 0.3; 5<em>1</em> +/- 5). Models of kinetics were constructed to interpret the results, and two of these were formally consistent with experimental results. Both models indicated that the variation of kcat(app) with concentrations of Factor Va reflects the formation of a Factor Va-Factor Xa binary complex. Analysis of kinetics indicated Kd values for this interaction of <em>1</em>.3 +/- 0.<em>1</em>, 3.0 +/- 0.5, and <em>1</em>.0 +/- 0.<em>1</em> microM for the three respective substrates. The models differed in the interpretation of Km. One indicated that Km reflects a binary interaction between Factor Xa and <em>prothrombin</em>, whereas the other indicated a binary interaction between Factor Va and <em>prothrombin</em>. Both indicated that two of the three possible binary interactions between the three components would be reflected in Km and kcat values but not the third. To distinguish these models, the binary interactions were studied by extrinsic fluorescence (Va.Xa), light-scattering (Factor Va.<em>prothrombin</em>), and competition kinetics (Xa.II). The first two interactions were detected and were characterized by Kd values of <em>2</em>.7 +/- 0.<em>1</em> microM (Va.Xa) and 8.8 +/- 0.8 microM (Factor Va.<em>prothrombin</em>). No active-site-dependent interaction between <em>prothrombin</em> and Factor Xa could be detected in the absence of Factor Va. The results of these studies suggest that Factor Va interacts with both Factor Xa and <em>prothrombin</em> and effectively presents one to the other in the formation of a ternary enzyme-substrate-cofactor complex. In addition, a comparison of the parameters of kinetics of conversion of <em>prothrombin</em> and its intermediates indicates that meizothrombin is the major intermediate of <em>prothrombin</em> activation in the absence, as well as in the presence of phospholipid.

BACKGROUND

The purpose of this study was to evaluate the relative importance of systemic hypercoagulability, preexisting and acquired risk factors, and specific injury patterns in the development of venous thromboembolism (VTE) after injury.

METHODS

Injured patients with an Injury Severity Score>> or = <em>1</em>5 were followed with lower extremity venous duplex ultrasonography, <em>prothrombin</em> <em>fragment</em> <em>1</em> + <em>2</em>, and quantitative D-dimer levels at <em>1</em> and 3 days and then weekly until discharge.

RESULTS

Among <em>1</em>0<em>1</em> patients with a mean Injury Severity Score of <em>2</em>7.3 +/- <em>1</em>0.5 followed for <em>1</em><em>2</em>.4 +/- 8.7 days, <em>2</em>8 (<em>2</em>7.7%) developed a lower extremity thrombosis, <em>2</em> (<em>1</em>.9%) sustained a pulmonary embolism, and <em>1</em> (0.9%) had a symptomatic upper extremity thrombosis. Although admission <em>fragment</em> <em>1</em> + <em>2</em> and D-dimer levels were elevated in 8<em>1</em>.4% and <em>1</em>00% of patients, respectively, mean levels were not significantly different in those with or without VTE. VTE was more common (p < 0.05) among those with obesity, age>> 40 years, immobilization for>> 3 days, spine fractures, and lower extremity fractures. However, only obesity (p = 0.004) and immobilization>> 3 days (p = 0.05) were independent predictors of VTE in a multivariate analysis.

CONCLUSIONS

Although elevated in seriously injured patients, neither markers of activated coagulation nor specific injury patterns are predictive of VTE. Associations with immobilization and obesity suggest that VTE after injury is a systemic hypercoagulable disorder with local manifestations of thrombosis related to lower extremity stasis.

BACKGROUND

Numerous interactions between the coagulation and complement systems have been shown. Recently, links between coagulation and mannan-binding lectin-associated serine protease-<em>1</em> (MASP-<em>1</em>) of the complement lectin pathway have been proposed. Our aim was to investigate MASP-<em>1</em> activation of factor XIII (FXIII), fibrinogen, <em>prothrombin</em>, and thrombin-activatable fibrinolysis inhibitor (TAFI) in plasma-based systems, and to analyse effects of MASP-<em>1</em> on plasma clot formation, structure and lysis.

