Longer Clot Lysis Time Research Articles

Elevated carbonylated proteins are associated with major cardiovascular events in patients with chronic coronary syndrome: A cohort study.

Protein carbonylation is reported in atherosclerosis, but its predictive value is unknown. We evaluated plasma carbonylated protein (PC) levels in association with clinical outcomes in coronary artery disease (CAD) in long-term follow-up. In patients with advanced stable CAD, we assessed plasma PC content along with fibrin clot properties, i.e., permeability (Ks) and clot lysis time, and its determinants: plasminogen activator inhibitor-1 (PAI-1) and thrombin-activatable fibrinolysis inhibitor. We recorded a composite of myocardial infarction, ischemic stroke, systemic embolism, and cardiovascular death during a follow-up of 8.3 (1.8) years. The analysis involved 178 patients aged 64.0 (57.0-70.0) years. The baseline PC content was 2.9 (2.2-3.7) nmol/mg protein and was elevated above the reference value obtained for a control group (2.03 nmol/mg protein) in 82.6% of patients. In linear regression models, high PC adjusted for age was associated with lower Ks, longer clot lysis time, and elevated PAI-1 and thrombin-activatable fibrinolysis inhibitor. Baseline PC was 48% higher in patients with the composite endpoint (n = 67, 37.6%) compared with others (P <0.001). Patients with PC in the highest quartile (3.7-5.1 nmol/mg protein) were more likely to develop the composite endpoint compared to the lowest quartile (hazard ratio [HR] 4.9; 95% confidence interval, 2.1-11.3; P <0.001). This is the first study showing that in CAD the extent of protein carbonylation, in part via its antifibrinolytic effects, predisposes to cardiovascular events in long-term follow-up, highlighting the role of persistent oxidative protein modifications in atherosclerotic vascular disease.

Formation of denser plasma fibrin clots is associated with coronary artery ectasias and aneurysms - preliminary results from the CARE-ANEURYSM study

Abstract Background Coronary artery ectasias and aneurysms (CAEAs) are defined as diffused or localized coronary artery dilations, that exceed diameter of the adjacent segment by at least 50%. In the Caucasian population, CAEAs are present in 2-5% of all-comer patients undergoing coronary angiography and are associated with increased risk of myocardial infarction and death. Purpose To determine whether CAEAs are associated with increased thrombin generation potential and unfavorably modified plasma fibrin clot properties. Methods In this prospective study 19 patients with CAEAs, 19 sex-, age-, fibrinogen- and diabetes mellitus-matched patients with coronary artery disease (CAD) but without CAEAs, and 20 healthy control subjects were assessed. Plasma fibrin clot permeability (Ks) reflecting an average pore size within fibrin network, plasma fibrinolytic potential expressed as clot lysis time (CLT), and thrombin generation capacity defined as endogenous thrombin potential (ETP) were assessed. The CARE-ANEURYSM study, founded by the Polish National Science Centre (no. UMO-2020/39/O/NZ5/02863), was approved by the Ethical Committee and conducted in accordance with the Declaration of Helsinki. Patients provided written informed consent. Results As many as 84.2% of CAEA patients were men, mean age was 69.6 years, and 73.7% individuals had CAD. On average, CAEA affected 3 coronary segments and exceeded vessels referral diameter by 2.15-fold. Patients with CAEA had 46.5% reduced Ks (median, 3.98 [3.30-4.63] vs 7.43 [6.15-8.88] x10-9cm2, p&amp;lt;0.001), a tendency to prolonged CLT (112 [103-116.5] vs 99 [89-114] min, p=0.07), and 51% higher ETP (1685 [1502-1786] vs 1119 [1025-1189] nM×min, p&amp;lt;0.001) compared to healthy control (Fig. 1A-C). Moreover, individuals with CAEA compared to CAD patients had 19.5% lower Ks (median 3.98 [3.30-4.63] vs 5.33 [4.86-6.78] x10-9cm2, p=0.016) and 21% longer CLT (112 [103-116.50] vs 92.52 [81.50-97.50] min, p=0.015) and (1685 [1502-1786] vs 1119 [1693-1966] nM×min, p&amp;lt;0.001) (Fig. 1A-C). In the CAEA group, Ks was associated with the CAEA maximal diameter (R=-0.595, p=0.007) and the ratio of CAEA maximal diameter to referral vessel diameter (R=0.743, p&amp;lt;0.001; Figu.1D). There were no associations between the number of coronary artery segments affected by the lesion and fibrin clot properties or ETP. However, CAEA patients in whom the ratio of the CAEA maximal diameter to the referral vessel diameter exceed 2 (n=9) had 24.5% lower Ks (3.45 [1.88-3.68] vs 4.57 [4.08-5.35] x10-9cm2, p=0.004) and 14.7% higher ETP (1779 [1685-1812] vs 1515 [1431-1674] nM×min, p=0.028) with no difference in CLT (p&amp;gt;0.05). Conclusion CAEAs are associated with denser fibrin clot structure and enhanced thrombin generation, which indicate that CAEA patients are characterized by prothrombotic state. Further studies are needed to elucidate whether prothrombotic clot phenotype may be associated with increased risk of thromboembolic events in CAEA patients.Figure 1.

