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How to prevent recurrences after a first venous thromboembolic (VTE) event in elderly patients is still an open issue, especially because of the high bleeding risk of anticoagulation in these patients. The placebo-controlled “Jason” study aims at assessing the efficacy and safety for secondary VTE prevention in elderly patients of oral Sulodexide (Vessel®) administration, a mixture of glycosaminoglycans (Alfasigma, Bologna, Italy) which proved effective against recurrences in a general population (SURVET study) without major bleeding (MB) complications. 1450 patients, aged ≥ 75 years, after at least 3 months of anticoagulation treatment for a first VTE episode, are double-blind randomized to receive for 12 months either sulodexide 500 lipasemic units (LSUs) twice daily, or sulodexide 250 LSU twice daily + indistinguishable placebo, or indistinguishable placebo. Primary outcomes for efficacy are the composite of death for VTE and recurrent VTE, and occurrence of MB for safety. Secondary outcomes include stroke, cardiovascular death and other thromboembolic events, and MB + clinically relevant non-MB. The first patient is scheduled to be randomized in May 2020. The study protocol has been approved by AIFA (Agenzia Italiana del Farmaco) and the Ethics Committee of the coordinating center. Written informed consent will be obtained from all patients prior to study participation. Jason study is an investigator-initiated trial, promoted by “Arianna Anticoagulazione” Foundation, Bologna, Italy, and supported by Alfasigma, Bologna, Italy. Study findings will be disseminated to participant centers, at research conferences and in peer-reviewed journals. Trial registration numbers NCT 04257487; EudraCT (2019–000570-33). Electronic supplementary material The online version of this article (10.1007/s11739-020-02381-5) contains supplementary material, which is available to authorized users.

Essentials Between-lab variations of cut-off values in lupus anticoagulant detection are unknown. Cut-off values were calculated in 11 labs each testing plasma from 120 donors with 3 platforms. Major variation was observed even within the same platform. Cut-off values determined in different labs are not interchangeable. SummaryBackground Cut-off values for interpretation of lupus anticoagulant (LA) detection are poorly investigated. Aims (i) To assess whether results from healthy donors were normally distributed and (ii) the between-laboratories differences in cut-off values for screening, mixing and LA confirmation when calculated as 99th or 95th centiles, and (iii) to assess their impact on the detection rate for LA. Methods Each of 11 laboratories using one of the three widely used commercial platforms for LA detection was asked to collect plasmas from 120 healthy donors and to perform screening, mixing and LA confirmation with two methods (activated partial thromboplastin time [APTT] and dilute Russell viper venom [dRVV]). A common set of LA-positive or LA-negative freeze-dried plasmas was used to assess the LA detection rate. Results were centralized (Milano) for statistical analysis. Results and conclusions (i) Clotting times or ratios for healthy subjects were not normally distributed in the majority of cases. The take-home message is that cut-off values should be determined preferably by the non-parametric method based on centiles. (ii) There were relatively large inter-laboratory cut-off variations even within the same platform and the variability was marginally attenuated when results were expressed as ratios (test-to-normal pooled plasma). The take-home message is that cut-off values should be determined locally. (iii) There were differences between cut-off values calculated as 99th or 95th centiles that translate into a different LA detection rate (the lower the centile the greater the detection rate). The take-home message is that cut-off values determined as the 95th centile allow a better LA detection rate.

Introduction: To measure direct factor Xa inhibitor (apixaban, edoxaban, rivaroxaban) concentrations, dedicated chromogenic anti-Xa assays are recommended as suitable methods to provide rapid drug quantification. Moreover, the high-performance liquid chromatography with ultraviolet detection (HPLC-UV) is reported as a reliable quantitative technique. We investigated seven anti-Xa assays and an HPLC-UV method for measurement of apixaban and rivaroxaban levels in patients enrolled in the START-Register. Methods: A total of 127 apixaban and 124 rivaroxaban samples were tested by HPLC-UV and the following anti-Xa assays: Biophen DiXaI and Heparin LRT (Hyphen BioMed), Berichrom and Innovance Heparin (Siemens), STA-Liquid Anti-Xa (Stago Diagnostics), Technochrom anti-Xa (Technoclone), and HemosIL Liquid Anti-Xa (Werfen). Each method was performed in one of the participating laboratories: Bologna, Cremona, Florence, and Padua. Results: Our data confirmed the overestimation of apixaban and rivaroxaban levels by the antithrombin-supplemented anti-Xa method (Berichrom). Performances and reproducibility of the six anti-Xa assays not supplemented with antithrombin and the HPLC-UV method were good, with limits of quantification from 8-39ng/mL (apixaban) and 15-33ng/mL (rivaroxaban). The six chromogenic methods showed good concordances with the quantitative HPLC-UV [bias: −26.9-22.3ng/mL (apixaban), −11.3-18.7ng/mL (rivaroxaban)]. Higher bias and wider range between limits of agreement were observed at higher concentrations [200ng/mL: bias −42.2-36.8ng/mL (apixaban) and −20.1-68.9ng/mL (rivaroxaban)]. Conclusion: Overall, the anti-Xa assays not supplemented with antithrombin and the HPLC-UV method proved to be suitable for apixaban and rivaroxaban quantification.

BackgroundD-dimer levels measured during and after vitamin K antagonist withdrawal may be used in clinical practice to assess the individual risk of recurrent venous thromboembolism. Currently, direct oral anticoagulants (DOACs) are frequently used in venous thromboembolism treatment; however, their pharmacokinetics and pharmacodynamics characteristics are completely different than vitamin K antagonists. The present study aimed at comparing the results of D-dimer levels during and after anticoagulation withdrawal in patients with venous thromboembolism treated with DOACs or warfarin.Material and methodsD-dimer levels were measured in 527 patients ("cases") during DOACs treatment (T0) and after 15 (T15), 30 (T30), 60 (T60) and 90 (T90) days after their discontinuation and in 527 patients ("controls") enrolled in the DULCIS study (all treated with warfarin), matched for sex, age (+/-3 y), type of D-dimer assay and site of venous thromboembolism. Both cases and controls received anticoagulant treatment after a first venous thromboembolism event that was unprovoked or associated with weak risk factors.ResultsThe rate of positive D-dimer results was significantly higher in cases than in controls at T0 (10.8% vs 5.1%, p = 0.002) and at T30 (18.8% vs 11.8%, p = 0.019), as well as at the other time-points, though not statistically significant.ConclusionD-dimer levels during and after stopping an anticoagulant treatment for a venous thromboembolism episode differ between patients treated with a DOAC than in those treated with warfarin. Specifically designed prospective studies are warranted to reassess the use of D-dimer as predictor of the risk of recurrent venous thromboembolism in patients treated with DOACs.

In the D-dimer and ULtrasonography in Combination Italian Study (DULCIS), serial D-dimer measurement in combination with assessment of residual thrombosis (in patients with deep vein thrombosis (DVT)) identified patients who safely discontinued anticoagulation after an unprovoked venous thromboembolism (VTE).In this subgroup analysis, the value of D-dimer tests was assessed in patients with isolated pulmonary embolism (PE) compared with those with DVT, with or without PE (DVT/PE). The DULCIS database was reanalysed in relation to this target.26.8% of the DULCIS patients had isolated PE as the index event; this was more prevalent in females (34.1%) than in males (21.1%; pversus 3.8% ppy; nonsignificant) who stopped anticoagulation for negative D-dimer, but it was markedly high (11.2% ppy; pversus two (7.4%) out of 27; p=0.0085).Serial D-dimer assessment can inform on the risk of recurrent VTE and help determine the duration of anticoagulation in patients with isolated PE.

Essentials Currently, DOACs are given at fixed doses and do not require laboratory monitoring. Direct oral anticoagulant-specific measurements were performed at trough and peak. Patients who developed bleeding events showed higher DOAC plasma levels at peak. This study suggests the need of a more accurate DOAC dose assessment. Background: Direct oral anticoagulants (DOACs) are administered at fixed dose. The aim of the study was to evaluate the relationship between DOAC C-trough or C-peak plasma levels and bleeding complications in patients with non-valvular atrial fibrillation (NVAF). Methods: Five hundred sixty five consecutive naive NVAF patients were enrolled. The DOAC measurements at C-trough and at C-peak (available in 411 patients) were performed at steady state, within the first month of treatment. Major bleeding (MB), clinically relevant non-major bleeding (CRNMB), and minor bleeding (MinB), occurring during 1year of follow-up after blood sampling, were recorded. For each DOAC, interval of C-trough and C-peak levels was subdivided into four equal classes and results were attributed to these classes; the median values of results were also calculated. Results: Two hundred eight patients were on apixaban, 185 on dabigatran, and 172 on rivaroxaban. For 1-[qqqdeletezzz]year follow up for all patients, we observed: 19 MB (3.36%), 6 CRNMB (1.06%), and 47 MinB (8.31%). The prevalence of bleeding patients with anticoagulant levels in the upper classes of C-peak activity (II + III + IV) was higher than that in the lowest class. Normalized results of C-peak levels were higher in patients with bleeding than in those without bleeding. Conclusions: Bleeding complications during DOAC treatment were more frequent among atrial fibrillation (AF) patients with higher C-peak anticoagulant levels. In addition to a previous study that showed an increased risk of thrombotic complications in the patients with low C-trough levels, this study seems to indicate that patients with NVAF on DOACs would need a more accurate definition of their optimal therapeutic window.

