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Antiphospholipid syndrome (APS) is an acquired thrombophilia with an uncertain role in the development of chronic thromboembolic pulmonary hypertension (CTEPH). We aimed to assess the association of APS with the clinical phenotype of CTEPH. We retrospectively reviewed data of CTEPH patients referred to our center. Clinical, angiographic, and hemodynamic data were available for all patients. APS was diagnosed in the presence of one or more positive antiphospholipid (aPL) tests confirmed more than 12 weeks apart. Data were compared between APS-positive and APS-negative patients. From May 2013 to December 2018, 297 patients with CTEPH were enrolled. Twenty-three (7.7%) were positive for laboratory tests exploring aPL antibodies. Among them, 17 patients (74%) had a triple positive aPL profile. When compared with the APS-negative group, APS patients were significantly younger (30.0 ± 11.1 vs. 55.6 ± 12.9 years, p  < 0.0001), had more frequently a history of pulmonary embolism (95.6% vs. 65.7%, p  = 0.003), and had more frequently associated autoimmune disease (43.5% vs. 2.9%, p  < 0.0001). In APS-positive patients, pulmonary artery lesions were more proximal and hemodynamic profiles were less compromised. Our results show that patients with APS are a unique group of CTEPH patients with well-defined clinic and hemodynamic characteristics., Competing Interests: None declared., (Georg Thieme Verlag KG Stuttgart · New York.)

The effect of additional treatments combined with conventional therapy on pregnancy outcomes was examined in high-risk primary antiphospholipid syndrome (PAPS) patients to identify the most effective treatment strategy. The study's inclusion criteria were (1) positivity to lupus anticoagulant alone or associated with anticardiolipin and/or anti-β2 glycoprotein I antibodies; (2) a history of severe maternal-foetal complications (Group I) or a history of one or more pregnancies refractory to conventional therapy leading to unexplained foetal deaths not associated with severe maternal-foetal complications (Group II). Two different additional treatments were considered: oral-low-dose steroids (10-20 mg prednisone daily) and/or 200 to 400 mg daily doses of hydroxychloroquine and parenteral-intravenous immunoglobulins at 2 g/kg per month and/or plasma exchange. The study's primary outcomes were live birth rates and pregnancy complications. A total of 194 pregnant PAPS patients attending 20 tertiary centres were retrospectively enrolled. Hydroxychloroquine was found to be linked to a significantly higher live birth rate with respect to the other oral treatments in the Group II patients. The high (400 mg) versus low (200 mg) doses of hydroxychloroquine ( p  = 0.036) and its administration before versus during pregnancy ( p  = 0.021) were associated with a significantly higher live birth rate. Hydroxychloroquine therapy appeared particularly efficacious in the PAPS patients without previous thrombosis. Parenteral treatments were associated with a significantly higher live birth rate with respect to the oral ones ( p  = 0.037), particularly in the Group I patients. In conclusion, some additional treatments were found to be safe and efficacious in high-risk PAPS pregnant women., Competing Interests: None., (Schattauer GmbH Stuttgart.)

The SAMe-TT2R2 score has recently been proposed to predict the quality of vitamin K antagonist (VKA) anticoagulation control in patients with atrial fibrillation. We aimed at investigating whether the score is effective also in patients with venous thromboembolism (VTE). Patients included in the START-Register because started VKA therapy for a recent VTE episode and with > 3 months follow-up were analyzed. The score was calculated using the baseline patient's characteristics present in the electronic database of the registry, where all INR results were also available and analysed to calculate the time in therapeutic range (TTR). A total of 1308 patients (53.4 % female, median age 68 years) were analysed. During 998 patient-years follow-up, the median TTR was 63 %. The maximum score in the patients was 4, with 70 % of them having 0-1. INR controls within range (2.0-3.0) were significantly less prevalent in patients with score ≥ 2 vs 0-1 score (58.5 ± 20 % vs 61.5 ± 19 %, respectively, p = 0.046). Patients with score ≥ 2 vs 0-1 had a highly significant lower TTR during the first 3 months of therapy (53 ± 26 % and 61 ± 26 %, respectively; p=0.0001), difference mainly due to more time spent below 2.0 INR (38 ± 28 % vs 31.3 ± 26.7 %, respectively; p=0.0001). In conclusion, the study proved, for the first time, that the SAMe-TT2R2 score is useful to predict among VTE patients those who will have good (score 0-1) or less good (score ≥ 2) VKA anticoagulation control. The score can help decision-making in everyday clinical practice, especially when choosing between VKA and non-vitamin K antagonists direct anticoagulants.

