Thrombin generation is pivotal to both physiological blood clot formation and pathological development of disseminated intravascular coagulation (DIC). In critical illness, extensive cell damage can release histones into the circulation, which can increase thrombin generation and cause DIC, but the molecular mechanism is not clear. Typically, thrombin is generated by the <em>prothrombin</em>ase complex, comprising activated factor X (FXa), activated cofactor V (FVa), and phospholipids to cleave <em>prothrombin</em> in the presence of calcium. In this study, we found that in the presence of extracellular histones, an alternative <em>prothrombin</em>ase could form without FVa and phospholipids. Histones directly bind to <em>prothrombin</em> <em>fragment</em> <em>1</em> (F<em>1</em>) and <em>fragment</em> <em>2</em> (F<em>2</em>) specifically to facilitate FXa cleavage of <em>prothrombin</em> to release active thrombin, unlike FVa, which requires phospholipid surfaces to anchor the classical <em>prothrombin</em>ase complex. In vivo, histone infusion into mice induced DIC, which was significantly abrogated when <em>prothrombin</em> F<em>1</em> + F<em>2</em> were infused prior to histones, to act as decoy. In a cohort of intensive care unit patients with sepsis (n = <em>1</em>44), circulating histone levels were significantly elevated in patients with DIC. These data suggest that histone-induced alternative <em>prothrombin</em>ase without phospholipid anchorage may disseminate intravascular coagulation and reveal a new molecular mechanism of thrombin generation and DIC development. In addition, histones significantly reduced the requirement for FXa in the coagulation cascade to enable clot formation in factor VIII (FVIII)- and FIX-deficient plasma, as well as in FVIII-deficient mice. In summary, this study highlights a novel mechanism in coagulation with therapeutic potential in both targeting systemic coagulation activation and correcting coagulation factor deficiency.

Activation of coagulation during hemodialysis (HD) is a relevant clinical problem, especially when patients at risk of bleeding are treated. However, little is known about the relative contribution of the various components of the circuit to the thrombotic process. Thus, an experimental model was developed that is aimed at evaluating biochemical markers of coagulation activation at different times and sites throughout the HD circuit. A HD blood-tubing set with integrated arterial and venous chambers (cartridge-line set) was used, which was added with the following sampling points: at the beginning of the arterial line (P<em>1</em>), before the blood pump (P<em>2</em>), after the blood pump (P3), and at the end of the venous line (P4). A bypass system allowed us to circulate the blood only into the blood lines for the first <em>2</em>0 min of the extracorporeal circulation. The extracorporeal circuit was rinsed with <em>1</em>.7 L of heparinized saline (<em>2</em>,500 IU/L) that was completely discarded before patient connection. A continuous administration of unfractionated heparin (500-800 IU/h) without a starting bolus was adopted as a low heparin extracorporeal treatment. Samples were collected before the start of the extracorporeal circulation from the fistula needle (T0P0), after 5 (T<em>1</em>), <em>1</em>0 (T<em>2</em>), and <em>2</em>0 min (T3) from P<em>1</em>, P<em>2</em>, P3, and P4. After <em>2</em>0 min, the blood was returned to the patient using only saline and HD was then started, circulating the blood through the dialyzer. Further samples were obtained from P<em>1</em> and P4 after 5 (T4) and <em>2</em><em>1</em>0 min (T5). Plasma levels of coagulation activation markers-thrombin-antithrombin complex (TAT) and <em>prothrombin</em> <em>fragment</em> <em>1</em> + <em>2</em> (F<em>1</em> + <em>2</em>)-were evaluated in all the samples in <em>1</em><em>2</em> stable HD patients. In each patient, the activated partial thromboplastin time (APTT) was measured at T0P0 and T<em>1</em>-T5 from P<em>1</em>. No significant changes were found at any time as far as F<em>1</em> + <em>2</em> is concerned. However, TAT levels increased over time only after the start of HD, suggesting that the latter test could be more useful in order to detect coagulation activation during HD. The same experiments performed with nonheparin-primed extracorporeal circuit showed similar results. The blood lines used did not significantly activate coagulation during the first <em>2</em>0 min, whereas only 5 min of blood circulation throughout the whole circuit increased TAT values, which still remained lower than previous reports, even after <em>2</em><em>1</em>0 min of treatment.

BACKGROUND

Lysinuric protein intolerance (LPI), a rare autosomal recessive transport disorder of cationic amino acids lysine, arginine and ornithine, affects intestines, lungs, liver and kidneys. LPI patients may display potentially life-threatening bleeding events, which are poorly understood.

OBJECTIVE

To characterize alterations in haemostatic and fibrinolytic variables associated with LPI.

METHODS

We enrolled <em>1</em>5 adult patients (8 female) and assessed the clinical ISTH/SSC-BAT bleeding score (BS). A variety of metabolic and coagulation assays, including fibrin generation test derivatives, clotting time (CT) and clot lysis time (CLT), thromboelastometry (ROTEM), and PFA-<em>1</em>00 and Calibrated Automated Thrombogram (CAT), were used.

