Evaluation of hypercoagulable state in patients with malignancies by using prothrombin fragment F1 + 2]

In order to elucidate the activation of the coagulation cascade in patients with malignant neoplasms, we measured the levels of plasma prothrombin fragment F1 + 2, which is liberated in the process of thrombin generation. Twenty healthy adults (Group A), 29 patients with malignancies not complicated with DIC (Group B) and 4 patients with DIC (Group C) were evaluated. The values of F1 + 2 in Group C (2.38 +/- 0.55 nmol/l) were significantly higher (p < 0.01) than those in Group A (0.52 +/- 0.19 nmol/l) and B (0.86 +/- 0.68 nmol/l). Many patients in Group B showed higher levels of F1 + 2 compared to normal subjects, however, no significant differences were found between Group A and B. With respect to other coagulation molecular markers such as TAT, D-Dimer and PIC, F1 + 2 levels revealed positive correlation to those levels. Concerning the clinical course of DIC, elevated levels of F1 + 2 normalized much rapidly than those of TAT and D-Dimer by continuous administration of heparin. In conclusion, the measurement of plasma F1 + 2 is important in monitoring the activation of coagulation system in patients with malignancies, especially with respect to early detection and treatment of DIC.     

Procoagulant activity in patients with sickle cell trait

Patients with sickle cell trait (STr) are usually considered to be asymptomatic. However, complications, including hypercoagulability, increased risk of venous thromboembolism and the exertional exercise syndrome with rhabdomyolysis and sudden death, have been described. The exact cause of these adverse events is unclear. We have investigated two patients, a set of monozygotic twins with STr, to establish their procoagulant activity status as a potential indicator of thrombotic risk. In-vivo thrombin generation was assessed by the measurement of prothrombin fragment 1 + 2 (F1 + 2) and thrombin-antithrombin complexes (TAT). D-dimer was used as a marker of fibrinolytic activity. The potential to generate thrombin was determined using an ex-vivo thrombin generation test (TGT). The impact of red blood cell (RBC)-derived microparticle shedding and RBC rheology were examined. TAT (>60 μg/l) and F1 + 2 (948 pmol/l) were markedly elevated in patient 2 but within the normal reference range in patient 1 (TAT = 2.5 μg/l; F1 + 2 = 138 pmol/l). D-dimer levels (0.9 mg/l FEU) were similarly elevated in both patients. TGT peak thrombin and endogenous thrombin potential (ETP) were elevated to similar degrees in both patients. Flow cytometric analysis for RBC-derived microparticles showed that both patients had elevated levels on two occasions. RBC deformability, blood viscosity and RBC aggregation were normal and similar in both patients. The results demonstrated different coagulation activity in the patients with one patient in a prothrombotic state, suggesting that there may be two levels of hypercoagulability in STr. Measurement of such differences would allow for separation of high and low-risk patients from serious complications.     

Prognostic significance of markers of thrombin generation in the acute and chronic phases of non cardioembolic ischemic stroke

BACKGROUND: Previous studies have shown there is activation of the hemostatic system, with thrombin generation, in the acute phase of stroke. Such an activation has unfavourable effects. It is still not known whether there is also a persistent hypercoagulability state in these patients as well as in subjects affected by acute coronary syndromes. METHODS: To know more about the issue, we measured plasma levels of the prothrombin fragment 1 + 2 (F1 + 2) and the thrombin-antithrombin III complex (TAT) in 40 consecutive patients with first ischemic non-cardioembolic stroke; 16 matched involutive cardiomyopathy patients served as the control group. TATs and F1 + 2 were also assessed six months after the onset of stroke symptoms. RESULTS: At baseline stroke patients had higher values than controls of both F1 + 2 and TAT (F1 + 2: 2.38 +/- 2.30 nmol/l vs 1.20 +/- 0.50; p < 0.03; TAT 16.11 +/- 19.60 ng/ml vs 5.51 +/- 4.29; p < 0.05) and these measurements were not related to the typical acute phase reactants. After 6 months F1 + 2 levels in stroke patients were still higher than controls (F1 + 2: 1.68 +/- 0.80 nmol/l vs 1.20 +/- 0.50; p < 0.05), but there were no differences from the baseline levels of F1 + 2 and TAT. According to survival curves mortality was significantly higher in patients with hypercoagulability (defined as F:1 + 2 and TAT levels more than two standard deviations above the mean). CONCLUSIONS: These data confirm that for stroke patients there is sustained activation of the blood coagulation system like in unstable angina and myocardial infarction these abnormalites may have unfavourable prognostic significance.     