Antiphospholipid antibodies (aPL) seem to induce a prothrombotic state by activating endothelium and platelets, but no studies have evaluated systematically the effects of aPL from patients with the antiphospholipid syndrome (APS) in quiescent versus catastrophic phase. Our aims were to evaluate the in vitro effects on platelet activation of anti-β2 glycoprotein I (anti-β2GPI) antibodiesisolated from APS patientin either quiescent or catastrophic phase and to investigate ex vivo platelet and endothelial activation in patients with quiescent or catastrophic APS. Anti-β2GPI antibodies were isolated from plasma of a pregnant woman in two different stages of APS (quiescent and catastrophic, respectively). They were co-incubated with washed platelets from healthy controls that were then challenged with TRAP-6 (thrombin receptor activating peptide 6) and the expression of P- selectin (P-sel) on platelets was assessed by flow cytometry. Moreover, plasma samples from six patients with quiescent, four with catastrophic APS and 10 controls were assessed for several markers of platelet and endothelial activation. The results showed that purified anti-β2GPI antibodies co-incubated with platelets enhanced TRAP-6- induced platelet P-sel expression. Notably, anti-β2GPI antibodies isolated during the catastrophic phase enhanced platelet P-sel expression more than antibodies isolated from the same patient in the quiescent stage of disease. Moreover, APS patients had significantly higher plasma levels of soluble (s) Psel, sCD40 ligand, soluble vascular cell adhesion molecule 1 and monocyte chemoattractant protein 1 than control subjects. In addition, sP-sel and von Willebrand factor activity were significantly higher during catastrophic than in quiescent phase.

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<0.0001), confirming the FVL paradox. The rate of FII mutation was similar among patients with isolated PE (11/176:6.2%) and patients with either DVT and PE (12/133:9%) or isolated DVT (52/563:9.2%). FVL and FII mutation were not significantly associated with RVO at the multivariate analysis in all patients, although data suggest that FVL and FII mutation may have a differential effect on RVO in the subgroups of patients with DVT and DVT plus PE patients. Male sex and isolated DVT were significantly associated with RVO in all patients. In conclusion, male sex and isolated DVT are associated with RVO, while FVL and FII mutations are not significantly associated with RVO in this study.

Dabigatran and other new oral anticoagulants (OAC) represent a step forward in stroke prevention in patients with atrial fibrillation (AF). They indeed have been shown to be an alternative to vitamin K antagonists (VKAs) without the burden of laboratory control. However, these new drugs compete with an effective and well-established therapy, thus bringing about a series of questions and doubts. In this report members of the board of the Italian Federation of Thrombosis Centers (FCSA) answer some questions every clinician might be confronted with.

The PROLONG randomised clinical trial showed that an abnormal D-dimer at one month after vitamin K antagonist (VKA) suspension for a first episode of unprovoked venous thromboembolism (VTE) is associated with a higher risk of recurrence. However, other patient characteristics, such as comorbidities, in combination with D-dimer could also influence the recurrence risk. It was the objective of this study to assess the predictive value of comorbidities and D-dimer in combination for recurrence after withdrawal of VKA in patients enrolled in the PROLONG study. On the day of VKA suspension, the presence of known (coronary, peripheral,cerebral) vascular disease, chronic inflammatory bowel disease, chronic obstructive pulmonary disease, autoimmune disease, diabetes, arterial hypertension, obesity and dyslipidaemias was registered. D-dimer was measured at 30 +/- 10 days afterwards. The primary outcome was recurrent objectively documented VTE. Mean follow-up was 2.55 years. An abnormal D-dimer was observed in 44% (135/309) of patients with comorbidities and in 29% (87/299) of patients without (p=0.0003). An on-treatment analysis was conducted in 483 patients in whom VKAs were not resumed. In patients with a normal D-dimer, recurrences were observed in 14.3% (24/168) of patients with comorbidities and 10.8% (22/203) of subjects without (p=ns). In patients with an abnormal D-dimer, recurrences were observed in 24.6% (16/65) patients with comorbidities and 21.3% (10/47) of patients without (p=ns). Although abnormal D-dimer levels were significantly more frequent in patients with comorbidities, D-dimer was an independent risk factor for recurrence and the presence of comorbidities did not increase the risk of recurrence associated with an abnormal post-anticoagulation D-dimer.

