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Background:  Women of childbearing age are exposed to venous thromboembolic risk mainly for pregnancy and use of oral contraceptives. The impact of risk factors (RF) on venous thromboembolism (VTE) in these circumstances is still unclear., Aim:  In the context of START registry, we aimed to investigate the weight of a series of RF on the occurrence of pregnancy- or combined oral contraceptive (COC)-associated VTE., Materials and Methods:  We selected all women included in the START for VTE occurred between 18-42 years and compared those with a first or recurrent pregnancy/postpartum- (group A) or COC-VTE (group B) with those who had VTE outside these circumstances (group C). Final analysis included a cohort of 532 women. Follow-up data showed that there were no significant differences between the groups in terms of thrombotic and haemorrhagic complications. As for pregnancy-associated VTE, the overall outcome was good in terms of both maternal and fetal prognosis., Results:  In a binary model of logistic regression, correcting for potential confounders, VTE family history conferred a significant and independent higher risk of COC-VTE compared with group C. Similarly, comparison between group A and C documented that family history significantly affected the risk of pregnancy-associated VTE. VTE in the group C was significantly associated with older age. Lastly, smoke was a significant risk factor for pregnancy/postpartum VTE when group A and group B were compared., Conclusion:  Present data suggest that in the setting of fertile women, family history of VTE has a greater role in predicting COC- and pregnancy/postpartum- VTE than outside these circumstances., Competing Interests: None declared., (Thieme. All rights reserved.)

Background  Isolated distal deep vein thromboses (IDDVT) are frequently diagnosed; however, their natural history and real risk of complications are still uncertain. Though treatment is still not well standardized, international guidelines recommend no more than 3 months of anticoagulation therapy. We investigated how Italian clinicians treat IDDVT patients in their real life in our country. Methods  Baseline characteristics and clinical history of the patients enrolled in the prospective, observational, multicenter START-Register for a first IDDVT or proximal DVT (PDVT) were analyzed. Results  Overall, 412 IDDVT patients were significantly younger, with better renal function, and more frequent major transient risk factors, when compared with 1,173 PDVT patients. The anticoagulation duration was >180 days in 52.7% of IDDVT patients (70.7% in PDVT). During treatment, bleeding occurred in 5.6 and 2.8% patient-years in IDDVT and PDVT, respectively ( p  = 0082). Bleeding was more frequent in IDDVT than PDVT patients treated with warfarin (6.8 vs. 3.2 patient-years, p  = 0.0228, respectively). Thrombotic complications occurred in 1.1 and 2.4% patient-years in IDDVT and PDVT patients, respectively. Analyzing together the two groups, 66.1% of bleeds and 86.1% thrombotic complications occurred after 90 days anticoagulation treatment. Conclusion  The large majority of IDDVT patients received anticoagulation for more than 3 months. Most bleeding and thrombotic complications occurred after the first 90 days of anticoagulation therapy. These results indicate that an extended anticoagulation beyond 90 days in IDDVT patients is associated with increased risk of complications. Whether an extended treatment may lower recurrences after anticoagulation withdrawal should be assessed by specifically designed studies., Competing Interests: Conflict of Interest The authors declare no potential conflicts of interest with respect to the research, authorship, and/or publication of this article. G.P. reports consulting fees from Alfasigma SpA., (The Author(s). This is an open access article published by Thieme under the terms of the Creative Commons Attribution License, permitting unrestricted use, distribution, and reproduction so long as the original work is properly cited. ( https://creativecommons.org/licenses/by/4.0/ ).)