Background High inter-individual variability of the anticoagulant plasma levels of the first three direct oral anticoagulants was previously reported. Aims of the present study were to evaluate edoxaban inter and intra-individual variability in patients with non valvular atrial fibrillation and to assess correlation between edoxaban plasma levels and coagulation screening test and renal function. Methods From January 31st 2017 to June 30th 2018, a total of 101 NVAF patients were enrolled: 48 patients were on edoxaban 60 mg and 53 on edoxaban 30 mg, once daily. Blood samples were taken at C-trough and at C-peak within the first month (15–25 days) of treatment and then at C-trough each three months. Prothrombin time (PT), activated partial thromboplastin time (aPTT), specific anti-FXa chromogenic test were performed. Creatinine clearance (CrCl) was calculated using the Cockcroft-Gault formula. Results Mean inter-individual variability expressed as overall coefficient of variation (CV%) values was lower at C-peak (CV% = 49) than at C-trough (CV% = 68). Mean CV% intra-individual variability was 26.5. No significant correlation was found between edoxaban plasma levels and CrCl (C-trough r/r2 = −0.007/0.000; C-peak r/r2 = 0.129/0.017). Correlations (r/r2), at C-trough and C-peak, between edoxaban levels and PT and aPTT, were 0.386/0.149–0.922/0.851 and 0.283/0.080–0.567/0.321, respectively. Significant discrepancies between PT or aPTT and edoxaban levels were found. Conclusions This study confirms also for edoxaban a high inter-individual variability in NVAF patients. PT and aPTT are not useful to measure this drug. As for the other two anti-FXa drugs, the absence of a significant correlation between CrCl and edoxaban plasma levels was observed.

Introduction Direct oral anticoagulant (DOAC) intra- and inter-individual variability was previously reported, but its magnitude is still considered negligible for patient management. Objective To evaluate inter- and intra-individual variability in real-world atrial fibrillation patients on dabigatran, rivaroxaban or apixaban in four Italian anticoagulation clinics and to assess the correlation between DOAC plasma concentration and creatinine-clearance (CrCl). Materials and Methods A total of 330 consecutive patients were enrolled, of which 160 were on dabigatran (70 and 90 taking 150 mg or 110 mg twice-daily, respectively), 71 on rivaroxaban (37 and 34 taking 20 mg or 15 mg once-daily) and 99 on apixaban (73 and 26 taking 5 mg or 2.5 mg twice-daily). Blood was taken at trough and peak within the first month (15–25 days) of treatment. Diluted-thrombin-time (dTT) calibrated for dabigatran and anti-FXa calibrated for rivaroxaban or apixaban was performed. Results Mean inter-individual variability expressed as overall CV values for all drugs was lower at peak (CV = 46%) than at trough (CV = 63%). Mean CV% intra-individual variability was 36.6% at trough and 34.0% at peak. Correlation with CrCl was poor for all drugs and only dabigatran at trough showed a significant correlation. Conclusion This multicenter study confirms high DOAC inter-individual variability that cannot be explained by the rate of renal clearance to which the three DOAC were subjected since the correlation with CrCl was relatively poor. This poor correlation suggests caution in using CrCl as the sole laboratory parameter to indirectly evaluate residual circulating DOAC.

Essentials Direct oral anticoagulants (DOACs) do not require laboratory monitoring currently. DOAC specific measurements were performed at trough in patients with atrial fibrillation. Patients who developed thromboembolic events showed lower DOAC plasma levels. This study supports the concept of measuring DOAC levels at steady state. Summary: Background Direct oral anticoagulants (DOACs) are administered at fixed doses without the need for dose adjustment according to laboratory testing. High interindividual variability in drug blood levels has been shown with all DOACs. To evaluate a possible relationship between DOAC C-trough anticoagulant levels and thromboembolic events, 565 consecutive naive patients with atrial fibrillation (AF) were enrolled in this study performed within the START Laboratory Registry. Methods DOAC-specific measurements (diluted thrombin time or anti-activated factor II calibrated for dabigatran; anti-activated FX calibrated for rivaroxaban or apixaban) at C-trough were performed locally at steady state within 15–25 days after the start of treatment. For each DOAC, the interval of C-trough levels, from the limit of quantification to the highest value, was subdivided into four equal classes, and results were attributed to these classes; the median values of results were also calculated. Thromboembolic complications occurring during 1 year of follow-up were recorded. Results Thromboembolic events (1.8%) occurred in 10 patients who had baseline C-trough levels in the lowest class of drug levels. The incidence of thromboembolic events among patients with DOAC C-trough levels in the lowest level class was 2.4%, and that in the remaining groups was 0%. The patients with thrombotic complications also had a higher mean CHA2DS2-VASc score than that of the total patient population: 5.3 (95% confidence interval [CI] 4.3–6.3 versus 3.0 (95% CI 2.9–3.1). Conclusion In this study cohort, thrombotic complications occurred only in DOAC-treated AF patients who had very low C-trough levels, with a relatively high CHA2DS2-VASc score. Larger studies are warranted to confirm these preliminary observations. © 2018 The Authors. Journal of Thrombosis and Haemostasis published by Wiley Periodicals, Inc. on behalf of International Society on Thrombosis and Haemostasis.

Background:Laboratory investigation with specific factor XIII (FXIII) assays plays a crucial role in diagnosis of FXIII deficiency. According to the International Society on Thrombosis and Hemostasis (ISTH), it is necessary a blank sample with iodoacetamide, provided by the kit or locally prepared, when the ammonia release assays are used, to avoid FXIII activity overestimation.Methods:In this study we set up a modification of the Berichrom FXIII chromogenic assay, in which iodoacetamide was added by the BCS analyzer in the reaction mixture of the blank sample, without modifications of the original reagents. We analyzed 100 plasma samples of outpatients with clinical symptoms suggestive of a bleeding diathesis (20 samples had FXIII activity Results:In all samples blank subtraction significantly reduced FXIII activity, mostly in the low activity range group (from 10.1% to 2.4%, pConclusions:Despite the low number of samples included in the study, the described automatic procedure seemed to decrease FXIII activity overestimation and, especially for low activity range samples (

Intracranial atherosclerotic disease (ICAD) is responsible for at least 10% of transient ischaemic attacks (TIA). Thrombin generation has been shown to be associated with several atherosclerotic conditions and may be relevant in the pathogenesis of TIA from ICAD. BACKGROUND: Intracranial atherosclerotic disease (ICAD) is responsible for at least 10% of transient ischaemic attacks (TIA). Thrombin generation has been shown to be associated with several atherosclerotic conditions and may be relevant in the pathogenesis of TIA from ICAD. OBJECTIVE: To evaluate the association between thrombin generation and ICAD in patients with TIA. MATERIALS AND METHODS: Consecutive patients with confirmed diagnosis of TIA by vascular neurologist were enrolled. Within 24h from diagnosis, all the patients underwent: blood samples including thrombin generation search, electrocardiography, brain CT scan, blood pressure (BP) measurement, supra-aortic echo-Doppler, transcranial Doppler (TCD) and standard echocardiogram. Thrombin generation was measured as endogenous thrombin potential (ETP) in platelet-rich plasma (PRP) and in platelet-poor plasma (PPP), in the presence and in the absence of thrombomodulin (TM). RESULTS: 120 patients (male 52.5%), aged 69±16years were enrolled. Ten patients on warfarin treatment had significantly lower ETP than the others. Among the remaining, ETP in the presence or absence of TM did not differ according to TOAST classification aetiology (large vessel vs. cardioembolic vs. lacunar vs. others). In PRP, ETP was similar in patients with ICAD and in those without (1748±160 vs. 1851±36nM·min, p=0.393), whereas, ETP measured in presence of thrombomodulin was higher in patients with than in those without ICAD (2045±99 vs. 1715±41nM·min, p=0.011). In PPP, ETP was similar in patients with ICAD and in those without, whereas thrombin peak was higher in patients with ICAD than in those without both in the presence (165±17 vs. 130±5nM, p=0.036) and in the absence of TM (178±19 vs. 142±5nM, p=0.037). CONCLUSION: ETP measured in presence of TM is enhanced in patients with ICAD, supporting that thrombomodulin-protein C pathways is relevant in TIA from ICAD. These hypothesis-generating data suggest that thrombin generation may be relevant in cerebral ischaemia from intracranial disease, and justify larger studies.