RESULTS

We used a FXIII incorporation assay and specific assays to measure the activation products <em>prothrombin</em> fragment F<em>1</em>+2, fibrinopeptide A (FPA), and activated TAFI (TAFIa). Clot formation and lysis were assessed by turbidimetric assay. Clot structure was studied by scanning electron microscopy. MASP-<em>1</em> activated FXIII and, contrary to thrombin, induced FXIII activity faster in the Val34 than the Leu34 variant. MASP-<em>1</em>-dependent generation of F<em>1</em>+2, FPA and TAFIa showed a dose-dependent response in normal citrated plasma (NCP), albeit MASP-<em>1</em> was much less efficient than FXa or thrombin. MASP-<em>1</em> activation of <em>prothrombin</em> and TAFI cleavage were confirmed in purified systems. No FPA generation was observed in <em>prothrombin</em>-depleted plasma. MASP-<em>1</em> induced clot formation in NCP, affected clot structure, and prolonged clot lysis.

CONCLUSIONS

We show that MASP-<em>1</em> interacts with plasma clot formation on different levels and influences fibrin structure. Although MASP-<em>1</em>-induced fibrin formation is thrombin-dependent, MASP-<em>1</em> directly activates <em>prothrombin</em>, FXIII and TAFI. We suggest that MASP-<em>1</em>, in concerted action with other complement and coagulation proteins, may play a role in fibrin clot formation.

BACKGROUND

It has been demonstrated that there is an increased risk of venous thromboembolism (VTE) during air travel on flights of long duration. Patients with COPD are also at increased risk of VTE, particularly during exacerbations, possibly because of a hypercoagulable state secondary to hypoxia and/or heightened systemic inflammation. We investigated the effects of hypoxia on indices of coagulation and systemic inflammation in patients with COPD.

METHODS

Twenty clinically stable patients with mild COPD were recruited. Patients were randomized to receive either medical air or <em>1</em>00% nitrogen through a 40% venturi mask at a flow rate of <em>1</em>0 L/min for <em>2</em> h. Blood was sampled for thrombin-antithrombin complex (TAT), <em>prothrombin</em> activation <em>fragments</em> <em>1</em> + <em>2</em> (F(<em>1</em> + <em>2</em>)), von Willebrand factor antigen (VWF:Ag), D-dimer, and interleukin-6 (IL-6) at baseline and after <em>2</em> h.

RESULTS

Patients in the hypoxia and control groups were similar in terms of age, sex, pack-years smoked, and severity of airflow obstruction. There was no difference in baseline TAT, F(<em>1</em> + <em>2</em>), VWF:Ag, D-dimer, or IL-6 levels between groups. In the control group, there was no change in markers of coagulation or systemic inflammation over the <em>2</em>-h study. In patients who underwent hypoxic challenge, there was an increase in TAT (P < .00<em>1</em>), F(<em>1</em> + <em>2</em>) (P < .0<em>1</em>), and IL-6 (P < .0<em>1</em>), whereas D-dimer and VWF:Ag levels were unchanged.

CONCLUSIONS

This study demonstrates that a <em>2</em>-h hypoxic challenge in patients with COPD results in coagulation activation in conjunction with an increase in systemic inflammation.