Purpose  The purpose of this study was to retrospectively evaluate the clinical outcome of revision anterior cruciate ligament (ACL) reconstruction with contralateral hamstring tendon autografts, specifically with regard to patient satisfaction, return to preinjury activity level, and postoperative functional outcomes. Methods  Between 2004 and 2011, 23 patients underwent revision ACL reconstruction with contralateral autogenous hamstring tendon grafts and were retrospectively reviewed at an average follow-up of 6.3 years. Subjective and functional evaluations were performed. The Tegner score, Knee Injury and Osteoarthritis Outcome Score (KOOS), and International Knee Documentation Committee (IKDC) Subjective Knee Form were used. Objective evaluation included range of motion, Lachman test, pivot-shift test, and KT-1000 instrumented laxity testing. Wilcoxon test was used to compare the preoperative and follow-up status. Differences with a p -value of <0.05 were considered statistically significant. Results  No major complications were reported. The mean KOOS significantly increased from a preoperative mean of 62.8 ± 8.3 to 85.8 ± 6.9 ( p < 0.001). IKDC subjective score significantly improved from 29.2 ± 10.4 to 72.8 ± 5.2 ( p < 0.001). The median Tegner activity score significantly improved from a preoperative mean of 6.5 (range: 4-10) to 7.5 (range: 7-10) ( p  < 0.001). Most of the patients increased or returned to the same activity level, with 61% of the patients returning to cutting and pivoting sports. Conclusion  The use of contralateral hamstring tendon autografts for ACL revision surgery represents a valid option following a failed primary ACL reconstruction and confirms subjective and objective clinical improvement 6 years after surgery. Level of Evidence  Level IV, therapeutic case series.

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.

Whether 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.

D-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 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.

Heparin-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 HIT-Ab(PF4-H) assay (sensitivity and negative predictive values = 100%; negative likelihood ratios <0.01). The specificity was higher for the HemosIL AcuStar HIT-IgG(PF4-H) in comparison with that of the HemosIL AcuStar HIT-Ab(PF4-H) (96.5% vs. 81.2%). Higher values of the HemosIL AcuStar HIT-IgG(PF4-H) were associated with increased probability of HIT. Patients with confirmed HIT and thrombotic complications had significantly higher levels of HemosIL AcuStar HIT-IgG(PF4-H) than those without thrombotic complications. The HemosIL AcuStar HIT-IgG(PF4-H) and HIT-Ab(PF4-H) assays showed a very high sensitivity, and therefore they can reliably be used to rule out HIT in suspected patients. The diagnostic specificity was greatly increased by using the HemosIL AcuStar HIT-IgG(PF4-H). Both the assays are reproducible (CVs <6%), rapid (turnaround time 30 minutes), automated, and semi-quantitative, and they can be run for single sample testing.

Few 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<0.001, respectively). The prevalence of FII 20210A allele was higher in patients with CLI vs. those with claudication and controls. Using a logistic model, FII G20210A mutation (odds ratio [OR] 19.8, confidence interval [CI] 4.5-87.1, p=0.001), LAC (OR 2.7, CI1.1-6.5, p=0.032), and fibrinogen (OR 1.01, CI 1.00-1.01, p=0.001) were associated with CLI, whereas homocysteine, FVIII, and FV R506Q mutation were not. CLI risk increased according to the number of thrombophilic alterations. In conclusion, altered levels of some important thrombophilic risk factors are independently associated with PAD severity. These data suggest that the presence of two or more thrombophilic risk factors raise the likelihood of PAD being more severe, justifying the need for larger longitudinal studies.

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.

D-dimer testing is widely applied for exclusion of deep-vein thrombosis (DVT) and pulmonary embolism (PE). We report on a multicenter performance evaluation of a new particle-enhanced immunoassay, Innovance D-Dimer. Innovance D-Dimer assay was performed in 1,543 frozen samples from outpatients suspected of DVT and/or PE enrolled in three management studies as well as in a routine clinical practice. Samples were assayed on BCS/BCS XP, BCT as well as Sysmex CA-7000, CA-1500 and CA-560 analyzers (cut-off on all analyzers: 0.5 mg/l). Stratus CS D-Dimer and Vidas D-Dimer Exclusion were used for comparison. The precision study indicated total coefficients of variation ranging from 2.1% to 8.4% depending on the analyzer and on the sample. Sensitivity and negative predictive values were above 99% and their lower 95% confidence interval were equal or above 97.4% and 98.6%, respectively. Specificity ranged from 38.2% to 40.4% and the respective lower 95% confidence intervals from 35.5% to 37.7%. Area under the curve was 0.90 for all assay systems except for Innovance D-Dimer with BCT (0.89). Two samples from patients with distal DVT tested negative with all assay systems. One patient with high pre-test clinical probability and proximal DVT tested negative with Vidas D-Dimer Exclusion. Our data indicate that the performances of Innovance D-Dimer, regardless of the analyzer, are similar to the reference methods, and that this assay can be used for the exclusion of venous thromboembolic disease.