It has been observed that elderly patients with nonvalvular atrial fibrillation (NVAF) benefit from standard [an international normalised ratio (INR) goal of 2.0-3.0] oral anticoagulant treatment (OAT). The hypothesis that lower-intensity anticoagulation therapy can offset the higher bleeding risk in this population has never been tested in an 'ad hoc' clinical trial. Patients over 75 years of age with NVAF were randomised to receive warfarin to maintain the INR at 1.8 (range 1.5-2.0) or at a standard target of 2.5 (range 2.0-3.0). There were 135 patients in the low-intensity and 132 in the standard-intensity groups. During a mean follow-up lasting 5.1 years, 59 primary outcome events (thromboembolism and major haemorrhage) were recorded, 24 (3.5 per 100 patient-years) in the low-intensity group and 35 (5.0 per 100 patient-years) in the standard-intensity group (HR=0.7, 95% CI 0.4-1.1, p=0.1). The reduction in the primary endpoint was mainly due to a diminution in major bleedings (1.9 vs. 3.0 per 100 patient-years; HR=0.6, 95% CI 0.3-1.2, p=0.1). The median achieved INR value was 1.86 in the low-intensity and 2.24 in the standard-intensity group (p<0.001). The frequency of INR testing was 26.1 +/- 13.5 vs. 24.3 +/- 11.6 days, p<0.0001). In this exploratory study we observed a low rate of stroke and major bleeding in elderly patients (>75) being managed in an anticoagulation clinic for primary stroke prevention with low-intensity anticoagulation (INR 1.5-2.0). However, further trials are needed to confirm the hypothesis generated by the present study.

In women diagnosed as having category I primary obstetric antiphospholipid syndrome, clinical characteristics and the risk of subsequent thromboembolic events and further unsuccessful pregnancy has not been clearly documented. Women with unexplained obstetric complications and no definite autoimmune systemic diseases were tested for lupus anticoagulant (LA), IgG/IgM anticardiolipin (aCL) and IgG/IgM anti-human beta2-Glycoprotein I (abeta2GPI) antibodies and diagnosed as having primary antiphospholipid syndrome (APS) in classification category I on the basis of more than one laboratory criteria present in any combination. Characteristics at the time of diagnosis and risk factors for subsequent clinical events during a mean follow-up of 6.3 years were evaluated. Fifty-three of 600 women studied were found to fulfil obstetric criteria and had more than one positive laboratory test at the time of diagnosis. All the women were aCL and abeta2GPI positive, and 16 were also LA positive. This latter group (triple positivity) had distinct features and had more frequently experienced previous thromboembolism (OR = 122.5, 95% CI 16-957, p < 0.001). They also had an increased rate of late pregnancy loss (OR = 16.2, 95%CI 0.9-292, p = 0.01), and a higher IgG abeta2GPI titer at diagnosis (median, 25(th) and 75(th) percentile were 118, 37-962, vs. 23, 18-32, respectively, p < 0.0001). During follow-up, the rate of thromboembolic events was significantly higher in the group of women with triple positivity and/ or previous thromboembolism (OR = 57.5, 95% CI 2.7-1160, p = 0.0004) which were the only independent predictors of TE in the multivariate model. Recurrent pregnancy loss took place in seven out of 47 women who had a new pregnancy. Triple positivity and/or previous thromboembolism were again the only independent markers (OR = 34.4, 95% CI 3.5-335.1, p = 0.003) of an unsuccessful new pregnancy. In conclusion, in primary APS with pregnancy morbidity in classification category I, quite different groups of patients may be identified on the basis of laboratory tests. Triple positivity and/or a history of thromboembolism predict new TE events and new unsuccessful pregnancies.