Comparison of prothrombin fragment 1 + 2 with thrombin-antithrombin III complex in plasma of patients with disseminated intravascular coagulation.

Plasma levels of prothrombin fragment 1 + 2 (F1 + 2) and thrombin-antithrombin III complex (TAT) are thought to be specific indicators of thrombin generation. To assess whether these two parameters behave similarly in vivo, we compared the plasma levels of F1 + 2 with TAT in 41 patients with disseminated intravascular coagulation (DIC). Both F1 + 2 and TAT were markedly elevated in patients with DIC compared to healthy subjects. Although a positive correlation was found between F1 + 2 and TAT (r = 0.585, P < 0.001) there was a large scatter among individuals. In addition, plasma concentrations of TAT were much lower than F1 + 2. The correlation between the TAT/F1 + 2 ratio and antithrombin III was weak (r = -0.268, P = 0.09). The TAT/F1 + 2 ratio was positively correlated with TAT (r = 0.481, P = 0.002), indicating that the difference in molar concentrations between F1 + 2 and TAT decreased as the TAT value increased. Serial determinations of these parameters showed that plasma TAT values changed roughly in parallel with F1 + 2 in the majority of patients. Although further studies are required to further clarify the observed differences between F1 + 2 and TAT, both parameters would be equally useful for the precise detection of haemostatic activation in patients with DIC.     

Corn trypsin inhibitor in fluorogenic thrombin-generation measurements is only necessary at low tissue factor concentrations and influences the relationship between factor VIII coagulant activity and thrombogram parameters.

The fluorogenic calibrated automated thrombin-generation assay is influenced by contact pathway activation in platelet-rich and platelet-poor plasma. This influence lessens with increasing tissue factor (TF) concentrations and is inhibited by corn trypsin inhibitor (CTI). CTI is expensive and at what TF concentration its influence becomes irrelevant is unclear. Spiking of factor VIII (FVIII)-depleted plasma with FVIII, in samples without CTI, shows a plateau of thrombin generation at low normal FVIII levels. Given the association with thrombosis at high levels, a continuing increase in thrombin generation would be expected. We studied the effect of CTI on this relation by spiking experiments up to 4.8 IU/ml at 1 pmol/l TF and compared the influence of CTI at 1 and 5 pmol/l in platelet-poor plasma. CTI significantly influences thrombin generation in platelet-poor plasma at 1 pmol/l TF (difference of means for endogenous thrombin potential of 232.5 nmol/l per min, P<0.0001) and peak of 48 nmol/l (P<0.0001)) but not at 5 pmol/l. Spiking experiments without CTI confirm the hyperbolic relation between FVIII coagulant activity (FVIII:C) and endogenous thrombin potential with a plateau at 0.70-1.40 IU/ml. With CTI, a near-linear response up to 1.0 IU/ml was found with a plateau at 2.4-4.8 IU/ml. For peak thrombin, no plateau was reached with CTI. The present study confirms and extends previous data on CTI and the relationship between FVIII:C and thrombin generation. CTI is not necessary at 5 pmol/l TF, and thrombin generation remains dependent on FVIII:C up to 4.8 IU/ml at 1 pmol/l with CTI. Higher levels than previously thought may be needed to normalize thrombin generation.     

Studies of the prothrombin activation pathway utilizing radioimmunoassays for the F2/F1 + 2 fragment and thrombin--antithrombin complex

