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Neuroinflammation is assumed as the critical pathophysiologic mechanism of white matter lesions (WMLs), and infiltrated peripheral monocyte-derived macrophages are implicated in the development of neuroinflammation. This study sought to explore the blood molecules that promote the migration of peripheral monocytes to the sites of WMLs. The serum protein expression profiles of patients and Sprague–Dawley rat models with WMLs were detected by data-independent acquisition (DIA) proteomics technique. Compared with corresponding control groups, we acquired 62 and 41 differentially expressed proteins (DEPs) in the serum of patients and model rats with WMLs respectively. Bioinformatics investigations demonstrated that these DEPs were linked to various Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways and Gene Ontology (GO) terms involved in neuroinflammation. Afterward, we identified thrombin-activatable fibrinolysis inhibitor (TAFI) as a shared and overexpressed protein in clinical and animal serum samples, which was further verified by enzyme-linked immunosorbent assay. Additionally, an upregulation of TAFI was also observed in the white matter of rat models, and the inhibition of TAFI impeded the migration of peripheral monocytes to the area of WMLs. In vitro experiments suggested that TAFI could enhance the migration ability of RAW264.7 cells and increase the expression of Ccr2. Our study demonstrates that neuroinflammatory signals can be detected in the peripheral blood of WMLs patients and model rats. TAFI may serve as a potential protein that promotes the migration of peripheral monocytes to WMLs regions, thereby providing a novel molecular target for further investigation into the interaction between the central and peripheral immune systems. [ABSTRACT FROM AUTHOR]

Background: Inflammatory and hemostatic disorders in diabetic microangiopathy (DMA) can be linked to thrombin-activatable fibrinolysis inhibitor (TAFI) and its own gene polymorphisms. Thus, the study aimed to investigate the associations of plasma TAFI and gene polymorphisms with DMA in Chinese patients with type 2 diabetes (T2D). Methods: Plasma TAFI of 223 patients with T2D was measured, and the genotypes and alleles of the 1040C/T, 438G/A, and 505G/A polymorphisms of the TAFI gene were analyzed. A ROC curve was constructed to evaluate the identifying power of TAFI between patients with T2D and DMA, and logistic regression analysis was used to observe the correlation of plasma TAFI and gene polymorphisms with the risk for DMA. Results: Plasma TAFI was higher in patients with DMA than in patients with only T2D (p < 0.05). TAFI exhibited the largest area under ROC in identifying diabetic kidney disease (DKD) from only T2D (0.763, 95% CI [0.674-0.853], p < 0.01), and adjusted multivariate analysis showed a high odds ratio (OR: 15.72, 95% CI [4.573-53.987], p < 0.001) for DKD. Higher frequencies of the CT genotype and T allele of the 1040C/T polymorphism were found in DKD compared with only T2D (respectively p < 0.05), and the CT genotype exhibited a high OR (1.623, 95% CI [1.173-2.710], p < 0.05) for DKD. DKD patients with the CT genotype had higher plasma TAFI levels, while T2D and DKD patients with CC/TT genotypes had lower plasma TAFI levels. Conclusion: Plasma TAFI and the CT genotype and T allele of the 1040C/T polymorphism are independent risk factors for DKD in Chinese T2D patients. [ABSTRACT FROM AUTHOR]

Thrombin-Activatable Fibrinolysis Inhibitor (TAFI) is a carboxypeptidase B-like proenzyme encoded by the CPB2 gene. After thrombin activation, TAFI downregulates fibrinolysis, thus linking the latter with coagulation. TAFI has been shown to play a role in venous and arterial thrombotic diseases, yet, data regarding the molecular mechanisms underlying its function have been conflicting. In this study, we focused on the prediction and functional enrichment analysis (FEA) of the TAFI interaction network and the microRNAs (miRNAs) targeting the members of this network in an attempt to identify novel components and pathways of TAFI-related thrombosis. To this end, we used nine bioinformatics software tools. We found that the TAFI interactome consists of 28 unique genes mainly involved in hemostasis. Twenty-four miRNAs were predicted to target these genes. Co-annotation analysis of the predicted interactors with respect to Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways and transcription factors (TFs) pointed to the complement and coagulation cascades as well as neutrophil extracellular trap formation. Cancer, stroke, and intracranial aneurysm were among the top 20 significant diseases related to the identified miRNAs. We reason that the predicted biomolecules should be further studied in the context of TAFI-related thrombosis. [ABSTRACT FROM AUTHOR]

[Display omitted] [ABSTRACT FROM AUTHOR]

Carboxypeptidase U (CPU, TAFIa, CPB2) is a potent attenuator of fibrinolysis that is mainly synthesized by the liver as its inactive precursor proCPU. Aside from its antifibrinolytic properties, evidence exists that CPU can modulate inflammation, thereby regulating communication between coagulation and inflammation. Monocytes and macrophages play a central role in inflammation and interact with coagulation mechanisms resulting in thrombus formation. The involvement of CPU and monocytes/macrophages in inflammation and thrombus formation, and a recent hypothesis that proCPU is expressed in monocytes/macrophages, prompted us to investigate human monocytes and macrophages as a potential source of proCPU. CPB2 mRNA expression and the presence of proCPU/CPU protein were studied in THP-1, PMA-stimulated THP-1 cells and primary human monocytes, M-CSF-, IFN-γ/LPS-, and IL-4-stimulated-macrophages by RT-qPCR, Western blotting, enzyme activity measurements, and immunocytochemistry. CPB2 mRNA and proCPU protein were detected in THP-1 and PMA-stimulated THP-1 cells as well as in primary monocytes and macrophages. Moreover, CPU was detected in the cell medium of all investigated cell types and it was demonstrated that proCPU can be activated into functionally active CPU in the in vitro cell culture environment. Comparison of CPB2 mRNA expression and proCPU concentrations in the cell medium between the different cell types provided evidence that CPB2 mRNA expression and proCPU secretion in monocytes and macrophages is related to the degree to which these cells are differentiated. Our results indicate that primary monocytes and macrophages express proCPU. This sheds new light on monocytes and macrophages as local proCPU sources. [ABSTRACT FROM AUTHOR]

