High thrombin-activatable fibrinolysis inhibitor expression in thrombi from stroke patients in elevated estrogen states.

BACKGROUND: The risk of acute ischemic stroke (AIS) associated with high estrogen states, including pregnant patients and those using oral contraceptives, has been well documented. We described the histological composition of thrombi collected in these cases. METHODS: From a prospective tissue registry (STRIP registry) of thrombi retrieved during mechanical thrombectomy for AIS, we identified 5 patients with high estrogen states: 1 post-partum patient, 1 undergoing hormone replacement therapy and 3 consuming oral contraceptive pills. Five male control patients were randomly chosen matched by age. Immunohistochemistry for CD42b (platelets), von Willebrand factor (vWF), thrombin-activatable fibrinolysis inhibitor (TAFI), fibrinogen and plasminogen activator inhibitor-1 (PAI-1) was performed. Expression was quantified using Orbit Image Software. Student's t-test was performed as appropriate. RESULTS: Mean TAFI content for the high estrogen state group was higher than controls (25.6 ± 11.9% versus 9.3 ± 9.0%, p = 0.043*). Mean platelet content for the high estrogen state group was lower than controls (41.7 ± 10.6% versus 61.8 ± 12.9%, p = 0.029*). No significant difference was found in vWF, fibrinogen and PAI-1 expression. Mean time to recanalize was higher in the high estrogen state group compared to the control group (57.8 ± 27.6 versus 22.6 ± 11.4 min, p = 0.0351*). The mean number of passes required was higher in the high estrogen group compared to controls 4.6 versus 1.2, p = 0.0261*). CONCLUSIONS: TAFI expression, a powerful driver of thrombosis, was significantly higher in stroke thrombi among patients with high estrogen states compared to controls.

Investigation of cofactor activities of endothelial microparticle-thrombomodulin with liposomal surrogate.

Thrombomodulin (TM) is a type I transmembrane glycoprotein mainly expressed on the endothelial cells, where it binds thrombin to form the thrombin-TM complex that can activate protein C and thrombin-activable fibrinolysis inhibitor (TAFI) and induce anticoagulant and anti-fibrinolytic reactions, respectively. Cell activation and injury often sheds microparticles that contain membrane TM, which circulate in biofluids like blood. However, the biological function of circulating microparticle-TM is still unknown even though it has been recognized as a biomarker of endothelial cell injury and damage. In comparison with cell membrane, different phospholipids are exposed on the microparticle surface due to cell membrane ''flip-flop'' upon cell activation and injury. Liposomes can be used as a microparticle mimetics. In this report, we prepared TM-containing liposomes with different phospholipids as surrogates of endothelial microparticle-TM and investigated their cofactor activities. We found that liposomal TM with phosphatidylethanolamine (PtEtn) showed increased protein C activation but decreased TAFI activation in comparison to liposomal TM with phosphatidylcholine (PtCho). In addition, we investigated whether protein C and TAFI compete for the thrombin/TM complex on the liposomes. We found that protein C and TAFI did not compete for the thrombin/TM complex on the liposomes with PtCho alone and with low concentration (5%) of PtEtn and phosphatidylserine (PtSer), but competed each other on the liposomes with higher concentration (10%) of PtEtn and PtSer. These results indicate that membrane lipids affect protein C and TAFI activation and microparticle-TM may have different cofactor activities in comparison to cell membrane TM.

Serum thrombin-activatable fibrinolysis inhibitor levels and its relation with pathogenesis and bleeding and prognosis in patients with Crimean Congo hemorrhagic fever.