We have evaluated the efficacy of utilizing radioimmunoassays (RIAs) for prothrombin activation fragments (F2/F1 + 2) and for thrombin-- antithrombin complex (TAT) in purified systems and in whole blood. During venipuncture, appropriate anticoagulants were employed in order to prevent the generation of thrombin and factor Xa. The RIAs were shown to be specific for F2/F1 + 2 as well as TAT and did not interact with other plasma components. Initially, thrombin generation was studied in a purified human system of prothrombin, antithrombin, factor Xa, and factor V as well as phospholipid and Ca++. Under these conditions, the kinetics of F2/F1 + 2 and TAT generation were virtually superimposable. However, when factor V was omitted from the reaction mixture, a significantly greater amount of F2/F1 + 2 as compared to TAT was observable. Subsequently, prothrombin activation was monitored during the spontaneous coagulation of freshly drawn blood. Throughout the entire course of thrombin generation, the observable rate of formation of F2/F1 + 2 was considerably greater than that of TAT. We have examined the levels of F2/F1 + 2 and TAT in normal individuals. Our studies indicate that the concentrations of F1 + 2 and TAT average 1.97 nM and 2.32 nM, respectively. We have also quantitated the concentrations of F2/F1 + 2 and TAT in patients with disseminated intravascular coagulation. In these individuals, the levels of both components are elevated. However, the ratio of F1 + 2 to TAT ranges from 2.37 to 5.55. Thus, we conclude that under in vivo conditions, prothrombin activation is characterized by the accumulation of a stable precursor, such as prethrombin-2, and that this phenomenon may be related to an alteration of factor V function.     

Low-density lipoprotein receptor-related protein polymorphisms in patients with elevated factor VIII coagulant activity and venous thrombosis.

Elevated factor VIII coagulant activity (FVIII:C) levels (>150 IU/dl) represent a prevalent independent risk factor for venous thromboembolism (VTE). Low-density lipoprotein receptor-related protein (LRP) is involved in factor VIII clearance in vivo, and elevated FVIII:C was a feature of the LRP knockout mouse model. Three coding polymorphisms of LRP1 (exon 3, C766T; exon 14, A217V; and exon 39, D2080N), together with an insertion/deletion polymorphism within the first intron of lipoprotein receptor-associated protein (LRPAP1), have been identified. In addition, LRP1 2080D was recently reported to be associated with increased plasma FVIII:C levels in normal individuals. In this study, we investigated the role of these four polymorphisms in patients with objectively confirmed VTE and elevated FVIII:C levels. In our control group, genotype distributions were consistent with previous reports. Neither the allele frequencies nor genotype distributions at LRP1 A217V, LRP1 D2080N and LRPAP1 intron 1 were significantly different between the elevated FVIII:C and control groups. In contrast to previous reports, we found no effect of LRP1 D2080N genotype on plasma FVIII:C levels in normal individuals. More importantly, prevalence of the LRP1 2080D allele was not increased in the group of patients with high FVIII:C and VTE. We conclude that LRP1 and LRPAP1 polymorphisms are not responsible for high FVIII:C levels in patients with VTE.     

Prothrombin fragment 1+2 and thrombin–antithrombin complex levels in patients with inherited APC resistance due to factor V Leiden mutation

Inherited activated protein C (APC) resistance is a newly described pathological condition associated with familial thrombophilia. A recent report on a family with APC resistance showed increased levels of prothrombin fragment 1+2 (F1+2) in the affected individuals. No data concerning thrombin–antithrombin complex (TAT) levels in patients with inherited APC resistance are presently available. The aim of this study was to assess the plasma levels of F1+2 and TAT in patients with inherited APC resistance due to factor V (F.V) Leiden mutation and to evaluate F1+2 and TAT levels in symptomatic and asymptomatic patients with the defect (‘carriers’) as compared to their family members having no evidence of F.V Leiden mutation (‘non-carriers’). One hundred and twenty-nine individuals belonging to 30 families with inherited APC resistance due to F.V Leiden mutation were studied. F1+2 and TAT levels were determined using two commercially available ELISA kits and cut-off values were defined as the higher limits of normal ranges obtained in healthy volunteers. Out of the 129 family members investigated, 36 were non-carriers, 85 were heterozygous and eight homozygous for F.V Leiden mutation. Thrombosis had occurred in 2/36 (6%) non-carriers, in 36/85 (42.3%) heterozygous and in 5/8 (63%) homozygous. Median levels of F1+2 and TAT were above cut-off values in carriers, whereas they were below in non-carriers. An overall percentage of 68.8% of carriers exhibited F1+2 levels above the cut-off value as compared to 38.9% of non-carriers. For TAT, an overall percentage of 63.4% of carriers presented with levels above the cut-off compared with 28% of non-carriers. In conclusion, patients with inherited F.V Leiden mutation may exhibit increased levels of F1+2 and TAT. There are no differences in F1+2 and TAT median levels among symptomatic and asymptomatic carriers. The percentage of carriers of F.V Leiden with levels of F1+2 and TAT above cut-off appears to be higher than that found in other clotting inhibitors defects and in this respect the defect might be considered different. However, these findings and the presence of a high percentage of non-carriers presenting with increased F1+2 and TAT levels may suggest the possible coexistence in these families of other unknown defects predisposing to thrombosis.     