The influence of phospholipid on thrombin-thrombomodulin-catalyzed activation of protein C has been studied by incorporating thrombomodulin into vesicles by dialysis from octyl glucoside-phospholipid mixtures. Thrombomodulin was incorporated into vesicles ranging from neutral (<em>1</em>00% phosphatidylcholine) to highly charged (30% phosphatidylserine and 70% phosphatidylcholine). Thrombomodulin is randomly oriented in vesicles of different phospholipid composition. Incorporation of thrombomodulin into phosphatidylcholine, with or without phosphatidylserine, alters the Ca<em>2</em>+ concentration dependence of protein C activation. Soluble thrombomodulin showed a half-maximal rate of activation at 580 microM Ca<em>2</em>+, whereas half-maximal rates of activation of liposome-reconstituted thrombomodulin were obtained between 500 microM Ca<em>2</em>+ and <em>2</em> mM Ca<em>2</em>+, depending on the composition (protein:phospholipid) of the liposomes. The Ca<em>2</em>+ dependence of protein C activation fits a simple hyperbola for the soluble activator, while the Ca<em>2</em>+ dependence of the membrane-associated complex is distinctly sigmoidal with a Hill coefficient greater than <em>2</em>.4. In contrast, the Ca<em>2</em>+ dependence of gamma-carboxyglutamic acid (Gla) domainless protein C activation is unchanged by membrane reconstitution (<em>1</em>/<em>2</em> max = 53 +/- <em>1</em>0 microM) and fits a simple rectangular hyperbola. Incorporation of thrombomodulin into pure phosphatidylcholine vesicles reduces the Km for protein C from 7.6 +/- <em>2</em> to 0.7 +/- 0.<em>2</em> microM. Increasing phosphatidylserine to <em>2</em>0% decreased the Km for protein C further to 0.<em>1</em> +/- 0.0<em>2</em> microM. Membrane incorporation has no influence on the activation of protein C from which the Gla residues are removed proteolytically (Km = 6.4 +/- 0.5 microM). The Km for protein C observed on endothelial cells is more similar to the Km observed when thrombomodulin (TM) is incorporated into pure phosphatidylcholine vesicles than into negatively charged vesicles, suggesting that the protein C-binding site on endothelial cells does not involve negatively charged phospholipids. In support of this concept, we observed that <em>prothrombin</em> and <em>fragment</em> <em>1</em>, which bind to negatively charged phospholipids, do not inhibit protein C activation on endothelial cells or TM incorporated into phosphatidylcholine vesicles, but do inhibit when TM is incorporated into phosphatidylcholine:phosphatidylserine vesicles. These studies suggest that neutral phospholipids lead to exposure of a site, probably on thrombomodulin, capable of recognizing the Gla domain of protein C.

OBJECTIVE

Epidemiological and laboratory studies suggest that increasing concentrations of plasma homocysteine (total homocysteine [tHcy]) accelerate cardiovascular disease by promoting vascular inflammation, endothelial dysfunction, and hypercoagulability.

METHODS

We conducted a randomized controlled trial in <em>2</em>85 patients with recent transient ischemic attack or stroke to examine the effect of lowering tHcy with folic acid <em>2</em> mg, vitamin B1<em>2</em> 0.5 mg, and vitamin B6 <em>2</em>5 mg compared with placebo on laboratory markers of vascular inflammation, endothelial dysfunction, and hypercoagulability.

RESULTS

At 6 months after randomization, there was no significant difference in blood concentrations of markers of vascular inflammation (high-sensitivity C-reactive protein [P=0.3<em>2</em>]; soluble CD40L [P=0.33]; IL-6 [P=0.77]), endothelial dysfunction (vascular cell adhesion molecule-1 [P=0.<em>2</em>7]; intercellular adhesion molecule-1 [P=0.08]; von Willebrand factor [P=0.9<em>2</em>]), and hypercoagulability (P-selectin [P=0.33]; prothrombin fragment 1 and <em>2</em> [P=0.81]; D-dimer [P=0.88]) among patients assigned vitamin therapy compared with placebo despite a 3.7-micromol/L (95% CI, <em>2</em>.7 to 4.7) reduction in total homocysteine (tHcy).

CONCLUSIONS

Lowering tHcy by 3.7 micromol/L with folic acid-based multivitamin therapy does not significantly reduce blood concentrations of the biomarkers of inflammation, endothelial dysfunction, or hypercoagulability measured in our study. The possible explanations for our findings are: (1) these biomarkers are not sensitive to the effects of lowering tHcy (eg, multiple risk factor interventions may be required); (<em>2</em>) elevated tHcy causes cardiovascular disease by mechanisms other than the biomarkers measured; or (3) elevated tHcy is a noncausal marker of increased vascular risk.

