Thrombin-activatable Fibrinolysis Inhibitor Levels Research Articles

Impaired Fibrinolysis in the Antiphospholipid Syndrome.

The pathogenesis of the antiphospholipid syndrome (APS) is complex and involves the persistent presence of antiphospholipid antibodies (aPL) in the bloodstream causing a prothrombotic condition. aPL induce excessive activation of the endothelium, monocytes, and platelets in consort with aberrations in hemostasis/clotting, fibrinolytic system, and complement activation. Impaired fibrinolysis has been found in APS patients with thrombotic as well as obstetric manifestations. Increased levels of plasminogen activator inhibitor-1 and thrombin-activatable fibrinolysis inhibitor, together with the presence of aPL against annexin-2, tissue-type plasminogen activator, and plasminogen contribute to the compromised fibrinolytic activity in these patients. Furthermore, unfavorably altered fibrin morphology, less amenable to fibrinolysis, has been proposed as a novel prothrombotic mechanism in APS. This review aims to summarize the present knowledge of the mechanisms involved in impaired fibrinolysis in APS patients. We also present a case from clinical practice as an illustration of fibrinolysis impairment in APS patients from a real-life setting.

Increased Thrombin-Activatable Fibrinolysis Inhibitor in Response to Sublingual Immunotherapy for Allergic Rhinitis.

The objective of this study was to determine the role of thrombin-activatable fibrinolysis inhibitor (TAFI) as a candidate biomarker for therapeutic efficacy of sublingual immunotherapy (SLIT) and to identify the role of TAFI in the pathogenesis of allergic rhinitis (AR). Retrospective cohort study and laboratory study. Serum was collected from patients with allergies to Japanese cedar pollen before, during, and after treatment with SLIT. We measured the levels of immunoreactive TAFI, C3a, and C5a in serum by enzyme-linked immunosorbent assay (ELISA) and assessed their relative impact on a combined symptom-medication score. We also examined the impact of TAFI on mast cells and fibroblasts in experiments performed in vitro. Serum levels of TAFI increased significantly in response to SLIT. By contrast, serum C3a levels decreased significantly over time; we observed a significant negative correlation between serum levels of TAFI versus C3a and symptom-medication score. Mast cell degranulation was inhibited in response to TAFI, as it was the expression of both CCL11 and CCL5 in cultured fibroblasts. High serum levels of TAFI may be induced by SLIT. TAFI may play a critical protective role in pathogenesis of AR by inactivating C3a and by inhibiting mast cell degranulation and chemokines expression in fibroblasts. 4 Laryngoscope, 131:2413-2420, 2021.

Inside the Thrombus: Association of Hemostatic Parameters With Outcomes in Large Vessel Stroke Patients.

Background: Actual clinical management of ischemic stroke (IS) is based on restoring cerebral blood flow using tissue plasminogen activator (tPA) and/or endovascular treatment (EVT). Mechanical thrombectomy has permitted the analysis of thrombus structural and cellular classic components. Nevertheless, histological assessment of hemostatic parameters such as thrombin-activatable fibrinolysis inhibitor (TAFI) and matrix metalloproteinase 10 (MMP-10) remains unknown, although their presence could determine thrombus stability and its response to thrombolytic treatment, improving patient's outcome.Methods: We collected thrombi (n = 45) from large vessel occlusion (LVO) stroke patients (n = 53) and performed a histological analysis of different hemostatic parameters [TAFI, MMP-10, von Willebrand factor (VWF), and fibrin] and cellular components (erythrocytes, leukocytes, macrophages, lymphocytes, and platelets). Additionally, we evaluated the association of these parameters with plasma levels of MMP-10, TAFI and VWF activity and recorded clinical variables.Results: In this study, we report for the first time the presence of MMP-10 and TAFI in all thrombi collected from LVO patients. Both proteins were localized in regions of inflammatory cells, surrounded by erythrocyte and platelet-rich areas, and their content was significantly associated (r = 0.41, p < 0.01). Thrombus TAFI was lower in patients who died during the first 3 months after stroke onset [odds ratio (OR) (95%CI); 0.59 (0.36–0.98), p = 0.043]. Likewise, we observed that thrombus MMP-10 was inversely correlated with the amount of VWF (r = −0.30, p < 0.05). Besides, VWF was associated with the presence of leukocytes (r = 0.37, p < 0.05), platelets (r = 0.32, p < 0.05), and 3 months mortality [OR (95%CI); 4.5 (1.2–17.1), p = 0.029]. Finally, plasma levels of TAFI correlated with circulating and thrombus platelets, while plasma MMP-10 was associated with cardiovascular risk factors and functional dependence at 3 months.Conclusions: The present study suggests that the composition and distribution of thrombus hemostatic components might have clinical impact by influencing the response to pharmacological and mechanical therapies as well as guiding the development of new therapeutic strategies.