Crimean-Congo hemorrhagic fever (CCHF) is a viral hemorrhagic fever, which is common in Turkey and globally. The pathogenesis of coagulation disorders, which is seen in viral hemorrhagic fevers remains to be elucidated. Thrombin-activatable fibrinolysis inhibitor (TAFI) has a key role in this process In this study, we aimed to evaluate whether TAFI levels contributed to bleeding and whether it is related to prognosis in CCHF patients. Eighty-four patients older than 15 years of age, who were admitted to our hospital who had positive immunoglobulin M (enzyme-linked immunosorbent assay [ELISA]) and/or polymerase chain reaction test results for CCHF between 2009 and 2010, were included in the study. The control group included 30 healthy adults. The plasma TAFI levels were compared between patients and controls, and also between patients with bleeding and no bleeding, and between patients with mild-moderate and severe disease. The mean TAFI levels were lower in patients (mean: 87.82 ng/ml, median: 61.69 ng/ml (interquartile range [IQR] 30.49-537.95) than controls (mean: 313.5 ng/ml with a median: 338.5 ng/ml (IQR 182-418). However, median TAFI levels were significantly higher in patients with bleeding compared to those without bleeding (78.99 and 50.28 ng/ml, respectively; p = 0.032). Median IQR TAFI levels were similar between patients with mild-moderate and severe disease (64.72 (41.37-113.85), and, 58.66 (42.44-118.93) ng/ml, respectively; p = 0.09) and survivors and nonsurvivors (86.14 ± 77.98 and 103.48 ± 69.92, respectively; p = 0.3). Although TAFI levels were lower in the patients with CCHF compared to healthy controls, it does not seem to be a major player in the prognosis.

Quantitation of thrombin-activatable fibrinolysis inhibitor in human plasma by isotope dilution mass spectrometry.

Measurement of Thrombin-activatable fibrinolysis inhibitor (TAFI) in human plasma is dependent on reproducible assays. To date, standards for measuring TAFI are frequently calibrated relative to pooled normal human plasma and arbitrarily assigned a potency of 100% TAFI, despite variation in TAFI concentrations between plasma pools. Alternatively, TAFI calibrators can be assigned a value in SI units but the approach used for value assignment is not consistent and furthermore, if purified TAFI is used to determine TAFI concentration in plasma, may be adversely affected by matrix effects. A TAFI plasma standard in mass units with traceability to the SI unit of mass is desirable. We report here the establishment of a quantitative mass spectrometry method for TAFI in plasma. Traceability is obtained by reference to calibrators that consist of blank plasma spiked with a defined amount of purified TAFI, value assigned by amino acid analysis. The calibrators are run alongside the samples, using the same preparation steps and conditions; an acetonitrile assisted tryptic digestion and multi-dimensional liquid chromatography (LC) separation followed by MRM-MS analysis. We measured the TAFI quantitatively in human plasma with reproducibility, reliability and precision, and demonstrated the applicability of this approach for value assigning a common reference standard.

Thrombin Activatable Fibrinolysis Inhibitor (TAFI): An Updated Narrative Review.

Thrombin activatable fibrinolysis inhibitor (TAFI), a proenzyme, is converted to a potent attenuator of the fibrinolytic system upon activation by thrombin, plasmin, or the thrombin/thrombomodulin complex. Since TAFI forms a molecular link between coagulation and fibrinolysis and plays a potential role in venous and arterial thrombotic diseases, much interest has been tied to the development of molecules that antagonize its function. This review aims at providing a general overview on the biochemical properties of TAFI, its (patho)physiologic function, and various strategies to stimulate the fibrinolytic system by interfering with (activated) TAFI functionality.

Plasma thrombin-activatable fibrinolysis inhibitor (TAFI) antigen levels in acromegaly patients in remission

Background/aim: Acromegaly is associated with increased morbidity andmortality, mostly due to cardiovascular complications.Plasma thrombin-activatable fibrinolysis inhibitor (TAFI) antigen levels are associated with coagulation/fibrinolysis and inflammation. Plasma TAFI may play a role in arterial thrombosis in cardiovascular diseases. In this study, it was aimed to evaluate the thrombin-activatable fibrinolysis inhibitor (TAFI) antigen and homocysteine levels in patients with acromegaly and healthy control subjects. Materials and methods: Plasma TAFI antigen and homocysteine levels in 29 consecutive patients with acromegaly and 26 age-matched healthy control subjects were measured. All patients included in the study were in remission. The TAFIa/ai antigen in the plasma samples was measured using a commercially available ELISA kit. Results: Routine biochemical parameters, fasting blood glucose, prolactin, thyroid stimulating hormone, total-cholesterol, low density lipoprotein cholesterol, triglyceride, and homocysteine levels were similar in the 2 groups (P > 0.05), whereas the plasma TAFI antigen levels were significantly elevated in the acromegalic patients (154.7 ± 94.0%) when compared with the control subjects (107.2 ± 61.6%) (P = 0.033). No significant correlation was identified by Pearson's correlation test between the plasma TAFI antigen and homocysteine levels (r = 0.320, P = 0.250). Conclusion: A significant alteration in the plasma TAFI antigen levels was detected in acromegaly. Increased plasma TAFI antigen levels might aggravate prothrombotic and thrombotic events in patients with acromegaly.