Plasmin-alpha<em>2</em>-antiplasmin complex (PAP) marks plasmin generation and fibrinolytic balance. We recently observed that elevated levels of PAP predict acute myocardial infarction in the elderly, yet little is known about the correlates of PAP. We measured PAP in 800 elderly subjects who were free of clinical cardiovascular disease in <em>2</em> cohort studies: the Cardiovascular Health Study and the Honolulu Heart Program. Median PAP levels did not differ between the Cardiovascular Health Study (6.05+/-<em>1</em>.46 nmol/L) and the Honolulu Heart Program (6.<em>1</em><em>1</em>+/-<em>1</em>.44 nmol/L), and correlates of PAP were similar in both cohorts. In CHS, PAP levels increased with age (r=0. 30), procoagulant factors (eg, factor VIIc, r=0.<em>1</em>5), thrombin activity (<em>prothrombin</em> <em>fragment</em> F<em>1</em>+<em>2</em>, r=0.<em>2</em>9), and inflammation-sensitive proteins (eg, fibrinogen, r=0.44; factor VIIIc, r=0.37). PAP was associated with increased atherosclerosis as measured by the ankle-arm index (AAI) (P for trend, </=0.00<em>1</em>). PAP was negatively related to factors associated with the insulin resistance syndrome (IRS) (eg, fasting insulin, r=-0.<em>2</em>6; body mass index, r=-0.<em>2</em>6), possibly reflecting an association with plasminogen activator inhibitor-<em>1</em> (r=-0.<em>2</em>9). Although our study did not have sufficient power to detect a significant interaction, PAP and AAI appeared to be more weakly associated in subjects with more manifestations of the IRS: PAP appeared more strongly associated with AAI in the subgroup with 0 or <em>1</em> metabolic disorders (P</=0.00<em>1</em>; slope estimate, -0.<em>1</em>4) compared with the subgroup with <em>2</em> or more metabolic disorders (P=0.<em>1</em>0; slope estimate, -0.08) and in those with non-insulin-dependent diabetes mellitus (P=0.46; slope estimate, -0.04). Although PAP reflects reactive fibrinolysis and is associated with subclinical atherosclerosis, this relationship may be weaker in populations with characteristics of the IRS, possibly reflecting the inhibitory effects of plasminogen activator inhibitor-<em>1</em> on PAP. Decreased fibrinolysis in the presence of subclinical disease in subjects with hyperinsulinemia or glucose intolerance is consistent with the premise that depressed plasmin generation may enhance the progression of atherosclerosis in these people.

DIC is a life-threatening complication of several disease states. It is characterized by systemic activation of the hemostasis system. In many instances the release of tissue factor (TF) from endothelial cells or other circulating cells triggers the system. Initially, the increased activation can be compensated for by the natural inhibitor systems, a state referred to as compensated DIC. As the trigger persists, inhibitors will be consumed leading to more coagulation. In this process many clotting factors, most notably fibrinogen and platelets are consumed, resulting eventually in a complete breakdown of the hemostasis system. This results in a profuse and diffuse bleeding tendency or decompensated DIC. The term consumptive coagulopathy denotes this process. Of crucial importance is the fate of fibrin that is formed from fibrinogen by thrombin. If the fibrinolytic system is insufficiently activated, fibrin will be deposited in the microcirculation leading to MODS. This will not occur if the fibrinolytic system is fully activated. The clinical suspicion of DIC must be confirmed by laboratory tests and decreasing fibrinogen levels and platelet counts support the diagnosis. The determination of D-dimer, fibrin(ogen) split products (FSP) and soluble fibrin monomer (FM) further support the diagnosis. FM suggest the presence of thrombin, FSP the generation of plasmin, and D-dimer, both thrombin and plasmin. While the tests are not specific for DIC, they can be helpful, in the proper clinical setting, to diagnose decompensated or acute DIC. The tests are not useful for the diagnosis of compensated DIC, except for D-dimer, FSP, and FM if elevated. Compensated DIC can be diagnosed by molecular markers of in vivo hemostasis activation, such as thrombin-antithrombin (TAT) complexes, <em>prothrombin</em> <em>fragment</em> <em>1</em> + <em>2</em> (F <em>1</em> + <em>2</em>), or plasmin-antiplasmin (PAP) complexes. For the treatment of DIC it is imperative to remove the triggering underlying disease. The consumption of coagulation constituents can be corrected by cryoprecipitate, platelet concentrates, and fresh frozen plasma, if needed. This may reduce the bleeding tendency. Arrest of the activated hemostasis system by heparins, either subcutaneous in low doses or intravenous in therapeutic doses, is only recommended in patients with compensated DIC. If the patient bleeds, heparins should not be given. The administration of concentrates of natural anticoagulants, i.e., antithrombin, protein C, or tissue factor pathway inhibitor are safer than heparins since they do not exacerbate the bleeding tendency. These concentrates were found to be very effective in animal models of DIC; human experience is still limited. Generally, the earlier treatment is initiated, the better the patient's prognosis.

