Altered clot formation and lysis are associated with increased fibrinolytic activity in ascites in patients with advanced cirrhosis.

Analysis of coagulation disorders and assessment of rebalanced hemostasis with the use of traditional coagulation assays is challenging in cirrhotic patients. Therefore, alternative tests are under investigation for the evaluation of coagulopathy in this specific setting. Aim of this study was to analyze the modifications of clot structure and function in cirrhotic patients with different degrees of severity. Cirrhotic patients referred to our Unit were consecutively enrolled. Global test measurements, including clot and lysis assays, clot lysis time, and determination of other fibrinolytic parameters, were performed. Analyses of clot formation, morphology, and lysis were performed with a turbidimetric clotting and lysis assay (EuroCLOT). Lysis of a tissue factor-induced clot by exogenous tissue plasminogen activator was analyzed by studying the modifications of turbidity during clot formation and the following lysis. We evaluated coagulative and fibrinolytic parameters in both plasma and ascites. Urokinase plasminogen activator (uPA) and gelatinase activity in ascites were also measured. We analyzed data from 33 cirrhotic patients (11 in Child-Pugh class A; 22 in class B or C and with ascites) and 21 healthy subjects (HS). In class B/C patients prolonged latency time, a decline in clotting absorbance, and decreased fibrin formation were observed in comparison with class A and HS. Generated curves and Thrombin-Activatable Fibrinolysis Inhibitor (TAFI) progressively declined from HS to class C patients, whereas levels of plasminogen activator inhibitor-1 and tissue plasminogen activator increased. D-dimer levels were markedly increased in ascites, together with significantly smaller levels of TAFI, αlfa2-antiplasmin, and plasminogen. Caseinolytic activity was also present. Class C patients showed smaller amount of uPA and significantly lower levels of matrix metallopeptidases (MMP)2 in ascites in comparison with Class B subjects. Clot formation and lysis are altered in cirrhosis and fibrinolysis is activated in ascites. Ascitic levels of uPA and MMP2 are reduced and inversely related to the severity of liver disease.

Residual perfusion defects in patients with pulmonary embolism are related to impaired fibrinolytic capacity.

BACKGROUND: Few studies investigated the relationship between fibrinolysis abnormalities and residual pulmonary perfusion defects after acute pulmonary embolism (PE). OBJECTIVE: To assess the fibrinolytic profile in patients with prior PE in relation to the extent of scintigraphically detectable residual perfusion abnormalities. PATIENTS AND METHODS: We studied 71 consecutive patients with a prior episode of PE, who were examined after one year of the incident embolic event, and at least one month after anticoagulation withdrawal. They underwent lung scintigraphy to assess the recovery of pulmonary perfusion, echocardiography and chest radiography to look for signs of pulmonary hypertension. Clot formation and lysis were evaluated by two turbidimetric methods: Clot and Lysis Assay and Clot Lysis Time. We also measured the in vitro plasmin-mediated lysis of fibrin from purified fibrinogen, and the circulating levels of fibrinolytic inhibitors. The sample was split in two categories based on the extent of residual perfusion defects: <10% (n=53), ≥ 10% (n=18). RESULTS: Patients with perfusion defects >10% had significantly longer lysis time (p<0.05), and higher levels of plasminogen activator inhibitor-1 (p<0.01) than those with perfusion defects <10%. The time interval between symptoms onset and PE diagnosis (time-to-diagnosis) was significantly longer in patients with perfusion defects >10% than in the others (p=0.005). In multivariate logistic regression, both lysis time and time-to-diagnosis were independently associated with perfusion defects >10% (p<0.001). None of the sampled patients had echocardiographic or radiologic signs of pulmonary hypertension. CONCLUSION: Prolonged time-to-diagnosis and fibrinolysis imbalance are independent predictors of incomplete perfusion recovery after acute PE.

Is tissue factor pathway inhibitor a marker of procoagulable status in healthy infertile women undergoing ovarian stimulation for assisted reproduction?