Impaired fibrinolysis in degenerative mitral and aortic valve stenosis.

Valvular heart disease is associated with an increased thromboembolic risk. Impaired fibrinolysis was reported in severe aortic stenosis (AS). Little is known about fibrinolysis in mitral stenosis (MS). We sought to compare fibrinolysis impairment in AS and MS. We studied 121 individuals scheduled for elective aortic valve (AV) or mitral valve (MV) surgery for AS (n = 76) or MS (n = 45), in order to compare fibrinolysis impairment. Fibrinolytic capacity was assessed by determination of clot lysis time (t50%) and fibrinolysis inhibitors, including plasma plasminogen activator inhibitor-1 (PAI-1) antigen (PAI-1:Ag) and activity, thrombin-activatable fibrinolysis inhibitor (TAFI) antigen and activity. Prolonged t50% (+ 29%), elevated TAFI activity (+ 12%), TAFI:Ag (+ 21%), and PAI-1:Ag (+ 84%) were observed in patients with MS, compared with those with AS. t50% Correlated with mean and maximal MV gradients (r = 0.43, p < 0.0001 and r = 0.39, p < 0.0001, respectively), but not with AV gradients. Mean and maximal MV gradients correlated with TAFI activity and PAI:Ag. Patients with permanent atrial fibrillation (AF; 35 with MS and 5 with AS) had longer t50% (by 22%, p = 0.0002) and higher PAI-1:Ag (by 74%, p < 0.0001) than the remainder. In the whole group, postoperative drainage volumes correlated inversely with PAI-1:Ag (r = - 0.22, p = 0.02). MS is associated with more pronounced impairment of global fibrinolytic capacity than AS at the stage of surgical intervention, which is in part driven by AF. Our findings suggest that hypofibrinolysis might be implicated in the progression of MS and its thromboembolic complications.

Activated thrombin-activatable fibrinolysis inhibitor (TAFIa) attenuates breast cancer cell metastatic behaviors through inhibition of plasminogen activation and extracellular proteolysis.

BACKGROUND: Thrombin activatable fibrinolysis inhibitor (TAFI) is a plasma zymogen, which can be converted to activated TAFI (TAFIa) through proteolytic cleavage by thrombin, plasmin, and most effectively thrombin in complex with the endothelial cofactor thrombomodulin (TM). TAFIa is a carboxypeptidase that cleaves carboxyl terminal lysine and arginine residues from protein and peptide substrates, including plasminogen-binding sites on cell surface receptors. Carboxyl terminal lysine residues play a pivotal role in enhancing cell surface plasminogen activation to plasmin. Plasmin has many critical functions including cleaving components of the extracellular matrix (ECM), which enhances invasion and migration of cancer cells. We therefore hypothesized that TAFIa could act to attenuate metastasis. METHODS: To assess the role of TAFIa in breast cancer metastasis, in vitro migration and invasion assays, live cell proteolysis and cell proliferation using MDA-MB-231 and SUM149 cells were carried out in the presence of a TAFIa inhibitor, recombinant TAFI variants, or soluble TM. RESULTS: Inhibition of TAFIa with potato tuber carboxypeptidase inhibitor increased cell invasion, migration and proteolysis of both cell lines, whereas addition of TM resulted in a decrease in all these parameters. A stable variant of TAFIa, TAFIa-CIIYQ, showed enhanced inhibitory effects on cell invasion, migration and proteolysis. Furthermore, pericellular plasminogen activation was significantly decreased on the surface of MDA-MB-231 and SUM149 cells following treatment with various concentrations of TAFIa. CONCLUSIONS: Taken together, these results indicate a vital role for TAFIa in regulating pericellular plasminogen activation and ultimately ECM proteolysis in the breast cancer microenvironment. Enhancement of TAFI activation in this microenvironment may be a therapeutic strategy to inhibit invasion and prevent metastasis of breast cancer cells.