Hypofibrinolysis is associated with the severity of Covid-19 infection: The role of obesity

Introduction: Obesity, by causing hypofibrinolysis and thrombotic complications, ought to be a risk factor in terms of disease severity and mortality in COVID-19 infection.We aim to investigate the effects of obesity on fibrinolytic system in coronavirus patients while studying the changes of major fibrinolytic inhibitors plasminogen activator inhibitor (PAI-1) and thrombin activatable fibrinolysis inhibitor (TAFI). Material and methods: Sixty-six patients and 21 healthy donors were investigated and observed until either their recovery or death. Body mass index (BMI) was calculated for each person and peripheral blood used for PAI-1 and TAFI measurements. PAI-1 antigen and TAFI activated/inactivated (TAFIa/i) were measured using ELISA kits. Results: PAI-1 antigen and TAFIa/i plasma levels are higher in patients than in control group (P<00001 for both).Patients needing ICU had higher TAFIa/i values than non-ICU patients. The 15 patients who died had higher TAFIa/i levels than those staying alive.Obese patients (BMI . 30) had higher PAI-1 levels than non-obese patients. TAFIa/i is associated with D-dimer, C-reactive protein (CRP), ferritin, creatinine and neutrophile count, whereas PAI-1 is associated only with the serum creatinine level. Conclusion: Our study shows that the levels of the major fibrinolytic inhibitors PAI 1 and TAFI increase in patients with COVID-19 infection, and proves for the first time that PAI 1 levels increase more in obese patients than in non-obese ones. It is also revealed that the plasma TAFI level is associated with the severity of COVID-19 infection, which suggests that the inhibitory treatments against TAFI should be effective in preventing thrombotic complications in the course of COVID-19 infection.

S62798, a TAFIa inhibitor, accelerates endogenous thrombolysis in a murine model of pulmonary thromboembolism

Abstract Pulmonary embolism (PE) is the third leading cause of cardiovascular death in western countries. The enhancement of fibrinolysis constitutes a promising approach to treat thrombotic diseases. In patients, venous thrombosis and thromboembolism risks are associated with increased plasma levels of TAFI (Thrombin Activatable Fibrinolysis Inhibitor) antigen as well as the active form TAFIa. S62798 is a competitive, selective and potent human TAFIa inhibitor (IC50±SD=11.2±0.4nM). It is however less potent on mouse TAFIa (IC50±SD=270±39nM). Here, we tested the ability of S62798 to enhance endogenous fibrinolysis in a mouse model of pulmonary thromboembolism. Human Tissue Factor (TF) was injected in C57Bl6 male mice. Ten minutes later, mice (n=4 to 14 per group) were treated (IV) with S62798 (from 0.01 to 100mg/kg) or vehicle (0.9% NaCl). Ten or twenty minutes (min) later, mice were anesthetized and lungs were collected, homogenized and pulmonary fibrin was quantified by ELISA. Results are expressed as ratio of geometric mean of pulmonary fibrin (μg/mL): tested treatment/ vehicle [95% confidence interval (CI)]. Ten minutes after S62798 treatment, pulmonary fibrin deposition was dose-dependently decreased with a Minimal Effective Dose of 0.04mg/kg [90% prediction interval 0.037 - 0.051] and an ED50 of 0.03mg/kg [95% CI: 0.01; 0.06]. Mice were then treated with 0.1mg/kg S62798 or vehicle (10 min after TF induction) and fibrin deposition in lungs was quantified 10 and 20 minutes post S62798 treatment. The level of pulmonary fibrin deposition was significantly decreased (p&amp;lt;0.0001) compared to vehicle group (ratio 0.31 [0.21; 0.45] at 10 min; 0.35 [0.24; 0.51] at 20 min). Finally, the effect of S62798 (1mg/kg) in combination with heparin was evaluated (n=10/group). When administered 10 min before TF injection, heparin (2000IU/kg) significantly (p&amp;lt;0.0001) decreased pulmonary fibrin level (20 min post TF: ratio 0.03 [0.01; 0.05]). When treatment was done in a curative setting (10 min post TF), heparin alone had no effect (p=0.85) on fibrin deposition (ratio 0.96 [0.65; 1.43]) whereas a similar significant (p&amp;lt;0.0001) decreased pulmonary fibrin deposition was observed in response to S62798 alone or associated with heparin (ratio 0.27 [0.18; 0.40] (S62798 alone) and 0.29 [0.20; 0.43] (S62798+heparin)). In this model, curative S62798 treatment, alone or associated to heparin, accelerated clot degradation by potentiating endogenous fibrinolysis and thus decreased pulmonary fibrin deposition. Due to its capacity to enhance endogenous fibrinolysis, S62798, which has completed phase I studies, is expected to be a therapeutic option for intermediate high risk PE patients on top of anticoagulants. With early recanalization, S62798 should rapidly reduce pulmonary artery pressure and resistance, with concomitant improvement in right ventricular function, preserving cardiac function, and reducing acute PE-related morbidity and mortality in these patients. Funding Acknowledgement Type of funding source: None