Increased serum estradiol levels occurred during ovarian stimulation for assisted reproduction. Tissue factor pathway inhibitor (TFPI) plays a relevant role in regulating haemostatic equilibrium, and its decrease has been documented in conditions in which blood coagulation occurs. We investigated TFPI concentrations and coagulative pathway in healthy infertile women undergoing ovarian stimulation. We investigated 27 healthy infertile women, median age 37 (25-41) years, undergoing ovarian stimulation, observed during the mid-luteal phase of cycle (T0) and on day 5 (T1), and between day 7 and 9 (T2) of ovarian stimulation. Coagulative pathway was assessed by a global test [endogenous thrombin potential, (ETP)] and TFPI concentrations. TFPI values progressively and significantly decreased throughout the ovarian stimulation procedure (P = 0.03), contemporarily estradiol levels progressively and significantly increased from baseline to T2 (P < 0.0001). A significant negative correlation between changes in estradiol and TFPI levels was observed (P = 0.03). As concerns ETP parameters a significant increase of ETP (mA) and Cmax (mA/min) throughout the ovarian stimulation cycle was found (P = 0.003 and P = 0.002, respectively). TFPI values progressively and significantly decreased throughout the ovarian stimulation, and negatively correlated with estradiol, thus suggesting that TFPI may represent one of the main 'actors' involved in the hypercoagulable status, occurring during assisted reproduction. The relationship between TFPI and estradiol levels might contribute to the knowledge of mechanisms able to modify a quite milieu into a prothrombotic status. Nevertheless, the small number of individuals investigated might influence the relevance of our results.

Hyperhomocysteinemia in patients with pulmonary embolism is associated with impaired plasma fibrinolytic capacity.

Hyperhomocysteinemia (HHcy) affects haemostasis and shifts its balance in favour of thrombosis. In vitro and in vivo studies suggested that HHcy may impair fibrinolysis either by influencing the plasma levels of fibrinolytic factors or by altering the fibrinogen structure. We investigated the influence of mild HHcy levels on plasma fibrinolytic potential by using clot lysis time (CLT) and fibrin susceptibility to plasmin-induced lysis in 94 patients with previous pulmonary embolism and no pulmonary hypertension. CLT was measured as lysis time of tissue factor induced clots exposed to exogenous tissue plasminogen activator (t-PA). The rate of in vitro plasmin-mediated cleavage of fibrin ß-chain was assessed over a 6-h period on fibrin clots, which were obtained by exposition to thrombin of purified fibrinogen. Homocysteine plasma levels were measured by Abbott Imx immunoassay and we considered as altered the values above 15 µmol/L according to the literature. In 68 patients homocysteine levels were below 15 µmol/L (NHcy) and in 26 they were above (HHcy). Significant differences were observed between the two groups regarding plasma fibrinolytic potential (p = 0.016), TAFIact (expressed as clot lysis ratio) (p = 0.02), t-PA (0.008) and PLG (0.037), but not for the other assessed components. The HHcy-patients had a threefold higher risk to have an impaired fibrinolysis. Instead, a multivariate logistic regression analysis adjusted for significances of univariate showed that HHcy (OR 5.2 95% CI 1.7-15.9; p = 0.003) and BMI (OR 5.0 95% CI 1.6-15.9; p = 0.006) resulted independently associated with impaired fibrinolytic activity. HHcy affects TAFI-mediated hypofibrinolysis but not fibrin(ogen) structure or function as documented by fibrin degradation analysis.

Fibrinolysis alterations in infertile women during controlled ovarian stimulation: influence of BMI and genetic components.