no abstract available

Essentials Predicting recurrences may guide therapy after unprovoked venous thromboembolism (VTE). We evaluated the DASH score in 827 patients with unprovoked VTE to verify prediction accuracy. A DASH score ≤ 1 had a cumulative recurrence risk at 1 year of 3.6%, as predicted by the model. The DASH score performed better in younger (< 65 years old) subjects. Summary: Background The DASH prediction model has been proposed as a guide to identify patients at low risk of recurrence of venous thromboembolism (VTE), but has never been validated in an independent cohort. Aims To validate the calibration and discrimination of the DASH prediction model, and to evaluate the DASH score in a predefined patient subgroup aged > 65 years. Methods Patients with a proximal unprovoked deep vein thrombosis (DVT) or pulmonary embolism (PE) who received a full course of vitamin K antagonist or direct oral anticoagulant (> 3 months) and had D-dimer measured after treatment withdrawal were eligible. The DASH score was computed on the basis of the D-dimer level after therapy withdrawal and personal characteristics at the time of the event. Recurrent VTE events were symptomatic proximal or distal DVT/PE, and were analyzed with a time-dependent analysis. Observed 12-month and 24-month recurrence rates were compared with recurrence rates predicted by the DASH model. Results We analyzed a total of 827 patients, of whom 100 (12.1%) had an objectively documented recurrence. As compared with the original DASH cohort, there was a greater proportion of subjects with a ‘low-risk’ (≤ 1) DASH score (66.3% versus 51.6%, P < 0.001). The slope of the observed versus expected cumulative incidence at 2 years was 0.71 (95% confidence interval 0.51–1.45). The c-statistic was lower for subjects aged > 65 years (0.54) than for younger subjects (0.72). Conclusions These results confirm the validity of DASH prediction model, particularly in young subjects. The recurrence risk in elderly patients (> 65 years) was, however, > 5% even in those with the lowest DASH scores.

Objectives Few data are available on thrombophilic risk factors and clinical outcome in patients undergoing percutaneous transluminal angioplasty (PTA) for peripheral arterial disease (PAD). We investigated the role of homocysteine, fibrinogen, Factor VIII (FVIII), lupus anticoagulant (LAC) and FII G20210A, and FV R506Q (FV Leiden) mutations as prognostic factors in 230 patients who underwent PTA for PAD (Fontaine’s stages: IIb through IV; aged 69 ± 1 years). Design and methods A prospective study. Major adverse cardiovascular events (MACE) were the composite ‘end’ point. Results During the follow-up (24.3 ± 1.5 months), 96 (41.7%) patients reached the ‘end’ point. According to Cox regression analysis, diabetes and critical limb ischaemia were predictors of MACE, whereas each single thrombophilic alteration was not. Thrombophilic alterations were more frequent in patients that reached the ‘end’ point, and the patients with two alterations (hazard ratio (HR) 2.55 confidence interval (CI): 1.20–5.46, p = 0.015) and those with three or more alterations (HR 2.91 CI: 1.31–6.45, p = 0.009) had an increased risk for MACE versus those without alterations. Thrombophilic alterations were not associated with limb loss during the follow-up. Conclusion The presence of multiple thrombophilic alterations in patients who underwent PTA for PAD is associated with increased risk of arterial thrombotic events.

SummaryD-dimer and residual venous obstruction (RVO) have been separately shown to be risk factors for recurrent venous thromboembolism (VTE) after a first episode of unprovoked proximal deep-vein thrombosis (DVT). It was the objective of this study to assess the predictive value of D-dimer and residual vein obstruction (RVO), alone and in combination, for recurrence after provoked DVT of the lower limbs. A total of 296 consecutive patients with a first episode of symptomatic provoked proximal DVT were evaluated at a university hospital in Bologna, Italy. On the day of anticoagulation withdrawal (T0), RVO was determined by compression ultrasonography. D-dimer levels (cut-off: 500 ng/ml) were measured at T0 and after 30 ±10 days (T1). The main outcome was recurrent VTE during a two-year follow-up. D-dimer was abnormal in 11.6% (32/276) and 31% (85/276) of subjects at T0 and at T1, respectively. RVO was present in 44.8% (132/294) of patients. Recurrence rate was 5.1% (15/296; 95% confidence interval [CI]: 3–8%; 3% patient-years; 95% CI: 2–5 %). An abnormal D-dimer either at T0 or at T1 was associated with an adjusted hazard ratio (HR) for recurrence of 4.2 (95% CI:1.2–14.2; p=0.02) and 3.8 (95%CI: 1.2–12.1; p=0.02), respectively, when compared with normal D-dimer. The HR for recurrence associated with RVO was not significant, and RVO did not increase the recurrence risk associated with an abnormal D-dimer either at T0 or T1. In conclusion, an abnormal D-dimer during vitamin K antagonist (VKA) treatment or at one month after VKA withdrawal is a risk factor for recurrence in patients with provoked DVT, while RVO at the time of anticoagulation withdrawal is not.

The PROLONG randomized trial showed that a normal D-dimer (D-d) 1 month after anticoagulation suspension for unprovoked venous thromboembolism (VTE) was associated with a low risk of late recurrences (4.4% patient years). However, it is unknown whether D-d changes subsequently. The aim of this prospective multicenter study was to assess D-d time course and its relation with late recurrences in patients with normal D-d 1 month after anticoagulation suspension for a first episode of unprovoked VTE. D-d was measured with a qualitative method (Clearview Simplify D-dimer; Inverness Medical Professional Diagnostics). Patients with a normal D-d 1 month after stopping anticoagulation repeated D-d testing every 2 months for 1 year. D-d was normal in 68% (243/355) of patients 1 month after anticoagulation suspension. Patients in whom D-d became abnormal at the third month and remained abnormal afterward had a higher risk of recurrence (7/31; 27% patient years; 95% confidence interval [CI]: 12-48) than patients in whom D-d remained normal at the third month and afterward (4/149; 2.9% patient years; 95% CI: 1-7; adjusted hazard ratio: 7.9; 95% CI: 2.1-30; P = .002). Repeated D-d testing after anticoagulation suspension for a first episode of unprovoked VTE could help tailor the duration of treatment. This trial is registered at http://clinicaltrials.gov as NCT00266045.

SummaryHeparin-induced thrombocytopenia (HIT) is a life-threatening complication of heparin treatment; the prognosis depends on early and accurate diagnosis, and prompt start of alternative anticoagulants. Because of high sensitivity, the commercially available immunologic assays are widely used, though not suited to be run on single samples and with a turnaround time of 2–3 hours. We evaluated two new, rapid, automated, semi-quantitative chemiluminescent immunoassays in HIT suspected patients: HemosIL® AcuStar HIT-IgG(PF4-H) (specific for IgG anti- PF4/heparin antibodies) and HemosIL® AcuStar HIT-Ab(PF4-H) (detecting IgG, IgM and IgA anti-PF4/heparin antibodies) (both from Instrumentation Laboratory). A total of 102 patients with suspected HIT were included; HIT was diagnosed in 17 (16.7%). No false negative cases were observed using either the HemosIL AcuStar HIT-IgG(PF4-H) or the HITAb(PF4-H) assay (sensitivity and negative predictive values = 100%; negative likelihood ratios

SummaryThe natural history of calf deep-vein thrombosis (DVT) is still uncertain and it is debated whether it warrants to be diagnosed and treated. We aimed to investigate the complication rate of untreated isolated calf DVT (ICDVT). Symptomatic outpatients were prospectively managed with serial compression ultrasonography (SCUS). Those without proximal DVT and with likely pre-test clinical probability (PCP) or altered D-dimer received immediate subsequent complete examination of calf deep veins (CCUS) by a different operator. The result of CCUS was kept blind both to the managing doctor and the patient and disclosed after three months. Primary outcome was the rate of venous thromboembolism at three months. We examined 431 subjects (196 males; median age 68.0 years) in whom five outcomes were recorded (1.2%; 95% confidence intervals [CI]: 0.4–2.7). If CCUS results had been available, outcomes would have been recorded in 3/424 patients (0.7%; 95% CI: 0.2–2.1) with two events in subjects negative at both serial and complete CUS. ICDVT was diagnosed in 65 subjects (15.3%; 95% CI: 12–19); of whom 59 remained uneventful (one was lost to follow-up). A significant higher rate of outcomes was recorded in subjects with than without ICDVT (5/64; 7.8%; 95% CI: 3–17 vs. 3/351; 0.8%; 95% CI: 0–2; p=0.003). However, after excluding two events picked at serial CUS in subjects with ICDVT, the difference became barely significant (3/64; 4.7%; 95% CI: 1–13; p=0.049). Thrombotic evolution of untreated ICDVT in high-risk subjects may be relevant. Larger studies are needed to address this issue.