Association of the metabolic syndrome with PAI 1act and clot lysis time over a 10-year follow up in an African population

Background and aimsThe association between the metabolic syndrome (MetS) and plasminogen activator inhibitor-1 (PAI-1) has been well established in cross-sectional studies. It is less clear whether this translates into decreased clot lysis rates and very little information is available on non-European populations. Little is known regarding prospective associations and whether clot lysis progressively worsens in MetS individuals over time. We determined the prospective association of MetS with PAI-1 activity (PAI-1act) and clot lysis time (CLT) over a 10-year period. Methods and resultsAs many as 2010 African men and women aged ≥30 years were stratified according to MetS status and number of MetS criteria (0–5). We also determined the contribution of the PAI-1 4G/5G polymorphism to these associations and identified which MetS criteria had the strongest associations with PAI-1act and CLT. Both PAI-1act and CLT remained consistently elevated in individuals with MetS throughout the 10-year period. PAI-1act and CLT did not increase more over time in MetS individuals than in controls. The 4G/5G genotype did not influence the association of PAI-1act or clot lysis with MetS. Increased waist circumference, increased triglycerides and decreased HDL-C were the main predictors of PAI-1act and CLT. ConclusionsBlack South Africans with MetS had increased PAI-1act and longer CLTs than individuals without MetS. The inhibited clot lysis in MetS did, however, not deteriorate over time compared to controls. Of the MetS criteria, obesity and altered lipids were the main predictors of PAI-1act and CLT and are thus potential targets for prevention strategies to decrease thrombotic risk.

An association of low-density lipoproteins and fibrinolysis in patients with severe aortic stenosis- valvular expression of fibrinolytic proteins

Abstract Background Aortic stenosis (AS) is associated with hyperlipidemia and hypofibrinolysis. The contribution of serum lipids to the valvular expression of fibrinolytic proteins is not known. Aims To evaluate whether fibrinolytic factors are present within stenotic aortic valves and to determine their potential associations with LDL cholesterol concentrations, their oxidized forms (oxPL-LDL), and AS severity. Methods We enrolled 75 patients with isolated severe AS aged 66±8 years (mean gradient 52 mmHg, max gradient 84 mmHg) undergoing valve replacement surgery. Aortic valve leaflets obtained from AS patients (n=30) and from age-matched healthy autopsy donors (n=3) were used to evaluate in loco the presence of plasminogen (PLG), tissue plasminogen activator (tPA), and fibrinolysis inhibitors (α2-antiplasmin and plasminogen activator inhibitor, PAI-1). PLG activity, plasma tPA and PAI-1 levels, along with cholesterol and oxPL-LDL were assayed. Clot lysis time (CLT) was performed to assess the overall fibrinolytic system activity. Results AS patients with LDL cholesterol ≥3.0 mM (n=25, 56% taking statin) compared to those with LDL cholesterol &amp;lt;3.0 mM (n=50, 86% taking statin) had higher oxPL-LDL levels (+80%, p=0.0014). Moreover, AS subjects with LDL cholesterol ≥3.0 mM, compared to those with LDL cholesterol &amp;lt;3.0 mM had longer CLT (+22.4%, p=0.0022) along with higher plasma tPA (+31.5%, p=0.004) and PAI-1 (+47.4%, p=0.01), while PLG activity tended to be higher (+10.8%, p=0.056). For the first time the valvular expression of fibrinolytic proteins was detected in AS patients, but not in healthy donors. The expression of studied proteins was observed mainly on the aortic side of the leaflets and only trace amounts were detectable in deeper layers of valves. The valve area positive for PLG was 4.4±1.8%, for tPA 1.6±0.8%, while the expression of α2-antiplasmin was 4.2±1.7% and 6.9±2.3% for PAI-1. In the whole AS group, oxPL-LDL, but not LDL cholesterol showed associations with mean (r=0.34, p=0.041) and max (r=0.3, p=0.016) gradients. Serum concentrations of LDL cholesterol correlated with CLT (r=0.42, p=0.0002), blood PLG activity (r=0.56, p=0.0012) and with tPA (r=0.43, p=0.0001) and PAI-1 (r=0.32, p=0.006) levels. Similarly, oxPL-LDL were associated with CLT, PLG activity, and tPA (r=0.34; r=0.59; r=0.47, all p&amp;lt;0.01). Valvular expression of fibrinolytic proteins did not correlate with LDL cholesterol, ox-PL-LDL or transvalvular gradients. Conclusions This study is the first to demonstrate that in AS patients hypercholesterolemia and enhanced lipid oxidation are associated with impaired systemic fibrinolysis, which supports the concept of beneficial effects of cholesterol-lowering therapy in mild-to-moderate AS. Moreover, valvular expression of fibrinolytic factors suggests that these factors are delivered with circulating blood but not synthesized de novo. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): National Science Centre (NCN)