RESULTS

All patients had mild-to-moderate renal insufficiency, and moderate bleeding tendency (BS 4) without spontaneous bleeds. Mild anaemia and thrombocytopenia occurred. Traditional clotting times were normal, but in contrast, CT in fibrin generation test, and especially ROTEM FIBTEM was abnormal. The patients showed impaired primary haemostasis in PFA, irrespective of normal von Willebrand factor activity, but together with lowered fibrinogen and FXIII. Thrombin generation (TG) was reduced in vitro, according to CAT-derived endogenous thrombin potential, but in vivo TG was enhanced in the form of circulating <em>prothrombin</em> <em>fragment</em> <em>1</em> and <em>2</em> values. Very high D-dimer and plasmin-α<em>2</em>-antiplasmin (PAP) complex levels coincided with shortened CLT in vitro.

CONCLUSIONS

Defective primary haemostasis, coagulopathy, fibrin abnormality (FIBTEM, CT and CLT), low TG in vitro and clearly augmented fibrinolysis (PAP and D-dimer) in vivo were all detected in LPI. Altered fibrin generation and hyperfibrinolysis were associated with the metabolic and renal defect, suggesting a pathogenetic link in LPI.

Thrombin and plasmin are the key enzymes involved in coagulation and fibrinolysis. A novel hemostasis assay (NHA) was developed to measure thrombin and plasmin generation in a single well by a fluorimeter. The NHA uses two fluorescent substrates with non-interfering fluorescent excitation and emission spectra. The assay was tested in vitro using modulators like heparin, hirudin, epsilon-aminocaproic acid, gly-pro-arg-pro peptide and reptilase and validated by measurement of <em>prothrombin</em> <em>fragment</em> <em>1</em>+<em>2</em> and plasmin-alpha<em>2</em>-antiplasmin levels. Intra- and inter-assay coefficients of variation were < 9% and 6-<em>2</em>5%, respectively. Interplay between coagulation and fibrinolysis was demonstrated by the effect of tissue-type plasminogen activator on thrombin generation and by the different responses of activated protein C and thrombomodulin on fibrinolysis. The last responses showed the linkage between coagulation and fibrinolysis by thrombin activatable fibrinolysis inhibitor. In conclusion, this strategy allows detection of coagulation, fibrinolysis and their interplay in a single assay.

BACKGROUND

Paroxysmal Nocturnal Hemoglobinuria is characterized by complement-mediated hemolysis and an increased thrombosis risk. Eculizumab, an antibody to complement factor C5, reduces thrombotic risk via unknown mechanisms. Clinical observations suggest that eculizumab has an immediate effect.

OBJECTIVE

A better understanding of the mechanism via which eculizumab reduces thrombotic risk by studying its pharmacodynamic effect on coagulation and fibrinolysis.

METHODS

We measured microparticles (MP), tissue factor (TF) activity, <em>prothrombin</em> <em>fragment</em> <em>1</em>+<em>2</em> (F<em>1</em>+<em>2</em>), D-dimer and simultaneously thrombin and plasmin generation in 55 PNH patients. In <em>2</em>0 patients, parameters were compared before and during eculizumab treatment (at <em>1</em> and <em>2</em>hours, <em>1</em>, 4 and≥<em>1</em><em>2</em>weeks after commencement).

RESULTS

Patients with a history of thrombosis had elevated D-dimers (p=0.0<em>2</em>) but not MP. Among patients on anticoagulants, those with thrombosis had higher F<em>1</em>+<em>2</em> concentrations (p=0.003). TF activity was undetectable in plasma MP. Unexpectedly, thrombin peak height and thrombin potential were significantly lower in PNH patients than in healthy controls. Fibrinolysis parameters were normal. During eculizumab treatment D-dimer levels significantly decreased after <em>1</em>hour (p=0.008) and remained decreased at≥<em>1</em><em>2</em>weeks (p=0.03). F<em>1</em>+<em>2</em> (p=0.03) and thrombin peak height (p=0.0<em>2</em>) in patients not on anticoagulants significantly decreased at≥week <em>1</em><em>2</em>. MP remained unchanged.

CONCLUSIONS

Eculizumab induces an immediate decrease of D-dimer levels but not of other markers. The decrease in thrombin peak height and F<em>1</em>+<em>2</em> suggests that eculizumab reduces thrombin generation. Elevated D-dimer levels in untreated PNH patients with a history of thrombosis suggest possible value in predicting thrombotic risk.

Thromboembolic complications rarely occur during infancy and childhood. It has been reported that increased capacity of cord plasma to inhibit thrombin due to elevated alpha(<em>2</em>)-macroglobulin (alpha(<em>2</em>)-M) levels may in part provide protection from thrombosis. In antithrombin (AT)-deficient plasma, alpha(<em>2</em>)-M exhibits anticoagulant action by complexing substantial amounts of generated free thrombin. It has been suggested that alpha(<em>2</em>)-M has the same impact on thrombin inhibition as AT, the most important thrombin inhibitor in adult plasma. The aim of our study was to examine this assumption by determining time-courses of free thrombin generation and <em>prothrombin</em> activation. Additionally, the amount of thrombin complexed to alpha(<em>2</em>)-M was assessed by comparing the heights of the end-level of amidolytic activity curves (AACs) after extrinsic activation of platelet poor plasma in the presence of different concentrations of AT or alpha(<em>2</em>)-M. Increasing the AT content by 30% resulted in significantly suppressed generation of free thrombin and <em>prothrombin</em> <em>fragment</em> <em>1</em>+<em>2</em> (F<em>1</em>+<em>2</em>) in cord and adult plasma. In contrast, increasing the alpha(<em>2</em>)-M content in plasma containing physiologic amounts of AT by the same percentage had no effect on free thrombin generation and on F<em>1</em>+<em>2</em> generation in both cord and adult plasma. In addition, the effect of AT supplementation on the end-level of the AACs was significantly higher compared to the effect of alpha(<em>2</em>)-M supplementation. Since alpha(<em>2</em>)-M, in contrast to AT, had no effect on free thrombin generation and <em>prothrombin</em> activation, our study suggests that the action between alpha(<em>2</em>)-M and thrombin might not be fast enough to prevent thrombin from its feedback activation in both cord and adult plasma and, therefore, in cord and adult plasma containing physiological amounts of AT alterations of the alpha(<em>2</em>)-M content had no effect on thrombin generation and inhibition.