SummaryFew data are available on thrombophilic risk factors and progression of atherosclerotic peripheral arterial disease (PAD). Thrombophilic alterations can be an aggravating factor when arterial stenoses are present. In a cross-sectional study, we evaluated the presence of the thrombophilic factors fibrinogen, homocysteine, factor (F)VIII, lupus anticoagulant (LAC), FII G20210A, and FV R506Q mutations in 181 patients with PAD at Fontaine’s stage II (claudication), in 110 patients with critical limb ischaemia (CLI), and in 210 controls. Fibrinogen was higher in patients with CLI vs. those with claudication and controls (427.9 ± 10.5 vs. 373.1 ± 5.2 vs. 348.9 ± 7.0 p=0.001, respectively). Homocysteine and FVIII were higher in patients with PAD than in controls, but were similar in patients with CLI and claudication. The prevalence of LAC increased in patients with CLI vs. those with claudication and controls (21.4% vs. 7.8% vs. 5.2% p

Background The optimal dose of low-molecular-weight-heparin (LMWH) to prevent venous thromboembolism (VTE) after bariatric surgery remains controversial. Aim The aim of this study was to evaluate the pharmacodynamic parameters of two doses of the LMWH parnaparin administered to patients undergoing bariatric surgery. Methods Patients were enrolled in a multicentre, open label, pilot study and were randomised to receive 4250 IU/day [n = 36; 30 females; median age: 38 years (23-56); median BMI: 46.7 Kg/m2 (36.5-58.8)] or 6400 IU/day [n = 30; 24 females; median age: 42 years (22-63); median BMI: 43.7 Kg/m2 (36.1-64.1)] of parnaparin s.c. for 7-11 days. The pharmacodynamic effects of parnaparin were analysed by measuring the anti Factor Xa activity on day 0 (12 hours after the first parnaparin injection), day 4 and day 6 after surgery (before and 4 hours after parnaparin administration). Results In 98.3% of patients receiving 4250 IU/day the peak anti-Xa levels were in the range of 0.1-0.4 IU/ml. Higher anti-Xa levels were observed in patients receiving 6400 IU/day: in 62.3% of these patients the peak anti-Xa levels were greater than 0.4 IU/ml. The anti-Xa levels measured 4 hours after injection on days 4 and 6 were not statistically correlated with BMI for either dose of-parnaparin (p = 0.077 and p = 0.401 for 4250 or 6400 IU/day, respectively). Conclusion The dose of 4250 IU/day seems adequate to achieve prophylactic anti-Xa levels in morbid obese patients undergoing bariatric surgery. Conversely, most of the patients receiving 6.400 IU/day show anti-Xa levels higher than the recommended prophylactic values.

Background and objectives: To assess the predictive value of D-dimer (D-d) and Factor VIII (FVIII) in combination for recurrent venous thromboembolism (VTE) after vitamin K antagonist (VKA) therapy suspension. Design and methods: Consecutive outpatients with a first episode of idiopathic proximal deep vein thrombosis of the lower limbs were enrolled on the day of VKA suspension. After 30+/−10 days, D-d (cut-off value: 500 ng/mL), chromogenic FVIII activity and inherited thrombophilia were determined. Follow-up was 2 years. Results: Overall recurrence rate was 16.4% (55/336; 95% CI:13–21%). The multivariate hazard ratio (HR) for recurrence was 2.45 (95% CI: 1.24–4.99) for abnormal D-d and 2.76 (95% CI:1.57–4.85) for FVIII above the 75th percentile (2.42 U/mL) after adjustment for age, sex, thrombophilia, VKA duration and residual venous obstruction. When compared with normal D-d and FVIII, the multivariate HR was 4.5 (95% CI: 1.7–12.2) for normal D-d with FVIII above 2.42 U/mL and 2.7 (95% CI: 1.2–6.6) and 7.1 (95% CI:2.8–17.6) for abnormal D-d with FVIII, respectively, below and above 2.42 U/mL. Interpretation and conclusions: D-d and FVIII at 30+/−10 days after VKA withdrawal are independent risk factors for recurrent VTE.

Essentials Prothrombin and partial thromboplastin time (PT/PTT) measure direct oral anticoagulants (DOACs). PT, PTT and specific tests for DOACs were performed on patients treated for atrial fibrillation. Normal PT/PTT don't exclude DOAC activity and their prolongation doesn't confirm DOAC action. The use of PT or PTT to evaluate DOAC activity could cause dangerous misinterpretations. Summary: Background Prothrombin time (PT) and activated partial thromboplastin time (APTT) have been proposed to measure the effect of oral anti-activated factor X (FXa) or anti-activated FII drugs, respectively. Aims To evaluate the relationships and responsiveness of PT and APTT versus direct oral anticoagulant (DOAC) concentrations measured with specific coagulation tests performed with different platforms in four Italian anticoagulation clinics. Methods Six hundred and thirty-five patients with atrial fibrillation participated in the study: 240 were receiving dabigatran, 264 were receiving rivaroxaban, and 131 were receiving apixaban. Blood was taken at trough and peak within the first month (15–25 days) of treatment. PT, APTT, diluted thrombin time (dTT) calibrated for dabigatran and anti-FXa calibrated for rivaroxaban or apixaban were determined. Results For dabigatran, the correlation between APTT and dTT ranged from r = 0.80 to r = 0.62. For rivaroxaban, the correlation between the anti-FXa assay and PT ranged from r = 0.91 to r = 0.73. For apixaban, the correlation between the anti-FXa assay and PT was lower than for the two other drugs (r = 0.81 to r = 0.54). Despite the above significant correlations, the responsiveness of PT or APTT was relatively poor. A discrepancy between global testing and DOAC plasma concentrations was shown in a considerable proportion of patients, depending on the platform and drug, with values ranging from 6% to 62%. Conclusions Overall, poor responsiveness of the screening tests to DOAC concentrations was observed. PT and APTT normal values cannot exclude DOAC anticoagulant activity, and PT or APTT prolongation is not always associated with DOAC anticoagulant effect as determined with specific tests.

Background The risk of heparin induced thrombocytopenia (HIT) associated with low molecular weight heparin (LMWH) for treatment of superficial vein thrombosis (SVT) is uncertain. As a result the necessity of platelet count monitoring is unclear in this setting. Aims To assess the risk of HIT in outpatients treated with LMWH for SVT. Methods In a prospective single centre study we included all symptomatic outpatients in whom a real-time B-mode and color Doppler ultrasonography examination revealed SVT without DVT. Patients treated with vitamin K antagonists or fondaparinux were excluded. Patients received full dose enoxaparin for 1 week followed by half therapeutic dose for 3 weeks or parnaparin 8500 UI aXa for 10 days followed by 6400 UI aXa once daily for 20 days. Platelet count was performed on the day of diagnosis (D0) and 7 (D7), and 14 (D14) days afterward. Primary outcomes were the rate of thromboembolic events and of HIT during a 3-month follow-up. Results 678 outpatients (age: 64.7 ± 16.2 years, male: 42.0%) were evaluated. During follow-up, 7 venous thrombo-embolic events were recorded (1.03% CI 95%: 0.50–2.11%), while no major bleeding was observed (0.0% CI 95%: 0.0–0.56%). Platelet count was 255 ± 93 × 109/L at D0, 245 ± 93 × 109/L at D7 (p = 0.204 vs. D0) and 261 ± 116 × 109/L at D14 (p = 0.405 vs. D0). No fall in platelet count > 50% and no case of HIT were recorded (HR 0.0% CI 95%: 0–0.56%). Conclusions A 4-week LMWH treatment for SVT is associated with an incidence of HIT lower than 0.6% and platelet count monitoring may be omitted in this setting.