A new pedigree with thrombomodulin‐associated coagulopathy in which delayed fibrinolysis is partially attenuated by co‐inherited TAFI deficiency

BackgroundThrombomodulin‐associated coagulopathy (TM‐AC) is a rare bleeding disorder in which a single reported p.Cys537* variant in the thrombomodulin gene THBD causes high plasma thrombomodulin (TM) levels. High TM levels attenuate thrombin generation and delay fibrinolysis. ObjectivesTo report the characteristics of pedigree with a novel THBD variant causing TM‐AC, and co‐inherited deficiency of thrombin‐activatable fibrinolysis inhibitor (TAFI). Patients/methodsIdentification of pathogenic variants in hemostasis genes by next‐generation sequencing and case recall for deep phenotyping. ResultsPedigree members with a previously reported THBD variant predicting p.Pro496Argfs*10 and chain truncation in TM transmembrane domain had abnormal bleeding and greatly increased plasma TM levels. Affected cases had attenuated thrombin generation and delayed fibrinolysis similar to previous reported TM_AC cases with THBD p.Cys537*. Coincidentally, some pedigree members also harbored a stop‐gain variant in CPB2 encoding TAFI. This reduced plasma TAFI levels but was asymptomatic. Pedigree members with TM‐AC caused by the p.Pro496Argfs*10 THBD variant and also TAFI deficiency had a partially attenuated delay in fibrinolysis, but no change in the defective thrombin generation. ConclusionsThese data extend the reported genetic repertoire of TM‐AC and establish a common molecular pathogenesis arising from high plasma levels of TM extra‐cellular domain. The data further confirm that the delay in fibrinolysis associated with TM‐AC is directly linked to increased TAFI activation. The combination of the rare variants in the pedigree members provides a unique genetic model to develop understanding of the thrombin‐TM system and its regulation of TAFI.

Is there thrombin-activatable fibrinolysis inhibitor in saliva?

The purpose of this study was to identify thrombin-activatable fibrinolysis inhibitor (TAFI) in saliva and to investigate the correlation between TAFI levels in saliva and plasma. Subjects included were healthy adults without diseases or medication that could affect coagulation. Samples of stimulated saliva and blood samples were obtained from 33 subjects. Levels of TAFI in saliva and plasma were analysed. The association between levels of TAFI in saliva and plasma was calculated using linear regression. Low levels of TAFIa/TAFIai were found in most saliva samples but only one sample had levels that were above the lower limit of detection of the assay used. TAFI (proenzyme) was not found in saliva, so no correlations could be calculated. In this study there was no indication that there is TAFI present in secreted saliva. Either TAFIa/TAFIai in saliva were much lower than in plasma and under the detection limit of the assay used, or there was no TAFIa/TAFIai in the saliva tested.

Hemostatic Alterations in Patients With Cirrhosis: From Primary Hemostasis to Fibrinolysis.

In the setting of liver cirrhosis (LC), profound hemostatic changes occur, which affect primary hemostasis, coagulation, and fibrinolysis. They involve prohemorrhagic and prothrombotic alterations at each of these steps. Patients with cirrhosis exhibit multifactorial thrombocytopenia and in vitro thrombocytopathy, counterbalanced by increased von Willebrand factor. The resultant shift is difficult to assess, but overall these changes probably result in a rebalanced primary hemostasis. Concerning coagulation, the reduced activity of coagulation factors is counterbalanced by an increase in factor VIII (produced by liver sinusoidal endothelial cells), a decrease of the natural anticoagulants, and complex changes, including changes in circulating microparticles, cell-free DNA, and neutrophil extracellular traps. Overall, these alterations result in a procoagulant state. As for fibrinolysis, increased tissue-type and urokinase-type plasminogen activators, a relatively decreased plasminogen activator inhibitor 1, and decreased levels of thrombin-activatable fibrinolysis inhibitor and α2-antiplasmin are counterbalanced by decreased plasminogen and a decreased fibrin clot permeability. Whether and how these changes shift fibrinolysis remains to be determined. Overall, the current consensus is that in patients with cirrhosis, the hemostasis is shifted toward a procoagulant state. We review the published evidence for the concept of LC as a prothrombotic state, discuss discordant data, and highlight the impact of the underlying cause of LC on the resultant imbalance.