Abstract Background Administration of recombinant human soluble thrombomodulin (rTM) is often used in Japan to treat septic disseminated intravascular coagulation (DIC). Thrombin-activatable fibrinolysis inhibitor (TAFI) is a fibrinolysis inhibitor activated by the thrombin-thrombomodulin complex, however, it is unknown whether circulating activated TAFI is increased after rTM administration in patients with DIC. Furthermore, the relationship between TAFI activation and the prognosis of septic DIC is not defined yet. Case presentation We report a series of 8 patient’s TAFI activation with septic DIC treated by rTM. We sought to investigate the effect of rTM on TAFI activation and the association of plasma activated TAFI (TAFIa/ai) levels with the prognosis of septic DIC. Using plasma samples from clinical studies conducted from May 2016–March 2017 on eight patients with septic DIC at Kagoshima University Hospital, we measured plasma levels of total TAFI, TAFIa/ai, thrombin-antithrombin complex (TAT), prothrombin fragment 1 + 2 (F1 + 2), soluble fibrin (SF), antithrombin (AT), protein C (PC), protein S (PS), and plasminogen activator inhibitor-1 (PAI-1) before and after intravenous rTM administration. Then, we evaluated the relationship of these marker levels to prognosis. The thrombin-rTM complex activated TAFI in vitro in plasma from a healthy volunteer. However, TAFIa/ai levels did not significantly increase over baseline in the septic DIC patients after intravenous rTM administration. Baseline TAFIa/ai levels in non-survivors were significantly higher than those in survivors. Conclusions Plasma TAFIa/ai did not increase with rTM administration. Elevated baseline TAFIa/ai concentration may be a negative prognostic indicator in septic DIC. Larger studies are needed to confirm the in vivo effect of rTM on TAFI activation.

The antifibrinolytic enzyme carboxypeptidase U (CPU, TAFIa, CPB2) is an appealing target for the treatment of acute ischemic stroke (AIS). Increased insights in CPU activation and inactivation during thrombolysis (rtPA) with or without endovascular thrombectomy (EVT) are required to develop CPU inhibitors as profibrinolytic agents with optimal benefits/risks. Therefore, CPU kinetics during ischemic stroke treatment were evaluated. AIS patients with documented cerebral artery occlusion receiving rtPA (N = 20) or rtPA + EVT (N = 16) were included. CPU activation during thrombolysis was measured by an ultrasensitive HPLC-based CPU activity method and by an ELISA measuring both CPU and inactivated CPU (CPU + CPUi). Intravenous blood samples were collected at admission and throughout the first 24 h. Additional in situ blood samples were collected in the rtPA + EVT cohort proximal from the thrombus. The NIHSS score was determined at baseline and 24 h. CPU activity and CPU + CPUi levels increased upon rtPA administration and reached peak values at the end of thrombolysis (1 h). High inter-individual variability was observed in both groups. CPU activity decreased rapidly within 3 h, while CPU + CPUi levels were still elevated at 7 h. CPU activity or CPU + CPUi levels were similar in in situ and peripheral samples. No correlation between CPU or CPU + CPUi and NIHSS or thrombus localization was found. The CPU system was rapidly activated and deactivated following thrombolysis and thrombectomy in stroke patients, suggesting that a CPU inhibitor would have to be administered during rtPA infusion and over the next few hours. The high CPU generation variability suggests that some patients may not respond to the treatment. EudraCT number 2017-002760-41. [ABSTRACT FROM AUTHOR]

Coronavirus disease 2019 (COVID-19) is a viral lower respiratory tract infection caused by the highly transmissible and pathogenic SARS-CoV-2 (severe acute respiratory-syndrome coronavirus-2). Besides respiratory failure, systemic thromboembolic complications are frequent in COVID-19 patients and suggested to be the result of a dysregulation of the hemostatic balance. Although several markers of coagulation and fibrinolysis have been studied extensively, little is known about the effect of SARS-CoV-2 infection on the potent antifibrinolytic enzyme carboxypeptidase U (CPU). Blood was collected longitudinally from 56 hospitalized COVID-19 patients and 32 healthy controls. Procarboxypeptidase U (proCPU) levels and total active and inactivated CPU (CPU+CPUi) antigen levels were measured. At study inclusion (shortly after hospital admission), proCPU levels were significantly lower and CPU+CPUi antigen levels significantly higher in COVID-19 patients compared to controls. Both proCPU and CPU+CPUi antigen levels showed a subsequent progressive increase in these patients. Hereafter, proCPU levels decreased and patients were, at discharge, comparable to the controls. CPU+CPUi antigen levels at discharge were still higher compared to controls. Baseline CPU+CPUi antigen levels (shortly after hospital admission) correlated with disease severity and the duration of hospitalization. In conclusion, CPU generation with concomitant proCPU consumption during early SARS-CoV-2 infection will (at least partly) contribute to the hypofibrinolytic state observed in COVID-19 patients, thus enlarging their risk for thrombosis. Moreover, given the association between CPU+CPUi antigen levels and both disease severity and duration of hospitalization, this parameter may be a potential biomarker with prognostic value in SARS-CoV-2 infection. [ABSTRACT FROM AUTHOR]

Background: Administration of recombinant human soluble thrombomodulin (rTM) is often used in Japan to treat septic disseminated intravascular coagulation (DIC). Thrombin-activatable fibrinolysis inhibitor (TAFI) is a fibrinolysis inhibitor activated by the thrombin-thrombomodulin complex, however, it is unknown whether circulating activated TAFI is increased after rTM administration in patients with DIC. Furthermore, the relationship between TAFI activation and the prognosis of septic DIC is not defined yet. Case presentation: We report a series of 8 patient's TAFI activation with septic DIC treated by rTM. We sought to investigate the effect of rTM on TAFI activation and the association of plasma activated TAFI (TAFIa/ai) levels with the prognosis of septic DIC. Using plasma samples from clinical studies conducted from May 2016–March 2017 on eight patients with septic DIC at Kagoshima University Hospital, we measured plasma levels of total TAFI, TAFIa/ai, thrombin-antithrombin complex (TAT), prothrombin fragment 1 + 2 (F1 + 2), soluble fibrin (SF), antithrombin (AT), protein C (PC), protein S (PS), and plasminogen activator inhibitor-1 (PAI-1) before and after intravenous rTM administration. Then, we evaluated the relationship of these marker levels to prognosis. The thrombin-rTM complex activated TAFI in vitro in plasma from a healthy volunteer. However, TAFIa/ai levels did not significantly increase over baseline in the septic DIC patients after intravenous rTM administration. Baseline TAFIa/ai levels in non-survivors were significantly higher than those in survivors. Conclusions: Plasma TAFIa/ai did not increase with rTM administration. Elevated baseline TAFIa/ai concentration may be a negative prognostic indicator in septic DIC. Larger studies are needed to confirm the in vivo effect of rTM on TAFI activation. [ABSTRACT FROM AUTHOR]