Summary. Background: The A > G polymorphism at position 19911 of the prothrombin gene is associated with increased plasma prothrombin levels but its role as a risk factor for venous thromboembolism (VTE) is not established. Objective: To investigate the role of prothrombin 19911 A > G polymorphism in the risk of VTE in patients with heterozygous prothrombin 20210GA or factor (F) V Leiden and in those without thrombophilia. Patients and methods: Case–control study of 793 patients with prothrombin 20210 GA (n = 167) or FV Leiden (n = 198), and without thrombophilia (n = 428), and of 795 healthy individuals with the corresponding coagulation profile, investigated for the presence of prothrombin 19911 A > G. Plasma prothrombin levels were measured in 342 individuals. Results: Prothrombin 19911 A > G did not increase the risk of VTE in carriers of prothrombin 20210 GA [odds ratio (OR) 1.2, 95% CI (95% CI) 0.8–1.8] but significantly increased the risk in carriers of FV Leiden (OR 2.1, 95% CI 1.3–3.4) and in patients without thrombophilia (OR 1.5, 95% CI 1.0–2.2). Higher plasma prothrombin levels in carriers of prothrombin 19911 A > G polymorphism than in non-carriers were found among individuals without thrombophilia (P =0.05) and with FV Leiden (P = 0.07), but not in carriers of prothrombin 20210 GA (P = 0.2). Conclusions: Prothrombin 19911 A > G polymorphism was independently associated with a 1.5-fold increased risk of VTE and increased 2-fold the risk of VTE associated with FV Leiden, both increases statistically significant. No effect was observed in carriers of prothrombin 20210 GA, perhaps because this polymorphism has a stronger influence on plasma prothrombin levels than the prothrombin 19911 polymorphism.

SummaryWe assessed the predictive value of D-dimer levels in combination with residual venous obstruction (RVO) for recurrent venous thromboembolism (VTE) in a prospective cohort of outpatients after oral anticoagulant therapy (OAT) suspension for a first episode of idiopathic proximal deep vein thrombosis of the lower limbs during a 2-year follow-up. Patients (n=400) were enrolled on the day of OAT suspension when RVO was determined by compression ultrasonography (present in 48.6% of patients). D-dimer (cut-off value: 500 ng/mL) was measured 30±10 days afterwards (abnormal in 56.4% of patients). The overall recurrence rate was 16.7% (67/400; 95% confidence intervals - CI -:13–21%). The multivariate hazard ratio (HR) for recurrence was 3.32 (95% CI:1.78–6.75; p > 0.0001) for abnormal D-dimer compared to normal D-dimer and 1.2 (95% CI:0.72–2.07; p>0.05) for RVO compared to absent RVO. The recurrence rate was 5.7% (95% CI:2–13%) and 10.4% (95% CI:6–18%), respectively, for normal D-dimer either without or with RVO, 22.9% (95% CI:14–33%) and 25.9% (95% CI: 18–35%), respectively, for abnormal D-dimer, either without or with RVO. When compared with normal D-dimer without RVO, the multivariate HR for recurrence was similar for abnormal D-dimer either with RVO (4.76 – 95% CI:1.78–12.8) or without RVO (4.3–95%:1.56–11.88). Abnormal D-dimer at one month after OAT withdrawal is an independent risk factor for recurrent VTE, while RVO at the time of OAT withdrawal, either with normal or abnormal D-dimer after one month, does not influence the risk of recurrence.

The accuracy of a new bedside, rapid and quantitative D-Dimer assay (Cardiac D-Dimer) was evaluated in outpatients with clinically suspected deep vein thrombosis (DVT); VIDAS test was used as reference method. Eighty consecutive outpatients with suspected DVT of a lower limb were included in the study. Patients were classified as DVT positive or negative according to results of objective test (serial CUS), pretest clinical probability and 3-month follow-up. DVT was diagnosed in 32/80 patients (40%). The performance of the two D-Dimer assays was comparable, as indicated by the areas under the ROC curves (0.89 and 0.88, for Cardiac D-Dimer and VIDAS, respectively) and the coefficient of correlation (r=0.91). The reproducibility of the test was acceptable (from 6.2% to 12.0%). The sensitivity and negative predictive values were 100% for both tests. The specificity (SP) and positive predictive values (PPV) were similar (SP: 50.0% and 52.0%, PPV: 57.1% and 58.2%, for Cardiac D-Dimer and VIDAS, respectively). The Cardiac D-Dimer test proved to be very accurate and produced results fully comparable to those obtained with the VIDAS test. Since the test can be directly performed in the emergency room within a few minutes, it seems to have great clinical potential. The place of this assay in the diagnostic strategy of DVT remains to be determined in prospective management studies.

Background: D-dimer role is well established in the diagnostic work-up for lower limb deep vein thrombosis (DVT), however it has not been formally tested for clinically suspected upper extremity DVT and/or superficial vein thrombosis (SVT). Aim: To ascertain D-dimer diagnostic accuracy for upper extremity DVT and/or SVT. Study design: We performed a single centre management study in outpatients referred by emergency or primary care physicians for clinically suspected upper extremity DVT. All patients underwent D-dimer testing (cut-off value: = 93\% for excluding DVT in symptomatic outpatients and it can be a useful test in the diagnostic work-up of suspected upper extremity DVT. (C) 2015 Elsevier Ltd. All rights reserved.

OBJECTIVES: The aim of the present investigation was to study the endogenous circadian clock phase rhythm in cirrhotic patients. METHODS: The study population comprised 13 patients with cirrhosis (seven in Child-Pugh class A and six in classes B/C) and nine healthy controls. Plasma melatonin, tissue plasminogen activator antigen, and plasminogen activator inhibitor 1 antigen were measured at 4-h intervals over a 24-h period. Multiple-components rhythmometry using population mean cosinor methods were employed to analyze the findings. RESULTS: All three variables were characterized in both patients and controls by a statistically significant circadian rhythm, with similar profiles. The peak times of tissue plasminogen activator and plasminogen activator inhibitor 1 antigens were practically identical in controls and cirrhotic patients, irrespective of Child-Pugh class (calculated peak at times 6:52, 6:56, and 7:20 for the inhibitor in controls and Child-Pugh class A and classes B/C patients, respectively; p = ns), whereas the peak of melatonin was delayed in classes B/C patients (at times 2:08, 1:56, and 4:00, respectively; p CONCLUSION: The similar circadian phases of plasminogen activator inhibitor antigen in controls and cirrhotic patients in the present investigation indicates that the output rhythm of the internal timekeeping system is not shifted in this pathological condition.

Among the so called antiphospholipid (aPL) antibodies Lupus Anticoagulant (LAC) is considered the strongest risk factor for thromboembolic events. In individuals without a previous thromboembolic event (carriers), LAC is a risk factor when associated with the presence of anticardiolipin (aCL) and aβ2-Glycoprotein I (aβ2GPI) antibodies. On the other hand, data on carriers of isolated LAC positivity are sparse and inconclusive. The aim of this study was to prospectively determine the incidence of thrombosis in a cohort of carriers of isolated LAC positivity. One-hundred seventy-nine carriers of LAC confirmed twelve weeks apart and in a reference laboratory were studied. During a total follow up of 552 person-years, there were seven thromboembolic events (1.3% person-y). All the seven patients had at least one adjunctive major risk factor for thrombosis. The cumulative incidence of thromboembolic events was 3.1% (95% CI 0.6-5.6) after 2years, and 5.9% (95% CI 1.2-10.6) after 5 and 10years. On a multivariate regression analysis considering age, sex, autoimmune disease, risk factors for arterial and venous thrombosis, use of aspirin, only age was found to be an independent predictor of thromboembolic events (HR=1.1, 95% CI 1.0-1.2, p=0.02). These data might be relevant in clinical practice and underline the importance of differentiating LAC carriers in terms of isolated positivity or positivity associated with the presence of antibodies to aCL and β2-glycoprotein I.

Very low factor IX (FIX) levels in patients during oral anticoagulant (OA) treatment, due to mutations in the FIX gene, with prolonged activated partial thromboplastin time (aPTT) and associated with bleeding complications have recently been described. We measured aPTT in 595 OA patients while being within therapeutic ranges. Patients were divided into increasing international normalized ratio (INR) classes, and FIX determined in those belonging to the first and fifth quintiles of the aPTT distribution in each INR class. Results obtained in patients of all the first aPTT quintiles were compared with those of all the fifth quintiles. While INR was not different (2.79 versus 2.77 INR), aPTT was longer (1.65 versus 1.21 ratio; P < 0.0001) and FIX lower (0.29 versus 0.44 IU/ml; P < 0.0001) in patients of the fifth quintiles. Only one patient had a markedly reduced FIX (0.03 IU/ml). Bleeding rate was 4.8 and 6.2% patient-years (not significant) in patients of the first and fifth quintiles, respectively. We therefore found that FIX levels vary greatly in spite of similar achieved anticoagulation intensity; very low FIX is, however, a rare condition. In conclusion, screening for the recently identified mutations in FIX gene does not seem justified, and identifying patients with disproportionately prolonged aPTT, to detect those with particularly low FIX levels, is difficult because of the effect of the achieved anticoagulation intensity. Therefore, aPTT measurement is indicated only in patients with increased bleeding during OA.