Venous thromboembolism (VTE) is a common problem in cancer patients. However, the rates of VTE vary widely between different types of malignancies. Furthermore, patient- and treatment-related risk factors contribute to the risk of VTE. The prediction of the individual risk of VTE and the identification of patients with cancer that might benefit from thromboprophylaxis is a major clinical challenge, because the pathogenesis of cancer-associated VTE is multifactorial. Therefore, recent studies have focused on identification of predictive biomarkers for VTE. As there is a close interrelation between cancer and the haemostatic system, which leads to activation of haemostasis and fibrinolysis, biomarkers of the haemostatic system seem to be promising in predicting risk of developing VTE in cancer patients. Some candidate biomarkers or laboratory tests of the haemostatic system including platelet count, soluble P-selectin, tissue factor (TF) and TF-bearing microparticles, coagulation factor VIII, D-dimer, <em>prothrombin</em> <em>fragment</em> <em>1</em>+<em>2</em> and the thrombin generation assay have been reported to predict cancer-associated VTE. In addition, it has been shown that risk-scoring models, incorporating clinical parameters and biomarkers, allow risk stratification of cancer patients into groups at high- and at low risk of VTE. The accuracy of a previously established risk scoring model can be improved when it is expanded and biomarkers of the haemostatic system are added. The benefit of a targeted thromboprophylaxis for primary prevention of VTE in cancer patients based on risk assessment by measuring biomarkers or applying risk scoring models has to be shown in interventional clinical trials.

Tumor progression is associated with aberrant hemostasis, and patients with malignant diseases have an elevated risk of developing thrombosis. A crosstalk among the vascular endothelium, components of the coagulation cascade, and cancer cells transforms the intravascular milieu to a prothrombotic, proinflammatory, and cell-adhesive state. We review the existing evidence on activation of the coagulation system and its implication in genitourinary malignancies and discuss the potential therapeutic benefit of antithrombotic agents. A literature review was performed searching the Medline database and the Cochrane Library for original articles and reviews. A second search identified studies reporting on oncological benefit of anticoagulants in genitourinary cancer. An elevated expression of procoagulatory tissue factor on tumor cells and tumor-derived microparticles seems to stimulate cancer development and progression. Several components of the hemostatic system, including D-dimers, von Willebrand Factor, thrombin, fibrin-/ogen, soluble P-selectin, and <em>prothrombin</em> <em>fragments</em> <em>1</em> + <em>2</em> were either overexpressed or overactive in genitourinary cancers. Hypercoagulation was in general associated with a poorer prognosis. Experimental models and small trials in humans showed reduced cancer progression after treatment with anticoagulants. Main limitations of these studies were heterogeneous experimental methodology, small patient numbers, and a lack of prospective validation. In conclusion, experimental and clinical evidence suggests procoagulatory activity of genitourinary neoplasms, particularly in prostate, bladder and kidney cancer. This may promote the risk of vascular thrombosis but also metastatic progression. Clinical studies linked elevated biomarkers of hemostasis with poor prognosis in patients with genitourinary cancers. Thus, anticoagulation may have a therapeutic role beyond prevention of thromboembolism.

BACKGROUND

Surgery for benign disease is associated with a low-risk of developing venous thromboembolism (VTE). Despite a relatively low incidence of postoperative VTE in patients after elective cholecystectomy and abdominal hernia repair there are data proving hypercoagulability in the early postoperative period. We focused on assessment of the systemic inflammatory response and coagulation status in these surgical patients after hospital discharge.

METHODS

Prospectively, patients who underwent surgery for benign disease were included. Two hundred sixteen patients were enrolled - 90 patients in laparoscopic cholecystectomy (LC) group and <em>1</em><em>2</em>6 patients in hernia surgery (HS) group. Risk assessment of VTE according to the Caprini risk assessment model was performed in all patients. Prevalence of VTE in postoperative period was observed. Markers of systemic inflammatory response (IL-6, CRP, α-<em>1</em>-acid glycoprotein, transferrin) and coagulation markers (PLT, fibrinogen, <em>prothrombin</em> <em>fragment</em> F<em>1</em> + <em>2</em> and D-dimer) were measured before surgery, on 7-<em>1</em>0th postoperative day and on <em>2</em>8-30th postoperative day.

RESULTS

Clinically apparent deep vein thrombosis was diagnosed in only one patient - 0.46%. Statistically significant elevation of inflammatory markers IL-6, CRP and α-<em>1</em>-acid glycoprotein (p < 0.00<em>1</em>; all) were proved in both groups of patients on 7-<em>1</em>0th postoperative day. Statistically significant elevation of coagulation markers PLT, fibrinogen, <em>prothrombin</em> <em>fragment</em> F<em>1</em> + <em>2</em> and D-dimer (p < 0.00<em>1</em>; all) were proved in LC and HS groups on 7-<em>1</em>0th postoperative day. No statistical difference was observed in IL-6, CRP and α-<em>1</em>-acid glycoprotein levels a month after surgery as compared with preoperative levels within each group. Statistically significant elevation of fibrinogen and <em>prothrombin</em> <em>fragment</em> F<em>1</em> + <em>2</em> levels (p < 0.00<em>1</em>; both) persisted on <em>2</em>8-30th postoperative day in both groups. Persisted elevation of D-dimer levels was proved only in HS group (p < 0.00<em>1</em>), not in LC group (p = 0.<em>1</em>38), a month after surgery.