Objective: Inflammation and migration of leukocytes to the brain parenchyma play a role in atherosclerosis and cerebral ischemic stroke. Migration occurs with the help of adhesion molecules on the surface of cerebral endothelial cells and leukocytes. P-selectin, an adhesion molecule, is present on the platelet and endothelial surface and allows leukocytes to loosely adhere to the endothelium, and its increase has been shown in acute ischemic stroke (AIS). Thrombin-activatable fibrinolysis inhibitor (TAFI) is a procarboxypeptidase molecule that can be another marker of AIS, which has been shown to increase the risk of thromboembolism and stroke 6-fold. Intima-media thickness (IMT) is thought to be associated with atherosclerotic diseases in carotid ultrasonography (USG) and increased risk of ischemic stroke has been found to be associated with increased carotid IMT. In this study, we investigated the relationship between P-selectin and TAFI levels, which have been shown to be effective for AIS via carotid IMT, and is considered significant for atherosclerosis. Materials and Methods: Forty patients with AIS and 22 healthy subjects were included in the study. In both groups, serum P-selectin and TAFI levels were studied at the time of presentation, and on day 7, day 14, and at one month; carotid IMT and stenosis rates were measured by Doppler USG. P-selectin and TAFI levels were compared with carotid IMT in both groups. Results: There was no significant difference between P-selectin levels and carotid IMT between the groups; TAFI levels were significantly higher in the patient group and were correlated with carotid IMT in both groups. Conclusion: TAFI increase has been suggested to be a marker of early atherosclerosis in asymptomatic atherosclerosis and ischemic stroke. A positive correlation between TAFI levels and carotid IMT and stenosis rates have been reported; however, the positive correlation between increased P-selectin levels in AIS and carotid IMT was not detected in our study.

Objectives/Hypothesis: 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). Study Design: Retrospective cohort study and laboratory study. Methods: 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. Results: 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. Conclusions: 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. Level of Evidence: 4 Laryngoscope, 131:2413–2420, 2021 [ABSTRACT FROM AUTHOR]

Sepsis is a life-threatening condition which develops as a dysregulated immune response in the face of infection and which is associated with profound hemostatic disturbances and in the most extreme cases disseminated intravascular coagulation (DIC). In addition, the fibrinolytic system is subject to alterations during infection and sepsis, and impaired fibrinolysis is currently considered a key player in sepsis-related microthrombus formation and DIC. However, we still lack reliable biomarkers to assess fibrinolysis in the clinical setting. Furthermore, drugs targeting the fibrinolytic system have potential value in sepsis patients with severe fibrinolytic disturbances, but these are still being tested in the preclinical stage. The present review provides an overview of key fibrinolytic changes in sepsis, reviews the current literature on potential laboratory markers of altered fibrinolysis in adult sepsis patients, and discusses future perspectives for diagnosis and treatment of fibrinolytic disturbances in sepsis patients. [ABSTRACT FROM AUTHOR]

The balance between coagulation and fibrinolysis is essential for a successful pregnancy. This study aimed to explore the genetic variant of +1040C/T in the coding region of thrombin-activatable fibrinolysis inhibitor (TAFI) gene in women with recurrent spontaneous abortion (RSA) and in unrelated healthy controls and to investigate the possible association between TAFI +1040C/T polymorphism and RSA. Peripheral blood samples were collected from 137 Chinese patients with RSA and 103 unrelated healthy Chinese controls. The TAFI +1040C/T polymorphism was analyzed using SNaPshot SNP typing after DNA extraction. The frequency of the C allele was lower in RSA patients compared with the controls (0.78 vs 0.84). A subanalysis of the TAFI +1040C/T polymorphism in the 2 populations of RSA women (groups 2RSA and >2RSA) showed that the +1040CT genotype was significantly higher and the +1040CC genotype was significantly lower than from that found in controls. The allele +1040C was associated with a reduced risk of RSA in both group 2RSA (OR = 0.418, 95%CI, 0.255-0.685) and group >2RSA (OR = 0.473, 95%CI, 0.274-0.819) compared with controls. Our data indicate a protective role for TAFI +1040C allele against RSA, and may be associated with the genetic susceptibility of RSA. [ABSTRACT FROM AUTHOR]

Introduction: Plasma‐derived FVIII/VWF complex was reported to be less sensitive to inhibitors than FVIII preparations devoid of VWF. Aim: To compare the efficacy of FVIII/VWF complex (Fanhdi) and five different VWF‐free FVIII preparations in restoring thrombin generation and activation of thrombin‐activatable fibrinolysis inhibitor (TAFI) in haemophilic plasma, with and without inhibitor, and in cell‐based models. Methods: Experiments were performed in haemophilic plasma supplemented with inhibitory IgG or in plasma samples obtained from haemophilia A patients without (n = 11) and with inhibitor (n = 12). Thrombin generation was evaluated by calibrated automated thrombography (CAT) under standard conditions, in the presence of activated protein C (APC) or thrombomodulin (TM), and in cell‐based models including endothelial cells, either alone or in combination with platelets or tissue factor‐expressing blood mononuclear cells. The kinetics of TAFI activation was determined by a two‐stage functional assay in the absence and in the presence of APC. Results: In haemophilic plasma without inhibitor, Fanhdi enhanced thrombin generation and TAFI activation as well as recombinant (2nd‐4th generation) and plasma‐derived FVIII preparations devoid of VWF. On the contrary, in plasma with inhibitor, Fanhdi displayed a greater ability to restore thrombin generation and TAFI activation under all tested conditions. Notably, in cell‐based models including endothelial cells, Fanhdi proved more efficient than all other preparations in improving thrombin generation even in the absence of inhibitor. Conclusion: The greater pro‐haemostatic activity of FVIII/VWF complex, either in haemophilic plasma with inhibitor or in the presence of endothelial cells, may offer therapeutic advantages. [ABSTRACT FROM AUTHOR]