INTRODUCTION: Ovarian stimulation protocols have been described to induce prothrombotic phenotype through alterations of both coagulation and fibrinolysis pathways. We investigated fibrinolytic changes during ovarian stimulation through a global test (CLT) and PAI-1 and TAFI concentrations at different times of ovarian stimulation procedure, and the influence of polymorphisms in genes encoding for fibrinogen chains (FGA, FGB, FGG), t-PA (PLAT), TAFI (CBP2), FXIII (FXIIA1, FXIIIB), plasminogen (PLG) and PAI-1 (PAI1) on their intermediate phenotype. MATERIALS AND METHODS: We evaluated fibrinolytic and genetic parameters in 110 infertile women undergoing ovarian stimulation procedure (in vitro fertilization, IVF or intracytoplasmic sperm injection, ICSI). All women were observed during the mid-luteal phase of cycle (T(0)) and on day 5 (T(1)), 7 (T(2)) and 9 (T(3)) of the ovarian stimulation. RESULTS: Significant changes in fibrinolytic parameters from T(0) to T(3) of ovarian stimulation were found (CLT p=0.003; TAFI p=0.009 and PAI-1 p=0.003). CLT values, TAFI and PAI-1 concentrations significantly increased from baseline to T(1) (p<0.0001, p=0.01, p=0.005, respectively)(,) and decreased at T(2,) but remained higher than those at T(0). Moreover, at baseline overweight women showed longer CLT, higher TAFI and PAI-1 concentrations than normal weight women, as well as at T(1) two-fold longer CLT and higher PAI-1 concentrations were observed (p=0.001 and p=0.05, respectively). Significant differences of TAFI and PAI-1 concentrations during ovarian stimulation according to TAFI and PAI1 polymorphisms were observed. CONCLUSIONS: This study shows alterations of fibrinolysis and suggests the contribution of TAFI and PAI1 genes in modulating fibrinolysis changes during the ovarian stimulation cycle.

A hypercoagulable and hypofibrinolytic state is detectable by global methods in patients with retinal vein occlusion.

The pathogenesis of retinal vein occlusion (RVO), has not been well understood. Recent data have shown the efficacy of an anticoagulant therapy with LMWHs in the treatment of acute RVO suggesting the presence of a hypercoagulable state in these patients. New global tests for detection of hypercoagulability and hypofibrinolysis have become available and their application might improve the knowledge of the pathophysiology of RVO and, potentially, its treatment. The aim of our study was to evaluate coagulation and fibrinolytic alterations by two global tests in RVO patients: Endogenous Thrombin Potential (ETP) and Clot Lysis Time (CLT), respectively. We studied 81 RVO patients (40 males; median age 61 years) and a control group matched for age and sex. The ETP was measured by functional chromogenic assay and expressed as the time until thrombin burst (LagTime), Time to peak (T(max)), Peak amount of thrombin generation (C(max)) and ETP. CLT was determined by a plasma-based, tissue factor-induced clot lysis assay. C(max), ETP and CLT values were significantly higher in RVO patients than in controls (C(max)p = 0.010; ETP p < 0.001; CLT p < 0.001) and remained significantly associated with the disease at the multivariate analysis adjusted for cardiovascular risk factors. Our results indicate that -beyond the assay of different parameters associated with clotting activation and lysis- global methods might allow us to easily detect the presence of hypercoagulability and hypofibrinolysis in RVO patients. Further studies should assess the possible clinical value of our data in the management of RVO patients.

Assessment of fibrinolytic activity by measuring the lysis time of a tissue-factor-induced clot: a feasibility evaluation.

A clot lysis time assay in which a tissue factor-induced fibrin clot is lysed by exogenously added tissue plasminogen activator has been recently reported. We evaluated the feasibility of clot lysis time in a routine hemostasis laboratory, and its correlation with thrombin activatable fibrinolysis inhibitor and plasminogen activator inhibitor-1 levels and changes with aging in 185 healthy participants. Clot lysis time was assessed by monitoring changes in turbidity during clot formation and subsequent lysis using a computerized kinetic spectrophotometric microtiter plate. After preliminary experiments, 100 and 160 ng/mL tissue plasminogen activator concentrations were chosen for the study. Clot lysis time was calculated by a new mathematical analysis of the lysis curve based on discrete derivative. Clot lysis time, thrombin activatable fibrinolysis inhibitor, and plasminogen activator inhibitor-1 plasma levels showed a normal distribution. For both concentrations of tissue plasminogen activator, clot lysis time progressively increased with increase in age (P < .0001) and was significantly correlated with thrombin activatable fibrinolysis inhibitor antigen, thrombin activatable fibrinolysis inhibitor activity, and plasminogen activator inhibitor-1 antigen (at least P < .01). During linear regression analysis, thrombin activatable fibrinolysis inhibitor and plasminogen activator inhibitor-1 antigen were found to significantly influence clot lysis time (at least P < .01). Clot lysis time determination has a good laboratory performance. Our new method of calculation is independent of the time of reading and allows a more accurate and consistent detection of both short and prolonged lysis times. Our data suggest the feasibility of the use of this test in the work of routine hemostasis laboratory.