Elevated Lactate Levels in Acute Pulmonary Embolism Are Associated with Prothrombotic Fibrin Clot Properties: Contribution of NETs Formation.

Background: Elevated plasma lactate levels correlate with high mortality rate in acute pulmonary embolism (PE) patients. We hypothesized that elevated lactate levels correlate with prothrombotic fibrin clot properties and enhanced neutrophil extracellular trap (NET) formation in acute PE. Methods: As many as 126 normotensive acute PE patients (aged 58 ± 14 years) were enrolled. Plasma fibrin clot permeability (Ks), clot lysis time (CLT), endogenous thrombin potential (ETP), citrullinated histone H3 (citH3), and plasminogen activator inhibitor-1 antigen (PAI-1), together with plasma L-lactate levels were evaluated on admission. Results: Lactate levels ≥2 mM were found in 70 (55.6%) patients in whom we observed 29% higher neutrophil count and 45% elevated plasma citH3 levels. Elevated lactate levels were associated with more prothrombotic fibrin properties as reflected by 11% reduced Ks, 13% longer CLT, along with 11% increased ETP. Lactate levels were positively associated with plasma citH3 concentrations, ETP, CLT, and PAI-1 (p < 0.05). An increase of lactate levels by 1 mM leading to the prolongation of CLT by 8.82 min was shown in the linear regression. Conclusions: Our findings suggest a new mechanism contributing to a negative impact of elevated lactate levels on prognosis in acute PE patients, in particular hypofibrinolysis, associated with enhanced NET formation.

Prothrombotic clot properties can predict venous ulcers in patients following deep vein thrombosis: a cohort study

Venous ulcers are the most severe manifestation of post-thrombotic syndrome (PTS). We have previously demonstrated that formation of compact fibrin clots resistant to lysis is observed in patients following deep-vein thrombosis (DVT) who developed PTS. The current study investigated whether unfavourable fibrin clot properties can predict post-thrombotic venous ulcers. In a cohort study on 186 consecutive patients following DVT, we determined plasma fibrin clot characteristics, including clot permeability and lysability, inflammatory markers, thrombin generation, fibrinolysis proteins at 3 months since the index event. Occurrence of PTS and venous ulcers was recorded during follow-up (median, 53; range 24 to 76 months). Fifty-seven DVT patients (30.6%) developed PTS, including 12 subjects (6.45%) with a venous ulcer (4 individuals with recurrent ulcers). Patients who developed ulcers compared with the remainder had at enrolment 13.0% lower clot permeability (Ks), 17.4% longer clot lysis time (CLT), 13.1% longer lag phase of clot formation, and 5.0% higher maximum absorbance, with no difference in fibrinogen, C-reactive protein, and thrombin generation. The baseline prothrombotic fibrin clot phenotype (Ks ≤ 6.5 × 10−9 cm2 and CLT > 100 min) was associated with a higher risk of ulcers [hazard ratio (HR), 5.37; 95% confidence interval (CI), 1.3–21.5]. A multivariate model adjusted for age, sex, and fibrinogen showed that independent predictors of the ulcer occurrence were body mass index (HR 1.53; 95% CI 1.30–1.86), CLT (HR 1.43; 95% CI 1.04–2.05), and α2-antiplasmin (HR 0.95; 95% CI 0.90–0.99). This study suggests that formation of denser fibrin clots with impaired fibrinolysis predisposes to post-thrombotic venous ulcers.

