Asthma is associated with enhanced thrombin generation and impaired fibrinolysis, which might contribute to thromboembolic events in this disease.

A and-much slower-B from the N -termini of the Aα and Bβ chains, respectively, resulting in the formation of fibrin monomer with exposed binding sites in the E domain. Fibrin monomers polymerize via noncovalent interactions between the D and E domains with subsequent lateral aggregation promoted mainly by intermolecular cross -linking of α chains and probably by interactions between α and γ chains.3,4 A half -staggered fibrin structure forms a protofibril.Fibrin resistance to plasmin degradation is determined largely by covalent cross -linking mediated by activated factor XIII (FXIII), a transglutaminase enzyme whose formation from zymogen FXIII is catalyzed by thrombin. Active FXIII catalyzes the formation of covalent bonds between γ -γ, γ -α, and α -α chains of contiguous fibrin polypeptide chains.5 FXIII also links α 2 -antiplasmin and plasminogen activator inhibitors to fibrin to ensure clot resistance to enzymatic degradation. 5Fibrinogen and fibrin specifically bind a variety Fibrin formation and structure Fibrin is the main protein component of a blood clot and intravascular thrombi in all locations. Efficient fibrin formation and its normal functions are essential for hemostasis.1 Fibrinogen, the soluble fibrin precursor synthesized in the liver, is a 340 -kDa glycoprotein composed of 3 paired polypeptide chains (AαBβγ) 2 that are cross -linked together by 29 disulfide bonds. Fibrinogen contains 3 main structural regions connected by α -helical coils: a central E domain composed of the N -termini of all 6 polypeptide chains and 2 outer D domains with C -termini of the Bβ and γ chains. The C -terminal of the Aα chain is a globular structure located near the central E domain. Approximately 10% of total plasma fibrinogen molecules contain γ' chain, whose presence may contribute to cardiovascular disease. [1][2][3] Fibrin formation is initiated by thrombin cleavage of the Aα and Bβ chains of fibrinogen. AbsTRACTFibrin formed as a result of fibrinogen polymerization is the main protein component of a clot in a test tube and intravascular thrombi in vivo. Fibrin clot structure characterized by fiber diameter and pore size differs between healthy persons and those with thromboembolic diseases, in part due to the quality and quantity of fibrinogen and the magnitude of thrombin generation. A key measure of plasma clot structure is its permeability, reflected by the Darcy constant (K s ). Reduced K s is a typical feature of the prothrombotic fibrin clot phenotype, which is associated with faster formation of denser fibrin mesh, relatively resistant to lysis. Low K s has been reported in patients with prior or acute myocardial infarction (MI), stroke, or venous thromboembolism (encompassing deep vein thrombosis [DVT] and pulmonary embolism [PE]), as well as in those with prothrombotic conditions (eg, in several thrombophilic states) and in the presence of cardiovascular risk factors (eg, obesity). Antithrombotic and anticoagulant agents, along with statins, have been shown to increase K s . Growing evidence indic...

Background Direct oral anticoagulants (DOACs) cause false positive lupus anticoagulant (LA) results. We assessed the impact of DOAC-Stop, reversing in vitro effects of DOACs, on LA testing in anticoagulated patients. Methods We assessed 75 venous thromboembolism patients aged 44.5±14.6 years. Blood samples were collected 2–28 h since intake of DOACs, including 50 patients on rivaroxaban, 20 on dabigatran and five on apixaban. LA testing was performed at baseline and after DOAC-Stop treatment. Positive LA was defined as the normalized (patient/standard plasma clotting time) LA screening and screening (LA1)/confirmation (LA2) ratios exceeding 1.2. Results LA diluted Russell’s viper venom time (dRVVT) normalized screening test revealed abnormal results in 73 (97.3%) and activated partial thromboplastin time (APTT)-LA in 49 (65.3%) patients. In six (8%) patients, antiphospholipid syndrome (APS) was diagnosed. dRVVT LA1/LA2 was abnormal in 35 (50.7%) patients taking DOACs. The APTT ratio was normal in all studied subjects. DOAC-Stop completely removed dabigatran and reduced by 98% rivaroxaban and by 92.3% apixaban concentrations (all p<0.05). After DOAC-Stop screening dRVVT remained prolonged in 34 (49.3%) patients (p<0.001), while dRVVT LA1/LA2 was abnormal in six (8.7%) subjects, with no association with DOAC concentrations at baseline and after DOAC-Stop. The APTT-LA screening test remained prolonged in five (7.2%) patients, while the APTT LA1/LA2 ratio was normal in those subjects. DOAC-Stop did not influence LA testing in APS patients. Conclusions Application of DOAC-Stop effectively reduced plasma DOAC concentrations leading to appropriate dRVVT results in up to 97% of VTE patients.