Inhibition of thrombin-activated fibrinolysis inhibitor decreases postoperative adhesion.

BACKGROUND: Postoperative adhesion is a problematic issue. The aim of this study is to identify the effect of a TAFI inhibitor (potato tuber carboxypeptidase inhibitor [PTCI]), novel agent, which reconciles formation and degradation of fibrosis, in the prevention of postoperative adhesions in rats. METHODS: A total of 48 rats were allocated in three groups; control group C, normal saline group N, and PTCI group P. After surgical adhesion formation under anesthesia, group N received 5 mL of normal saline, group P; 5 mg of PTCI in 5 mL normal saline was instilled in the peritoneum. Four weeks after surgery, rats were sacrificed and adhesions were assessed in gross macroscopic, microscopic, and immunohistologic aspects, and scoring was performed in each category. RESULTS: Besides two rats, one from each group N and C who died on the third postoperative day, no adverse events were found among the groups throughout the study. In gross adhesions, group P had significantly lower adhesion score than all other groups (P < 0.001). Group P had a significantly lower fibrosis and inflammation in microscopic assessments (P < 0.05) and immunohistochemistry (P < 0.001) compared with groups C and N. CONCLUSIONS: In conclusion, TAFI pathway inhibition resulted in reduction of postoperative adhesion number and severity. Clinical and microscopic findings show that TAFI inhibition plays a role in modulating adhesion formation.

Higher thrombin activatable fibrinolysis inhibitor levels are associated with inflammation in attack-free familial Mediterranean fever patients.

BACKGROUND: Coagulation abnormalities have been reported in familial Mediterranean fever (FMF) patients with amyloidosis and nephrotic syndrome; but there is not enough data about the continuity of the thrombogenic activity in FMF patients in clinical remission. The purpose of this study was to assess thrombin activatable fibrinolysis inhibitor (TAFI) levels and its relationship with fibrinolytic activity and also evaluate relationships between mutations and clinical signs in attack-free patients without amyloidosis. METHODS: Seventy-nine FMF patients and 40 healthy adults were included. The study group was divided into five groups as follows: first group, homozygote M694V; second group, homozygote M680I; third group, M694V in one allele, the other allele have other mutations or not; fourth group, other mutations; and fifth group, no mutation. RESULTS: Serum TAFI levels were significantly increased in patients compared with healthy individuals (116.64 ± 21.8 vs. 78.48 ± 19.7 µg/mL, p < 0.001) and a positive correlation was detected between TAFI antigen level and erythrocyte sedimentation rate and C-reactive protein levels (r = 0.247, p = 0.029 and r = 0.252, p = 0.032, respectively). Mean fibrinogen and TAFI levels were significantly higher in Group 1 than the other groups (p = 0.04 and p = 0.001, respectively) and in Group 3 it was higher than Groups 2, 4 and 5 (p = 0.04 and p = 0.001, respectively). CONCLUSIONS: High level of TAFI antigen in attack-free period of FMF disease shows ongoing subclinical inflammation and hypercoagulability. Clinicians should be careful about thrombosis even in patients at clinical remission. Also, genetic tests must be considered to predict clinical outcome and to reduce complications of FMF disease.

Evaluation of thrombomodulin/thrombin activatable fibrinolysis inhibitor function in plasma using tissue-type plasminogen activator-induced plasma clot lysis time.