Patients with severe sepsis vary markedly in their ability to generate activated protein C

Activated protein C (APC) supplementation significantly reduces mortality in patients with severe sepsis, presumably by down-regulating coagulation, inflammation, and apoptosis. In vivo, endogenous APC is generated from protein C (PC) "on demand" in response to elevated thrombin levels. Thrombomodulin and endothelial cell protein C receptor are endothelial receptors required to generate APC endogenously. Since these receptors may be down-regulated in sepsis, we measured plasma markers of APC generation in 32 patients with severe sepsis to determine whether APC generation is impaired and whether markers of APC generation correlate with 28-day mortality. Relative to normals, all patients had elevated F1 + 2 and thrombin-antithrombin complex (TAT) levels (markers of thrombin generation and inhibition, respectively), and 28 of 32 patients had reduced PC levels. In 20 patients, APC levels paralleled elevated F1 + 2 levels, whereas 12 patients had low APC levels despite elevated F1 + 2 levels, suggesting that APC generation is impaired in the latter. No significant differences exist between survivors and nonsurvivors with respect to baseline PC levels, F1 + 2 levels, and APACHE II (acute physiology and chronic health evaluation) scores. Baseline APC levels were higher in survivors (P = .024), and baseline F1 + 2/APC ratios were lower in survivors (P = .047). Larger studies are warranted to establish whether APC generation profiles aid in managing sepsis.     

Hemostasis activation in thrombophilic subjects with or without a history of venous thrombosis.

Thrombophilia is considered to increase the risk of venous thrombosis (VT) due to hemostasis activation. To determine the level of hemostasis activation in thrombophilic subjects with or without a history of VT, hemostasis activation markers prothrombin fragment 1 and 2 (F1+2), thrombin-antithrombin complex (TAT), and cross-linked fibrin degradation products (D-dimer) were measured in 94 subjects with (patients) and 101 subjects without a history of VT (controls). A total of 34.8% of patients and 14.8% of controls (P= .002) had at least 1 thrombophilic defect (protein C deficiency, activated protein C [APC] resistance, presence of lupus anticoagulants, or prothrombin G20210A polymorphism). The subjects were divided into 4 subgroups: patients with (TF(+) patients) and without (TF(-) patients) thrombophilia, and controls with (TF(+) controls) and without (TF(-) controls) thrombophilia. Hemostasis activation was comparable between all patients and controls (TAT: 2.1 vs 2.6 microg/L; F1+2: 1.0 vs 0.9 nmol/L; D-dimer: 36 vs 37 microg/L, respectively) and between TF(+) and TF(- ) patients. However, TF(+) controls had a significantly higher prevalence of increased hemostasis activation markers compared with TF(-) controls (TAT>4.4 microg/L, 38.4 vs 7.3%; F1+2>1.1 nmol/L, 53.8 vs 22.0%; D-dimer >78 microg/L, 30.7 vs 8.8% of subjects, respectively; all P< .05). After stratification for thrombophilic defects, hemostasis activation was associated with APC resistance in controls and with protein C deficiency in patients. To conclude, thrombophilia was associated with hemostasis activation in controls. We assumed that, in patients, the differences in hemostasis activation between subjects with or without thrombophilia were blurred due to undetermined and unidentified thrombophilic defects.     

Coagulopathy in disseminated intravascular coagulation due to abdominal sepsis: determination of prothrombin fragment 1 + 2 and other markers.