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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 (K s ), 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 K s (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 K s , and reduced maximum rate of increase in D-dimer levels in the lysis assay (all P<0.05). Conclusions-Altered

Background: Accumulation of advanced glycation end products (AGEs) leads to chronic glycation of proteins and tissue damage, particularly in patients with diabetes mellitus (DM). We aimed to evaluate whether increased accumulation of AGEs in patients with aortic stenosis (AS) and concomitant type 2 diabetes (DM) is associated with AS severity. Methods: We prospectively enrolled 76 patients with severe AS (47.1% males; nonDM), aged 68 [66-72] years, and 50 age-matched DM patients with a median blood glucose level of 7.5 [5.9-9.1] mM and glycated hemoglobin (HbA1c) of 6.8 [6.3-7.8]%, scheduled for aortic valve replacement. Valvular expression of AGEs, AGEs receptor (RAGE), interleukin-6 (IL-6), and reactive oxygen species (ROS) induction were evaluated ex vivo by immunostaining and calculated as the extent of positive immunoreactive areas/total sample area. Plasma levels of AGEs and soluble RAGE (sRAGE) were assessed by ELISAs. Results: Subjects with DM had increased valvular expression of both AGEs (6.6-fold higher, 15.53 [9.96-23.28]%) and RAGE (1.8-fold higher, 6.8 [4.9-8.45]%) compared to nonDM patients (2.05 [1.21-2.58]% and 2.4 [1.56-3.02]%, respectively; both p < 0.001). Plasma levels of AGEs (12-fold higher) and sRAGE (1.3-fold higher) were elevated in DM patients, compared to nonDM (both p < 0.0001). The percentage of valvular ROS-positive (2.28 [1.6-3.09] vs. 1.15 [0.94-1.4]%, p < 0.0001) but not IL-6-positive areas was higher within DM, compared to nonDM valves. In DM patients, the percentage of valvular AGEs-and RAGE-positive areas correlated with HbA1c (r = 0.77, p < 0.0001 and r = 0.30, p = 0.034). Similarly, plasma AGEs and sRAGE levels were associated with HbA1c in the DM group (r = 0.32, p = 0.024 and r = 0.33, p = 0.014, respectively). In all DM patients, we found an association between the amount of valvular AGEs and the disease severity measured as aortic valve area (AVA; r = 0.68, p < 0.0001). Additionally, in DM patients with HbA1c > 7% (n = 24, 48%) we found that valvular expression of AGEs correlated with mean transvalvular pressure gradient (PG mean ; r = 0.45, p = 0.027). Plasma AGEs levels in the whole DM group correlated with AVA (r = − 0.32, p = 0.02), PG mean (r = 0.31, p = 0.023), and PG max (r = 0.30, p = 0.03). Conclusions: Our study suggests that poorly-controlled diabetes leads to increased AGEs and RAGE valvular accumulation, which at least partially, might result in AS progression in DM patients.

Venous thromboembolism is associated with formation of denser fibrin clots resistant to lysis. We investigated whether prothrombotic plasma clot properties are associated with the severity of acute pulmonary embolism (PE). We enrolled 126 normotensive acute PE patients (aged 58 ± 14 years) and 25 age- and sex-matched healthy controls. Plasma fibrin clot permeability (K s ), clot lysis time (CLT), endogenous thrombin potential (ETP), plasminogen activator inhibitor-1 (PAI-1), and citrullinated histone H3 (citH3) were evaluated on admission. PE patients compared to controls had 370% higher citH3 levels, 41% higher ETP, 16.5% reduced K s , and 25.6% prolonged CLT. Patients with intermediate-high (n = 29) and intermediate-low (n = 77) PE mortality risk had reduced K s and prolonged CLT, increased PAI-1 and ETP as compared to low-risk PE (n = 20) patients. Prolonged CLT was predicted by PAI-1 and citH3, while low K s by C-reactive protein. During a 12-month follow-up 9 (7.1%) patients who had 24% higher ETP, 45% higher citH3 levels, and 18% prolonged CLT at baseline died. High ETP combined with elevated citH3 levels and prolonged CLT was associated with eightfold increased risk of PE-related death. Prothrombotic fibrin clot properties and enhanced neutrophil extracellular traps formation are associated with higher early mortality risk in acute PE patients, which suggests a prognostic role of these biomarkers.

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.