Background: Thrombin activatable fibrinolysis inhibitor (TAFI) is one of the most important physiological fibrinolysis inhibitors. Its inhibitory efficacy under physiological conditions remains uncertain. Objectives: Elucidate the role of soluble thrombomodulin (sTM)/TAFI axis in the regulation of fibrinlysis. Methods: Since thrombin is required to generate activated TAFI (TAFIa) that targets the C-terminal lysine of partially digested fibrin, a clot lysis assay is suitable for evaluating its function. Using tissue-type plasminogen activator-induced plasma clot lysis time (tPA-PCLT) together with TAFIa inhibitor and recombinant sTM (rsTM), we evaluated the specific function of TM/TAFI in the plasma milieu. Results: tPA-PCLT values were significantly shortened by the TAFIa inhibitor. rsTM supplementation prolonged tPA-PCLT, which was shortened by the TAFIa inhibitor to a time similar to that obtained without rsTM and with the TAFIa inhibitor. Plasma obtained from patients treated with rsTM showed prolonged tPA-PCLT, which was shortened by the TAFIa inhibitor but not further prolonged by rsTM. However, no significant correlation was observed between tPA-PCLT and parameters of TM/TAFI system in the plasma. Conclusion: The role of the TM/TAFI system in regulating fibrinolysis was successfully evaluated using TAFIa inhibitor and rsTM. Trace amounts of soluble TM in normal plasma appeared sufficient to activate TAFI and inhibit fibrinolysis. Further, a therapeutic dose of rsTM appeared sufficient to activate TAFI and regulate fibrinolysis in the plasma milieu.

Increased thrombin activatable fibrinolysis inhibitor activity is associated with hypofibrinolysis in dogs with sepsis.

Introduction: Disorders of coagulation are well-recognized in dogs with sepsis, but data regarding fibrinolysis disorders are limited. We aimed to characterize fibrinolysis in dogs with sepsis compared to healthy controls. We hypothesized that dogs with sepsis would be hypofibrinolytic, and that hypofibrinolysis would be associated with non-survival. Methods: This was a prospective observational cohort study. We enrolled 20 client-owned dogs with sepsis admitted to the Cornell University Hospital for Animals and 20 healthy pet dogs. Coagulation and fibrinolytic pathway proteins including antiplasmin activity (AP), antithrombin activity (AT), thrombin activatable fibrinolysis inhibitor activity (TAFI), D-dimer concentration, fibrinogen concentration, and plasminogen activity were measured and compared between groups. Overall coagulation potential, overall fibrinolysis potential, and overall hemostatic potential were calculated from the curve of fibrin clot formation and lysis over time. Results: Compared to healthy controls, dogs with sepsis had lower AT (P = 0.009), higher AP (P = 0.002), higher TAFI (P = 0.0385), and higher concentrations of fibrinogen (P < 0.0001) and D-dimer (P = 0.0001). Dogs with sepsis also had greater overall coagulation potential (P = 0.003), overall hemostatic potential (P = 0.0015), and lower overall fibrinolysis potential (P = 0.0004). The extent of fibrinolysis was significantly negatively correlated with TAFI. No significant differences were observed between survivors and non-survivors. Discussion: Dogs with sepsis were hypercoagulable and hypofibrinolytic compared to healthy dogs, suggesting potential utility of thromboprophylaxis in this patient population. The association between high TAFI and low overall fibrinolysis potential might provide a potential mechanism for this hypofibrinolysis.

Glycemic Control and Plasma Levels of Pro-Inflammatory and Pro-Thrombotic Biomarkers in Diabetic Patients Presenting with Acute Pulmonary Embolism.

Type I and type II diabetes are closely associated with a pro-inflammatory state and to a pro-thrombotic state. The role of glycemic control in pulmonary embolism (PE) is poorly understood and requires additional investigation. The aim of this study is to investigate the relationship between glycemic control and thrombo-inflammatory biomarkers in a PE patient cohort compared to normal samples. Demographic and clinical information for 86 diabetic patients and 106 non-diabetic patients presenting with acute PE was collected via retrospective chart review. Plasma levels of pro-inflammatory (C-reactive protein [CRP], tumor necrosis factor-alpha [TNF-α], interleukin-6 [IL-6]) and pro-thrombotic (d-dimer, plasminogen activator inhibitor-1 [PAI-1], tissue plasminogen activator [tPA], thrombin activatable fibrinolysis inhibitor [TAFI], von-Willebrand factor [vWF], endogenous glycosaminoglycans [GAGs]) biomarkers were drawn within 24 hours of diagnosis of acute PE. Data was also obtained for a population of healthy adult controls. All the pro-inflammatory and pro-thrombotic biomarkers were elevated in diabetic PE patients in comparison to healthy controls. None of the biomarkers were elevated in diabetic PE patients when compared to non-diabetic PE patients. There was no difference in the levels of the pro-inflammatory biomarkers according to glycemic control. The plasma level of TAFI was elevated in diabetic patients with poor glycemic control. Diabetic patients were more likely to have a more severe PE. These studies demonstrate that thrombo-inflammatory biomarkers are elevated in diabetic PE patients with associated comorbidities in comparison to normal individuals. However, there is no difference between the PE cohort alone in comparison to PE with diabetes. The role of TAFI within the continuum of diabetic vascular disease warrants additional investigation.