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.

Post-Traumatic Sepsis Is Associated with Increased C5a and Decreased TAFI Levels.

Background: Sepsis frequently occurs after major trauma and is closely associated with dysregulations in the inflammatory/complement and coagulation system. Thrombin-activatable fibrinolysis inhibitor (TAFI) plays a dual role as an anti-fibrinolytic and anti-inflammatory factor by downregulating complement anaphylatoxin C5a. The purpose of this study was to investigate the association between TAFI and C5a levels and the development of post-traumatic sepsis. Furthermore, the predictive potential of both TAFI and C5a to indicate sepsis occurrence in polytraumatized patients was assessed. Methods: Upon admission to the emergency department (ED) and daily for the subsequent ten days, circulating levels of TAFI and C5a were determined in 48 severely injured trauma patients (injury severity score (ISS) ≥ 16). Frequency matching according to the ISS in septic vs. non-septic patients was performed. Trauma and physiologic characteristics, as well as outcomes, were assessed. Statistical correlation analyses and cut-off values for predicting sepsis were calculated. Results: Fourteen patients developed sepsis, while 34 patients did not show any signs of sepsis (no sepsis). Overall injury severity, as well as demographic parameters, were comparable between both groups (ISS: 25.78 ± 2.36 no sepsis vs. 23.46 ± 2.79 sepsis). Septic patients had significantly increased C5a levels (21.62 ± 3.14 vs. 13.40 ± 1.29 ng/mL; p < 0.05) and reduced TAFI levels upon admission to the ED (40,951 ± 5637 vs. 61,865 ± 4370 ng/mL; p < 0.05) compared to the no sepsis group. Negative correlations between TAFI and C5a (p = 0.0104) and TAFI and lactate (p = 0.0423) and positive correlations between C5a and lactate (p = 0.0173), as well as C5a and the respiratory rate (p = 0.0266), were found. In addition, correlation analyses of both TAFI and C5a with the sequential (sepsis-related) organ failure assessment (SOFA) score have confirmed their potential as early sepsis biomarkers. Cut-off values for predicting sepsis were 54,857 ng/mL for TAFI with an area under the curve (AUC) of 0.7550 (p = 0.032) and 17 ng/mL for C5a with an AUC of 0.7286 (p = 0.034). Conclusion: The development of sepsis is associated with early decreased TAFI and increased C5a levels after major trauma. Both elevated C5a and decreased TAFI may serve as promising predictive factors for the development of sepsis after polytrauma.

Levels of TAFI, TFPI and ADAMTS-13 in inflammatory bowel disease.

There is an increased tendency for thrombosis and thromboembolic complications in patients with inflammatory bowel disease (IBD). The aim of the present study was to determine the serum concentrations of thrombin-activatable fibrinolysis inhibitor (TAFI), tissue factor pathway inhibitor (TFPI) and a disintegrin and metalloproteinase with thrombospondin motif-13 (ADAMTS-13) in patients with IBD and to assess their possible role in the etiopathogenesis of the disease. Thirty-four patients with IBD (23 ulcerative colitis and 11 Crohn's disease) and 20 healthy controls were included in the present study. TAFI, TFPI, and ADAMTS-13 concentrations were determined by enzyme-linked immunosorbent assay. Mean TAFI, TFPI, and ADAMTS-13 concentrations in the patient group were 17.75 ng/ml, 72.10 ng/ml, and 14.90 U/l, respectively. In the control group, these values were 117.10 ng/ml, 300 ng/ml, and 191.55 U/l, respectively. TAFI, TFPI, and ADAMTS-13 values were significantly lower in the patient group than in the control group (all p<0.01). TAFI, TFPI, and ADAMTS-13 levels were significantly lower in the patient group. These findings indicate the presence of a clear, multifactorial imbalance in the coagulation-fibrinolytic system in the patient group. It is also possible that this imbalance in the coagulation and fibrinolytic system may play a role in the still unclear etiopathogenesis of the disease.