To estimate the degree of coagulopathy in abdominal sepsis, we measured the plasma levels of prothrombin fragment 1 + 2 (F1 + 2), thrombin-antithrombin III complex (TAT) and plasmin-alpha 2-plasmin inhibitor complex (PIC) by the enzyme-linked immunosorbent assay in 38 patients with disseminated intravascular coagulation (DIC). In 20 patients with DIC due to abdominal sepsis, plasma levels of F1 + 2, TAT and PIC were 2.6 nmol/l, 27.9 micrograms/l and 1.5 micrograms/ml, respectively, with a mean antithrombin III (AT III) activity of 41.7%. F1 + 2, TAT, PIC and AT III levels were 4.7 nmol/l, 75.8 micrograms/l, 8.8 micrograms/ml and 70.9% in 18 patients with DIC as the result of malignancy. Though AT III levels in DIC due to sepsis were lower than those in DIC due to malignancy, the levels of F1 + 2, TAT and PIC in the former were not significantly more increased than those in the latter. The plasma levels of F1 + 2 were positively correlated with TAT and PIC in DIC patients with malignancy; however, there was no correlation between F1 + 2 and TAT or PIC in DIC patients with sepsis. In addition, the levels of serum albumin in the two groups were similar. These results suggest that activation of coagulation and fibrinolytic systems may not be so prominent in cases of DIC due to abdominal sepsis, compared to related events in DIC due to malignancy. It is also suggested that the depletion of AT III in cases of sepsis is not only caused by a consumption related to intravascular coagulation or to an alternate distribution of protein.(ABSTRACT TRUNCATED AT 250 WORDS)     

Activated Factor XI is Increased in Plasma in Response to Surgical Trauma but not to Recombinant Activated FVII-Induced Thrombin Formation

Aim: Feedback activation of factor XI (FXI) by thrombin is believed to play a critical role in the amplification phase of thrombin generation and to contribute to thrombosis development and hemostasis. However, the activation of FXI by thrombin has been shown in vitro to require a cofactor. In this study, the role of thrombin in activated FXI (FXIa) formation in vivo is investigated. Methods: The study population comprised probands in whom coagulation activation was triggered by low-dose (15 µg/kg) recombinant activated factor VII (rFVIIa, n =89), of whom 34 with (VTE+) and 45 without a history of venous thromboembolism (VTE−), and patients undergoing major orthopedic surgeries ( n =45). FXIa was quantified via an enzyme capture assay using a monoclonal FXI-specific antibody. Thrombin formation was monitored using an oligonucleotide-based enzyme capture assay and the thrombin activation markers prothrombin fragment 1＋2 (F1＋2) and thrombin–antithrombin complex (TAT). Results: In the rFVIIa cohort, FXIa and thrombin remained below their lower limit of quantification of 3.48 and 1.06 pmol/L, respectively. By contrast, during the surgeries, median FXIa levels increased from 3.69 pmol/L pre-operatively to 9.41 pmol/L mid-operatively ( P =4·10 ⁻⁴ ) and remained significantly elevated 24 h thereafter, with 9.38 pmol/L ( P =0.001). Peak levels of F1+2 were comparable in the VTE+, VTE−, and surgery cohort (235, 268, and 253 pmol/L), whereas peak TAT levels were higher in the surgery cohort (53.1, 33.9, and 147.6 pmol/L). Conclusions: Under in vivo conditions, the activation of FXI requires specific local features that are present at the wounded site including potential cofactors of thrombin.     

Thrombin generation in acute promyelocytic leukemia

Six patients with disseminated intravascular coagulation (DIC) in association with acute promyelocytic leukemia (APL) were studied with sensitive radioimmunoassays that are able to quantitate the extent of thrombin generation within the human circulation. The levels of prothrombin activation fragment, F1 + 2, and thrombin-antithrombin complex (TAT) were obtained at clinical presentation and were then followed serially in several patients during induction chemotherapy. The antileukemic therapy often resulted in a rise in the plasma levels of these molecular species. Simultaneous measurements of fibrinopeptide A (FPA) were also obtained, and the concentrations of this polypeptide were correlated with the levels of F1 + 2 and TAT in patients who were not receiving heparin. Nine individuals with other morphological subtypes of acute nonlymphocytic leukemia (ANLL) were investigated and were usually found to have increased levels of F1 + 2, TAT, and FPA at clinical presentation. However, the magnitude of the elevations was considerably greater and the correlation between TAT and FPA levels was stronger in APL than in ANLL. These studies provide direct evidence that patients with APL, as well as ANLL, generate excessive amounts of thrombin within their vascular system. Furthermore, the data suggest that the concentrations of F1 + 2, compared with the levels of FPA, may be a more sensitive indicator of hemostatic system hyperactivity in individuals with DIC.     