Unfavourably altered plasma clot properties in patients with primary Raynaud\u2019s phenomenon: association with venous thromboembolism

Associations of Raynaud’s phenomenon (RP) with venous thromboembolism (VTE) are unclear. We investigated the occurrence of RP together with prothrombotic state markers and fibrin clot properties in VTE patients. In this prospective cohort study we enrolled 360 patients free of known autoimmune disease. D-dimer, von Willebrand factor (vWF), plasma clot permeability (Ks), clot lysis time (CLT) along with fibrinolysis activators and inhibitors were determined at least 3 months since the VTE event. The presence/absence of RP was diagnosed at least 6 months before VTE. Primary RP occurred in 57 subjects (17%) with a 3.6-fold higher prevalence among women. Patients with RP had 11% higher fibrinogen, 16% higher vWF, 5% lower Ks, and 10% longer CLT (all p < 0.05). Females with RP (21%) had 6.6% lower Ks, 11.2% longer CLT, and 18.5% higher vWF (all p < 0.05) compared with men. CLT was predicted by PAI-1 and vWF levels. Regression analysis showed that RP was a predictor of prolonged CLT in the whole patient group (OR 3.46, 95% CI 1.92–6.24) and in women following VTE (OR 2.75, 95% CI 1.31–5.78). Primary RP patients tend to form denser plasma fibrin clots displaying impaired lysability and increased endothelial damage. RP might be a novel risk factor for VTE, especially in women.

Altered Plasma Clot Properties and Trauma-Related Venous Thromboembolism despite Thromboprophylaxis.

Prothrombotic clot phenotype may characterize patients developing deep vein thrombosis (DVT) despite pharmacological thromboprophylaxis. We studied the role of fibrin clot properties and its potential determinants in individuals who experienced DVT after lower limb injury. In a case-control study, we assessed 50 patients who developed DVT despite prophylactic use of low-molecular-weight heparins (the failed thromboprophylaxis group) after a lower limb injury, and three age- and sex-matched control groups, 50 patients each: (1) patients with trauma-related DVT without prior thromboprophylaxis; (2) individuals with unprovoked DVT; (3) patients without history of DVT (the no-DVT controls). Fibrin clot properties, along with thrombin concentration and α2-antiplasmin, were assessed following 3 months of anticoagulation in all DVT patients. Compared with the no-DVT controls, the failed thromboprophylaxis group exhibited denser fibrin networks (12.8% lower clot permeability [Ks], p = 0.0008) and impaired fibrinolysis (46.2% longer clot lysis time [CLT], p = 0.0001 and 8% lower rate of D-dimer release from clots, p = 0.0008). In the unprovoked DVT, similar Ks and 14.9% shorter CLT (p = 0.02) were reported compared with the failed thromboprophylaxis group. The failed thromboprophylaxis patients had higher odds of having elevated peak thrombin generation (>241.5 nM, 90th percentile in the no-DVT controls; odds ratio [OR]: 3.62; 95% confidence interval [CI], 1.86-7.06; p = 0.002), and higher odds of having elevated α2-antiplasmin (>115.05%; OR: 3.38; 95% CI, 1.64-6.98; p = 0.001). Patients who experienced DVT despite thromboprophylaxis following lower limb trauma display a strongly prothrombotic fibrin clot phenotype, including increased clot density and hypofibrinolysis associated with higher plasma α2-antiplasmin.

Impaired fibrinolysis and lower levels of plasma \u03b12-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.

Elevated NT-proBNP is associated with unfavorably altered plasma fibrin clot properties in atrial fibrillation

BackgroundDense fibrin clot formation and hypofibrinolysis have been reported in atrial fibrillation (AF). It is unclear which factors affect fibrin clot properties in AF. Methods and resultsWe investigated plasma fibrin clot permeability (Ks), clot lysis time (CLT), endogenous thrombin potential (ETP) as well as other coagulation and fibrinolysis parameters along with N-terminal pro-B-type natriuretic peptide (NT-proBNP) in 160 AF patients (median age, 70.5years). Previous stroke (n=15; 9.4%) was associated with decreased Ks (P=0.04) and longer CLT (P=0.005), together with higher antiplasmin (P=0.03) and lower tissue-type plasminogen activator (P=0.01). Lower Ks (P=0.04) and tendency towards longer CLT (P=0.10) were observed in patients with a left atrium diameter>40mm. Patients with a CHA2DS2-VASc score of 3 or more (82.5%) were characterized by higher thrombin-activatable fibrinolysis inhibitor antigen (P=0.009). Ks was inversely correlated with log NT-proBNP (r=−0.34, P<0.0001), plasminogen activator inhibitor-1 (PAI-1) antigen (r=−0.24, P=0.002) and C-reactive protein (r=−0.18, P=0.02), while CLT was positively correlated with log NT-proBNP (R=0.61, P<0.0001) and ETP (r=0.37, P<0.0001), which were interrelated (r=0.59, P<0.0001). After adjustment for potential confounders, PAI-1 (odds ratio [OR]: 1.14; 95% confidence interval [CI]: 1.02–1.26) was the only independent predictor of low Ks (the lowest quartile,≤6×10−9cm2), while NT-proBNP (OR: 1.21; 95% CI: 1.12–1.31) and PAI-1 (OR: 1.30; 95% CI: 1.12–1.51) both predicted prolonged CLT (the top quartile,≥109min). ConclusionIn AF patients prothrombotic fibrin clot properties assessed ex vivo are determined by PAI-1 and NT-proBNP and this phenotype is associated with prior ischemic stroke.