Among the so called 'antiphospholipid antibodies', the presence of Lupus Anticoagulant (LA) is associated with thrombosis-related events and defines the antiphospholipid syndrome. The role of anti-cardiolipin (aCL) antibodies and anti-human beta2-glycoprotein I (abeta2GPI) antibodies is less striking. Since the problem of standardization for these tests is far from resolved, we evaluated whether the combination of results (antiphospholipid laboratory profiles) could help to better classify these patients. Over a 6-year period, 618 consecutive subjects (55% of whom had previous documented thrombosis-related events) were referred to our clinic for Antiphospholipid antibody detection. LA was detected according to internationally accepted recommendations. ACL and abeta2GPI antibodies were detected by Enzyme-Linked-Immunosorbent Assay (ELISA). Patients' records were reviewed for the presence of previous thromboembolic events or obstetric complications according to Sapporo's clinical criteria for the diagnosis of antiphospholipid syndrome (APS) and each patient underwent a physical examination. When individual tests were considered in a multivariate analysis which took into account age, gender, the presence of SLE or other autoimmune diseases and established risk factors for venous and arterial thromboembolism, LA (Odds Ratio 4.4, Confidence Interval 1.5-13.3) and abeta2GPI antibodies (Odds Ratio 2.9, Confidence Interval 1.1-7.5) but not aCL antibodies (Odds Ratio 1.2, Confidence Interval 0.5-2.7) were found to be independent risk factors for thrombosis-related events. When antiphospholipid antibody profiles instead of individual test positivity were analyzed in the above mentioned model, triple positivity resulted a strong independent risk factor (Odds Ratio 33.3, Confidence Interval 7.0-157.6), retaining its significance when the association with venous or arterial thromboembolism was considered. Double positivity with negative LA was close to significance for thrombosis-related events (Odds Ratio 2.2, Confidence Interval 1.0-5.2, p=0.056) and highly significant risk factor for obstetric complications (Odds Ratio 10.8, Confidence Interval 2.9-40.8). Other combinations did not reach statistical significance. The mean level of IgG abeta2GPI antibodies was statistically higher in triple positive profile and might account for positive LA. As compared to a single test, the analysis of a complete antiphospholipid antibody profile can better determine patients at risk.

Mitral stenosis (MS) in association with atrial fibrillation (AF) is a clinical condition at high risk for systemic thromboembolism. Although oral anticoagulants greatly reduce the incidence of thromboembolism in these patients, the optimal intensity of treatment has never been tested in specific clinical trials, and current recommendations are derived from studies of nonrheumatic AF. In this study we tested the effectiveness of two different intensities. The study design was carried out as an open randomized prospective study in an anticoagulation clinic. We randomized 103 patients with MS and AF to a low (target INR = 2) or moderate (target INR = 3) anticoagulation regimen. The primary end points were systemic thromboembolism, major bleeding and vascular death. During a mean follow-up of 4.5 years, 1 systemic embolism occurred in the low intensity group (0.41 per 100 pt/yrs, CI 0.01-2.3), and 1 minor stroke occurred in the moderate intensity group (0.40 per 100 pt/yrs, CI 0.01-2.3; p = ns). Major bleeding occurred in 8 patients, with 3 in the low intensity (1.25 per 100 pt/yrs) and 5 in the moderate intensity group (2.0 per 100 pt/yrs, Incidence Rate Ratio 0.6, CI 0.1-3.1; p = ns). Total events (systemic embolism, major bleeding and vascular death) occurred in 7 low intensity patients and 8 moderate intensity patients. As expected, minor bleeding was more frequent in the moderate intensity group of patients, who actually had more intense treatment and required closer monitoring of oral anticoagulant treatment. These data suggest that low intensity anticoagulation, as performed in an anticoagulation clinic, is effective and safe in high risk patients with MS and AF.