Deep vein thrombosis (DVT) requires a prompt and accurate diagnosis to avoid the potentially fatal consequences of a delay in adequate treatment. The diagnostic strategy of suspected venous thromboembolism is an example of the application of Bayes' theorem. As a result, the available diagnostic tools, both noninvasive (such as ultrasonography, D-dimer, and computed tomography) and invasive (such as venography), are usually integrated in diagnostic strategies that require the evaluation of the a priori clinical probability of disease (pretest probability). These strategies also vary according to the patient characteristics that allow the definition of five different categories: symptomatic outpatients, symptomatic inpatients, high-risk asymptomatic subjects, patients with suspected recurrence, and pregnant women. Some areas are still a matter of debate, such as the clinical relevance of isolated distal DVT and the diagnosis of suspected ipsilateral recurrence.

We investigated the psychological impact of testing for the presence of thrombophilic alterations. The enrolled subjects received counseling from a physician before blood sampling and after receiving results, with a view to provide clear information about the aim of thrombophilia screening (TS). Participants were requested to complete a pre-test questionnaire during this interview and a post-test questionnaire 20 days after receiving the TS results. One hundred ninety-seven subjects completed the pretest questionnaire and 140/197 (71.1%) returned the post-test one. The TS results were altered in 36 (25.7%, R506Q mutation n = 19; G20210A mutation n = 9; antithrombin deficiency n = 1; LAC phenomenon n = 4; hyperhomocysteinemia n = 3) and normal in 104 subjects. We assessed: perceived health status (PHS), state of anxiety, health fears, depressive reactions, moods, perceived well-being, and perceived daily-life stress. For both groups, both at pre- and post-test, none of the psychological variable scores showed significant worsening, regardless of whether TS resulted altered or normal. Anxiety significantly (p < or = 0.05) decreased at post-test in the altered group and a non-significant improvement in PHS after TS result communication was recorded in both groups. Age was an important factor in mediating psychological impact. In conclusion, diagnosis of thrombophilic alterations seems to be well accepted in the short term and TS should not be discouraged for potential adverse psychological effects. However, the psychological impact over a longer period of receiving altered results needs to be further investigated. The relationship between absence of adverse psychological reactions and quality of counseling program provided before and after TS should also be investigated.

We 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.

Elevated factor VIII (FVIII) levels are a recognized risk factor for venous thrombosis. Recently, family studies suggested that the G allele of the 3951C/G (D1241E) FVIII polymorphism is associated to lower FVIII activity. We investigated in case-control studies both biological effects (FVIII levels and activated protein C sensitivity ratio) and clinical associations (venous thromboembolism) of the D1241E change. Among 145 healthy and 150 thrombotic women, not carriers of known thrombophilic defects, the 1241E allele was associated with 11% reduced (t-test, P<0.05) FVIII levels. The effect on activated protein C sensitivity ratio was not statistically significant. Carriership of the 1241E allele, potentially conferring protection from thrombosis, was found in 22.8% of controls and in 15.3% of cases. In an additional cohort of factor V Leiden carriers (n=283), carriership of the 1241E allele was 25.2% among 143 asymptomatic subjects and 17.1% among 140 thrombotic patients. Our data do not indicate a specific interaction with factor V Leiden. These genotype distributions suggest a mild protective effect from venous thrombosis conferred by 1241E FVIII, masked by other genetic and/or environmental components, and detectable only in very large population studies. Our findings point toward the presence of genetic determinant of coagulation factor levels with a biologically significant role, but with a poor predictive value to estimate thrombotic risk beyond established risk factors.

The factor V Leiden mutation (FVL), associated with reduced sensitivity to activated Protein C (APC), is a risk factor for venous thromboembolism (VTE) and displays a strong interaction with oral contraceptives (OC). The aim of this study was to evaluate the risk of VTE in OC users with reduced APC sensitivity unrelated to the FVL. APC sensitivity was measured by an original aPTT-based test (without sample pre-dilution in factor V-deficient plasma) in 195 women who suffered from VTE in reproductive age and in 487 healthy women with results being expressed as normalized ratio. Subjects with currently known clinically relevant thrombophilic alterations were excluded. APC normalized ratios were stratified into quartiles. The adjusted ORs of subjects in the lower quartile (>/= 0.90) was 2.46 (95%CI: 1.02-5.95). Of the 195 patients, 89 had suffered VTE during OC. The 181 healthy women who had used OC for at least 6 months in the two year period before presentation but who had stopped OC at least 3 months before blood sampling were considered OC users. The risk of VTE in subjects using OC with APC normalized ratio in the lower quartile was increased 4.9-fold (95% CI: 1.92-12.6). In conclusion, our results showed that altered APC resistance in women not carrying the FVL significantly increased the VTE risk, albeit to a lesser extent than in women also carrying the FVL. Our data also showed that OC use in women with altered APC resistance further increased the risk of VTE in a way that exceeded the additive expectation.