Objective: Inflammation and migration of leukocytes to the brain parenchyma play a role in atherosclerosis and cerebral ischemic stroke. Migration occurs with the help of adhesion molecules on the surface of cerebral endothelial cells and leukocytes. P-selectin, an adhesion molecule, is present on the platelet and endothelial surface and allows leukocytes to loosely adhere to the endothelium, and its increase has been shown in acute ischemic stroke (AIS). Thrombin-activatable fibrinolysis inhibitor (TAFI) is a procarboxypeptidase molecule that can be another marker of AIS, which has been shown to increase the risk of thromboembolism and stroke 6-fold. Intima-media thickness (IMT) is thought to be associated with atherosclerotic diseases in carotid ultrasonography (USG) and increased risk of ischemic stroke has been found to be associated with increased carotid IMT. In this study, we investigated the relationship between P-selectin and TAFI levels, which have been shown to be effective for AIS via carotid IMT, and is considered significant for atherosclerosis. Materials and Methods: Forty patients with AIS and 22 healthy subjects were included in the study. In both groups, serum P-selectin and TAFI levels were studied at the time of presentation, and on day 7, day 14, and at one month; carotid IMT and stenosis rates were measured by Doppler USG. P-selectin and TAFI levels were compared with carotid IMT in both groups. Results: There was no significant difference between P-selectin levels and carotid IMT between the groups; TAFI levels were significantly higher in the patient group and were correlated with carotid IMT in both groups. Conclusion: TAFI increase has been suggested to be a marker of early atherosclerosis in asymptomatic atherosclerosis and ischemic stroke. A positive correlation between TAFI levels and carotid IMT and stenosis rates have been reported; however, the positive correlation between increased P-selectin levels in AIS and carotid IMT was not detected in our study. [ABSTRACT FROM AUTHOR]

Fibrinolysis is regulated by the thrombin/thrombin-activatable fibrinolysis inhibitor (TAFI) system. Thus, anticoagulants and inhibitors of TAFI are expected to accelerate fibrinolysis. The combined effects of an anticoagulant and a TAFIa inhibitor on fibrinolysis remain unknown. The aim of this study was to evaluate the combined effect of edoxaban, an oral direct factor Xa (FXa) inhibitor, and a TAFIa inhibitor, potato tuber carboxypeptidase inhibitor (PCI) on tissue-type plasminogen activator (t-PA)-induced clot lysis in human plasma in vitro. Pooled human plasma (containing 180 ng/mL t-PA and 0.1 nM thrombomodulin) was mixed with edoxaban and/or PCI. Clot formation was induced by 2.5 pM tissue factor and 4 µM phospholipids and clot lysis time was examined. Plasma plasmin-α2 antiplasmin complex (PAP) concentration was measured as a marker of plasmin generation. Edoxaban or PCI alone significantly shortened the t-PA-induced clot lysis time and plasma PAP concentration. The combination of these compounds significantly accelerated the clot lysis compared with the inhibitors alone. Addition of PCI (0.3, 1, and 3 μg/mL) to 75 ng/mL edoxaban increased plasma PAP concentration compared with edoxaban alone; however, compared with PCI alone only the combination of 0.3 μg/mL PCI + 75 ng/mL edoxaban showed the significant increase in PAP concentration. Concomitant use of an oral direct FXa inhibitor, edoxaban, and a TAFIa inhibitor, PCI, significantly accelerate fibrinolysis via enhancement of plasmin generation. These results suggest that the combination of edoxaban and a TAFIa inhibitor might be beneficial for the treatment of thromboembolic diseases. [ABSTRACT FROM AUTHOR]

DS‐1040, a low‐molecular‐weight imidazole derivative, inhibits the enzymatic activity of thrombin‐activatable fibrinolysis inhibitor (TAFIa), enhancing endogenous tissue plasminogen activator–triggered fibrinolysis. This first‐in‐human, randomized, placebo‐controlled, phase 1 study evaluated the safety, pharmacokinetics (PK), and pharmacodynamics (PD) of an oral formulation of DS‐1040. Healthy adults (aged 20‐45 years; N = 56) were randomized 3:1 to receive DS‐1040 orally administered as single ascending doses (50, 100, 200, or 400 mg) or placebo, or DS‐1040 multiple ascending doses (100 mg once daily, 200 mg once daily, or 150 mg twice daily) or placebo for 14 days. Safety, PK, and PD parameters were assessed. All doses of DS‐1040 were well tolerated; no serious/severe adverse events (AEs) or discontinuations due to AEs occurred. DS‐1040 had no effect on coagulation parameters, and no treatment‐related trends in the bleeding time were observed. DS‐1040 exposure (peak concentration and area under the concentration‐time curve) increased in a dose‐proportional manner across the single‐dose range. With multiple doses, steady state was achieved by day 7 with minimal accumulation (mean accumulation ratio 1.15‐1.25), and the PK was time‐independent. After 72 hours, approximately 10% of the DS‐1040 400‐mg single dose was recovered in urine as intact parent drug. The mean terminal half‐life ranged from 17.2 to 24.9 hours, which was similar to previous intravenous administration data. Dose‐dependent inhibition of total TAFIa activity was observed following single and multiple doses of oral DS‐1040. The safety and PK/PD profiles of oral DS‐1040 in healthy subjects support further clinical development. [ABSTRACT FROM AUTHOR]

A direct oral anticoagulant, edoxaban, is as effective as vitamin K antagonists for the treatment of venous thromboembolism (VTE). However, the mechanism underlying the treatment effect on VTE remains to be determined. The aims of this study were to evaluate the effect of edoxaban on tissue plasminogen activator (t-PA)-induced clot lysis in human plasma and to determine the roles of plasmin and thrombin-activatable fibrinolysis inhibitor (TAFI) in the profibrinolytic effect by edoxaban. Pooled human normal plasma or TAFI-deficient plasma (containing 180 ng/mL t-PA and 0.1 nM thrombomodulin) was mixed with edoxaban or an activated TAFI inhibitor, potato tuber carboxypeptidase inhibitor (PCI). Clot was induced by adding tissue factor and phospholipids. Clot lysis time and plasma plasmin-α2 antiplasmin complex (PAP) concentration were determined. Clot structure was imaged with a scanning electron microscope. In normal plasma, edoxaban at clinically relevant concentrations (75, 150, and 300 ng/mL) and PCI significantly shortened clot lysis time. PCI increased PAP concentration and a correlation between PAP concentration and percent of clot lysis was observed. Edoxaban also dose-dependently elevated PAP concentration. In TAFI-deficient plasma, the effects of edoxaban and PCI on clot lysis and PAP concentration were markedly diminished as compared with normal plasma. Fibrin fibers were thinner in clots formed in the presence of edoxaban. In conclusion, edoxaban at clinically relevant concentrations accelerates t-PA-induced fibrinolysis via increasing plasmin generation in human plasma. The effects of edoxaban is mainly dependent on TAFI. The profibrinolytic effect of edoxaban might contribute to the efficacy for the treatment of VTE. [ABSTRACT FROM AUTHOR]