Prevention of Bleeding-Induced Vascular Abnormalities in the Hemophilic Mouse Joint By Increasing TAFI Levels or Inhibiting uPA Activity

Hemophilic arthropathy is characterized by vascular abnormalities in the joint soft tissue that may cause rebleeding and progressive joint destruction. Defective thrombin generation in hemophilia leads to impaired activation of Thrombin Activatable Fibrinolysis Inhibitor (TAFI). Activated TAFI (TAFIa) is a carboxypeptidase that cleaves C-terminal Lysine from fibrin or cell plasminogen receptors, thereby diminishing fibrinolysis or pericellular plasmin generation. In addition, TAFIa inactivates pro-inflammatory (i.e. C3a, C5a, bradykinin) and anti-angiogenic (i.e. VEGF-A, SDF-1α) substrates. We have recently shown that TAFI-gene deficiency causes vascular abnormalities in the mouse joints that phenocopy those in hemophilic arthropathy, either after joint bleeding or spontaneously with aging. Based on these mechanistic insights, we evaluated treatment with TAFI for its ability to mitigate bleeding-induced vascular abnormalities and associated bleeding susceptibility in the FVIII KO mouse joint. Knee joint bleeding in FVIII KO mice was induced by a subpatellar puncture injury that was standardized to cause severe bleeding as defined by a hematocrit (Hct) drop from 47 ± 2 % at baseline (BL) to 29 ± 11 % two days after injury. The vascular abnormalities formed in the hemophilic mouse joint 2 weeks after injury capture those in patients with hemophilic arthropathy: i.e. increased vascular permeability and abnormal vessel enlargement. Hydrodynamic gene delivery was used in mice for continuous treatment with human wild type TAFI (hTAFI-WT). To compensate for lower TAFI activation in hemophilia, two TAFI mutants were tested with extended enzymatic stability: hTAFI-CYQ (8-fold increased) and hTAFI-CIIYQ (180-fold increased). The TAFI constructs had comparable antigen expression profiles (reaching a similar level as in human plasma at day 2 and gradually returning to BL by week 2). Treatment efficacy to reduce vascular permeability, however, differed according to TAFIa's enzymatic stability in FVIII KO mice 2 weeks after injury. Without treatment, Evans blue-BSA extravasation in the joints was increased (1.8-fold compared to the contralateral joint; p= 0.0001 vs BL). hTAFI-WT overexpression initiated 2 days after injury had no effect, whereas hTAFI-CYQ and hTAFI-CIIYQ overexpression reduced the vascular permeability to 1.5-fold (n= 10; p= 0.10 vs untreated) and 1.3-fold (n= 8; p= 0.0032 vs untreated), respectively, indicating that hTAFI-CIIYQ was most effective. Histology at week 2 confirmed that hTAFI-CIIYQ reduced the number of excessively enlarged CD31+ vessels (with ≥ 20 µm diameter) by 3-fold (n= 6; p= 0.0011 vs untreated). To determine whether the protective effect of TAFI overexpression on bleeding-induced vascular abnormalities was associated with inhibition of the plasminogen activation system, continuous administration of tranexamic acid (TXA; 50 mg/ml in drinking water) or antibody-mediated blockage of uPA's proteolytic activity (mU1; 60 mg/kg; 2x weekly) was initiated in FVIII KO mice 2 days after injury. Histology at week 2 showed mU1, but not TXA, reduced vessel enlargement to the same extent as hTAFI-CIIYQ. Finally, the joint bleeding model was adopted to appreciate functional consequences of hypervascularity to changes in bleeding susceptibility. Aged TAFI KO mice (8 months) were used for their spontaneously (3-fold) increased number of enlarged vessels (≥ 40 µm diameter; p= 0.0076) in the joint that correlates with Power Doppler (PD) signal. A mild injury was optimized to cause minimal bleeding and showed that PD signals in joints of aged TAFI KO mice correlated negatively with the Hct 2 days after mild injury with injection of a FVIII inhibitor (r= -0.53, p= 0.024; n= 18), supporting hypervascularity as a major contributor to hemophilic joint bleeding. Furthermore, the same mild injury caused minimal bleeding in naive FVIII KO mice with low BL vascularity (median Hct 46 % vs 48 %; p= 0.81 vs BL; n= 7), but excessive bleeding in FVIII KO mice injured 2 weeks previously with vascular abnormalities (median Hct 38 %; p= 0.021 vs BL; n= 7). hTAFI-CIIYQ effectively reduced bleeding severity upon re-injury (median Hct 47 %; p&amp;gt; 0.99 vs BL; n=5). Our data provide an experimental link between vascular abnormalities and bleeding susceptibility in hemophilic joints and conclude that the vascular abnormalities were alleviated by increasing TAFI levels or inhibiting uPA activity. Disclosures von Drygalski: University of California San Diego: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Patents &amp; Royalties; UniQure, Bayer, Bioverativ/Sanofi, Pfizer, Novo Nordisk, Biomarin, Shire, CSL Behring: Consultancy; Hematherix Inc.: Membership on an entity's Board of Directors or advisory committees, Other: Founder. Mosnier:The Scripps Research Institute: Membership on an entity's Board of Directors or advisory committees, Patents &amp; Royalties.