CONCLUSIONS

Activated systemic inflammatory response and hypercoagulable condition were verified in patients after laparoscopic cholecystectomy and hernia surgery after their hospital discharge. Hypercoagulability persisted even a month after surgery. Nevertheless, we observed very low prevalence of clinically apparent VTE in patients with in-hospital postoperative VTE prophylaxis.

BACKGROUND

Trials of the Czech Ministry of Health No. RVO-VFN64<em>1</em>65 and NT <em>1</em>3<em>2</em>5<em>1</em>-4 .

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.

METHODS

Sixteen healthy male or female volunteers with a mean age of <em>2</em>6 ± 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 <em>1</em>0 mg rivaroxaban. Circadian rhythms of <em>prothrombin</em> <em>fragment</em> <em>1</em> + <em>2</em>, 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.

RESULTS

Concentrations of rivaroxaban were higher <em>1</em><em>2</em> h after evening intake of rivaroxaban than <em>1</em><em>2</em> h after morning intake (53.3 ng mL(-<em>1</em>) [95% confidence interval 46.0-67.8] vs. <em>2</em>3.3 ng mL(-<em>1</em>) [<em>1</em>9.4-<em>2</em>9.<em>1</em>, respectively]). Rivaroxaban intake in the evening reduced morning F<em>1</em>+<em>2</em> concentrations better at 8:00 AM than did administration on awakening (85 ± <em>2</em>5 nmol L(-<em>1</em>) vs. <em>1</em>06 ± 34 nmol L(-<em>1</em>) , CI: 9.4-3<em>2</em>.<em>1</em>). 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.

BACKGROUND

The pharmacological inhibition of blood-foreign surface interactions is an attractive strategy for reducing the morbidity associated with cardiopulmonary bypass. We compared the inhibitory effects of nafamostat mesilate (a broad-spectrum synthetic protease inhibitor) and minimal-dose aprotinin on blood-surface interactions in clinical cardiopulmonary bypass.

METHODS

Eighteen patients undergoing coronary surgery were divided into three groups: (<em>1</em>) the control group (heparin, 4 mg/kg; n = 6), (<em>2</em>) the nafamostat mesilate group (heparin plus nafamostat, 0.<em>2</em> mg/kg bolus followed by <em>2</em>.0 mg/kg/h during cardiopulmonary bypass; n = 6), and (3) the aprotinin group (heparin plus aprotinin, <em>2</em>.0 x <em>1</em>0(4) KIU/kg; n = 6). Platelet count, platelet aggregation, beta-thromboglobulin, <em>prothrombin</em> <em>fragment</em> F<em>1</em>.<em>2</em>, thrombin-antithrombin complex, plasminogen activator inhibitor-<em>1</em>, alpha<em>2</em>-plasmin inhibitor-plasmin complex, D-dimer, neutrophil elastase, and interleukin-6 were measured before, during, and after bypass. Bleeding times and blood loss were recorded.

RESULTS

There were no significant differences between groups in platelet count, beta-thromboglobulin, plasminogen activator inhibitor-<em>1</em>, interleukin-6, bleeding times, or blood loss. Platelet aggregation was better preserved at <em>1</em><em>2</em> hours after surgery in the nafamostat and aprotinin groups than in the control group. Prothrombin <em>fragment</em> F<em>1</em>.<em>2</em>, thrombin-antithrombin complex and neutrophil elastase levels were significantly reduced by aprotinin, but not by nafamostat as compared with the control group. The alpha<em>2</em>-plasmin inhibitor-plasmin complex and D-dimer were significantly lower with either of the drugs. Aprotinin showed better control of D-dimer than did nafamostat.

CONCLUSIONS

Nafamostat mesilate fails to reduce thrombin formation and neutrophil elastase release, whereas minimal-dose aprotinin inhibits both. Neither nafamostat nor aprotinin inhibits platelet activation. Nafamostat reduces fibrinolysis during cardiopulmonary bypass, although its effect is not as potent as aprotinin.

Replacement therapy with missing factor (F) VIII or IX in haemophilia patients for bleed management and preventative treatment or prophylaxis is standard of care. Restoration of thrombin generation through novel mechanisms has become the focus of innovation to overcome limitations imposed by protein replacement therapy. Tissue factor pathway inhibitor (TFPI) is a multivalent Kunitz-type serine protease inhibitor that regulates tissue factor (TF)-induced coagulation through a FXa-dependent feedback inhibition of the TF.FVIIa complex in plasma and on endothelial surfaces. Concizumab is a monoclonal, humanised antibody, specific for the second Kunitz domain of TFPI that binds and inhibits FXa, abolishing the inhibitory effect of TFPI. Concizumab restored thrombin generation in FVIII and FIX deficient plasmas and decreased blood loss in a rabbit haemophilia model. Phase <em>1</em> single and multiple dose escalation studies in haemophilia patients demonstrated a dose dependent decrease in TFPI levels and a pro-coagulant effect with increasing d-dimers and <em>prothrombin</em> <em>fragment</em> <em>1</em> + <em>2</em>. A dose dependent increase in peak thrombin and endogenous thrombin potential was observed with values in the normal range when plasma TFPI levels were nearly undetectable. A few haemophilia patients in the highest dose cohorts with complete inhibition of plasma TFPI showed a decreased fibrinogen concentration with normal levels of anti-thrombin and platelets and no evidence of thrombosis. Pharmacokinetic parameters were influenced by binding to the target (TFPI), demonstrating target mediated drug disposition. A trend towards decreasing bleeding tendency was observed and this preventative effect is being studied in Phase <em>2</em> studies with additional data gathered to improve our understanding of the therapeutic window and potential for thrombosis.