Identification of Genes and miRNAs Associated with TAFI-Related Thrombosis: An in Silico Study.

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.

Platelet Microvesicles, Inflammation, and Coagulation Markers: A Pilot Study.

BACKGROUND: Platelet "Microvesicles" (MVs) are studied for their role in blood coagulation and inflammation. The study aimed to establish if MVs are related to age, plasma levels of inflammation, coagulation, and fibrinolysis markers in healthy individuals. METHODS: We prospectively enrolled volunteers aged over 18 years. MVs, plasma levels of C-reactive protein (CRP), Interleukin 6 (IL-6), Interleukin 10 (IL-10), Interleukin 17 (IL-17), and transforming growth factor ß (TGF-ß), fibrinogen, plasminogen activator inhibitor-1 (PAI-1), von Willebrand factor (VWF), homocysteine, factor VII (FVII), thrombin activatable fibrinolysis inhibitor (TAFI), and Protein S were tested. RESULTS: A total of 246 individuals (median age 65 years ("IQR"54-72)) were evaluated. Both univariate analysis and logistic regression models showed that MVs positively correlate with age, CRP, IL-6, IL-10, IL-17, TGF-ß, fibrinogen, PAI-1, VWF, FVII, and homocysteine, while inversely correlating with TAFI and Protein S. The ROC curve analysis performed to identify a cut off for MV values (700 kMP) showed a good accuracy with over-range cytokines fibrinolysis factor and coagulation markers. CONCLUSIONS: To the best of our knowledge, this study is the first to correlate MVs with an entire panel of cardiovascular risk factors in healthy individuals. A future possible role of MVs in screening exams is suggested.

Pre-administration of a carboxypeptidase inhibitor enhances tPA-induced thrombolysis in mouse microthrombi: Evidence from intravital imaging analysis.

INTRODUCTION: Thrombolysis using recombinant tissue-type plasminogen activator (rt-PA) is the pharmacological treatment of choice in acute thrombotic events. However, a narrow therapeutic window and bleeding complications limit its use. We describe the role of carboxypeptidase inhibitor from potato tuber (PTCI), an inhibitor of activated thrombin-activatable fibrinolysis inhibitor (TAFIa), on Glu-plasminogen accumulation and microthrombus dynamics in vivo and demonstrate its influence on rt-PA-mediated thrombolysis. MATERIALS AND METHODS: In conjunction with real-time intravital two-photon excitation fluorescence microscopy, we produced and imaged laser-induced microthrombi in the mesenteric venules of Green Fluorescent Protein (GFP)-expressing mice. We examined microthrombus dynamics and thrombolysis patterns in vivo by measuring the changes in the fluorescence intensity of labeled Glu-plasminogen following administration of epsilon aminocaproic acid (EACA), PTCI, and rt-PA. RESULTS: PTCI enhanced Glu-plasminogen accumulation at the core of the thrombus by inhibiting TAFIa, while EACA inhibited this process. Exogenous rt-PA effectively triggered Glu-plasminogen activation within the thrombus and promoted thrombolysis. Administration of PTCI and rt-PA together showed no significant benefit on thrombolysis compared to rt-PA administration alone. However, early-phase systemic administration of PTCI before thrombolytic therapy by rt-PA expedited clot lysis as evidenced by significantly faster time to reach peak Glu-plasminogen fluorescence intensity and shorter time to achieve near-complete clot lysis (P = 0.014 and P = 0.003, respectively). CONCLUSIONS: PTCI potentiates rt-PA-mediated thrombolysis when administered early in acute thrombotic events. Further studies are warranted to explore the potential of TAFI inhibitors as adjunct agents in thrombolysis or thromboprophylaxis.