ATIII is decreased in sepsis and/or shock and its baseline value correlates with mortality. The efficacy of ATIII therapy on mortality was assessed in a selected group of patients admitted to the intensive care unit (ICU) in a double-blind, randomized, multicenter study.120 patients admitted to the ICU with an ATIII concentration70% were randomized to receive ATIII (total dose 24000 units) or placebo treatment for 5 days; 56 patients had septic shock.ATIII concentrations in the treated group remained constant throughout the treatment period (range 97-102%). The Kaplan-Meier analysis showed no difference in overall survival between the two groups: 50 and 46% for ATIII and placebo, respectively. Septic shock and hemodynamic support were unbalanced in the two groups at admission. Therefore the Cox analysis was carried out after adjusting for these two variables. Treatment with ATIII decreases the risk of death with an odds ratio (OR) of 0.56. Of the covariates analyzed, septic shock and the baseline multiple organ failure score were negatively associated with survival and plasma activity level was positively associated with survival with an OR of 0.97 for each 1% increase in the ATIII plasma concentration at baseline.The results of ATIII treatment in this population of patients suggests that replacement therapy reduces mortality in the subgroup of septic shock patients only.

Summary Introduction The use of adapted cut-off values in the elderly, combined with clinical probability (PTP), increases the proportion of patients in whom venous thromboembolism (VTE) can be safely excluded, compared with the conventional cut-off value of 500 μg/L fibrinogen equivalent units (FEU). We evaluated the clinical performance of three different approaches to establish cut-off values for a D-dimer assay whose results are expressed in D-dimer units (D-DU). Methods HemosIL D-dimer HS assay (Instrumentation Laboratory) was performed in 279 consecutive outpatients with suspected deep venous thrombosis (DVT) and nonhigh PTP. Results Considering patients >60 years, the number of negative D-dimer results increased using the modified (376 ng/mL if ≥60 years) and the age-adjusted cut-off (age years × 5 ng/mL if >50 years) compared to the conventional one (230 ng/mL for all patients; 54.6%, 58.2%, and 25.0%, respectively), with no false-negative results. The higher increase was observed in patients >80 years (43.9%, 56.1%, and 8.8%, respectively). Conclusion For the HemosIL D-dimer HS, the use of specific cut-off values in older subjects with suspected DVT and nonhigh PTP increases the number of patients in whom DVT can be safely excluded.

Background Post-thrombotic syndrome (PTS) is the most common complication of deep vein thrombosis (DVT), but few data are available on the risk factors for PTS. Aims To assess whether the time-course of D-dimer, FVIII, and thrombotic burden are related to PTS development. Methods Patients (n = 59) with proximal DVT of the lower limbs (age 64; range:20-88 years; male 56%) were enrolled on the day of diagnosis (D0) and all received heparin for 5-7 days, overlapped and followed by vitamin K antagonists (VKA) for 3 months. Whole-leg compression ultrasound examination was conducted on D0 and 7 (D7), 30 (D30), and 90 (D90) days afterwards, when blood samples were also taken for D-dimer (STA Liatest) and FVIII (chromogenic assay) testing. Thrombotic burden was defined at each time point according to a score, which considered thrombosis extent and occlusion degree. Villalta score was evaluated at D30, D90, and D180. Results At D90, 12 patients developed PTS (Villalta score ≥ 5) and the median Villalta score was 1 (IQR 0.3-3.0) and was not correlated with either D-dimer or FVIIII time course. At D180, 13 patients had PTS and they had similar thrombotic score at D0, D30 to those without PTS, but higher at D90 (7.6 ± 5.1 vs. 3.2 ± 3.6; p = 0.011). Thrombotic score at D90 was correlated with Villalta score at D90 (rho = 0.374, p = 0.009) and at D180 (rho = 0.436, p = 0.006). Conclusions Thrombotic burden after 90 days of VKA is correlated with PTS.

Inherited and acquired thrombophilic alterations in patients with superficial vein thrombosis of lower limbs

Background/Aims: To evaluate risk factors for recurrent events in patients enrolled in the SteFlux (Superficial Thromboembolism Fluxum) clinical trial which compared different doses and duration of low molecular weight heparin (parnaparin) for superficial vein thrombosis (SVT). Materials and Methods: Outpatients with acute SVT of at least 4 cm in length of the internal or external saphenous veins or their collaterals were randomized in a double blind fashion to receive either parnaparin 8500 UI aXa od for ten days followed by placebo for 20 days or 8500 UI aXa od for ten days followed by 6400 UI aXa od for 20 days or 4250 UI aXa od for 30 days. Outcomes were the composite of symptomatic and asymptomatic deep vein thrombosis, pulmonary embolism and SVT recurrence or extension in the first 30+/-3 days with a 60+/-3 day follow-up. Results: 98 outcomes (14.7\%) were recorded during 93 days among 664 patients (M/F: 246/418, mean age 65). After correction for treatment, outcomes during 33 days were associated with previous venous thromboembolism (VTE) and/or SVT and/or family history of VTE (odds ratio-OR: 2.5; 95\% confidence interval -CI: 1.4-4.8; p = 0.003). After stopping LMWH treatment, only the absence of varicose veins (OR: 2.5; 95\% CI 1.3-5.0; p = 0.004) and previous VTE and/or SVT and/or family history of VTE (OR: 1.9; 95\% CI: 1.0-3.7; p = 0.048) were significantly associated with outcomes. Conclusions: SVT patients with these factors may deserve a higher intensity and/or longer anticoagulant treatment. (C) 2013 Elsevier Ltd. All rights reserved.

Introduction Treatment with B-vitamins and betaine reduces the high risk of thrombosis in patients with homocystinuria, a metabolic syndrome that is characterized by severe hyperhomocysteinemia (HHcy). In contrast, there is no clear demonstration that B-vitamins reduce the risk of thrombosis in patients with mild HHcy: for this reason, many question the clinical utility of measuring total Hcy (tHcy) in patients with thrombosis. However, thrombosis may be the first clinical manifestation of homocystinuria in patients reaching adulthood without signs and symptoms of the syndrome. Aim 1) to measure the prevalence of severe, previously undiagnosed, HHcy among patients with thrombosis 2) to profile these patients on the basis of their characteristics. Methods Six Italian Thrombosis Centers completed a first questionnaire, reporting tHcy levels in patients with thrombosis who underwent thrombophilia screening, and a second questionnaire, reporting the characteristics of patients with severe HHcy (tHcy>100μmol/L). Results Of 19,678 cross-sectionally collected patients with thrombosis who underwent thrombophilia screening in the last 12.5years (median value, range 6-17), 38 had severe HHcy (0.2%). Their median age at diagnosis was 47years (range 19-83) and the median level of tHcy was 130μmol/L (range 101-262). Venous thromboembolism (71%) was more frequent than arterial thromboembolism (26%); recurrent thrombosis occurred in 42% of cases. Conclusions Measurement of tHcy in adult patients with thrombosis may reveal the presence of severe HHcy. Since treatment of patients with severe HHcy decreases the risk of thrombosis, measurement of tHcy in patients with thrombosis may prove clinically useful.

Summary Background A successful outcome of pregnancy requires an efficient uteroplacental vascular system. Since this system may be compromised by disorders of haemostasis associated with a prothrombotic state, we postulated that maternal thrombophilia might be a risk factor for fetal loss. We studied the relation between heritable thrombophilic defects and fetal loss in a cohort of women with factor V Leiden or deficiency of antithrombin, protein C, or protein S. Methods We studied 1384 women enrolled in the European Prospective Cohort on Thrombophilia (EPCOT). Of 843 women with thrombophilia 571 had 1524 pregnancies; of 541 control women 395 had 1019 pregnancies. The controls were partners of male members of the EPCOT cohort or acquaintances of cases. We analysed the frequencies of miscarriage (fetal loss at or before 28 weeks of gestation) and stillbirth (fetal loss after 28 weeks of gestation) jointly and separately. Findings The risk of fetal loss was increased in women with thrombophilia (168/571 vs 93/395; odds ratio 1 35 [95% CI.1 01–1·82]). The odds ratio was higher for stillbirth than for miscarriage (3·6 [14–9·4] vs 1·27 [0·94–1·71]). The highest odds ratio for stillbirth was in women with combined defects (14·3 [2·4–86·0]) compared with 5·2 (1·5–18·1) in antithrombin deficiency, 2·3 (0·6–8·3) in protein-C deficiency, 3·3 (1·0–11·3) in protein-S deficiency, and 2·0 (0·5–7·7) with factor V Leiden. The corresponding odds ratios for miscarriage in these subgroups were 0·8 (0·2–3·6), 17 (10–2·8), 14 (0·9–2·2), 12 (0·7–19), and 0·9 (0·5–15). Significantly more pregnancy terminations had been done in women with thrombophilia than in controls (odds ratio 2·9 [1·8–4·8]); this discrepancy was apparent in nine of 11 participating centres and for all thrombophilia subgroups. Interpretation Women with familial thrombophilia, especially those with combined defects or antithrombin deficiency, have an increased risk of fetal loss, particularly stillbirth. Our findings have important implications for therapy and provide a rationale for clinical trials of thromboprophylaxis for affected women with recurrent fetal loss.

