Michal Zabczyk

Asthma is associated with enhanced thrombin formation and impaired fibrinolysis

There is evidence that altered blood coagulation and fibrinolysis are involved in the pathogenesis of asthma. Increased thromboembolic risk has been reported in asthmatics.

Prothrombotic Fibrin Clot Phenotype Is Associated With Recurrent Pulmonary Embolism After Discontinuation of Anticoagulant Therapy.

Objective— Pulmonary embolism (PE) is a life-threatening manifestation of venous thromboembolism with a high recurrence rate after anticoagulation cessation. Recently, we have reported that prothrombotic clot phenotype in venous thromboembolism patients is associated with an increased risk of recurrent deep-vein thrombosis. Approach and Results— We tested whether abnormal clot properties are predictive of recurrent PE. We investigated 156 consecutive white patients aged 18 to 65 years after the first-ever provoked or unprovoked PE (n=89), with or without deep-vein thrombosis. Plasma fibrin clot permeability (Ks), turbidity measurements, calibrated automated thrombography, and efficiency of fibrinolysis using clot lysis time, maximum D-dimer levels, and rate of increase in D-dimer levels were evaluated at ≥3 months of anticoagulant therapy, at least 4 weeks since the anticoagulation withdrawal. The primary end point was recurrent PE during a median follow-up of 50 months. Recurrent PE was diagnosed in 23 (14.7%; 5%/yr) patients. Recurrent PE was associated with formation of denser fibrin networks reflected by lower Ks (P=0.007) and impaired fibrinolysis, as evidenced by prolonged clot lysis time (P=0.012) and reduced maximum rate of increase in D-dimer levels in the lysis assay (P=0.004). Patients with recurrent PE had higher plasma D-dimer (P<0.001) and thrombin peak (P=0.007) compared with the remainder, whereas turbidity measurements and maximum D-dimer levels did not differ in the recurrence. Multivariate model showed that independent predictors of recurrent PE were female sex, unprovoked venous thromboembolism, higher plasma D-dimer, reduced Ks, and reduced maximum rate of increase in D-dimer levels in the lysis assay (all P<0.05). Conclusions— Altered fibrin clot properties including formation of more compact clots displaying impaired susceptibility to lysis may predispose to recurrent PE.

Increased Plasma Clot Permeability and Susceptibility to Lysis Are Associated with Heavy Menstrual Bleeding of Unknown Cause: A Case-Control Study

Background Formation of compact and poorly lysable clots has been reported in thromboembolic disorders. Little is known about clot properties in bleeding disorders. Objectives We hypothesized that more permeable and lysis-sensitive fibrin clots can be detected in women with heavy menstrual bleeding (HMB). Methods We studied 52 women with HMB of unknown cause and 52 age-matched control women. Plasma clot permeability (Ks), turbidity and efficiency of fibrinolysis, together with coagulation factors, fibrinolysis proteins, and platelet aggregation were measured. Results Women with HMB formed looser plasma fibrin clots (+16% [95%CI 7–18%] Ks) that displayed lower maximum absorbancy (-7% [95%CI -9 – -1%] ΔAbsmax), and shorter clot lysis time (-17% [95%CI -23 – -11%] CLT). The HMB patients and controls did not differ with regard to coagulation factors, fibrinogen, von Willebrand antigen, thrombin generation markers and the proportion of subjects with defective platelet aggregation. The patients had lower platelet count (-12% [95%CI -19 – -2%]), tissue plasminogen activator antigen (-39% [95%CI -41 – -29%] tPA:Ag), and plasminogen activator inhibitor-1 antigen (-28% [95%CI -38 – -18%] PAI-1:Ag) compared with the controls. Multiple regression analysis upon adjustment for age, body mass index, glucose, and fibrinogen showed that decreased tPA:Ag and shortened CLT were the independent predictors of HMB. Conclusions Increased clot permeability and susceptibility to fibrinolysis are associated with HMB, suggesting that altered plasma fibrin clot properties might contribute to bleeding disorders of unknown origin.

Impaired fibrinolysis and lower levels of plasma α 2 -macroglobulin are associated with an increased risk of severe asthma exacerbations

Recently we have reported that asthma is associated with enhanced plasma thrombin formation, impaired fibrinolysis and platelet activation. In the present study we investigated whether described prothrombotic blood alterations might predispose to thromboembolic events or asthma exacerbations. In 164 adult asthmatics we assessed clinical events during 3-year follow-up and analyzed their associations with measured at baseline prothrombotic blood parameters. Data were obtained from 157 (95.7%) of the asthma patients. We documented 198 severe asthma exacerbations (64/year), which occurred in 53 subjects (34%). These patients were older (p = 0.004), had worse asthma control (p = 0.02) and lower spirometry values (p = 0.01), at baseline. Interestingly, this subgroup had longer clot lysis time (CLT), as well as lower α2-macroglobulin (p = 0.038 and p = 0.04, respectively, after adjustment for potential confounders). Increased CLT and lower α2-macroglobulin were demonstrated as independent predictors of asthma exacerbation in multiple regression model. Moreover, we documented two episodes of deep vein thrombosis (1.3%), and eight acute coronary syndromes (5.1%). Patients who experienced thromboembolic events (n = 10, 6.4%, 2.1%/year) had lower α2-macroglobulin (p = 0.04), without differences in efficiency of fibrinolysis and thrombin generation. Impaired fibrinolysis and lower levels of α2-macroglobulin might predispose to a higher rate of asthma exacerbations, suggesting new links between disturbed hemostasis and asthma.