PP.16.14] OBSTRUCTIVE SLEEP APNEA BUT NOT RESISTANT HYPERTENSION INFLUENCES PLASMA FIBRIN CLOT PROPERTIES IN PATIENTS WITH HYPERTENSION AT HIGH CARDIOVASCULAR RISK

Objective: Both obstructive sleep apnea (OSA) and resistant hypertension (RHT) are related with high cardiovascular (CV) risk. Formation of dense and poorly lysable plasma fibrin clots is observed in clinical settings characterized by high CV risk. We investigated plasma fibrin clot properties in high-risk hypertensive patients with and without OSA or RHT but without clinically overt CV disease Design and method: We studied 88 hypertensive patients (64 M, 24F, mean age 48.6 ± 8.9years) at high CV risk (2007 ESH/ESC criteria) suspected of OSA. Patients with diagnosed atherosclerotic CV disease were excluded. Fibrin clot properties including clot permeability (Ks), clot lysis time (CLT), and turbidimetric parameters of clot formation (the lag phase of fibrin formation and maximum absorbency (δAbsmax) at 405 nm) were determined. All patients underwent PSG - the apnea/hypopnea index (AHI) > 15 events/h defined clinically important moderate-to-severe OSA. RHT was defined as uncontrolled HT despite using 3 antihypertensive drugs including diuretic. Patients were divided into 4 groups based on the presence of OSA or RHT: OSA-RHT-, OSA-RHT+, OSA+RHT−, OSA + RHT+ Results: There were no significant differences in gender, age and BMI between the groups. Patients in the OSA+ groups were characterized by lower Ks (5.88 ± 0.86 and 6.09 ± 0.96 vs 7.11 ± 0.90 and 7.58 ± 1.27 10 - 9 cm2; p < 0.001) and by longer CLT (108.85 ± 13.88 and 102.12 ± 11.34 vs 96.46 ± 12.20 and 89.48 ± 15.67 min; p < 0.001 for OSA + RHT+ and OSA + RHT− vs OSA-RHT+ and OSA-RHT- groups respectively) as compared with the groups without OSA. In both OSA groups clots formed faster (lag phase, 41.39 ± 3.82 and 40.71 ± 4.81 vs 41.15 ± 4.20 and 44.60 ± 5.31 s; p = 0.018) and tended to had higher δAbsmax (0.86 ± 0,05 and 0.86 ± 0.05 vs 0.85 ± 0.08 and 0.83 ± 0.07 min; p = 0.12 for OSA + RHT+ and OSA + RHT− vs OSA-RHT+ and OSA-RHT- groups respectively). Ks and CLT correlated with AHI but now with office BP and ambulatory BP levels Conclusions: In hypertensive patients at high CV risk, the presence of OSA but not of RHT was associated with unfavorable, prothrombotic changes in the structure and function of fibrin clots. The study may indicate the involvement of prothrombothic mechanism in which OSA may has an impact on CV risk

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. To investigate whether enhanced thrombin generation and impaired fibrinolysis occur in asthmatics. Plasma thrombin generation profile together with a computational assessment of thrombin dynamics and fibrinolytic capacity expressed as clot lysis time (CLT) were determined in 164 consecutive patients with stable asthma and 72 controls matched for age, gender, weight and smoking. Asthma patients had 20.2% increased endogenous thrombin potential (ETP), 41.4% higher peak thrombin concentration, 61% higher maximal prothrombin conversion rate, 15.5% faster rate of thrombin formation (all, P < 0.0001) and 10% lower thrombin decay capacity (P = 0.0004) compared with controls. Asthmatics had also 14.4% longer CLT (P = 0.001) associated with 21.3% higher plasminogen activator inhibitor-1 (PAI-1) (P < 0.0001), and 13% higher plasma α2 -macroglobulin (P = 0.0002). Using ETP and CLT above 75th percentile of the control values as the cut-off levels, we found increased risks of enhanced thrombin generation and hypofibrinolysis in asthmatics, also after correction for potential confounders. ETP and CLT were associated inversely with forced expiratory volume in 1 s/vital capacity (FEV1 /VC) index, after adjustment for age and body mass index. Non-allergic asthma (n = 70, 42.6%) was characterized by 17.5% longer CLT (P = 0.02), which positively associated with PAI-1. Thrombin generation profile was not affected by allergy. Asthma is associated with enhanced thrombin generation and impaired fibrinolysis, which might contribute to thromboembolic events in this disease.