It was our aim to assess whether factor V Leiden (FVL) and G20210A prothrombin (FII) mutation are associated with the presence of residual vein obstruction (RVO) after a standard course of anticoagulation for a first episode of idiopathic proximal deep-vein thrombosis (DVT) of the lower limbs, with or without symptomatic pulmonary embolism (PE). Patients were enrolled in two prospective multicentre studies: PROLONG and PROLONG II. RVO was detected by compression ultrasonography according to the method of Prandoni on the day of anticoagulation withdrawal. Patients were also screened for FVL and FII mutation. The presence of FVL and/or FII mutation was determined in 872/963 (90.5%) patients, in 753 of whom RVO was assessed. FVL was significantly less frequent among subjects with isolated PE (7/176:4%) than among patients with either DVT and PE (15/133:11.3%; p=0.0018) or isolated DVT (89/563:15.8%; p

In family studies, the risk for venous thromboembolism (VTE) in relatives with factor V Leiden (FVL) or G20210A prothrombin (PT20210A) gene polymorphisms may differ according to genotype and clinical presentation of the proband. To address this hypothesis, a retrospective cohort family study was carried out on 192 kindreds with at least one member with homozygous FVL or PT20210A, for a total of 886 relatives. The proband of the family was heterozygous in 68 and homozygous or with both polymorphisms in 124 kindreds. Twenty-three probands were asymptomatic, 11 had had arterial thrombosis, 7 obstetrical complications, and 151 venous thrombosis (122 VTE and 29 superficial vein thrombosis). The incidence of VTE (per 1000 patient-years) in relatives was higher when the proband had heterozygous rather than homozygous polymorphism (1.25 [95\% confidence interval (CI), 0.73-1.91] vs 0.44 [0.19-0.78]) and when the proband had had VTE instead of other or no clinical manifestations (0.95 [0.57-1.42] vs 0.50 [0.19-0.96]). Compared with relatives belonging to kindreds with homozygous probands without VTE, the adjusted hazard ratio of VTE for relatives selected from kindreds with heterozygous probands with VTE was 4.14 (95\% CI, 1.17-14.71). The genotype and clinical presentation of the proband influence the risk for VTE in relatives with FVL or PT20210A.

The assessment of new and more sensitive serum markers for hepatocellular carcinoma (HCC) represents a useful contribution to the diagnosis of small liver tumors, still amenable by surgery. We evaluated the efficacy of the tumor markers proposed during recent years for the study of HCC: alpha fetoprotein (AFP), carcinoembryonic antigen (CEA), serum ferritin (SF), tissue polypeptide antigen (TPA), and, finally, the more recently proposed des-gamma-carboxy prothrombin (DCP). Of the 227 patients included in this retrospective study, 111 had HCC, and 85 of these were also cirrhotic. The remaining 116 patients, considered as the control group, included 23 patients with liver metastases from colorectal cancer, 26 with benign hepatic lesions, 20 with tumors other than HCC without hepatic metastases, and 47 with other liver diseases. For each single tumor marker, the sensitivity, specificity, positive and negative predictive values, diagnostic accuracy, and Younden index were assessed. AFP and DCP proved to be the most effective, with sensitivity, specificity, and diagnostic accuracy of 54.9%, 97.4%, and 76.6% and of 53.3%, 88.1%, and 71.1%, respectively. The same parameters evaluated for combined use of the two markers were 74.2%, 87.2%, and 80.9%, respectively. Analysis of the other markers produced no further significant contribution. Of the 111 patients with HCC, 35 (33.3%) were positive for both AFP and DCP, 43 (41%) were positive for one of them, and 27 (25.7%) were completely negative. In the 44 patients who underwent liver resection or transplantation, DCP correlated significantly with the histological presence of microvascular thrombosis, the major factor determining long-term survival after curative surgery. As a tumor marker for HCC, DCP is at least as effective as AFP; the combined use of AFP and DCP significantly improves the chances of identifying HCC by serodiagnosis.

BackgroundD-dimer testing to rule out deep vein thrombosis is less useful in older patients because of a lower specificity. An age-adjusted D-dimer cut-off value increased the proportion of older patients (>50 years) in whom pulmonary embolism could be excluded. We retrospectively validated the efficacy of this cut-off combined with clinical probability for the exclusion of deep vein thrombosis.Design and MethodsFive management study cohorts of 2818 consecutive outpatients with suspected deep vein thrombosis were used. Patients with non-high or unlikely probability of deep vein thrombosis were included in the analysis; four different D-dimer tests were used. The proportion of patients with a normal D-dimer test and the failure rates were calculated using the conventional (500 mg/L) and the age-adjusted D-dimer cut-off (patient's age x 10 mg/L in patients >50 years).ResultsIn 1672 patients with non-high probability, deep vein thrombosis could be excluded in 850 (51%) patients with the age-adjusted cut-off value versus 707 (42%) patients with the conventional cut-off value. The failure rates were 7 (0.8; 95% confidence interval 0.3-1.7%) for the age-adjusted cut-off value and 5 (0.7%, 0.2-1.6%) for the conventional cut-off value. The absolute increase in patients in whom deep vein thrombosis could be ruled out using the age-adjusted cut-off value was largest in patients >70 years: 19% among patients with non-high probability.ConclusionsThe age-adjusted cut-off of the D-dimer combined with clinical probability greatly increases the proportion of older patients in whom deep vein thrombosis can be safely excluded.

Background: Pretest clinical probability with the Wells rule and D-dimer have been widely investigated for the diagnosis of symptomatic proximal deep vein thrombosis (DVT) of the lower limbs, but they have not been formally tested for symptomatic isolated distal DVT diagnosis. Objective: To evaluate the diagnostic accuracy of the Wells rule and D-dimer for isolated distal DVT. Design, Setting, and Patients: This was a single-center, cross-sectional study including 873 consecutive outpatients with suspected DVT, in whom pretest clinical probability determination, D-dimer determination (STA Liatest; cut-off of 95% for isolated distal DVT. not available

Introduction Though thrombin-activatable fibrinolysis inhibitor (TAFI) may contribute to hypercoagulability during pregnancy, limited data are available on the role of TAFI in women with recurrent fetal loss. Material/Methods We performed a case–control study aimed at evaluating any possible association between TAFI levels and early recurrent fetal loss (≥ 3, or 2 with at least one normal fetal karyotype, before the 10th week of gestation). 140 women with early recurrent fetal loss and 140 age-matched healthy controls with at least one normal pregnancy were included. The number of miscarriages was 2.59 and occurred at gestational age 6.89 weeks. TAFI levels were determined by a chromogenic assay measuring total potential activatable TAFI. Results TAFI levels were significantly lower in early recurrent fetal loss women (12.2 ± 2.3 μg/ml vs 13.2 ± 2.6 μg/ml in healthy controls, p = 0.001). ORs of early recurrent fetal loss (crude and adjusted for possible confounding variables) were calculated after stratification of TAFI levels into quartiles. 25/140 (17.8%) early recurrent fetal loss women had TAFI levels above 14.0 μg/ml (4th quartile) vs 44/140 (31.3%) in healthy women (p = 0.014). Crude and adjusted ORs of early recurrent fetal loss in women with TAFI levels in the 4th quartile vs those in the reference category (1st quartile = below 11.0 μg/ml) were 0.42 (95%CI: 0.22-0.82) and 0.39 (95%CI: 0.19-0.80), respectively. Conclusions Our study provides evidence that high TAFI levels are associated with reduced risk of early recurrent fetal loss. Further studies are needed to better understand the actual role of TAFI in recurrent fetal loss.