Elevated factor VIII levels and risk of venous thrombosis

Modern thrombophilia testing fails to identify any underlying prothrombotic tendency in a significant number of patients presenting with objectively confirmed venous thromboemboembolism (VTE). This observation has led to a search for other novel inherited or acquired human thrombophilias. Although a number of putative mechanisms have been described, the evidence behind many of these candidates remains weak. In contrast, an increasing body of work supports the hypothesis that increased plasma factor VIII (FVIII) levels may be important in this context. An association between elevated plasma FVIII levels and VTE was first described in the Leiden Thrombophilia Study (LETS). Subsequently, these conclusions have been supported by an increasing number of independent case-control studies. Cumulatively, these studies have clearly demonstrated that high FVIII levels constitute a prevalent, dose-dependent risk factor for VTE. Furthermore, more recent studies have shown that the risk of recurrent venous thrombosis is also significantly increased in patients with high FVIII levels. In this review, we present the evidence supporting the hypothesis that elevated FVIII levels constitute a clinically important thrombophilia. In addition, we examine the biological mechanisms that may underlie persistently elevated FVIII levels, and the pathways through which high FVIII may serve to increase thrombotic risk.     

Thrombin-antithrombin complex and the prothrombin fragment in arterial and venous blood of patients with peripheral arterial disease

BACKGROUND: Thrombin generation has recently been recognized as an important factor in the development of arterial occlusive disease in all vascular provinces. Several reports concerning markers of thrombin generation have been published with however, conflicting results. It has been demonstrated in vitro that accelerated blood flow velocity causes increased thrombin generation via higher shear rates. In recent articles TAT and F1 + F2 concentrations are reported significantly higher in arterial than in venous blood. A correlation with the severity of atherosclerosis or specially with the stage of PAD was expected. In the present investigation we additionally collected blood from the femoral vein. PATIENTS AND METHODS: In 11 patients with Fontaine stages IIb to IV and two healthy subjects TAT and F1 + F2 concentrations were determined in blood samples from the femoral artery, the femoral vein (diseased leg) and cubital vein. In all cases and at all puncture sites exactly the same atraumatic technique of venipuncture was used. RESULTS: The concentrations of TAT and F1 + F2 were significantly elevated in patients with PAD. There was no significant difference between the concentrations of TAT and F1 + F2 in arterial (femoral artery) and venous (femoral vein and cubital vein) blood. CONCLUSION: The results from previous investigations have been confirmed only partially. Differences in the puncture techniques to collect arterial or venous blood result in an increased scattering of the data and a systematic error.     

Effects of ximelagatran,an oral direct thrombin inhibitor,r-hirudin and enoxaparin on thrombin generation and platelet activation in healthy male subjects

OBJECTIVES: The effects of ximelagatran, an oral direct thrombin inhibitor (DTI), recombinant hirudin (r-hirudin) and enoxaparin on thrombin generation and platelet activation were studied in humans. BACKGROUND: Recombinant hirudin (parenteral DTI) and enoxaparin (low molecular weight heparin) have been demonstrated to be clinically effective in acute coronary syndromes. Ximelagatran is currently under investigation for the prevention and treatment of thromboembolism. The shed blood model allows for the study of thrombin generation and platelet activation in humans in vivo. METHODS: This was an open-label, parallel-group study involving 120 healthy male volunteers randomized to receive one of three oral doses of ximelagatran (15, 30 or 60 mg), r-hirudin (intravenous) or enoxaparin (subcutaneous) at doses demonstrated to be clinically effective in acute coronary syndromes, or to serve as a control. Thrombin generation (prothrombin fragment 1+2 [F1+2] and thrombin-antithrombin complex [TAT]) and platelet activation (beta-thromboglobulin [beta-TG]) biomarkers were studied using a shed blood model involving blood collection from skin incisions made using standardized bleeding time devices. RESULTS: Oral ximelagatran, intravenous r-hirudin and subcutaneous enoxaparin rapidly and significantly (p < 0.05) decreased F1+2, TAT and beta-TG levels in shed blood, indicating inhibition of thrombin generation and platelet activation. Statistically significant concentration (melagatran, the active form of ximelagatran)-response relationships for F1+2 (p = 0.005), TAT (p = 0.005) and beta-TG (p < 0.001) levels, with IC(50)s of 0.376 (F1+2), 0.163 (TAT) and 0.115 (beta-TG) micromol/l, were detected. Melagatran showed dose-proportional pharmacokinetics with low variability. All drugs were well tolerated. CONCLUSIONS: Oral administration of the DTI ximelagatran resulted in a rapid inhibition of both thrombin generation and platelet activation in a concentration-dependent manner using a human shed blood model. The inhibition of thrombin generation by 60 mg ximelagatran was comparable to that observed with doses of r-hirudin and enoxaparin demonstrated to be effective for the treatment of acute coronary syndromes.     