Aim: As angiogenesis is an essential step for chorionic villi formation. Vascular endothelial growth factor (VEGF) is essential for endothelial cell proliferation. Endothelial nitric oxide synthase (eNOS) is a powerful playmaker in hypoxia‐induced angiogenesis. Thrombin‐activatable fibrinolysis inhibitor (TAFI) regulates both fibrinolysis and inflammation. Genetic alterations of these factors may lead to recurrent spontaneous abortion (RSA). We aimed to investigate the combined genetic variants of VEGF G‐1154A and two eNOS genetic variants: T‐786C promoter region and intron 4 variable number of tandom repeats in addition to TAFI C‐1040T among RSA patients. Methods: The study included 50 patients with RSA and 50 healthy controls. Polymerase chain reaction and restriction fragment length polymorphism were used for genotyping. Results: Both genetic alterations of eNOS confirmed at least a sixfold increase of RSA risk. Interestingly, they were associated with TAFI C‐1040Tgenetic variant in 21 patients, eight of them had both studied eNOS genetic alterations and TAFI C‐1040Tgenetic variant, while each eNOS genetic variant associated with TAFI C‐1040Tconfirmed an almost one and half fold increase risk of RSA. Conclusion: These findings highlighted the role of eNOS and nitric oxide metabolism in RSA and opened the gate to investigate the interaction of vasoconstrictive and fibrinolytic inhibitor systems. [ABSTRACT FROM AUTHOR]

Background: Inflammatory and hemostatic disorders in diabetic microangiopathy (DMA) can be linked to thrombin-activatable fibrinolysis inhibitor (TAFI) and its own gene polymorphisms. Thus, the study aimed to investigate the associations of plasma TAFI and gene polymorphisms with DMA in Chinese patients with type 2 diabetes (T2D)., Methods: Plasma TAFI of 223 patients with T2D was measured, and the genotypes and alleles of the 1040C/T, 438G/A, and 505G/A polymorphisms of the TAFI gene were analyzed. A ROC curve was constructed to evaluate the identifying power of TAFI between patients with T2D and DMA, and logistic regression analysis was used to observe the correlation of plasma TAFI and gene polymorphisms with the risk for DMA., Results: Plasma TAFI was higher in patients with DMA than in patients with only T2D ( p < 0.05). TAFI exhibited the largest area under ROC in identifying diabetic kidney disease (DKD) from only T2D (0.763, 95% CI [0.674-0.853], p < 0.01), and adjusted multivariate analysis showed a high odds ratio (OR: 15.72, 95% CI [4.573-53.987], p < 0.001) for DKD. Higher frequencies of the CT genotype and T allele of the 1040C/T polymorphism were found in DKD compared with only T2D (respectively p < 0.05), and the CT genotype exhibited a high OR (1.623, 95% CI [1.173-2.710], p < 0.05) for DKD. DKD patients with the CT genotype had higher plasma TAFI levels, while T2D and DKD patients with CC/TT genotypes had lower plasma TAFI levels., Conclusion: Plasma TAFI and the CT genotype and T allele of the 1040C/T polymorphism are independent risk factors for DKD in Chinese T2D patients., Competing Interests: The authors declare that they have no competing interests., (© 2023 Huang et al.)

Background: The optimal treatment of intermediate-risk pulmonary embolism (PE) in hemodynamically stable patients remains unknown. Fibrinolytics reduce the risk of hemodynamic deterioration but increase bleeding risk. DS-1040, an inhibitor of thrombin-activatable fibrinolysis inhibitor, enhanced endogenous fibrinolytic activity without increasing bleeding risk in preclinical studies., Objectives: To evaluate the tolerability and explore the efficacy of DS-1040 in patients with acute PE., Methods: In this multicenter, randomized, double-blind, placebo-controlled study, ascending doses of intravenous DS-1040 (20-80 mg) or placebo were added to enoxaparin (1 mg/kg twice daily) in patients with intermediate-risk PE. The primary endpoint was the number of patients with major or clinically relevant nonmajor bleeding. The percentage change in thrombus volume and right-to-left ventricular dimensions, assessed using quantitative computed tomography pulmonary angiography, at baseline and after 12 to 72 hours were used to explore the efficacy of DS-1040., Results: Of 125 patients with all available data, 38 were randomized to placebo and 87 to DS-1040. The primary endpoint occurred in 1 patient in the placebo group (2.6%) and 4 patients who received DS-1040 (4.6%). One subject experienced major bleeding (DS-1040 80 mg group); no fatal or intracranial bleeding occurred. Thrombus volume was 25% to 45% lower after infusion, with no differences between the DS-1040 and placebo groups. There was no difference in the change from baseline right-to-left ventricular dimensions between the DS-1040 and placebo groups., Conclusion: In patients with acute PE, adding DS-1040 to standard anticoagulation was not associated with an increase in bleeding but did not improve thrombus resolution or right ventricular dilation., (Copyright © 2023 International Society on Thrombosis and Haemostasis. All rights reserved.)