Impaired plasminogen binding in patients with venous thromboembolism : association with protein carbonylation

INTRODUCTION Venous thromboembolism (VTE) is associated with hypofibrinolysis. Its mechanisms are poorly understood. We investigated plasminogen-fibrin interaction and its association with fibrinolytic capacity and protein oxidation/carbonylation in VTE patients. MATERIALS AND METHODS Plasma-purified plasminogen conversion to plasmin and surface plasmon resonance employed for plasminogen-fibrin interactions were individually evaluated in all healthy controls and non-anticoagulated patients following VTE, 10-23months since the event. We also assessed plasma fibrin clot permeability (Ks), clot lysis time (LT), activators and inhibitors of fibrinolysis together with oxidation/carbonylation markers. RESULTS VTE patients had impaired plasminogen binding to fibrin (apparent Kd, +290%, p=0.002), reduced rate of plasmin generation (-4.7%, p=0.001), and longer LT (+18.6%, p<0.001) compared with controls. Fibrinogen and Ks were similar in both groups. Apparent Kd correlated with LT (r=0.43, p=0.037), tissue plasminogen activator-plasminogen activator inhibitor 1 (tPA-PAI-1) complexes (r=0.63, p=0.012), and active PAI-1 (r=0.49, p=0.03). Compared with controls, VTE patients had higher thiobarbituric acid reactive substances (TBARS), total protein carbonyl content (PC), and lower total antioxidant capacity (all p<0.001), that all were associated with LT (r=0.61, r=0.56, and r=-0.47, respectively, all p<0.05). Impaired plasminogen binding to fibrin reflected by apparent Kd positively correlated with TBARS (r=0.48, p=0.032) and PC (r=0.54, p=0.013) in the whole group. CONCLUSIONS Plasminogen-fibrin interactions are altered in young and middle-aged VTE patients, without known thrombophilias, except increased factor VIII. The mechanisms underlying these phenomena remain to be established.

Reduced plasma fibrin clot permeability is associated with recurrent thromboembolic events in patients with antiphospholipid syndrome

Objectives APS is associated with arterial and venous thrombosis. The unfavourable fibrin clot phenotype, including formation of dense and poorly lysable clots, has been reported in thrombotic APS. We investigated whether abnormal plasma clot properties are predictive of recurrent thromboembolism in APS. Methods We followed 126 consecutive patients with thrombotic APS and 105 control subjects, without APS, matched for thrombotic events. Plasma fibrin clot permeability (Ks), turbidity measurements and clot lysis time were evaluated ⩾5 months after a thrombotic event. The primary composite end point was symptomatic recurrent venous thromboembolism, ischaemic stroke and/or myocardial infarction. Results During follow-up (median, 62 months; range 46-74 months; 1183.2 patient-years), the primary outcome was observed in 33 (26.2%) APS patients and 16 (15.2%) controls, including 25 (19.8%) and 14 (13.3%) subjects with recurrent venous thromboembolism, respectively. Reduced Ks and prolonged clot lysis time predicted recurrent thromboembolic events in APS patients [per 1 × 10-9 cm2: hazard ratio (HR) = 0.37; 95% CI: 0.24, 0.56; and per 10 min: HR = 1.20; 95% CI: 1.01, 1.40, respectively] and in controls (per 1×10-9 cm2: HR = 0.23; 95% CI: 0.11, 0.42; and per 10 min: HR = 1.51; 95% CI: 1.08, 2.16, respectively). A multivariate analysis showed that positive IgG and IgM anti-β2 glycoprotein I antibodies, withdrawal of anticoagulation, lower platelet count and reduced Ks predicted thromboembolic events in APS patients. Conclusion Formation of denser fibrin networks could be a novel risk factor for recurrent thromboembolism in APS, which highlights the importance of fibrin phenotype in thrombotic disorders.

Altered structure and function of fibrinogen after cleavage by Factor VII Activating Protease (FSAP)

Factor VII Activating Protease (FSAP) is a plasma protease affecting both coagulation and fibrinolysis. Although a role in hemostasis is still unclear, the identification of additional physiologic substrates will help to elucidate its role in this context. FSAP has been reported to cleave fibrinogen, but the functional consequences of this are not known. We have therefore undertaken this study to determine the implications of this cleavage for fibrin-clot formation and its lysis. Treatment of human fibrinogen with FSAP released an N-terminal peptide from the Bβ chain (Bβ1-53) and subsequently the fibrinopeptide B; within the Aα chain a partial truncation of the αC-region by multiple cleavages was seen. The truncated fibrinogen showed a delayed thrombin-catalyzed polymerization and formed fibrin clots of reduced turbidity, indicative of thinner fibrin fibers. Confocal laser scanning and scanning electron microscopy of these clots revealed a less coarse fibrin network with thinner fibers and a smaller pore size. A lower pore size was also seen in permeability studies. Unexpectedly, FSAP-treated fibrinogen or plasma exhibited a significantly faster tPA-driven lysis, which correlated exclusively with cleavage of fibrinogen and not with activation of plasminogen activators. Similar observations were also made in plasma after activation of endogenous zymogen FSAP, but not in plasma of carrier of the rare Marburg I single nucleotide polymorphism. In conclusion, altering fibrin clot properties by fibrinogenolysis is a novel function of FSAP in the vasculature, which facilitates clot lysis and may in vivo contribute to reduced fibrin deposition during thrombosis.