Oral anticoagulants (OA) are the drug of choice for stroke prevention in patients with non-rheumatic atrial fibrillation (NRAF). This clear benefit/risk ratio comes from several randomized clinical trials (RCT) in which highly selected patients were strictly monitored. The aim of this study was to ascertain whether the safety of OA was also obtained outside the setting of clinical trials in consecutive patients starting treatment and routinely followed at Italian anticoagulation clinics. A total of 433 patients with NRAF were enrolled in the ISCOAT study and followed up for a mean of 1.4 years. Two patients (0.3% per year) suffered from a complete non-fatal ischemic stroke, 8 patients (1.3% per year) died of thrombosis-related vascular death, and 11 patients (11 events, 1.8% per year) suffered from major bleedings (2 fatal). Major bleeding occurred more frequently in patients >75 years of age (6 events, 5.1% per year) than in younger patients (5 events, 1.0% per year). The cumulative incidence of major bleeding in patients over 75 years of age (10.8%; 95% CI, 1.8-19.8) was significantly higher than in younger patients (2.8%; 95% CI, 0.3-5.3, p = 0.006). Major primary bleeding unrelated to organic lesions (7 patients, 1 male and 6 females) occurred in 5 elderly patients (>75 years old) with a cumulative incidence (9.6%; 95% CI 0.8-18.4) significantly higher than in younger patients (1.2%; 95% CI, 0-3.0, p = 0.0003). Univariate analysis revealed a higher frequency of major primary bleeding in females, in diabetic patients and in in those who had suffered a previous thromboembolic event. Multivariate analysis revealed that only age grater than 75 years was independently related to major primary bleedings (RR 6.6; 95% CI 1.2-37, p = 0.032). Minor bleedings (n = 27) were not more frequent in elderly patients (6% vs 4% per year, p = ns). Patients were kept at optimal intensity of treatment for 63% of the time. These data confirm the efficacy of OA but identify elderly patients as a high risk group of major bleeding.

Contrary to infective anticardiolipin (aCL) antibodies, autoimmune aCL antibodies react with phospholipids (PL) mainly via binding to the plasma glycoprotein cofactor beta2-Glycoprotein I (beta2GPI). While there is a well-documented link between the risk of thrombosis and the presence of beta2GPI-dependent anticardiolipin antibodies, the pathological impact of other antiphospholipid antibodies is less clear. By means of cardiolipin affinity-chromatography, we isolated and identified 3 CL-binding proteins, complement component C4, complement factor H and a kallikrein-sensitive glycoprotein, and tested for the presence of autoantibodies against these proteins in patients with antiphospholipid syndrome (APS), systemic lupus erythematosus (SLE) and other autoimmune diseases. High titers of autoantibodies to C4 as compared to age- and sex-matched healthy controls were present in 3 of 26 patients with APS, and weak titers were found in 2 of 26 patients with SLE and in none of 26 patients with other autoimmune diseases. Autoantibodies to complement factor H were found in 4 APS, 3 SLE and none of the other autoimmune patients. Autoantibodies to kallikrein-sensitive glycoprotein were detected in 6 APS patients, 1 SLE patient, and 1 patient with another autoimmune disease. A close relationship between these antibodies was found, suggesting their origin from a common macromolecular complex. However, no relationship with anti-beta2GPI antibodies was found, with the three patients with higher levels of autoantibodies having a low titer of anti-beta2GPI antibodies. In conclusion, some patients with APS harbor circulating antibodies to other CL-binding proteins which might be useful to further characterize these patients.

The optimal long-term treatment of acute venous thromboembolism (VTE) in patients with malignancy remains undefined. In particular, based on current evidence, it is uncertain whether secondary prophylaxis using standard intensity oral anticoagulant therapy is associated with higher risks of bleeding and recurrent thrombosis in patients with cancer than in those without cancer. This study compared the outcome of anticoagulation courses in 95 patients with malignancy with those of 733 patients without malignancy. All patients were participants in a large, nation-wide population study and were prospectively followed from the initiation of their oral anticoagulant therapy. Based on 744 patient-years of treatment and follow-up, the rates of major (5.4% vs 0.9%), minor (16.2% vs 3.6%) and total (21.6% vs 4.5%) bleeding were statistically significantly higher in cancer patients compared with patients without cancer. Bleeding was also a more frequent cause of early anticoagulation withdrawal in patients with malignancy (4.2% vs. 0.7%; p <0.01; RR 6.2 (95% CI 1.95-19.4). There was a trend towards a higher rate of thrombotic complications in cancer patients (6.8% vs. 2.5%; p = 0.058; RR 2.5 [CI 0.96-6.5]) but this did not achieve statistical significance. In the group of patients with cancer, the bleeding rate was high across the different INR categories and was independent of the temporally associated International Normalized Ratio (INR). In contrast, the bleeding rate was increased only with INR values greater than 4.5 in the group of patients without cancer. The rate of thrombotic events was significantly higher in both cohorts when the INR was less than 2.0. In conclusion, patients with malignancy treated with oral anticoagulants have a higher rate of bleeding and possibly an increased risk of recurrent thrombosis compared with patients without malignancy. Safer and more effective anticoagulant therapy is needed for this challenging group of patients.