Abstract Introduction Coronary stenosis is a consequence of atherosclerotic plaque progression that is associated with impaired fibrinolysis. Thrombin-activatable fibrinolysis inhibitor (TAFI) and plasminogen activator inhibitor 1 (PAI-1) are fibrinolysis inhibitors whose levels are influenced by acquired conditions and by polymorphisms. This study therefore aimed to investigate the association of TAFI and PAI-1 gene polymorphisms with severity of coronary stenosis in subjects with stable coronary artery disease (CAD). Materials and methods A total of 327 subjects suspected with CAD who underwent a coronary angiogram were recruited. Gensini score was applied to stratify the severity of coronary stenosis. Based on the Gensini score, the subjects were categorized into low-medium (<20) or high (≥20) groups. The study polymorphisms included TAFI Ala147Thr (505G/A), Thr325Ile (1040C/T), +1542C/G, +1583T/A and PAI-1 -675 4G/5G. Most polymorphisms were genotyped by allele-specific polymerase chain reaction, except for TAFI Thr325Ile that was genotyped by polymerase chain reaction-restriction fragment length polymorphism. Results A significant increase in the Gensini score was found in TAFI 505A and +1583A allele carriers. Binary regression analysis revealed the independent association of the TAFI 505G/A and +1583T/A polymorphisms with a high Gensini score [adjusted OR = 1.67 (95% CI: 1.03, 2.73) and 1.69 (95% CI: 1.04, 2.76), respectively]. Neither the homozygous PAI-1 -675 4G/4G nor the heterozygous 4G/5G was associated with a high Gensini score. Conclusions The results indicated the contribution of TAFI polymorphisms to atherosclerosis progression and severity of coronary stenosis in stable CAD. Highlights • First study of association of TAFI polymorphisms with coronary stenosis severity • Significant association between TAFI 505G/A and +1583T/A and high Gensini score • TAFI polymorphisms seem to contribute to atherosclerotic plaque progression. [ABSTRACT FROM AUTHOR]

Thrombin-activatable fibrinolysis inhibitor (TAFI) gene polymorphisms have been proposed as a predisposing factor for cerebral venous thrombosis (CVT). We analyzed the association between CVT and TAFI single-nucleotide polymorphisms (rs3742264, rs2146881, and rs1926447) compared to healthy controls. Mexico Mestizo confirmed cases with CVT and age- and sex-matched controls with no history of venous thrombotic events were recruited from July 2006 to July 2015. Demographic, clinical, and imaging information was included in the analysis. Genotyping single-nucleotide polymorphisms were performed by allele-specific polymerase chain reaction. Allelic univariate analysis, haplotype association, and Hardy-Weinberg equilibrium were assessed. A total of 113 CVT cases (94 females [83.2%]; median age 35 years [interquartile range 27-43 years]) and 134 age- and sex-matched controls were included. The main risk factors for CVT were pregnancy/puerperium (30.9%), oral contraceptive use (19.5%), and hereditary thrombophilia (7.1%). We found no significant association for heterozygous and homozygous models for rs3742264 (P = .30 and P = .69, respectively), rs2146881 (P = .90 and P = .17, respectively), or rs1926447 (P = .40 and P = .52, respectively) compared to controls; these findings were consistent in subgroup and haplotype analyses. In conclusion, TAFI rs3742264, rs2146881, and rs1926447 polymorphisms do not increase the risk of CVT in comparison to healthy controls. [ABSTRACT FROM AUTHOR]

An activated thrombin-activatable fibrinolysis inhibitor (TAFIa) attenuates fibrinolysis by removing C-terminal lysine/arginine residues from partially degraded fibrin. We have identified a novel low-molecular-weight inhibitor of TAFIa, DS-1040, to be potentially useful for treating thrombotic diseases. In this study, we investigated its in vitro pharmacological profile and in vivo effects in animal models of microthrombosis and bleeding. DS-1040 inhibited human TAFIa and carboxypeptidase N (CPN) in vitro with IC 50 values of 5.92 and 3.02 × 10 6  nmol/L, respectively, suggesting that DS-1040 is highly selective for TAFIa over CPN. DS-1040 did not affect platelet aggregation and coagulation time. In a tissue factor-induced rat microthrombosis model, intravenously administered DS-1040 reduced existing fibrin clots in the lung, whereas post-treatment with enoxaparin had limited effect. Both intravenously and orally administered DS-1040 elevated plasma D-dimer levels with similar plasma exposures of DS-1040. DS-1040 significantly augmented plasma D-dimer level on top of silent dose of recombinant tissue-plasminogen activator (t-PA), suggesting DS-1040 enhances fibrinolytic activity of t-PA. In addition, DS-1040 did not prolong the tail bleeding time beyond its efficacy dose. These results indicate that DS-1040 is a potent, selective, intravenously/orally available inhibitor of TAFIa with minimum risk of bleeding. DS-1040 is a potential novel fibrinolysis enhancer useful in treating thrombotic diseases. [ABSTRACT FROM AUTHOR]

Pro-carboxypeptidase B2 (pro-CPB2) or thrombin-activatable fibrinolysis inhibitor (TAFI) is a glycoprotein encoded by the CPB2 gene and deregulated in several cancer types, including breast cancer. Thrombin binding to thrombomodulin (TM), encoded by THBD , is important for TAFI activation. CPB2 gene expression is influenced by genetic polymorphism and cytokines such as interleukin 10 (IL-10). Our previous results showed that tumor infiltrating monocytes/macrophages (CD14 + /CD16 + ) isolated from inflammatory breast cancer (IBC) patients’ secrete high levels of IL-10. The aim of the present study is to test genetic polymorphism and expression of CPB2 in healthy breast tissues and carcinoma tissues of non-IBC and IBC patients. Furthermore, to investigate whether IL-10 modulates the expression of CPB2 and THBD in vivo and in-vitro . We tested CPB2 Thr325Ile polymorphism using restriction fragment length polymorphism, (RFLP) technique in healthy and carcinoma breast tissues. The mRNA expression of CPB2 , THBD and IL10 were assessed by RT-qPCR. Infiltration of CD14 + cells was assessed by immunohistochemistry. In addition, we investigated the correlation between infiltration of CD14 + cells and expression of IL10 and CPB2 . Furthermore, we correlated IL10 expression with the expression of both CPB2 and THBD in breast carcinoma tissues. Finally, we validated the role of recombinant IL-10 in regulating the expression of CPB2 and THBD using different breast cancer cell lines. Our results showed that CPB2 genotypes carrying the high-risk allele [Thr/Ile (CT) and Ile/Ile (TT)] were more frequent in both IBC and non-IBC patients compared to control group. CPB2 genotypes did not show any statistical correlation with CPB2 mRNA expression levels or patients’ clinical pathological properties. Interestingly, CPB2 and IL10 expression were significantly higher and positively correlated with the incidence of CD14 + cells in carcinoma tissues of IBC as compared to non-IBC. On the other hand, THBD expression was significantly lower in IBC carcinoma versus non-IBC tissues. Based on molecular subtypes, CPB2 and IL10 expression were significantly higher in triple negative (TN) as compared to hormonal positive (HP) carcinoma tissues of IBC. Moreover, CPB2 expression was positively correlated with presence of lymphovascular invasion and the expression of IL10 in carcinoma tissues of IBC patients. Furthermore, recombinant human IL-10 stimulated CPB2 expression in SUM-149 (IBC cell line) but not in MDA-MB-231 (non-IBC cell line), while there was no significant effect THBD expression. In conclusion, carcinoma tissues of IBC patients are characterized by higher expression of CPB2 and lower expression of THBD . Moreover, CPB2 positively correlates with IL10 mRNA expression, incidence of CD14 + cells and lymphovascular invasion in IBC patients. IL-10 stimulated CPB2 expression in TN-IBC cell line suggests a relevant role of CPB2 in the aggressive phenotype of IBC. [ABSTRACT FROM AUTHOR]