SummaryWhether the administration of low-molecular-weight heparin (LMWH) during pregnancy is effective in preventing obstetric complications and pregnancy-related venous thromboembolism (VTE) in women who are carriers of factor V Leiden (FVL) and/or prothrombin variant G20210A (PTm) is controversial. This observational study investigated the possible efficacy of pharmacological treatment with LMWH ± aspirin (ASA) in pregnancy outcomes in 1,011 pregnancies of 416 women with thrombophilia (FVL and/or PTm). Most patients were chosen on the basis of previous obstetrical complications (36%), or because of familial or personal history of venous/arterial thromboembolism (28% and 18%, respectively); 74 patients (18%) were incidentally identified. The outcome was evaluated according to the type of treatment and of the period of pregnancy when the treatment was started. After adjustment for observation before and after diagnosis of thrombophilia, previous miscarriages and VTE, parity, age and centre, we observed that LMWH had a protective effect on miscarriages (odds ratio [OR] 0.52, 95% confidence interval [CI] 0.29–0.94) and VTE (OR 0.05, 95% CI 0.01–0.21). ASA appeared to have no effect on the prevention of obstetric complications and VTE. A nested analysis performed in 116 women with two or more obstetric complications confirmed that the highest number of live births was recorded in the group under LMWH prophylaxis (OR 0.19, 95% CI 0.05–0.75). These results suggest that LMWH prophylaxis reduces the risk of obstetric complications in carriers of FVL and/or PTm, particularly in those with previous obstetric events. Furthermore, LMWH prophylaxis reduces the risk of pregnancy-related VTE.

SummaryThe occurrence of a “rebound hypercoagulable state” in patients after discontinuation of oral anticoagulants is still a matter of debate and no definite recommendation can be made on the best procedure for anticoagulant withdrawal. The present study investigated the changes in the levels of markers of activated blood coagulation in 32 patients (pts) in whom warfarin treatment (for venous thromboembolic disease) was randomly withdrawn abruptly (n = 17, group A) or gradually (n = 15, group B: ⅔ of initial dose the 1st week, ⅓ the 2nd weeks and nothing from the 3rd week on). Blood was sampled at baseline, once a week for the first three weeks and after 2 months. At the 1st week group A had significantly higher F1+2 and TAT values (p

Warfarin dosing is affected by clinical and genetic variants, but the contribution of the genotype associated with warfarin resistance in pharmacogenetic algorithms has not been well assessed yet. We developed a new dosing algorithm including polymorphisms associated both with warfarin sensitivity and resistance in the Italian population, and its performance was compared with those of eight previously published algorithms.Clinical and genetic data (CYP2C9*2, CYP2C9*3, VKORC1 -1639 GA, and VKORC1 3730 GA) were used to elaborate the new algorithm. Derivation and validation groups comprised 55 (58.2% men, mean age 69 years) and 40 (57.5% men, mean age 70 years) patients, respectively, who were on stable anticoagulation therapy for at least 3 months with different oral anticoagulation therapy (OAT) indications.Performance of the new algorithm, evaluated with mean absolute error (MAE) defined as the absolute value of the difference between observed daily maintenance dose and predicted daily dose, correlation with the observed dose and R(2) value, was comparable with or slightly lower than that obtained using the other algorithms. The new algorithm could correctly assign 53.3%, 50.0%, and 57.1% of patients to the low (≤25 mg/week), intermediate (26-44 mg/week) and high (≥ 45 mg/week) dosing range, respectively. Our data showed a significant increase in predictive accuracy among patients requiring high warfarin dose compared with the other algorithms (ranging from 0% to 28.6%).The algorithm including VKORC1 3730 GA, associated with warfarin resistance, allowed a more accurate identification of resistant patients who require higher warfarin dosage.

The Prolong study shows that continuing vitamin K antagonists (VKA) in patients with abnormal D: -dimer (evaluated by a qualitative assay, Clearview Simplify D: -dimer) results in a significant reduction of venous thromboembolism (VTE) recurrence. The present study retrospectively analyzes a subgroup of patients enrolled in the Prolong study with a view to calculate cut-off values for six quantitative D: -dimer methods to predict the risk of VTE recurrence. We measured D: -dimer levels by VIDAS D: -dimer Exclusion (bioMerieux), STA Liatest D: -dimer (DiagnosticaStago), HemosIL D: -dimer and HemosIL D: -dimer HS (Instrumentation Laboratory), Innovance D: -dimer (Siemens) and AutoDimer (Trinity Biotech) in frozen plasma aliquots sampled 30 ± 10 days after VKA cessation in 390 patients enrolled in the Prolong study. During follow-up (562.7 years), 28 patients had recurrent VTE (7.2%, 5.0% person-years). Since D: -dimer levels are positively correlated with age and significantly lower in men, we calculated method-specific cut-off values according to age and gender. The HRs for VTE recurrence calculated using method-specific cut-off values based on age and gender are higher than those using cut-off values indicated by the manufacturers for VTE exclusion in symptomatic outpatients. These data suggest that method-specific cut-off values calculated according to patient age and gender can be more accurate in identifying patients at a higher risk for VTE recurrence. These method-specific cut-off values are being evaluated in the ongoing prospective management multicenter DULCIS study.

BACKGROUND: This randomized crossover study investigated the effects of unfractioned heparin (UFH) and low-molecular-weight heparin (LMWH) on intra- and post-dialytic blood levels of osteoprotegerin (OPG), receptor activator of nuclear factor kappa B ligand (RANKL) and inflammatory cytokines. METHODS: Forty patients on haemodialysis for at least 12 months were selected. UFH or LMWH was randomly assigned and maintained for 1 month, and then, in the following month, each patient was switched to the other form of heparin. In the mid-week session, we determined the changes in anti-Xa activity, OPG, RANKL, IL-1β, IL-6 and TNF-α values before heparin administration and after 15 min, 4, 8 and 24 h (T0, T1, T2, T3 and T4 respectively). Since these parameters at the various experimental times showed a non-normal distribution, log transformation was applied in order to run parametric ANOVA, with Bonferroni correction for multiple comparisons. RESULTS: The changes in anti-Xa activity over time were similar but not the same for the UFH and LMWH. A highly significant (P < 0.001) increase in anti-Xa activity was detected at T1, regardless of the type of heparin, as confirmed in the comparison of T0 vs T1 using one-way ANOVA. Moreover, with both heparins, significant differences were found in the comparisons of anti-Xa activity at T1 vs T2 (both P < 0.001) and at T2 vs T3 (P = 0.0003 with UFH; P < 0.001 with LMWH). Conversely, the difference in anti-Xa activity at T3 vs T4 was still significant with UFH (P = 0.0186) but not significant with LMWH (P = 0.728). When comparing anti-Xa activity at T4 vs T0, no significant differences were found either with UFH (P = 0.1996) or with LMWH (P = 0.7470), thus indicating that 24 h after heparin infusion, anti-Xa activity returned back to the pre-infusion values. When we analysed the changes in OPG levels over time, we found that the administration of heparin, regardless of the type, determined an increase in circulating OPG with a zenith at 15 min (T1), with a return back to the baseline levels within the 24th hour post-infusion. One-way ANOVA revealed significant differences in OPG blood levels at T0 vs T1 with both UFH (P = 0.0112) and LMWH (P = 0.0288), whereas no significant difference was observed in the comparisons of OPG levels at T1 vs T2, T2 vs T3, T3 vs T4 and T4 vs T0, either with UFH or with LMWH. The circulating levels of RANKL, IL-1β, IL-6 and TNF-α at the different intra- and post-dialytic times did not show significant variations following heparin administration, either with UFH or with LMWH. One-way ANOVA performed on the log-transformed values of RANKL, IL-1β, IL-6 and TNF-α at the various experimental times (T0 vs T1, T1 vs T2, T2 vs T2, T3 vs T4 and T4 vs T0) revealed no significant intra- and post-dialytic changes in their blood levels, thus confirming that heparin infusion did not affect their blood levels. CONCLUSIONS: These results suggest that heparin-regulated cyclic increases of OPG might play a role in the vascular pathology of haemodialysis patients.

Persistent antiphospholipid (aPL) antibodies are occasionally found in subjects without prior history of thromboembolic events (TEs), raising the dilemma of whether to initiate or not a primary thromboprophylaxis. A first TE is considered rare in aPL carriers, but previous studies did not consider the aPL profile nor was the test positivity confirmed in a reference laboratory. In this study, 104 subjects with high-risk aPL profile (positive lupus anticoagulant, anticardiolipin, and anti-β2–glycoprotein I antibodies, triple positivity) confirmed in a reference laboratory, were followed up for a mean of 4.5 years. There were 25 first TEs (5.3% per year): the cumulative incidence after 10 years was 37.1% (95% confidence interval [CI], 19.9%-54.3%). On multivariate analysis, male sex (hazard ratio = 4.4; 95% CI, 1.5-13.1, P = .007) and risk factors for venous thromboembolism (hazard ratio = 3.3; 95% CI, 1.3-8.5, P = .01) were independent predictors for TEs. Aspirin did not significantly affect the incidence of TE. In conclusion, the occurrence of a first TE in carriers of high-risk aPL profile is considerable; it is more frequent among male subjects and in the presence of additional risk factors for venous TE. These data can help in the decision to initiate primary thromboprophylaxis in these subjects.