Circulating Plasminogen Activator Inhibitor-1 (PAI-1) Is Reduced By In Vivo Thrombin Generation and Subsequent Formation of Activated Protein C (APC)

Background: APC has been suggested to contribute to a hyperfibrinolytic state in various acquired coagulation disorders, including coagulopathies induced by trauma and sepsis. However, in vivo evidence for the proposed underlying mechanism of proteolytic degradation of PAI-1 by APC remains inconclusive. Recently, we have shown increased APC generation in response to in vivo thrombin formation in carriers of the factor V Leiden mutation (FVL). In this approach of stimulated hemostasis activity pattern evaluation (SHAPE), in vivo thrombin formation was triggered by low-dose administration of recombinant activated factor VII (rFVIIa). Aim of the present study was to investigate the resulting effects on the fibrinolytic system. Methods: The study population consisted of 30 FVL carriers (thereof 13 with a history of venous thromboembolism) and 15 healthy controls. Blood samples were collected immediately before and during a period of 8 hours following injection of 15 µg/kg rFVIIa. Plasma levels of APC were quantified using an oligonucleotide-based enzyme capture assay (OECA). Other monitored parameters included plasma levels of prothrombin activation fragment 1+2 (F1+2), tissue-type plasminogen (t-PA), PAI-1, plasminogen, α2-antiplasmin, plasmin-α2-antiplasmin complexes (PAP), soluble fibrin monomers, d-dimer, and thrombin-activatable fibrinolysis inhibitor (TAFI). Results: At baseline, FVL carriers showed higher median levels of APC in comparison to the controls (1.39 vs. 0.86 pmol/L, P=.001), higher PAI-1 levels (30.1 vs. 15.3 ng/mL, P=.002) and lower plasminogen levels (94.8 vs. 110.4%, P=6·10-4). The other baseline parameters did not differ significantly. In both cohorts a comparable increase of F1+2 was observed after administration of rFVIIa, whereas APC increased more (P=.004) in FVL carriers (by 6.40 pmol/L) than in healthy controls (by 2.14 pmol/L), Concurrently, median PAI-1 levels decreased more (P=.007) in FVL carriers (by 19.8 pmol/L) than in healthy controls (by 8.0 pmol/L) (Figure 1). TAFI levels decreased temporarily, from 104.8 to 94.4% (P=.007) in FVL carriers and from 105.0 to 92.1% (P=2·10-4) in healthy controls. PAP levels increased from 164 to 209 ng/mL (P&lt;10-4) in FVL carriers and from 154 to 189 ng/mL (P=.023) in the control group. The extent of these changes did not differ between the two cohorts. D-dimer level increased only in FVL carriers, from 0.34 to 0.41 mg/L (P=.008). t-PA and the other parameters did not show significant changes after rFVIIa administration. Conclusion: Increased APC formation rates in FVL carriers were associated with a greater decline of PAI-1 levels in the absence of interfering changes in t-PA levels. These data provide further in vivo evidence that APC down-regulates PAI-1. Overall, the SHAPE approach utilized here does not induce a significant profibrinolytic response, even in patients with thrombophilia. Disclosures Reda: Grifols: Honoraria; Shire: Honoraria. Winterhagen:SOBI: Honoraria. Berens:Pfizer: Honoraria; Shire: Honoraria; Biotest: Honoraria; CSL Behring: Honoraria; Novo Nordisk: Honoraria; Baxter: Honoraria. Müller:NovoNordisk: Membership on an entity's Board of Directors or advisory committees; Bayer: Membership on an entity's Board of Directors or advisory committees; Siemens: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees. Oldenburg:CSL Behring: Consultancy, Research Funding, Speakers Bureau; NovoNordisk: Consultancy, Honoraria, Research Funding; Bayer: Consultancy, Research Funding, Speakers Bureau; Swedish Orphan Biovitrum: Consultancy, Speakers Bureau; Grifols: Consultancy, Speakers Bureau; Takeda (Shire): Consultancy, Research Funding, Speakers Bureau; Chugai: Consultancy, Speakers Bureau; Roche: Consultancy, Speakers Bureau; Biotest: Consultancy, Research Funding, Speakers Bureau; Octapharma: Consultancy, Research Funding, Speakers Bureau; Pfizer: Consultancy, Speakers Bureau. Rühl:CSL Behring: Honoraria; Bayer: Honoraria; Shire: Honoraria; SOBI: Honoraria; Sanofi Genzyme: Honoraria; Grifols: Honoraria.