Apo(a), the distinguishing protein component of lipoprotein(a) [Lp(a)], exhibits sequence similarity to plasminogen and can inhibit binding of plasminogen to cell surfaces. Plasmin generated on the surface of vascular cells plays a role in cell migration and proliferation, two of the fibroproliferative inflammatory events that underlie atherosclerosis. The ability of apo(a) to inhibit pericellular plasminogen activation on vascular cells was therefore evaluated. Two isoforms of apo(a), 12K and 17K, were found to significantly decrease tissue-type plasminogen activator-mediated plasminogen activation on human umbilical vein endothelial cells (HUVECs) and THP-1 monocytes and macrophages. Lp(a) purified from human plasma decreased plasminogen activation on THP-1 monocytes and HUVECs but not on THP-1 macrophages. Removal of kringle V or the strong lysine binding site in kringle IV10 completely abolished the inhibitory effect of apo(a). Treatment with carboxypeptidase B to assess the roles of carboxyl-terminal lysines in cellular receptors leads in most cases to decreases in plasminogen activation as well as plasminogen and apo(a) binding; however, inhibition of plasminogen activation by apo(a) was unaffected. Our findings directly demonstrate that apo(a) inhibits pericellular plasminogen activation in all three cell types, although binding of apo(a) to cell-surface receptors containing carboxyl-terminal lysines does not appear to play a major role in the inhibition mechanism.

The host response to infection involves complex interplays between inflammation, coagulation, and fibrinolysis. Deregulation of hemostasis and fibrinolysis are major causes of critical illness and important determinants of outcome in severe sepsis. The hemostatic responses to infection vary widely between individuals, and are in part explained by polymorphisms in genes responsible for the protein C and fibrinolytic pathway. This review gives an overview of genetic polymorphisms in the protein C and fibrinolytic pathway associated with susceptibility and severity of pediatric sepsis. In addition, genetic polymorphisms associated with adult sepsis and other pediatric thromboembolic disorders are discussed, as these polymorphisms might be candidates for future molecular genetic research in pediatric sepsis. [ABSTRACT FROM AUTHOR]

Background: Statins efficiently lower cholesterol and also exert pleiotropic effects that extend beyond lipid lowering. In a recent pilot study, valuable information on the carboxypeptidase U (CPU) system in hyperlipidemia and the effect of statin therapy was collected. It was shown that proCPU levels are increased in hyperlipidemic patients. Statins significantly decreased proCPU levels and improved plasma fibrinolysis. Furthermore, it was suggested that patients with high baseline proCPU levels are most likely to benefit from statin therapy., Objectives: We aimed to further substantiate the effect of hyperlipidemia and statin therapy on CPU-related parameters in a larger cohort of hyperlipidemic and statin-treated individuals., Methods: Blood was collected from 141 individuals treated with different dosages of atorvastatin (10-80 mg), 38 normolipidemic, and 37 hyperlipidemic controls. Lipid parameters and markers of fibrinolysis (proCPU and clot lysis time) were determined and compared between the groups., Results: Pilot study results of high proCPU concentrations in hyperlipidemic patients and the proCPU-reducing effect of atorvastatin were confirmed. Accordingly, an improvement in plasma fibrinolytic potential was seen under the influence of atorvastatin. High interindividual variation in proCPU concentrations was observed in the hyperlipidemic cohort, with up to 80% higher proCPU levels compared with normolipidemic controls. Furthermore, proCPU concentration and the dosage of atorvastatin were inversely correlated., Conclusions: This study clearly shows that plasma proCPU concentrations and its expected effect on the fibrinolytic rate (as measured by clot lysis time) are increased in hyperlipidemic patients and that these effects can be normalized (and even further reduced compared with normolipidemic patients) by atorvastatin treatment., Competing Interests: Declaration of competing interests There are no competing interests to disclose., (Copyright © 2023 International Society on Thrombosis and Haemostasis. Published by Elsevier Inc. All rights reserved.)

Background An imbalance between coagulation and fibrinolytic system plays an important role in the pathogenesis of arterial thrombosis. It has been identified that elevated plasma thrombin-activatable fibrinolysis inhibitor (TAFI) concentration, an anti-fibrinolytic factor, is associated with an increased risk of cardiovascular disease (CVD). But the effect of genetic variations in TAFI gene on the risk of CVD is inconclusive. Objectives To investigate the associations between two variants Ala147Thr(rs3742264) and Thr325Ile(rs1926447) in TAFI and the risk of CVD. Methods Systematic review and meta-analysis of eligible studies published before January 2014. Coronary heart disease(CHD) and stroke are regarded as end-points of CVD. Results A total of 18 articles including 23 studies were enrolled. Among these articles were 19 studies of Ala147Thr and 15 of Thr325Ile variants, comprising 4,977 CVD patients and 8,082 controls together with 4,890 cases and 8,311 controls, respectively. There were no significant associations between Ala147Thr variant and CVD under allele, dominant, recessive genetic models. Similar results were observed when end-point, ethnicity, sample size, genotyping method were taken into account. Likewise, meta-analysis of Thr325Ile variant did not show significant associations with CVD under three genetic models. Nevertheless, in sub-analysis based on end-point, the TT(Ile/Ile) genotype was associated with a 25% higher risk of coronary heart disease(CHD) (OR = 1.25, 95%CI, 1.02-1.54; P = 0.03) compared with TC + CC(Thr/Ile + Thr/Thr) genotype(recessive model). Conclusions The present meta-analysis failed to confirm the influence of Ala147Thr and Thr325Ile variants on the susceptibility to CVD. However, potentially increased risk of CHD was detected in Ile325 allele carriers under recessive model. [ABSTRACT FROM AUTHOR]