Uncontrolled activation of matrix metalloproteinases (MMPs) can result in tissue injury and inflammation, yet little is known about the activation of MMPs during orthotopic liver transplantation (OLT). OLT is associated with increased fibrinolytic activity due to elevated plasmin generation. The serine-protease plasmin not only causes degradation of fibrin clots but is also thought, amongst others, to play a role in the activation of some matrix metalloproteinases. We therefore studied the evolution of MMP-2 and -9 plasma concentrations during OLT and the effect of serine-protease inhibition by aprotinin on the level and activation of these MMPs. In a group of 24 patients who participated in a randomized, double-blind, placebo-controlled study we determined serial MMP-2 and MMP-9 plasma levels during transplantation using ELISA (total MMP), activity assays (activatable MMP) and zymography. In addition, the MMP-inhibitors TIMP-1 and TIMP-2 were assessed by ELISA. The putative regulating factors tumor necrosis factor alpha (TNF-alpha) and tissue-type plasminogen activator (t-PA) were assessed as well. Patients were administered high-dose aprotinin, regular-dose aprotinin or placebo during surgery. Plasma TIMP-1, TIMP-2 and MMP-2 level gradually decreased during transplantation. Approximately two-thirds of total MMP-2 appeared to be in its activatable proMMP form. No release of MMP-2 from the graft could be detected. In contrast, plasma levels of MMP-9 increased sharply during the anhepatic and postreperfusion periods. Peak MMP-9 levels of about eight times above baseline were found at 30 minutes after reperfusion. Most MMP-9 appeared to be in its active/inhibitor-complexed form. No significant differences were observed between the three treatment groups. However, in patients with more severe ischemia/reperfusion (I/R) injury the MMP-9 concentration, particularly of the active/inhibitor-complexed form, remained high at 120 minutes postreperfusion compared to patients with no or mild I/R injury. The decrease in plasma levels of MMP-2, TIMP-1 and TIMP-2 during OLT occurred irrespective of the severity of the I/R injury. There was a significant correlation between MMP-9 and t-PA levels, but not with TNF-alpha. In conclusion, OLT is associated with a sharp increase of MMP-9 during the anhepatic and postreperfusion periods, which coincided with the changes in t-PA. MMP-2, TIMP-1 and TIMP-2 gradually decreased during OLT. The composition of these MMPs was not altered by the use of aprotinin, suggesting that serine-protease/plasmin-independent pathways are responsible for MMP regulation during OLT. In addition, only MMP-9 seems to be involved in I/R injury during human liver transplantation.

Activated protein C resistance (APCR) is a major risk factor for venous thromboembolism (VTE). Although the factor V (FV) Leiden mutation accounts for the vast majority of APCR cases, other polymorphisms may contribute to the APCR phenotype. Genetic components of APCR and thrombophilia were investigated by two dinucleotide repeats, characterized in introns 2 and 11 of the FV gene. Only the intron 11 marker was genetically stable and thus suitable for further analysis. Its allelic frequencies were found to differ significantly (P=0.003) between subjects selected for increased APCR in the absence of the FV R506Q mutation (n=70, normalized ratios /=1.31). Genotype differences were also found (P=0.017) between FV R506Q heterozygotes (n=100) who had experienced previous VTE and those (n=100), who were still asymptomatic for VTE. Significance was mostly driven by the relative over-representation of the 12R allele and to a minor extent by the under-representation of the 15R allele among the symptomatic versus the asymptomatic FV Leiden carriers. Two SNPs (4070A/G and 2391A/G) were found to underlie the 12R and 15R alleles respectively, and marked extended haplo-types, previously (HR2) or newly (HT2) identified. Only the FV HR2 differed (P=0.002) in frequency between the two groups of FV R506Q heterozygotes, suggesting that it represents the most relevant FV genetic component of APCR or VTE detectable by this experimental and clinical approach. Our analysis indicates that frequent FV genetic components might contribute to shape the risk for VTE in FV Leiden carriers.