<AbstractText>Comparative studies on the restoration of hemostasis with different reversal agents after dabigatran therapy have not been performed. We compared the efficacy and prothrombotic potential of the specific antidote idarucizumab with that of previously recommended non-specific procoagulant concentrates.</AbstractText><p><div><b>STUDY DESIGN AND METHODS</b></div>We explored the in vitro effects of dabigatran (<em>1</em>84 ng/mL) on fibrin and platelet-aggregate formation onto a damaged vessel under flow conditions (600 s^-<em>1</em> ). The reversal mechanisms and efficacy of idarucizumab (0.3-3 mg/mL) were compared with that of the non-specific procoagulant concentrates aPCC (<em>2</em>5-75 U/Kg), PCC (70 U/Kg), or rFVIIa (<em>1</em><em>2</em>0 μg/Kg). Generation of thrombin and <em>prothrombin</em> <em>fragment</em> (F<em>1</em> + <em>2</em>), and thromboelastometry parameters of clot formation were measured.</p><AbstractText>Dabigatran caused pronounced reductions in fibrin (87%) and platelet interactions (36%) with damaged vessels (p < 0.0<em>1</em>) and significantly impaired thrombin generation and thromboelastometric parameters (delayed dynamics and reduced firmness). Idarucizumab completely normalized rates of fibrin and platelet coverage to baseline values in flow studies; and reversed the alterations in thrombin generation, F<em>1</em> + <em>2</em> and thromboelastometry parameters produced by dabigatran. In comparison, aPCC and PCC only partially compensated for the dabigatran-induced alterations in fibrin deposition, but were unable to fully restore them to baseline values. Reversal with aPCC or PCC improved the majority of alterations in coagulation-related tests, but tended to overcompensate thrombin generation kinetics and significantly increased F<em>1</em> + <em>2</em> levels.</AbstractText><AbstractText>Idarucizumab antagonizes alterations of direct and indirect biomarkers of hemostasis caused by dabigatran. In our studies, idarucizumab was clearly more efficacious than strategies with non-specific procoagulant concentrates and devoid of the excessive procoagulant tendency observed with the latter.</AbstractText>

BACKGROUND

ST-elevated myocardial infarction (STEMI) is most frequently caused by coronary occlusion due to formation of an intracoronary thrombus in reaction to rupture of atherosclerotic plaques. Little is known about kinetics of coagulation markers after STEMI in patients treated according to current guidelines. We aimed to investigate kinetics of important coagulation markers in percutaneous coronary intervention (PCI)-treated STEMI patients.

METHODS

60 consecutive PCI-treated STEMI patients were prospectively included. Blood samples were collected immediately after as well as <em>1</em>, 4 and 7 days following PCI. Samples collected 90 days after PCI served as baseline values. ADAMTS<em>1</em>3 activity, VWF (von Willebrand factor) activity, VWF antigen, VWF propeptide, fibrinogen antigen, D-dimer, alpha<em>2</em>-antiplasmin (α<em>2</em>AP), plasmin-alpha<em>2</em>-antiplasmin complex (PAP), <em>prothrombin</em> <em>fragment</em> F<em>1</em>+<em>2</em> (F<em>1</em>+<em>2</em>), <em>prothrombin</em> time (PT), activated partial thromboplastin time (aPTT), and anti-factor Xa (anti-Xa) were measured. Cardiac magnetic resonance (CMR) was performed at 4-6 and 90 days after PCI in 49 patients and left ventricular ejection fraction (LVEF), infarct size and microvascular injury (MVI) were determined.

RESULTS

Immediately after PCI, ADAMTS<em>1</em>3 activity, fibrinogen antigen and α<em>2</em>AP levels were significantly decreased and VWF activity, VWF antigen and VWF propeptide levels were significantly elevated, compared to baseline. Individual coagulation markers and different combinations thereof were not related to LVEF or infarct size at 90 days, or the occurrence of MVI at 4-6 days after PCI.

CONCLUSIONS

Coagulation parameters show a very dynamic profile in the early days after STEMI. However, individual coagulation parameters or combinations thereof do not predict CMR-defined LVEF, infarct size or MVI.