Thrombin-activatable fibrinolysis inhibitor (TAFI) is a procarboxypeptidase, which is synthesised in liver and activated by thrombin and the thrombin-thrombomodulin complex. TAFI suppresses fibrinolysis by removing carboxy-terminal lysine residues from partially degraded fibrin. In this study we aimed to assess the circulating levels of TAFI antigen, 'a fibrinolytic parameter' in women with gestational diabetes (GDM). Thirty-four pregnant women with GDM and 50 pregnant women with normal glucose tolerance were included in the study. Plasma TAFI antigen levels were significantly higher in pregnant women with GDM when compared with controls. Increased TAFI levels may contribute to the decreased fibrinolytic potency, causing a thrombophilic state. GDM is regarded as a specific form of diabetes, and it could in addition be a predictor of type 2 diabetes mellitus in the future and the risk of complications due to hypercoagulability increases in this disease. Increased TAFI levels may also have a role in increased risk of hypercoagulability. [ABSTRACT FROM AUTHOR]

Objective:We determined the profile of coagulation/fibrinolytic and vascular endothelial cell function parameters including plasminogen activator inhibitor (PAI) and thrombin-activatable fibrinolysis inhibitor (TAFI), tissue factor pathway inhibitor (TFPI), thrombomodulin (TM), and tissue plasminogen activator (tPA) levels in children with hypothyroidism. Methods: Forty children with hypothyroidism aged 0-16 months who presented for the first time to our hospital and 29 age-and sex-matched healthy controls were enrolled in the study. All coagulation tests were performed with ELISA method. One year after Na-L-thyroxine treatment, the study parameters were re-evaluated in 25 euthyroid children out of the 40 patients diagnosed with hypothyroidism. Results: Although no significant effect was detected regarding PAI antigen (Ag) and tPA Ag, the levels of TAFI, TM, and TFPI were consistent with subclinical hypercoagulability and hypofibrinolysis. There was a significant increase in TAFI Ag levels and a significant decrease in TFPI Ag and TM Ag levels in hypothyroid patients compared to healthy controls. As a result of correlation tests, the largest impact of hypothyroidism on coagulation system was on TFPI. In accordance with these findings, TAFI Ag levels decreased and TFPI Ag and TM Ag levels increased with hormonal replacement therapy. Conclusions: Increased TAFI and decreased TFPI and TM in patients with hypothyroidism may indicate a potential hypercoagulable and hypofibrinolytic state as well as possible endothelial dysfunction, which may increase the risk of atherosclerotic and atherothrombotic complications. Thyroid hormone levels should also be checked in patients with a predisposition to coagulation, and thyroid replacement therapy should be initiated. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the levels of thrombin-activatable fibrinolysis inhibitor (TAFI) activity and also its relationship with other homeostasis markers in breast cancer patients. Subjects and Methods: Forty-two female patients with breast cancer and 24 healthy women (controls) were enrolled in the study and fasting blood samples of all cases were drawn from a large antecubital vein for assay of TAFI and other homeostasis tests. Results: The TAFI levels were 79.5 ± 15.5 and 39.3 ± 12.1 in patients and controls, respectively, and the difference was statistically significant (p < 0.001). In the patient group, the serum fibrinogen level was 504.9 ± 224.8, while in the control group it was 393.9 ± 100.5, and the difference was also statistically significant (p < 0.001). Conclusion: The data showed that increased levels of TAFI are a contributing factor of thrombotic disorders in breast cancer patients. Copyright © 2011 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

BACKGROUND Recurrent implantation failure (RIF) following embryo transfer (ET) is a major continuing problem in IVF. Women with haemostatic defects may be at increased risk of miscarriage and preclinical pregnancy loss. The fibrinolytic system is considered, at present, the key to new thrombotic pathogenic mechanisms. Patients with unexplained recurrent miscarriage have an impairment of fibrinolysis, as demonstrated by prolonged clot lysis time (CLT) in association with increased plasma levels of thrombin-activatable fibrinolysis inhibitor (TAFI). In this study, we investigated fibrinolytic potential in patients with RIF. METHODS Three groups of patients were studied: 30 women with RIF (RIF group), 60 patients undergoing a first successful IVF–ET cycle (IVF group) and 60 healthy fertile women (FER group). Plasma CLT was measured using a global fibrinolysis assay. TAFI antigen plasma levels and polymorphisms in the TAFI gene (+505A/G and +1542C/G) were analysed using enzyme-linked immunosorbent assay and allele-specific PCR, respectively. RESULTS CLT was significantly longer (P< 0.0001 and P< 0.0009, respectively) and TAFI antigen levels were significantly higher (both P< 0.0001) in the RIF versus the IVF and FER groups. A direct relationship between CLT and TAFI antigen levels (r = 0.40; P = 0.001) was detected in the whole study population. There were no differences in distribution of TAFI polymorphisms between groups. CONCLUSIONS Patients with RIF have reduced plasma fibrinolytic potential, as shown by a prolonged CLT, and this may be explained, at least in part, by increased TAFI antigen levels. [ABSTRACT FROM AUTHOR]

The CPB2 gene encodes thrombin-activatable fibrinolysis inhibitor (TAFI), a hepatically secreted zymogen acting as a molecular link among coagulation, fibrinolysis, and inflammation. Variants in CPB2 have been associated with several human conditions. We resequenced and analyzed the two regions carrying previously known nonsynonimous single-nucleotide polymorphisms (Ala147Thr and Ile325Thr) and variants affecting transcript stability. Our data indicate that whereas the gene portion extending from exon 9 to the 3′ untranslated region fits a model of neutral evolution, variants in the region encompassing exons 6–7 have been maintained by balancing selection. Indeed, we verified that the region displays high nucleotide diversity, many intermediate frequency variants, and an excess of polymorphism compared with interspecific divergence. Consistently, haplotype analysis indicated the presence of two major haplotype clades separated by deep branches. Transcript analysis revealed that in both HepG2 cells and human liver samples, CPB2 exon 7 undergoes haplotype-preferential skipping. Therefore, we indicate that balancing selection has been maintaining functional variants that promote alternative exon 7 splicing. Although transcripts lacking exon 7 represent a minority of total CPB2 products, the effect on antifibrinolytic activity might be much greater as the intrinsic instability of TAFI is a major determinant of its antifibrinolytic potential. [ABSTRACT FROM PUBLISHER]