Reduced plasma fibrin clot permeability and susceptibility to lysis are associated with increased risk of postthrombotic syndrome

The postthrombotic syndrome (PTS) is a severe complication of deep vein thrombosis (DVT). Reduced plasma clot permeability and lysability have been linked to DVT and residual vein obstruction. OBJECTIVES We investigated whether altered fibrin clot properties are associated with the occurrence of PTS. Plasma fibrin clot permeability (Ks ) and lysability were investigated in a cohort of 197 consecutive patients aged 18 to 65 years recruited 3 months following the first-ever DVT. Patients with severe thrombophilia or comorbidities known to adversely affect clot phenotype were ineligible. During a 1-year follow-up PTS developed in 48 (24%) patients, who were characterized by lower Ks , prolonged fibrin clot lysis time (CLT) and slower release of D-dimer from clots (D-Drate ), together with higher plasma D-dimer, C-reactive protein and thrombin-activatable fibrinolysis inhibitor (TAFI). No PTS-associated differences in fibrinogen, thrombin generation, factor VIII, other fibrinolysis proteins and the quality of anticoagulation were observed. Ks (r = -0.71), CLT (r = 0.45), D-Drate (r = -0.30) and TAFI activity (r = 0.38) were associated with the Villalta scale (all P < 0.05). Recurrent VTE occurred also more commonly in PTS patients during follow-up and the 26 (13.2%) patients had lower Ks , longer CLT and lower D-Drate (all P < 0.05). A multivariate model adjusted for age, body mass index, fibrinogen and glucose showed that independent predictors of PTS were idiopathic DVT, plasma D-dimer, Ks , D-Drate , tissue plasminogen activator and TAFI activity. This study demonstrates that formation of more compact fibrin clots displaying impaired susceptibility to lysis predisposes to PTS.

Altered fibrin clot structure/function in patients with antiphospholipid syndrome: association with thrombotic manifestation

We tested the hypothesis that plasma fibrin clot structure/function is unfavourably altered in patients with antiphospholipid syndrome (APS). Ex vivo plasma clot permeability, turbidity and susceptibility to lysis were determined in 126 consecutive patients with APS enrolled five months or more since thrombotic event vs 105 controls. Patients with both primary and secondary APS were characterised by 11% lower clot permeability (p<0.001), 4.8% shorter lag phase (p<0.001), 10% longer clot lysis time (p<0.001), and 4.7% higher maximum level of D-dimer released from clots (p=0.02) as compared to the controls. Scanning electron microscopy images confirmed denser fibrin networks composed of thinner fibres in APS. Clots from patients with "triple-antibody positivity" were formed after shorter lag phase (p=0.019) and were lysed at a slower rate (p=0.004) than in the remainder. Clots from APS patients who experienced stroke and/or myocardial infarction were 8% less permeable (p=0.01) and susceptible to lysis (10.4% longer clot lysis time [p=0.006] and 4.5% slower release of D-dimer from clots [p=0.01]) compared with those following venous thromboembolism alone. Multivariate analysis adjusted for potential confounders showed that in APS patients, lupus anticoagulant and "triple-positivity" were the independent predictors of clot permeability, while "triple-positivity" predicted lysis time. We conclude that APS is associated with prothrombotic plasma fibrin clot phenotype, with more pronounced abnormalities in arterial thrombosis. Molecular background for this novel prothrombotic mechanism in APS remains to be established.

Acute CO Poisoning is Associated with Impaired Fibrinolysis and Increased Thrombin Generation

Carbon monoxide (CO) poisoning is a leading cause of unintentional poisoning deaths in many countries. In ex vivo studies, CO released from carbon monoxide-releasing molecules has been shown to attenuate fibrinolysis via increased alpha-2-antiplasmin activity. Hypofibrinolysis is associated with coronary ischaemia, which is also commonly observed in CO poisoning. We examined fibrin clot properties in acutely poisoned CO patients. Ex vivo plasma fibrin clot permeability, turbidimetry and efficiency of fibrinolysis were investigated in 48 patients and controls matched for age and sex. CO-poisoned patients had 11.6% longer clot lysis time than the controls (p < 0.0001). No intergroup differences in clot permeability or turbidimetric variables were observed. Plasma tissue-type plasminogen antigen (tPA), plasminogen activator inhibitor-1 (PAI-1) antigen and activity and F1.2 prothrombin fragments were higher in the patients than in the controls (all p < 0.0001). Plasma tPA activity was lower in the CO-poisoned group. Multiple linear regression showed that a thrombin generation marker, F1.2, is the strongest predictor of clot lysis time, followed by PAI-1 activity and carboxyhaemoglobin levels. In conclusion, this report is the first to demonstrate that acute CO poisoning in human beings is linked to increased thrombin generation and impaired fibrinolysis, which might contribute to ischaemic complications.