The aims of this study were to investigate on the general adhesion of the patients to oral anticoagulant therapy, and particularly on the quality of life of our patients, the doctor-patient relationship and the Centre-patient relationship. For this purpose we administered a questionnaire containing 17 main questions each with a maximum of 4 secondary questions. The questionnaire was administered to two groups of 127 and 137 oral anticoagulated patients (127 males and 137 females, mean age 55 +/- 19 years), followed at two Anticoagulation Clinics, in two Italian cities, Cagliari (Sardinia) and Padua (North East Italy). The cities differed in the number of patients monitored and the management modalities of anticoagulation. The results show that oral anticoagulant therapy does not limit the life-style of the patients. Only 11% of the patients complain of limitations to their daily life. Fifty-two percent believe their health has improved, and 87% are not afraid of negative consequences. The doctor-patient relationship is considered very important by 96% of patients. Seventy-eight percent refer to the Anticoagulation Clinic also for other health problems, 93% consider it important to be assessed by the doctor at the Anticoagulation Clinic, while 83% believe the doctor should always hand out the results personally. We conclude that in general oral anticoagulant therapy is accepted by the majority of patients, in spite of the need for periodic monitoring. The doctor-patient relationship should be taken into account, even in the case of a monitored, computer-assisted method of dose-adjustment.

Anti-beta2-glycoprotein I (beta2-GPI) antibodies behave as classical Lupus Anticoagulants (LA), as they inhibit phospholipid-dependent coagulation reactions and their activity disappears in the presence of excess exogenous phospholipids (PLs). We have recently shown that a certain amount of PLs in the dilute Russell Viper Venom Time (dRVVT) test system is required to express LA activity of anti beta2-GPI antibodies. We have now extended this observation to two other tests, i.e., Kaolin Clotting Time (KCT) in which PLs are not added, and Tissue Thromboplastin Inhibition test (TTI) in which PLs are extremely diluted. In fact, affinity-purified antibody preparations from 5 patients with antiphospholipid syndrome did not express or only weakly expressed anticoagulant activity in both tests; the mean ratios of coagulation times obtained with purified antibodies and that of control buffer were 1.11 and 1.0 for KCT and TTI, respectively. On the contrary, the mean ratios in dRVVT were 1.31 and 1.49 at a PLs dilution of 1:8 and 1:64, respectively. Therefore, the presence of LA activity due to autoantibodies to beta2-GPI is characterized by a positive dRVVT and negative or only weakly positive KCT and TTI.

The paper reports on rate and type of thrombotic events occurring during the observational, prospective, inception-cohort, multicenter ISCOAT study. 2,745 unselected, daily practice patients, consecutively referring to 34 Italian anticoagulation clinics to monitor the oral anticoagulant treatment, were included in the study from beginning of their first anticoagulant course. During a total follow-up of 2,011 patient-years of treatment 70 thrombotic events (3.5 per 100 patient years) were recorded in 67 patients: 20 fatal (1%), 39 major (1.9%) and 11 minor (0.6%). 34/70 events occurred within the first 90 days of treatment (relative risk - at multivariate analysis - of < or =90 days vs. >90 = 20.6, C.I. 12.7-33.5; p <0.0001). The risk was higher in patients aged > or =70 y (1.62, C.I. 1.0-2.61; p <0.05), and when indication for anticoagulant treatment was peripheral/cerebral arterial disease (1.84, C.I. 1.01-3.36; p <0.05). The frequency of thrombotic events was 17.5% when international normalised ratio (INR) levels were < 1.5, decreasing to 2.3% for INRs within the 2-2.99 category (relative risk of INRs <2.0 vs. > or =2 = 1.88, C.I. 1.16-3.07; p <0.05). The recorded rate of thrombotic events was lower than that reported in the few available studies. A greater risk should be expected during the first 90 days of treatment, when anticoagulation levels are <2.0 INR, in patients > 70 years and in those with cerebrovascular/peripheral arterial disease.