Increased thrombin-activatable fibrinolysis inhibitor levels in patients with chronic rhinosinusitis with nasal polyps

Chronic rhinosinusitis (CRS) is a heterogeneous chronic inflammatory disease subdivided based on the presence or absence of nasal polyps (NPs). Histologic features of chronic rhinosinusitis with nasal polyps (CRSwNP) include inflammatory cell infiltration and excessive fibrin deposition in NPs. Thrombin-activatable fibrinolysis inhibitor (TAFI) is an enzyme that plays an antifibrinolytic role in the body. The significance of TAFI has been documented in patients with chronic inflammatory diseases, including chronic lung disease; however, it has not been evaluated in the pathogenesis of NPs. The objective of this study was to evaluate the potential role of TAFI in the pathogenesis of NPs. Nasal lavage fluid was collected from control subjects and patients with CRS. We measured levels of thrombin/anti-thrombin complex (TATc) and TAFI protein using an ELISA. TATc levels in nasal lavage fluid were significantly increased in patients with CRSwNP and patients with chronic rhinosinusitis without nasal polyps (CRSsNP) compared with control subjects, and TAFI levels in nasal lavage fluid were also significantly increased in patients with CRSwNP compared with those in control subjects and patients with CRSsNP. There was a significant correlation between TATc and TAFI levels in nasal lavage fluid. Interestingly, patients with CRS and asthma showed increased TATc and TAFI levels in nasal lavage fluid compared with those in patients with CRS without asthma, especially patients with CRSwNP. Increased TATc and TAFI levels in nasal passages of patients with CRSwNP might participate in fibrin deposition in NPs and might play a role in the pathogenesis of CRSwNP and asthma.

Recent Advances in the Understanding of Thrombosis.

Recent Advances in the Understanding of Thrombosis.

Effect of android fat distribution on thrombin-activatable fibrinolysis inhibitor levels in women with polycystic ovary syndrome.

Central obesity in women with polycystic ovary syndrome, assessed clinically and by dual‐energy X‐ray absorptiometry, may directly affect the hemostatic parameter thrombin‐activatable fibrinolysis inhibitor, suggesting a state of hypercoagulability.

KADAR PENGHAMBAT PENGURAIAN FIBRIN OLEH TROMBIN YANG TERGIATKAN (THROMBIN ACTIVATABLE FIBRINOLYSIS INHIBITOR) DI PASIEN KEGEMUKAN DENGAN DAN TANPA DIABETES MELLITUS

Obesity will cause the development of insulin resistance and endothelial dysfunction through fat metabolism hormones and cytokines products. The metabolic disorders is hallmarked by the decrease of insulin function and will cause disorders of coagulation andfibrinolysis. Thrombin-activatable fibrinolysis inhibitor (TAFI) is a newly found glycoprotein that has a role in the balance of coagulationand fibrinolytic process. The aim of this study was to analyze the TAFI level in obese subjects with and without diabetes mellitus. A crosssectional study was carried out by collecting data from TAFI level measurements at Prodia Clinical Laboratory Jakarta and Makassarduring January-March 2008. The data was analyzed using t test with SPSS version 12.0. From 80 samples that match with the criteria,was found in four (4) diabetes mellitus patients and 76 subjects without diabetes mellitus. The mean TAFI level in obese subjects withdiabetes mellitus was 7.67 ug/ml and in obese subjects without diabetes mellitus was 8.10 ug/ml. The result of t test showed p = 0.58.There was no significant difference between TAFI level in obese subjects with and without diabetes mellitus.

Platelet Drop and Fibrinolytic Shutdown in Patients With Sepsis.