Altered fibrin clot properties in patients with premature peripheral artery disease

It has been shown that formation of denser and poorly lysable fibrin clots is observed in elderly patients with peripheral artery disease (PAD). The aim of the study was to test the hypothesis that premature PAD is associated with more prothrombotic fibrin clot phenotype. Ex‑vivo plasma fibrin clot permeability, turbidity, and susceptibility to lysis were evaluated in 31 premature PAD patients (median ankle brachial index [ABI], 0.75; interquartile range, 0.5-0.8) aged 55 or less and 32 PAD patients (ABI, 0.66; 0.56-0.76) aged over 55 years. Subjects without PAD matched for age and sex (n = 40) served as controls. Premature PAD patients were characterized by 32% lower clot permeability (Ks) (P <0.001), 7% longer clot lysis time (t50%) (P = 0.004), and 31% higher maximum D-dimer levels released from fibrin clots (D-Dmax) (P <0.001) compared with controls. These differences remained significant after adjustment for risk factors and medications. None of the fibrin clot parameters differed between premature and older PAD patients. There were correlations between fibrin clot parameters and CRP in premature PAD patients and with ABI in older PAD patients. In a multiple regression model, premature PAD and ABI were independent predictors of Ks, and premature PAD and plasma fibrinogen of the maximum absorbance of a fibrin gel. Plasma fibrin clots show similarly abnormal prothrombotic phenotype in premature and older PAD patients. However, different factors influence fibrin clot parameters in these patient groups. Premature PAD was an independent predictor of clot permeability and maximum absorbance of a fibrin gel.

Thyroid Dysfunction and Fibrin Network Structure: A Mechanism for Increased Thrombotic Risk in Hyperthyroid Individuals

Hyperthyroidism is associated with increased thrombosis risk, and fibrin clot structure determines susceptibility to vascular thrombotic events. Our objective was to investigate clot formation and lysis in hyperthyroidism using observational and interventional studies. Ex vivo fibrin clot structure/fibrinolysis and plasma levels of thrombotic/inflammatory markers were investigated in hyperthyroid individuals (n = 24) and matched controls (n = 19), using turbidimetric assays, ELISA, and confocal and electron microscopy. The effects of normalizing thyroid function were analyzed (n = 19) and the role of short-term exogenous hyperthyroidism in healthy volunteers studied (n = 16). Hyperthyroid subjects displayed higher clot maximum absorbance compared with controls (0.41 ± 0.03 and 0.27 ± 0.01 arbitrary units, respectively; P < 0.01), and longer clot lysis time (518 ± 23 and 461 ± 18 sec, respectively; P < 0.05), which correlated with free T(4) levels. Plasma levels of fibrinogen and plasminogen activator inhibitor-1 were significantly higher in patients compared with controls. Normalizing thyroid function in 19 subjects was associated with lower maximum absorbance and shorter lysis time, accompanied by reduction in fibrinogen, plasminogen activator inhibitor-1, and D-dimer levels. Complement C3, but not C-reactive protein, levels were higher in hyperthyroid subjects compared with controls (0.92 ± 0.05 and 0.64 ± 0.03 g/liter, respectively; P < 0.01), correlated with clot structure parameters, and decreased after intervention. Confocal and electron microscopy confirmed more compact clots and impaired fibrinolysis during hyperthyroidism. Exogenous hyperthyroidism in healthy volunteers had no effect on any of the clot structure parameters. Endogenous hyperthyroidism is associated with more compact clots and resistance to fibrinolysis ex vivo, related to the degree of hyperthyroidism and C3 plasma levels, and these changes are modulated by achieving euthyroidism. Altered clot structure/lysis may be one mechanism for increased thrombotic risk in hyperthyroidism.