In some patients with previous venous thromboembolism (VTE) D-dimer levels (D-Dimer) tend to increase after oral anticoagulant therapy (OAT) is stopped. The aim of our study was to evaluate the predictive value of D-Dimer for the risk of VTE recurrence after OAT withdrawal. After a first episode of deep vein thrombosis (DVT) of the lower limbs and/or pulmonary embolism (PE), 396 patients (median age 67 years, 198 males) were followed from the day of OAT discontinuation for 21 months. D-dimer was measured on the day of OAT withdrawal (T1), 3-4 weeks (T2) and 3 months (+/- 10 days, T3) thereafter. The main outcome events of the study were: objectively documented recurrent DVT and/or PE. D-dimer was found to be increased in 15.5%, 40.3% and 46.2% of the patients at T1, T2 and T3, respectively. In 199 (50.2%) patients, D-dimer levels were elevated in at least one measurement. During a follow-up of 628.4 years, 40 recurrences were recorded (10.1% of patients; 6.4% patient-years of follow-up). D-dimer was increased in at least one measurement in 28 of these cases, but remained normal in 11 subjects (three of whom had recurrent events triggered by circumstantial factors, three with malignancy-associated factors) (in one subject D-dimer was not measured). The negative predictive value (NPV) of D-dimer was 95.6% (95% CI 91.6-98.1) at T3 and was even higher (96.7%; 95% CI 92.9-98.8) after exclusion of the six recurrences due to circumstantial factors. Only five idiopathic recurrences occurred in the 186 patients with consistently normal D-dimer. In conclusion, D-dimer has a high NPV for VTE recurrence when performed after OAT discontinuation.

Cytomegalovirus encephalitis is rarely diagnosed in immunocompetent patients. It is more frequent among immunocompromised hosts. We present a case of a young male patient without evidence of immunosupression who had a cytomegalovirus encephalitis. The diagnosis was made by detection of viral DNA in cerebral tissue obtained by biopsy. Neuropathologic and ultrastructural changes are described and compared with the descriptions in immunocompromised patients.

Homozygous carriers of factor V Leiden have an approximately 80-fold increased risk of venous thrombosis. Also double heterozygous carriers of both the factor V Leiden and the prothrombin gene mutations are at high thrombotic risk. The magnitude of the risk of venous thrombosis in pregnant women with the two severe thrombophilic conditions has not been estimated so far. We performed a multicenter retrospective family study in women with homozygous factor V Leiden, double heterozygous factor V Leiden and the prothrombin gene mutation, and women with normal coagulation. Only relatives of index patients with thrombosis formed the study cohort. Fifteen homozygous and 39 double heterozygous women were compared to 182 women with normal coagulation. Venous thrombosis occurred in 3 of 19, 2 of 50 and 1 of 221 pregnancies, respectively. One thrombotic episode occurred in the third trimester, the remaining 5 in the postpartum. The prevalence of venous thrombosis was 15.8% (95% CI 3.4-39.6) for homozygotes. 4.0% (95% CI 0.5-13.7) for double heterozygotes and 0.5% for women with normal coagulation. The relative risk of pregnancy-related venous thrombosis was 41.3 (95% CI 4.1-419.7) for homozygous and 9.2 (95% CI 0.8-103.2) for double heterozygous carriers. In conclusion, homozygous carriers of factor V Leiden and, to a lesser extent, double heterozygous carriers of factor V Leiden and of the prothrombin mutation have an increased risk of venous thrombosis during pregnancy, particularly high during the postpartum period. On the basis of these findings we recommend that these women receive anticoagulant prophylaxis at least in the postpartum, that should perhaps be extended to the whole pregnancy in homozygous carriers.

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.

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 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.