OBJECTIVE

To determine the roles of tissue factor and thrombin on the systemic inflammatory response syndrome (SIRS) in posttrauma patients, as well as to investigate the relationship between SIRS and sepsis.

METHODS

Prospective, cohort study.

METHODS

General intensive care unit of a tertiary care emergency department.

METHODS

Forty trauma patients were classified into subgroups, according to the duration of SIRS: non-SIRS patients (n = 9); patients with SIRS for < <em>2</em> days (n = 15); and patients with SIRS for>> 3 days (n = 16).

METHODS

None.

RESULTS

Tissue factor antigen concentration, prothrombin fragment F1+<em>2</em>, thrombin antithrombin complex, fibrinopeptide A, and cross-linked fibrin degradation products (D-dimer) were measured on the day of admission, and on days 1 through 4 after admission. Simultaneously, the number of SIRS criteria that the patients met and the disseminated intravascular coagulation score were determined. The results of these measurements, frequency of acute respiratory distress syndrome (ARDS), multiple organ dysfunction syndrome, sepsis, and outcome were compared among the groups. The values of all five hemostatic molecular markers in the patients with SIRS for>> 3 days were significantly more increased than those molecular marker values measured in the other groups on the day of admission. These values continued to be markedly high up to day 4 of admission. The occurrence rates of disseminated intravascular coagulation in these patient groups were significantly higher than those rates in the other two groups (p = .0001), and the disseminated intravascular coagulation scores did not improve during the study period. The occurrence rates of ARDS (p < .05) and multiple organ dysfunction syndrome (p < .01) were higher in patients with SIRS for>> 3 days compared with those rates in the other groups, and the patients with SIRS for>> 3 days had a poor outcome. No significant difference was noted in the frequency of sepsis among the groups.

CONCLUSIONS

Sustained SIRS is the main determinant for ARDS, multiple organ dysfunction syndrome, and outcome in posttrauma patients. Disseminated intravascular coagulation associated with massive thrombin generation and its activation is involved in the pathogenesis of sustained SIRS. Sepsis has a small role in early posttrauma multiple organ dysfunction syndrome.

OBJECTIVE

To compare the ultrastructure and protein content, particularly <em>prothrombin</em> <em>fragment</em> <em>1</em> and osteopontin, of calcium oxalate monohydrate (COM) and calcium oxalate dihydrate (COD) crystals precipitated from human urine, and their susceptibility to proteolysis, to try to clarify the role of intracrystalline proteins in urolithiasis, as differences between these types of crystal may determine whether calcium oxalate crystals nucleated in urine progress to stone formation.

METHODS

Sodium dodecyl sulphate gel electrophoresis and Western blotting were used to analyse demineralized extracts of COM and/or COD crystals deposited from the same centrifuged and filtered urine (which contains abundant urinary proteins) by adjusting the calcium concentration to 2 and 7 mmol/L, respectively. Similar analyses were performed on COM and COD crystals deposited from ultrafiltered urine (which contains only proteins of < <em>1</em>0 kDa) and then incubated in centrifuged and filtered urine, as well as crystals generated in the presence of increasing concentrations of proteins derived from the organic matrix of urinary calcium oxalate crystals. Field-emission scanning electron microscopy was used to assess effects of proteinase K and cathepsin D on internal and superficial crystal structure.

RESULTS

Osteopontin was undetectable in COM extracts, but clearly visible in COD. Prothrombin <em>fragment</em> <em>1</em> was abundant in COM, but present in COD in lesser amounts than osteopontin. The selectivity was also the same with crystals from ultrafiltered urine that were incubated in centrifuged and filtered urine: <em>prothrombin</em> <em>fragment</em> <em>1</em> binding was favoured by low calcium concentration, while osteopontin bound at higher levels. Scanning electron microscopy of COM and COD digested with proteinase K and cathepsin D revealed superficial and internal texture, as wells as surface erosion, in crystals from centrifuged and filtered urine, thus confirming the presence of intracrystalline proteins. Such features were absent from crystals precipitated from ultrafiltered urine.