Obesity and exercise constitute important factors for cardiovascular disease risk, but the long-term effects of different exercise modalities on haemostatic biomarkers are not well elucidated. We investigated the effects of 6 months of active commuting or leisure-time exercise on measures of fibrin turnover in individuals who are overweight and obese. Ninety younger (<em>2</em>0-40 years), sedentary, healthy women and men who are overweight and obese (BMI: <em>2</em>5-35 kg/m<em>2</em>) were randomised to 6 months of habitual lifestyle (CON, n=<em>1</em>6), active commuting (BIKE, n=<em>1</em>9), or leisure-time exercise of moderate (MOD, ∼50% VO<em>2</em>peak reserve, n=3<em>1</em>) or vigorous intensity (VIG, ∼70% VO<em>2</em>peak reserve, n=<em>2</em>4). Fasting blood samples (baseline and 3 and 6 months) were analysed for cholesterols and triglycerides, thrombin generation, <em>prothrombin</em> <em>fragment</em> <em>1</em> + <em>2</em>, D-dimer, fibrin clot properties, and fibrinolytic activity. We observed no differences between CON, BIKE, MOD, and VIG during the intervention and no time effects for any of the variables measured despite increased VO<em>2</em>peak in all exercise groups. We found no difference between CON and all exercise groups combined and no gender-specific effects of exercise. Our findings suggest that thrombin generation capacity, coagulation activation, fibrin clot structure, and lysability are unaffected by long-term active commuting and leisure-time exercise in women and men who are overweight and obese.

OBJECTIVE

Coagulation changes in pediatric trauma patients are not well defined. To fill this gap, we tested the hypothesis that trauma evokes a hypercoagulable response.

METHODS

A prospective observational study was conducted in hospitalized patients (age 8months to <em>1</em>4years) admitted for trauma or elective surgery. Informed consent was obtained from the parents and informed assent was obtained in patients 7years of age or older. Coagulation changes were evaluated on fresh whole blood using thromboelastography (TEG) and on stored plasma using assays for special clotting factors.

RESULTS

Forty three patients (<em>2</em><em>2</em> trauma, median injury severity score =9; and <em>2</em><em>1</em> uninjured controls) were evaluated. For trauma vs control, <em>prothrombin</em> time (PT) was higher by about <em>1</em>0% (p<0.00<em>1</em>), but activated partial thromboplastin time was not altered. TEG clotting time (R;p=0.005) and fibrin cross-linking were markedly accelerated (K time, alpha angle; p<0.00<em>1</em>) relative to the control patients. d-Dimer, <em>Prothrombin</em> <em>Fragment</em> <em>1</em>+<em>2</em>, and Plasminogen Activator Inhibitor-<em>1</em> were all elevated, whereas Protein S activity was reduced (all p<0.0<em>1</em>). Importantly, a large fraction of TEG values and clotting factor assays in the pediatric control group were outside the published reference ranges for adults.

CONCLUSIONS

A hypercoagulable state is associated with minor trauma in children. More work is needed to determine the functional significance of these changes and to establish normal pediatric reference ranges.

This study was to investigate the relationship between circadian blood pressure (BP) variation and circadian variation of neurohumoral factors during the acute phase of stroke. We studied <em>1</em>7 patients with cerebral infarction in <em>1</em>6 and cerebral hemorrhage in one. We performed <em>2</em>4-hour ambulatory BP monitoring and examined plasma renin activity (PRA), catecholamine, atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), endothelin <em>1</em> (ET<em>1</em>) and <em>prothrombin</em> <em>fragment</em> <em>1</em>+<em>2</em> (PT F<em>1</em>+<em>2</em>) and urinary catecholamine. Our result showed that the circadian variation of BP, neurohumoral and coagulation factors were diminished. There were significant relationships between BP levels and plasma BNP levels, nocturnal urinary adrenalines and ET<em>1</em>s. There were also significant relationships between night/day ratio of BP and plasma ET<em>1</em> level. In conclusion the abnormal patterns of circadian BP rhythm were frequently observed during the acute phase of stroke. The cause of this abnormality may result from the diminished circadian rhythms of neurohumoral factors.

Many circulating haemostatic markers have been investigated in relation to the abdominal aortic aneurysm (AAA) size, growth as well as intraluminal thrombus (ILT) size. However, the results of these studies seem to be uncertain and inconsistent. The first aim of the present study was to compare the haemostatic parameters of fibrinolysis and some of thrombotic markers in patients with AAA and controls. We also examined the relationship between those parameters and both maximum aneurysm diameter and intraluminal thrombus thickness. Tissue plasminogen activator (t-PA), plasminogen activator inhibitor (PAI-<em>1</em>), fibrinogen (Fb), D-dimer, <em>prothrombin</em> <em>fragments</em> <em>1</em> and <em>2</em> (F<em>1</em>+<em>2</em>), thromboxane B<em>2</em> (TXB<em>2</em>) and lipids profile were measured in 36 patients with AAA and 30 controls. The mean maximum aortic diameter in patients with the AAA was 59±<em>1</em><em>2</em> mm (range 4<em>2</em>-<em>1</em>00). The mean ILT thickness was 3<em>2</em>±<em>1</em>0 mm (range 8-56). Among haemostatic factors, t-PA and D-dimer levels, but not PAI-<em>1</em>, were significantly higher in subjects with the AAA. There was a strong positive correlation between thickness of intraluminal thrombus and maximum aneurysm size (r=0.69, p<0.000<em>1</em>), and the negative relationship between t-PA and ILT thickness (r= -0.53, p=0.00<em>1</em>) as well as aneurysm diameter (r= -0.38, p=0.0<em>2</em>3). Higher plasma concentrations of t-PA and D-dimer support the hypothesis that the secondary fibrinolysis plays an important role in the pathogenesis of the aortic abdominal aneurysm formation. In addition, the negative correlation between t-PA plasma level and ILT thickness suggests that thrombotic/fibrinolysis imbalance may favour accelerated formation of intraluminal thrombus and possibly aneurysm progression.