A history of early stent thrombosis is associated with prolonged clot lysis time

It has been demonstrated that formation of compact plasma fibrin clots resistant to plasmin-mediated lysis characterises patients following in-stent thrombosis (IST). The relationship between defective fibrinolysis, reflected as prolonged clot lysis time (CLT) and IST is unclear. We sought to investigate whether patients with acute and subacute IST have impaired fibrinolytic capacity. We studied 41 definite IST patients, including 15 with acute and 26 with subacute IST experienced 2-73 months prior to enrollment, versus 41 controls matched for demographics, cardiovascular risk factors, concomitant treatment and angiographic/stent parameters. CLT, reflecting lysis of a tissue factor-induced plasma clot by exogenous tissue plasminogen activator, together with plasminogen activator inhibitor-1 (PAI-1) antigen and activity, thrombin-activatable fibrinolysis inhibitor (TAFI) antigen and activity, thrombomodulin (TM), plasminogen and α2-antiplasmin (α2AP) were measured. There were no inter-group differences in angiographic parameters, indication to the first PCI, culprit vessel or a type of stent. Patients with IST had 11% longer CLT (p=0.005) and 13% higher PAI-1 antigen (p=0.04) compared to controls. There were positive correlations in both groups between CLT and PAI-1 antigen and TAFI activity (all p<0.001). Multiple regression analysis showed that CLT (odds ratio [OR]=1.04 per 1 minute, 95% CI 1.01-1.08, p=0.02) and platelet count (OR=1.01 per 1,000/μl, 95% CI 1.00-1.02, p=0.034) were independent predictors of IST (R(2)=0.28, p<0.05). Concluding, impaired fibrinolytic potential, that is in part determined by plasma PAI-1 antigen and TAFI activity, characterises patients with a history of acute and subacute IST, which might help identify patients at higher risk of IST.

Thromboembolic events are associated with prolonged clot lysis time in patients with permanent atrial fibrillation

Atrial fibrillation (AF) is associated with a prothrombotic state. We evaluated associations of previous thromboembolic events with fibrinolytic parameters in patients with AF. We studied 62 consecutive patients with permanent AF (27 men, 35 women, aged 46-89 years [median, 78 years]). Patients receiving warfarin or acenocoumarol on a long-term basis were eligible. We determined plasma fibrin clot lysis time (CLT), plasminogen activator inhibitor-1 (PAI-1) antigen, thrombin-activatable fibrinolysis inhibitor (TAFI) activity and antigen, plasminogen, α2-antiplasmin (α2AP), and soluble thrombomodulin (sTM). There were 19 subjects (30.6%) with a history of thrombotic events (stroke in 11, myocardial infarction in 8, and pulmonary embolism in 3 patients). They had longer CLT (P = 0.0035 for patients with previous stroke and P = 0.001 for patients with any previous thrombotic event), together with higher PAI-1 (P = 0.025 and P = 0.016, respectively), TAFI activity (P = 0.002 and P = 0.011, respectively), sTM (P = 0.0023 and P = 0.012, respectively), and α2AP (P = 0.007 and P = 0.0006, respectively) than the remaining subjects. AF patients with previous stroke had also higher TAFI antigen than the remainder (P = 0.04). CLT (P = 0.024), PAI-1 (P = 0.022), TAFI activity (P = 0.048), and sTM (P = 0.032, all P for trend) increased with higher CHA2DS2-VASc scores. CLT was not associated with time from thrombotic event to enrollment. Patients taking oral anticoagulants (n = 46) had only slightly higher sTM levels (3.6 [2.9-6.3] vs. 2.9 [2.2-4.1] ng/ml, P = 0.049) than the remaining subjects. Stroke or other thromboembolic event in AF patients is associated with impaired lysability of fibrin clots combined with elevated PAI-1, TAFI, sTM, and α2AP.

Clot lysis time and thrombin activatable fibrinolysis inhibitor in severe preeclampsia with or without associated antiphospholipid antibodies

We investigated clot lysis time, thrombin activatable fibrinolysis inhibitor antigen (TAFI) levels and TAFI gene polymorphisms in pregnant patients with severe preeclampsia, with or without associated antiphospholipid syndrome (APS). The study groups included 82 pregnant patients without antiphospholipid antibodies with severe preeclampsia (PE group) and 10 pregnant APS patients who developed severe preeclampsia (APS-PE group). Controls included 76 primary pregnant APS patients (APS group) and 89 healthy pregnant patients (NOR group) with uneventful term pregnancy and delivery. Patients in the APS-PE, APS and NOR groups were sampled during each trimester of pregnancy and at 4-6 months and 12 months after delivery. Patients in the PE group were sampled during the third trimester and after delivery. Significantly prolonged clot lysis time after delivery was found in the PE, APS-PE and APS groups compared to the NOR group. The PE and APS-PE groups had longer clot lysis time than the APS group. Levels of TAFI were found to be higher after delivery in patients of the PE and APS-PE groups compared to the APS and NOR groups. Allele distribution of the TAFI gene polymorphisms was similar among the four study groups. We conclude that increased TAFI antigen levels and impaired fibrinolysis are pathogenetic factors in preeclampsia, regardless of whether or not preeclampsia is associated with the presence of antiphospholipid antibodies.