CONCLUSIONS

Binding of osteopontin and <em>prothrombin</em> <em>fragment</em> <em>1</em> to calcium oxalate is dictated primarily by ambient calcium concentration. Each protein may inhibit urolithiasis by inhibiting crystallization of its preferred crystal habit, and by facilitating the intracellular disintegration and dissolution of crystals attached to and internalized by renal epithelial cells.

BACKGROUND

Early classification of ischemic stroke subtype is important for secondary stroke prevention and may guide further investigations.

METHODS

Levels of coagulation activation [fibrinopeptide A (FPA), <em>prothrombin</em> <em>fragment</em> <em>1</em>+<em>2</em> (F<em>1</em>+<em>2</em>), thrombin-antithrombin complex (TAT)] and fibrinolysis activation [plasmin-alpha(<em>2</em>)-antiplasmin complex (PAP), D-dimers] markers were measured in 98 consecutive patients with a first-ever acute ischemic stroke admitted within <em>1</em><em>2</em> h after symptom onset.

RESULTS

Median age was 67 years and 44% were women. Median time from symptom onset to blood sampling was 4 h. Stroke subtype was classified as 'cardioembolic' (54%), 'large-artery atherosclerosis' (<em>1</em><em>1</em>%), 'small-vessel disease' (5%), 'other determined' (9%) or 'undetermined etiology' (<em>2</em>0%). Patients with cardioembolic stroke suffered more often from coronary artery disease than patients with other stroke etiologies (40 vs. <em>2</em><em>2</em>%, p = 0.0<em>1</em>9). There were no differences in age, sex, stroke severity, time to blood sampling, frequency of hypertension, diabetes mellitus or current smoking. D-dimers (medians) were higher in patients with cardioembolic strokes than in those with other etiologies (6<em>1</em>5 vs. 3<em>2</em><em>2</em> microg/l, p < 0.00<em>1</em>). No differences in F<em>1</em>+<em>2</em>, FPA, TAT or PAP levels were found. After multivariate analysis, higher D-dimer levels remained independently associated with cardioembolic stroke (p = 0.0<em>2</em><em>2</em>). When measured within 6 h, D-dimers below 300 microg/l excluded cardioembolic stroke with a sensitivity of <em>1</em>00% and a specificity of 5<em>2</em>%.

CONCLUSIONS

Low D-dimer levels in the first few hours make a cardioembolic stroke unlikely, and may be useful to guide further investigations. Other coagulation markers were not useful in differentiating between different stroke etiologies.

Hypercoagulability observed in patients with inflammatory bowel diseases (IBD) may lead to thromboembolic events (TE), which affect the venous and arterial systems alike and are an important factor in patients' morbidity and mortality. The risk of TE in IBD patients has been demonstrated to be approximately three-fold higher as compared to the general population. The pathogenesis of thrombosis in IBD patients is multifactorial and not fully explained. The most commonly listed factors include genetic and immune abnormalities, disequilibrium between procoagulant and anticoagulant factors, although recently, the role of endothelial damage as an IBD-triggering factor is underlined. Several studies report that the levels of some coagulation enzymes, including fibrinogen, factors V, VII, VIII, active factor XI, tissue factor, <em>prothrombin</em> <em>fragment</em> <em>1</em> + <em>2</em> and the thrombin-antithrombin complex, are altered in IBD patients. It has been demonstrated that there is a significant decrease of tissue plasminogen activator level, a marked increase of plasminogen activator inhibitor type <em>1</em> and thrombin-activable fibrinolysis inhibitor, a significantly lower level of antithrombin III and tissue factor pathway inhibitor. IBD patients have been also observed to produce an increased amount of various anticoagulant antibodies. Hyperhomocysteinemia, which is a potential risk factor for TE was also observed in some IBD patients. Further studies are necessary to assess the role of coagulation abnormalities in IBD etiology and to determine indications for thromboprophylactic treatment in patients at high risk of developing TE.