During orthotopic liver transplantation (OLT) excessive bleeding is the main cause of death and graft failure. The acute bleeding tendency that accompanies OLT, particularly during the anhepatic period and after reperfusion of the graft, is due to the depletion or functional abnormalities of several hemostasis components caused by the enhanced activity of enzymes such as plasmin, trypsin and leukocyte proteases. We surmised that enhanced proteolysis might also cause abnormalities of von Willebrand factor (vWF), and that these abnormalities are implicated in the bleeding tendency that develops during OLT. Therefore, the pattern of vWF proteolysis was studied with 16 patients with chronic liver disease, in serial blood samples obtained before OLT, during the anhepatic stage, after graft reperfusion and at the end of the surgical procedure. vWF became markedly degraded during the anhepatic and reperfusion stages, as shown by the partial loss of high molecular weight multimers, the relative decrease of the intact 225 kD subunit and the increase of the native proteolytic fragments of 176 and 140 kD. Novel proteolytic fragments also became detectable. Using monoclonal antibody epitope mapping, it could be demonstrated that some of the proteolytic fragments corresponded in apparent molecular mass to those produced in vitro by incubating purified vWF with plasmin or elastase, but other fragments could not be attributed to these proteases. During the anhepatic and reperfusion stages there was a significant correlation between the degree of vWF degradation and the total amount of blood components transfused to replace blood losses. To evaluate whether or not vWF degradation could be controlled by the administration of a broad-spectrum protease inhibitor such as aprotinin, 5 patients were given a bolus dose of 500,000 U before surgery followed by 100,000 U/h during surgery, 5 were given a 2,000,000 U bolus followed by 500,000 U/h, and no aprotinin was given to the remaining 6 patients. There were no differences in the patterns or degrees of vWF degradation between patients treated with aprotinin or not. In conclusion, there is a marked degradation of a key hemostasis protein during OLT. These alterations may be of clinical significance, because they are correlated with the transfusion requirements.

305 patients with juvenile thromboembolic episodes were screened for the presence of heparin cofactor II deficiency. The heterozygous deletion of two bases was found in the exon 5 of the heparin cofactor II gene in two unrelated patients, very likely due to a founder effect. This molecular lesion, causing a frameshift and elongated translation, affects the core of the molecule and should cause the complete unfolding of the protein, which is in accordance with the observed type I deficiency. The corresponding region of antithrombin III gene is affected by a cluster of frameshift mutations suggesting that heparin cofactor II and antithrombin III could share similar mutational patterns. The heparin cofactor II gene alteration was associated with, in one patient, the factor V Leiden mutation and, in the other, type I protein C deficiency. The tracing of the single defects in several family members indicated that the mutations became clinically manifest only when present in the doubly heterozygous condition. This study provides two examples, based on molecular findings, of the interplay of risk factors which is potentially useful to define a role for heparin cofactor II deficiency in inherited thrombophilia.

The occurrence of a "rebound hypercoagulable state" in patients after dicontinuation 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: 2/3 of initial dose the 1st week, 1/3 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 < 0.001); at the 2nd week F1+2 levels remained higher (p < 0.05) though INR values were not different from those of group B. After baseline, higher than normal F1+2 levels were recorded in 32/66 (48%) controls in group A vs 15/60 (25%) in group B (p < 0.01); at the 2nd week, 10/17 (59%) patients in group A vs 1/15 (7%) in group B still had higher than normal F1+2 levels (p < 0.01). The values of areas under curve (AUC) and maximum concentrations of all variables were not statistically different in the two groups; however, very high levels were observed in a few cases of group A. Thrombotic events (one DVT recurrence and one thrombophlebitis in a varicose vein) occurred in 2 pts of group A, both with high F1+2 and TAT AUC values.(ABSTRACT TRUNCATED AT 250 WORDS)

Twenty-one thyroprival patients, previously submitted to total thyroidectomy for tumours, were investigated during stabilized L-thyroxine supplementation and at the end of a 20-25 day "no-therapy" period, necessary for a 131I total body scintigraphy. During supplementation therapy a lower than normal mean beta-thromboglobulin (beta-TG) release level was found, the other blood clotting and platelet function tests being normal. After substitution therapy withdrawal, platelet function tests showed reduced adrenalin aggregation, increased collagen threshold aggregating concentrations, decreased beta-TG release values and reduced aggregation to ristocetin, whereas blood clotting tests showed prolonged aPTT values and reduced levels of factor VIII:C and vWf:Ag. We conclude that in acquired hypothyroidism the lowering of factor VIII:C and vWf:Ag (acquired von Willebrand disease) is associated with impaired platelet reactivity not only to ristocetin but also to collagen and especially adrenalin. In the patients investigated these changes were almost completely corrected by substitutive therapy with L-thyroxine at clinically effective doses.