Assessment of markers of thrombin generation in patients with acute myocardial infarction complicated by ventricular fibrillation

BACKGROUND: In most cases, sudden cardiac death is triggered by ischemia-related ventricular tachyarrhythmias and accounts for 50% of deaths from cardiovascular disease in developed countries. Chronic elevation of indicators of coagulation activation has been found in patients with coronary heart disease, but a role of coagulation activation as a potential risk factor for ventricular fibrillation (VF) during acute myocardial infarction (MI) has not been investigated. METHODS: We enrolled 50 patients with a history of MI, of whom 26 presented with VF in the acute phase of myocardial ischemia; 24 patients had an acute MI without ventricular tachyarrhythmias. Levels of thrombin-antithrombin complexes (TAT), prothrombin fragment F1 + 2 (F1 + 2), fibrinopeptide A (FPA), plasmin-antiplasmin complexes (PAP), protein C, antithrombin, activated partial thromboplastin time (aPTT), thromboplastin time, D-Dimer, fibrinogen, and high-sensitivity C-reactive protein (hs-CRP) were measured in plasma samples of all patients. Blood collection was obtained sequentially in two separate settings. Patients were studied at a median of 351 days after the acute coronary event. RESULTS: Higher levels of TAT complexes (13.4 +/- 22.2 vs. 3.03 +/- 4.3 microg/l; p = 0.02), FPA (79.7 +/- 132.3 vs. 24.04 +/- 41.3 ng/ml; p = 0.04), and F1+2 (1.89 +/- 1.3 vs. 1.16 +/- 0.5 nmol/l; p = 0.01) were observed in patients with VF compared with patients without ventricular tachyarrhythmias during the acute phase of MI. D-Dimer levels displayed a trend without reaching statistical significance (0.69 +/- 0.48 vs. 0.48 +/- 0.24 mg/l; p = 0.06). No differences were found in hs-CRP (3.25 +/- 4.5 vs. 4.4 +/- 8.8 mg/l; p = 0.5) and fibrinogen (2.8 +/- 0.9 vs. 2.7 +/- 0.9 g/l; p = 0.6) measurements. Repeat assessment of markers of coagulation activation at a median of 847 days revealed a highly significant decrease in patients with VF. CONCLUSIONS: Markers of thrombin generation are transiently increased in patients with VF during the acute phase of MI. These findings have implications for risk assessment and genetic screening of patients prone to VF during acute myocardial ischemia.     

Procoagulant activity induced by vascular injury determines contribution of elevated factor VIII to thrombosis and thrombus stability in mice

Studies have correlated elevated plasma factor VIII (FVIII) with thrombosis; however, it is unclear whether elevated FVIII is a proinflammatory biomarker, causative agent, or both. We raised FVIII levels in mice and measured the time to vessel occlusion (TTO) after ferric chloride-induced injury. Compared with control (saline-infused) mice, elevated FVIII had no effect after longer (3-minute) carotid artery injury, but it shortened the TTO after shorter (2-minute) injury (P < .008). After injury, circulating thrombin-antithrombin (TAT) complexes were lower after short versus long injury (P < .04), suggesting short treatment produced less coagulation activation. TAT levels in FVIII-infused mice were higher than in controls after short, but not longer, injury. Accordingly, elevated FVIII had no effect on in vitro thrombin generation or platelet aggregation triggered by high tissue factor, but it increased thrombin generation rate and peak (2.4- and 1.5-fold, respectively), and it accelerated platelet aggregation (up to 1.6-fold) when initiated by low tissue factor. Compared with control mice, elevated FVIII stabilized thrombi (fewer emboli) after short injury, but it had no effect after longer injury. TTO and emboli correlated with TATs. These results demonstrate dependence of FVIII activity on extent of vascular injury. We propose elevated plasma FVIII is an etiologic, prothrombotic agent after moderate but not extensive vascular damage.     