Pregnancy is associated with substantial changes in the haemostatic system and a six-fold higher incidence of venous thromboembolism. Conventional global tests, such as <em>prothrombin</em> time and activated partial thromboplastin time, do not definitely detect this hypercoagulable condition. We investigated whether the changes in haemostatic system during pregnancy are reflected in the calibrated automated thrombography (CAT). Thrombin generation was measured in platelet-poor plasma (PPP) of <em>1</em>50 healthy pregnant women without any pregnancy associated diseases by means of CAT. In addition, <em>prothrombin</em> (FII), antithrombin (AT), protein S, protein C, tissue factor pathway inhibitor (TFPI), plasminogen activator inhibitor-<em>1</em> (PAI-<em>1</em>), thrombin-antithrombin complex (TAT), and <em>prothrombin</em> <em>fragments</em> <em>1</em>+<em>2</em> (F<em>1</em>+<em>2</em>) were measured. Endogenous thrombin potential (ETP) and peak of thrombin generation increased significantly with gestational weeks, while lag time and time to peak remained unchanged. A significant increase of PAI-<em>1</em>, TFPI, F<em>1</em>+<em>2</em> and TAT as well as a significant decrease of free protein S, protein S antigen, and protein S activity was observed. Levels of AT and protein C remained stable during pregnancy. Division of population in trimester of pregnancy and analysis of differences between the trimesters showed rather similar results. Our study shows that endogenous thrombin potential does increase with duration of normal uncomplicated pregnancy. Whether parameters of continuous thrombin generation will correlate with thrombembolic disease remains to be shown.

Normal pregnancy is associated with alterations of the hemostatic system towards a hypercoagulable state and an increased risk of venous thromboembolism. The risk of venous thrombosis is higher in pregnant women with factor V Leiden (FVL) than in those with wildtype factor V. Routine laboratory assays are not useful to detect hypercoagulable conditions. A prospective and systematic evaluation of hemostatic system activation in women with and without FVL during an uncomplicated pregnancy employing more sensitive markers of hypercoagulability, such as <em>prothrombin</em> <em>fragment</em> <em>1</em>+<em>2</em> (F<em>1</em>+<em>2</em>), thrombin-antithrombin complex (TAT), D-Dimer, or the endogenous thrombin potential (ETP), an indicator of the plasma's potential to generate thrombin, has not been performed. We prospectively followed <em>1</em><em>1</em>3 pregnant women with (n = <em>1</em><em>1</em>) and without (n = <em>1</em>0<em>2</em>) FVL and measured F<em>1</em>+<em>2</em>. TAT, D-Dimer and the ETP at the <em>1</em><em>2</em>th, <em>2</em><em>2</em>nd and 34th gestational week as well as 3 months after delivery (baseline) in each subject. None of the women developed clinical signs of venous thromboembolism during pregnancy or postpartum. Pregnant women with and without FVL exhibited substantial activation of the coagulation and fibrinolytic system as indicated by a gradual increase of F<em>1</em>+<em>2</em>, TAT and D-Dimer throughout uncomplicated pregnancy up to levels similar to those found in acute thromboembolic events (p < 0.000<em>1</em> by analysis of variance for each parameters). Levels of F<em>1</em>+<em>2</em> and TAT were comparable between women with and without FVL, but levels of D-Dimer were significantly higher in women with FVL than in those without the mutation (p = 0.0005). The ETP remained unchanged in both women with and without FVL at all timepoints. Our data demonstrate a substantial coagulation and fibrinolytic system activation in healthy women with and without FVL during uncomplicated pregnancy. An elevated F<em>1</em>+<em>2</em>, TAT or D-Dimer level during pregnancy is not necessarily indicative for an acute thromboembolic event. The normal ETP in both women with and without FVL suggests that the capacity of the plasma to generate thrombin after in vitro activation of the clotting system is not affected by pregnancy. Higher levels of D-Dimer in women with FVL than in women with wildtype factor V at baseline as well as during pregnancy indicate increased fibrinolytic system activation in carriers of the mutation.