This study assessed hemostatic effects of an HMC-CoA reductase inhibitor, atorvastatin, on different parameters in 3<em>2</em> hypercholesterolemic patients of both sexes. In the patients and in <em>2</em>5 control subjects, plasma levels of tissue-type plasminogen activator, plasminogen activator inhibitor (PAI-<em>1</em>), D-dimer, <em>prothrombin</em> <em>fragment</em> <em>1</em> + <em>2</em> (F<em>1</em> + <em>2</em>), total cholesterol, triglycerides and fibrinogen had been measured. All these parameters were evaluated in patients after 6 and <em>1</em><em>2</em> months of treatment with atorvastatin at a dosage of <em>2</em>0 mg/day. This treatment significantly lowered the total cholesterol level in all patients. Moreover, after 6 months of atorvastatin treatment, PAI-<em>1</em> and F<em>1</em> + <em>2</em>, which were both increased at baseline, were significantly reduced. This reduction continued after <em>1</em><em>2</em> months. The present results show that a reduction of hemostatic abnormalities, which exist in hypercholesterolemia, may be another important effect of the atorvastatin therapy.

BACKGROUND

Even though thrombocytopenia following transcatheter aortic valve implantation (TAVI) has been described, further investigation of this phenomenon is needed.

OBJECTIVE

To determine which factors may explain the fall in platelet count that occurs after implantation of a TAVI device, including markers of platelet and blood coagulation activation.

METHODS

3<em>2</em> patients without previous indications for dual antiplatelet therapy (mean age 78.5±7.9 years, 6<em>2</em>% females) with severe aortic valve stenosis (mean gradient 54.6±<em>1</em>6.9mmHg) who qualified for TAVI procedure (Edwards Sapien XT) were prospectively analyzed. Platelet counts were analyzed before the surgery, on the day of the procedure and for the three following postoperative days (POD <em>1</em> to 3). To assess platelet activation P-selectin (PS, serum) and platelet factor 4 (PF-4, CTAD plasma) were measured, whereas for the evaluation of coagulation activation <em>prothrombin</em> <em>fragments</em> <em>1</em>+<em>2</em> (F<em>1</em>+<em>2</em>, plasma) were assessed before the procedure, on POD-<em>1</em> and POD-3 (ELISA).

RESULTS

During the postoperative period a significant platelet count drop, the most evident on POD-<em>2</em>, was observed followed by a platelet count raise. The platelet count drop correlated directly with the amount of iodinated contrast agent (r=0.4<em>2</em>, p=0.0<em>1</em>6) and inversely with baseline mean platelet volume (r=-0.37, p=0.046). Neither clinical nor perioperative parameters, except contrast medium, influenced platelet count decrease. No significant differences regarding the concentration of the evaluated markers in patients with and without thrombocytopenia were found. PF-4 and F<em>1</em>+<em>2</em> significantly changed during the study (p<0.05). Greater acute PF-4 decrease correlated with greater acute platelet count drop (r=0.48, p=0.043), and during the study slower PF-4 increase correlated with higher platelet count increase on POD-3 (r=-0.505, p=0.03<em>2</em>). Lower baseline PS correlated with lower baseline platelet count and higher platelet count increase on POD-3 (r=0.45, p=0.04 and =-0.55, p=0.0<em>2</em>, respectively). No significant correlations between F<em>1</em>+<em>2</em> concentrations and platelet count changes have been found.

CONCLUSIONS

Platelet reduction shortly after TAVI procedure is related to the amount of contrast agent applied during the procedure. Platelet activation and blood coagulation along with impaired baseline platelet renewal might be the mechanisms of thrombocytopenia following TAVI procedure.

BACKGROUND

Hypercoagulation was suggested to be involved in preterm birth etiology; however, the coagulation state of preterm parturients remains unelucidated. The study aim was to evaluate the haemostatic system of pregnant women with premature uterine contractions (PUC).

METHODS

The cohort study population consisted of 76 healthy pregnant women admitted with regular PUC. The study group included 38 women who experienced preterm birth; <em>1</em>4 of them had preterm premature rupture of membranes (PPROM). The control group included 38 women who eventually had term delivery. Groups were matched for maternal age, number of births and gestational age at admission. Blood samples were tested for haemostatic parameters and coagulation activation markers.

RESULTS

Significantly shorter PT and aPTT were documented in the study compared to control group (<em>2</em>5.7±<em>2</em> vs. <em>2</em>7.4±<em>2</em>.7seconds, P=0.003, and 9.96±0.5 vs. <em>1</em>0.<em>1</em>±0.4seconds, P=0.05, respectively), although differences in absolute values were small. There was no significant difference between the two groups in levels of: fibrinogen, D-dimer, protein C-global, free protein S antigen, factor VIII activity, Von Willebrand factor, plasminogen activator inhibitor-<em>1</em>, <em>prothrombin</em> <em>fragments</em> F<em>1</em>+<em>2</em> (PT F<em>1</em>+<em>2</em>), tissue factor and tissue factor pathway inhibitor. Women with PPROM had significantly lower PT F<em>1</em>+<em>2</em> levels compared to those who had preterm delivery with intact membranes (35<em>1</em>±99 vs. 56<em>1</em>±<em>2</em>4<em>2</em>pmol/L, P=0.003).