Prothrombin fragments F1+2, thrombin-antithrombin III complexes, fibrin monomers and fibrinogen in patients with coronary atherosclerosis

We determined the plasma levels of prothrombin fragment F1+2, thrombin-antithrombin III complexes (TAT), fibrin monomers (FM), D-dimers (DD) and fibrinogen in 57 patients with angiographically verified graded coronary artery disease (CAD) free of concomitant peripheral atherosclerosis, cerebrovascular disease or diabetes mellitus and a group of 21 apparently healthy controls. Blood was collected from the antecubital vein through atraumatic venipuncture prior to the angiographic procedure. Plasma levels of hemostatic markers were related to the presence and graded severity of CAD. The levels of prothrombin fragment F1+2 (1.74+/-0.11 vs. 1.0+/-0.07 nmol/l, P<0.001), FM (41.6+/-5.5 vs. 7.42+/-3.05 nmol/l, P<0.001), TAT (15.6+/-2.7 vs. 2.96+/-0.32 microg/l, P<0.001) and fibrinogen (3.64+/-1.3 vs. 3.08+/-0.33 g/l, P<0.01) were significantly higher in patients with CAD compared to controls, while there was no difference regarding the fibrinolytic system represented by DD (441.6+/-58.9 vs. 337.4+/-42.05 microg/l, n.s.). Within the CAD group, patients with extensive coronary atherosclerosis (> or =2 vessel disease) had significantly higher values for prothrombin fragment F1+2 (1.89 vs. 1.57 nmol/l, P = 0.04), FM (50.7 vs. 29.8 nmol/l, P = 0.03), and a trend to significance was noted for fibrinogen (3.9 vs. 3.3 g/l, P = 0.07) suggesting that blood coagulability was related to the severity of the disease and that hemostatic markers of thrombin activity represent a useful tool to identify patients with a latent hypercoagulable state with a higher susceptibility to sustain coronary thrombosis.     

Plasma levels of factor XIIa and factor VIIa are increased but not related in primary hyperlipidaemia.

The lipolysis of triglyceride-rich lipoproteins may provide a surface that supports the activation of factor XII (FXII) with subsequent activation of factor VII (FVII). Plasma levels of activated FVII (FVIIa) but not activated FXII (FXIIa) are increased in the post-prandial state when there is a transient increase in triglyceride levels. We compared plasma levels of FXIIa antigen in control subjects (n = 33) and in patients with chronically elevated lipids (primary hyperlipidaemia, n = 49), with FVIIa and markers of thrombin generation. Results are given as median (first and third quartiles). Plasma levels of FXIIa [2.34 (1.68-3.32) ng/ml versus 1.53 (0.93-1.86) ng/ml, P = 0.0002], FVIIa [3.02 (2.15-4.64) ng/ml versus 2.20 (1.66-2.56) ng/ml, P = 0.0004], thrombin-antithrombin complexes [3.08 (2.16-5.54) microg/I versus 2.13 (1.46-2.84) microg/l, P = 0.005] and prothrombin fragment 1 + 2 (Pro F1 + 2) [1.28 (1.08-1.50) nmol/l versus 0.92 (0.65-1.08) nmol/l, P = 0.0001] were increased compared with controls irrespective of the type of hyperlipidaemia. In hyperlipdaemic subjects, levels of Pro F1 + 2 were correlated with FVIIa (r = 0.56, P = 0.0002) and FXIIa (r = 0.31, P = 0.03). These results suggest increased activation of both FVII and FXII in hyperlipidaemic subjects, which correlates with increased thrombin generation. Given the lack of correlation between levels of FXIIa and FVIIa, it remains to be established whether the increase in FXIIa is responsible for increased FVIIa activity in this subject group.