Thrombocytopenia is the most common hemostatic disorder during sepsis and is associated with high mortality. We examined whether fibrinolytic changes precede incident thrombocytopenia and predict outcome in patients with severe sepsis. Nested study from the multicenter, randomized, controlled trial on the efficacy of albumin replacement in severe sepsis or septic shock (the Albumin Italian Outcome Sepsis trial). Forty ICUs in Italy. Three groups of patients were selected: 1) patients with platelet count less than or equal to 50 × 10/L at study entry (n = 85); 2) patients with baseline platelet count greater than or equal to 100 × 10/L who developed thrombocytopenia (≤ 50 × 10/L) within 28 days (n = 100); 3) patients with platelet count always more than or equal to 100 × 10/L (n = 95). Fibrinolytic variables, including fibrinolysis inhibitors and in vivo markers of plasmin generation, were measured on day 1. Patients with early thrombocytopenia (group 1) and those who developed it later (group 2) had similar illness severity and 90-day mortality, whereas patients without thrombocytopenia (group 3) had milder disease and lower mortality. Fibrinolysis was markedly (and similarly) depressed in groups 1 and 2 as compared with group 3. Major fibrinolytic changes included increased levels of plasminogen activator inhibitor 1 and extensive activation/consumption of thrombin activatable fibrinolysis inhibitor. Most fibrinolytic variables were significantly associated with mortality in univariate models. However, only thrombin activatable fibrinolysis inhibitor level and in vivo markers of fibrinolysis activation, namely plasmin-antiplasmin complex, and D-dimer, were independently associated with mortality after adjustment for Simplified Acute Physiology Score-II score, sex, and platelet count. Furthermore, the coexistence of impaired fibrinolysis and low platelets was associated with an even greater mortality. Impaired fibrinolysis, mainly driven by plasminogen activator inhibitor-1 increase and thrombin activatable fibrinolysis inhibitor activation, is an early manifestation of sepsis and may precede the development of thrombocytopenia. Thrombin activatable fibrinolysis inhibitor level, in particular, proved to be an independent predictor of mortality, which may improve risk stratification of patients with severe sepsis.

Thrombin‐activatable fibrinolysis inhibitor in human abdominal aortic aneurysm disease

Background Intra-luminal thrombosis is a key factor in growth of abdominal aortic aneurysms (AAAs). Patients with AAA form dense clots that are resistant to fibrinolysis. Thrombin-activatable fibrinolysis inhibitor (TAFI) has been shown to influence AAA development in murine models. Objective The aim of this study is to characterize the role of TAFI in human AAA. Methods Plasma levels of TAFI, TAFI activation peptide (TAFI-AP), activated/inactivated TAFI (TAFIa/ai) and plasmin-α2-antiplasmin complex were measured by ELISAs in patients with AAA (n=202) and controls (n=188). Results TAFIa/ai and TAFI-AP levels were higher in patients than controls (median [IQR], 20.3 [14.6-32.8]ngmL-1 vs. 14.2 [11.2-19.3]ngmL-1 and 355.0 [232.4-528.1]ngmL-1 vs. 248.6 [197.1-328.1]ngmL-1 ). TAFIa/ai was positively correlated with TAFI-AP (r=0.164). Intact TAFI levels were not different between patients and controls (13.4 [11.2-16.1]μgmL-1 vs. 12.8 [10.6-15.4]μgmL-1 ). Plasmin-α2-antiplasmin was higher in AAA patients than controls (690.0 [489.1-924.3]ngmL-1 vs. 480.7 [392.6-555.3]ngmL-1 ). Conclusions The increase in TAFIa/ai and TAFI-AP suggests an increased TAFI activation in patients with AAA. Prospective studies are required to further elucidate the role of TAFI and fibrinolysis in AAA pathogenesis.

Reduced thrombin activatable fibrinolysis inhibitor and enhanced proinflammatory cytokines in acute coronary syndrome

ObjectiveA study was made of the changes in the serum levels of thrombin activatable fibrinolysis inhibitor (TAFI), proinflammatory cytokines and acute phase proteins in the acute stage of acute coronary syndrome (ACS), in order to explore the possibility of using TAFI as a biomarker for ACS risk assessment. MethodsA total of 211 patients with ACS were enrolled, and healthy subjects were used as controls. Blood samples were taken within 24h after admission. Serum TAFI levels were determined by immunoturbidimetry. Serum levels of interleukin-1β (IL-1β), interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) were determined by enzyme linked immunosorbent assay (ELISA). Procalcitonin (PCT) and C-reactive protein (CRP) levels were measured by gold-immunochromatographic assay. ResultsSerum TAFI levels in ACS patients were significantly decreased versus the controls. The IL-1β, IL-6, TNF-α, PCT and CRP levels were markedly higher in the ACS patients than in the controls. Correlation analysis revealed a strong negative correlation between TAFI concentration and the IL-1β, IL-6, TNF-а, PCT and CRP levels in ACS patients and in controls. Multivariate logistic regression analysis suggested decreased serum TAFI to be an independent risk factor for ACS (OR 9.459; 95% CI 2.306–38.793; P=0.002). The area under the receiver operating characteristic (ROC) curve for TAFI was 0.872 (95% CI 0.787–0.909; P<0.001). The optimum TAFI cutoff point for the prediction of ACS was 24μg/ml, with a sensitivity of 75.83% and a specificity of 72.57%. ConclusionThese findings suggest that TAFI can be useful as a potential biomarker for ACS risk assessment.