Background: The long-term administration of oral anticoagulants to patients with mechanical heart valve prostheses is generally accepted. However, the appropriate intensity of oral anticoagulant treatment in these patients is still controversial., Methods and Results: From March 1991 to March 1994, patients referred to the Padova Thrombosis Center who had undergone mechanical heart valve substitution at least 6 months earlier were randomly assigned to receive oral anticoagulants at moderate intensity (target INR = 3) or moderate-high intensity (target INR = 4). Principal end points were major bleeding, thromboembolism and vascular death. Minor bleeding was a secondary end-point. A total of 104 patients were assigned to the target 3 group and 101 to the target 4 group; they were followed for from 1.5 years to up 4.5 years (mean, 3 years). Principal end-points occurred in 13 patients in the target 3 group (4 per 100 patient-years) and in 20 patients in the target 4 group (6.9 per 100 patient-years). Major hemorrhagic events occurred in 15 patients, 4 in the target 3 group (1.2 per 100 patient-years) and 11 in the target 4 group (3.8 per 100 patient-years) (p = 0.019). The 12 recorded episodes of thromboembolism, 4 of which consisted of a visual deficit, were all transient ischemic attacks, 6 in the target 3 group (1.8 per 100 patient-years) and 6 in the target 4 group (2.1 per 100 patient-years). There were 3 vascular deaths in each group (0.9 and 1 per 100 patient-years for target 3 and target 4 groups, respectively). Minor bleeding episodes occurred 85 times (26 per 100 patient-years) in the target 3 group and 123 times (43 per 100 patient-years) in the target 4 group (p = 0.001)., Conclusions: Mechanical heart valve patients on anticoagulant treatment who had been operated on at least 6 months earlier experienced fewer bleeding complications when maintained on a moderate intensity regimen (target INR = 3) than those on a moderate-high intensity regimen (target INR = 4). The number of thromboembolic events and vascular deaths did not differ between the two groups.

Lupus anticoagulant (LA) is a general term to define immunoglobulins interfering with phospholipid-dependent coagulation tests. It is now clear that the phospholipid-dependence of some LA is related to the presence of the phospholipid-binding plasma protein beta 2-glycoprotein I (beta 2-GPI) and that autoantibodies to beta 2-GPI might represent a specific category of LA. To verify this hypothesis we have purified IgG autoantibodies to beta 2-GPI from plasma of 6 patients with antiphospholipid antibody syndrome, by means of agarose-immobilized human beta 2-GPI. All 6 preparations tested positive in anti-beta 2-GPI IgG antibody ELISA and showed a marked LA activity by prolonging dilute Russell Viper Venom Time (dRVVT) from a minimum of 5.3 s in patient # 1 to a maximum of 41.1 s in patient # 3. These IgG preparations behaved as typical LA, with this activity tending to disappear in the presence of increasing phospholipid (PL) concentrations. Moreover, the LA activity of the IgG preparations was not detectable in the absence of PL, in which case the ratio between dRVVT obtained in the presence and absence of IgG autoantibodies to beta 2-GPI was close to 1. This pattern was confirmed by using plasma from patients with antiphospholipid antibody syndrome testing positive for anti-beta 2-GPI IgG antibodies. These findings suggest that dRVVT performed both in the presence and absence of PL might constitute a sensitive screening test to detect specific antibodies with LA activity.