CONCLUSIONS

Shortened PT and aPTT, reflecting increased thrombotic activity in maternal plasma, could serve as a marker of real preterm labor in women with premature uterine contractions. Further prospective studies in a larger cohort are warranted to validate these findings.

Our previous studies have shown that platelets play a crucial role in the development of endometriosis, and women with endometriosis appear to be in a state of hypercoagulability. However, a recent study could only replicate part of our previous finding, casting doubts on this notion. We further investigated this question through a cross-sectional study by measuring additional coagulation factors in women with and without endometriosis. To this end, we conducted a cross-sectional study of <em>1</em>00 women with laparoscopically and pathologically diagnosed ovarian endometriomas (OMA) and another <em>1</em>00 women without endometriosis. The platelet count; platelet activation rate; maximum platelet aggregation rate; plasma levels of D-dimer, fibrinogen, fibrin degradation products (FDPs), plasma soluble P-selectin (sP-sel), and <em>prothrombin</em> <em>fragment</em> <em>1</em>+<em>2</em> (F<em>1</em>+<em>2</em>); <em>prothrombin</em> time; thrombin time (TT); and activated partial thromboplastin time were measured before surgery and 3 months after surgery, and their clinical data were recorded. These measurements were also performed in control patients. We found that, compared with controls, women with OMA had a significantly higher platelet activation rate and platelet aggregation rate, elevated plasma D-dimer, fibrinogen, FDPs, sP-sel, and F<em>1</em>+<em>2</em> levels as well as shortened TT. Remarkably, TT was prolonged, and all the other coagulation measurements, except plasma fibrinogen level, were significantly reduced 3 months after surgical removal of endometriotic lesions. Thus, our study provides another piece of evidence that endometriosis is a hypercoagulable disease, and anticoagulation therapy may hold promises in treating endometriosis.

BACKGROUND

Patients undergoing cardiac surgery requiring cardiopulmonary bypass develop a systemic inflammatory reaction. Antithrombin III (AT) has anticoagulant effects but also shows evidence of anti-inflammatory activity. The aim of this study was to examine whether exogenous AT could reduce white blood cell activation (CD<em>1</em><em>1</em>b up regulation or elastase release), in addition to inhibiting platelet (PLT) activation and fibrin generation, during simulated cardiopulmonary bypass (sCPB), undertaken in the absence of endothelium.

METHODS

sCPB was carried out with minimally heparinized (<em>2</em> U/mL) human blood for 90 minutes in controls and with supplementation by low-dose (<em>1</em> U/mL) and high-dose (5 U/mL) AT.

RESULTS

High-dose AT blunted thrombin generation during sCPB (prothrombin fragment <em>1</em>.<em>2</em>); both doses significantly inhibited thrombin activity (fibrinopeptide A). Complement activation (C3a and C5b-9) was unaffected by AT. High-dose AT inhibited PLT activation (P-selectin expression and P-selectin-dependent monocyte-PLT conjugate formation). AT supplementation at the higher dose significantly abrogated monocyte and neutrophil CD<em>1</em><em>1</em>b up regulation and neutrophil elastase release.

CONCLUSIONS

In addition to anticoagulant and anti-PLT effects, pharmacologic AT doses significantly blunted monocyte and neutrophil CD<em>1</em><em>1</em>b up regulation and neutrophil elastase release during sCPB, independent of endothelial effects. These data provide evidence for the direct anti-inflammatory activity of AT that has clinical relevance for CPB complications.

OBJECTIVE

The pro-inflammatory cytokine interleukin (IL)-6 is involved in the pathogenesis of both rheumatoid arthritis (RA) and cardiovascular events. We evaluated the correlation of prothrombotic biomarkers, in particular those of thrombin generation, with inflammatory and clinical parameters in RA patients treated with tocilizumab, an IL-6 receptor (IL-6R) inhibitor. Naïve and maintenance patients were compared.

METHODS

We studied 15 RA patients undergoing tocilizumab infusions at a University Outpatient Clinic. Eight received tocilizumab for the first time and were evaluated at baseline. Seven were in maintenance therapy (9 to 77 months). All 15 patients were evaluated four weeks after the last administration of tocilizumab. At each time, we assessed disease activity score <em>2</em>8 (DAS<em>2</em>8), erythrocyte sedimentation rate (ESR) and plasma levels of C-reactive protein (CRP), IL-6, soluble (s)IL-6R, tumour necrosis factor-alpha (TNF-alpha), <em>prothrombin</em> <em>fragment</em> F1+<em>2</em> and fibrin <em>fragment</em> D-dimer. Forty healthy subjects served as basal controls.

RESULTS

At baseline, RA patients showed a moderate-to-high disease activity and median ESR of 51 mm/1(st) hour (interquartile range <em>2</em>5-63). Plasma levels of CRP (p=0.0001), IL-6 (p=0.043), sIL-6R (p=0.003), TNF-alpha (p=0.0001), F1+<em>2</em> (p=0.0001) and D-dimer (p=0.00<em>2</em>) were higher than those of healthy controls. After four weeks we observed reduction of DAS<em>2</em>8 (p=0.0001), ESR (p=0.0001), CRP (p=0.014), TNF-alpha (p=0.006), F1+<em>2</em> (p=0.009) and D-dimer (p=0.04). No differences were observed between naïve and maintenance patients.

CONCLUSIONS

The reduction of prothrombotic biomarkers parallels the reduction of inflammatory parameters and clinical symptoms in RA patients treated with tocilizumab, both four weeks after the first administration and during maintenance therapy.