Anti-phospholipid (aPL) antibodies are defined as antibodies detected in systems employing phospholipids (PL). This general definition is misleading as it comprises a large group of autoimmune phospholipid-reactive antibodies that are directed against specific phospholipid-binding plasma proteins, such as beta 2-glycoprotein I (beta 2GPI) and prothrombin. Definition of phospholipid-reacting antibodies according to the plasma protein against which they are directed appears more appropriate and could be useful in understanding clinical events and pathogenic mechanisms. Using ELISA systems we have studied the presence of antibodies directed against specific phospholipid-binding proteins in a series of 22 patients with thrombosis and phospholipid-reactive antibodies of the IgG isotype. High levels of anti-beta 2 GPI IgG were detected in all 22 patients. Normal values were calculated on the basis of OD values at 405 nm (OD405) obtained for 22 age- and sex-matched healthy subjects (cut off value = 0.401). Levels of anti-beta 2 GPI antibodies were linearly correlated with those of cardiolipin-reactive (aCL) antibodies. Eleven out of 22 patients (50%) had values of anti-prothrombin antibodies exceeding the cut-off value of 0.250. No relationship was found between the levels of anti-beta 2GPI and anti-prothrombin antibodies. Tests for antibodies against two natural inhibitors of blood coagulation, protein C and protein S, revealed elevated levels of anti-protein C IgG and anti-protein S IgG in 4 and 12 patients, respectively. A highly significant correlation between anti-protein C IgG and anti-protein S IgG values as well as between antibody titers against the two studied natural coagulation inhibitors and anti-prothrombin IgG was found. When comparing patients positive for aCL and presence or absence of a previous thrombotic episode (aCL+/T+ vs aCL+/T-), the positivity of anti-beta 2GPI IgG was found to be statistically associated with thrombosis. Conversely, among patients with previous thromboembolism with or without aCL (aCL+/T+ vs aCL-/T+ vs aCL-/T+) the positivity of anti-beta 2GPI IgG was strictly associated with the positivity of aCL, thus identifying the aPL antibody syndrome. These data demonstrate that anti-beta 2GPI antibodies are a marker of "autoimmune" thrombosis. Anti-prothrombin antibodies are not a marker of thrombosis and are closely associated with antibodies to protein C and protein S.

The antiphospholipid antibodies (aPL) present in autoimmune disorders are associated with thromboembolic episodes, and their binding to phospholipids (PL) is mediated by a plasma cofactor, beta 2-glycoprotein I (beta 2GPI). Both PL and beta 2GPI seem necessary for binding, thus indicating that the two components comprise the epitope against which aPL are directed. Using an anti-beta 2GPI antibody ELISA with the antigen adsorbed onto polyvinylchloride (PVC) plates, we detected high antibody titres in 12 out of 12 plasma from patients with the antiphospholipid syndrome. No or very low positivity was obtained when the same ELISA was carried out in polystyrene (PST) plates, while an increasing positivity was found when processed (i.e. more hydrophilic) or COOH-surface PST plates were used. When beta 2GPI dependent IgG-aPL were purified using agarose-immobilized cardiolipin, 4 out of 4 preparations were highly positive in anti-beta 2GPI antibody ELISA using PVC plates, while beta 2GPI was not fully recognized by aPL-IgG when adsorbed onto PST plates. These findings demonstrate that aPL are, in fact, anti-beta 2GPI antibodies directed against a cryptic epitope which is expressed when beta 2GPI is bound to anionic phospholipid, or another suitable surface.

In vivo platelet activity was studied in 58 patients with AMI on admittance to the Coronary Care Unit, in 48 of these patients after 1 week, in 30 after 1 month and in 24 patients after 6 months. Patients were carefully selected and excluded if they had associated diseases known to increase platelet activity. In vivo activation was studied by evaluating the plasma concentration of beta-thromboglobulin (beta-TG) and platelet factor 4 (PF4) in the same blood sample. On admittance (x = 58.5) and on day 7 (x = 52.9) beta-TG values were significantly higher than those obtained in the control group (x = 29). beta-TG values were moderately elevated after 1 month (x = 37.7) and then returned to values similar to those of the control group after 6 months (x = 27.9). The simultaneous assessment of PF4 shows a beta-TG/PF4 ratio indicative for in vitro release (less than or equal to 2.5) in many patients on days 1 and 7. Moreover, the beta-TG/PF4 ratio in patients with AMI tends towards 2 when beta-TG values are high. These results may indicate a greater tendency to an in vitro platelet release in the acute phase of AMI.

A brief contact between native whole blood and ADP promotes a dose-dependent release of platelet alpha-granules without a fall in the platelet number. We assessed the "ex vivo" effect of three widely used antiplatelet drugs, aspirin dipyridamole and ticlopidine, on this system. Aspirin (a single 800 mg dose) and dipyridamole (300 mg/die for four days) had no effect, while ticlopidine (500 mg/die for four days) significantly reduced the alpha-granules release for an ADP stimulation of 0.4 (p less than 0.02), 1.2 (p less than 0.01) and 2 microM (p less than 0.01). No drug, however, completely inhibits this early stage of platelet activation. The platelet release of alpha-granules may be related to platelet shape change of the light transmission aggregometer and may be important "in vivo" by enhancing platelet adhesiveness and by liberating the platelet-derived growth factor.