A Case-Control Study of the Dose-Response Relationship Between Thrombin Activatable Fibrinolysis Inhibitor and Acute Myocardial Infarction.

Background: Acute myocardial infarction (AMI) is considered an acute coronary syndrome (ACS), which is caused by the death of myocardial cells after prolonged ischemia, and there is a high risk of sudden death during AMI. Therefore, the purpose of this study is to explore the relationship between thrombin activatable fibrinolysis inhibitor (TAFI) and AMI and provide evidence for their association and potentially the prevention of AMI. Methods: There were 228 subjects included in this retrospective study, which included 78 AMI patients and 150 controls. The immune turbidimetry was used to measure TAFI concentration in the serum. Mann-Whitney U test was used to compare serum TAFI levels. The logistic regression analysis was used to construct a model of influencing factors of AMI. The dose-response relationship between serum TAFI level and AMI was explored by using the restricted cubic spline (RCS) functions combined with logistic regression analysis. Results: The serum TAFI levels of the AMI group were higher than the control group's (P = 0.003). The risk of AMI in the high-TAFI level group was 2.24 times higher than the low-TAFI level group (P = 0.007) and it was 2.74 times higher after adjustment of other risk factors (P = 0.025). According to the dose-response curve, the risk of AMI increased significantly with an increase of serum TAFI concentration (P = 0.0387). Conclusion: Acute myocardial infarction patients had higher serum TAFI levels, and TAFI was an independent risk factor for AMI patients. Serum TAFI levels demonstrated a dose- dependent response to the risk of AMI. Our study provides evidence that TAFI could be used for risk stratification of AMI patients.

PP2A is activated by cytochrome c upon formation of a diffuse encounter complex with SET/TAF-Iß.

Intrinsic protein flexibility is of overwhelming relevance for intermolecular recognition and adaptability of highly dynamic ensemble of complexes, and the phenomenon is essential for the understanding of numerous biological processes. These conformational ensembles-encounter complexes-lack a unique organization, which prevents the determination of well-defined high resolution structures. This is the case for complexes involving the oncoprotein SET/template-activating factor-Iß (SET/TAF-Iß), a histone chaperone whose functions and interactions are significantly affected by its intrinsic structural plasticity. Besides its role in chromatin remodeling, SET/TAF-Iß is an inhibitor of protein phosphatase 2A (PP2A), which is a key phosphatase counteracting transcription and signaling events controlling the activity of DNA damage response (DDR) mediators. During DDR, SET/TAF-Iß is sequestered by cytochrome c (Cc) upon migration of the hemeprotein from mitochondria to the cell nucleus. Here, we report that the nuclear SET/TAF-Iß:Cc polyconformational ensemble is able to activate PP2A. In particular, the N-end folded, globular region of SET/TAF-Iß (a.k.a. SET/TAF-Iß ΔC)-which exhibits an unexpected, intrinsically highly dynamic behavior-is sufficient to be recognized by Cc in a diffuse encounter manner. Cc-mediated blocking of PP2A inhibition is deciphered using an integrated structural and computational approach, combining small-angle X-ray scattering, electron paramagnetic resonance, nuclear magnetic resonance, calorimetry and molecular dynamics simulations.

Human TAF-Iα promotes oncogenic transformation via enhancement of cell proliferation and suppression of apoptosis.

Template activating factor-I (TAF-I) is a multifunctional protein involved in various biological processes including the inhibition of histone acetylation, DNA replication, cell cycle regulation, and oncogenesis. Two main TAF-I isoforms with different N-termini, TAF-Iα and TAF-Iß (SET), are expressed in cells. There are numerous data about functional properties of TAF-Iß, whereas the effects of TAF-Iα remain largely unexplored. Here, we employed focus formation and cell proliferation assays, TUNEL staining, cytological analysis, and RT-qPCR to compare the effects of human TAF-Iα and TAF-Iß genes, transiently expressed in Rat2 cells and in Misgurnus fossilis loaches. We found that both TAF-I isoforms possessed equal oncogenic potential in these systems. Furthermore, an overexpression of human TAF-Iα and TAF-Iß in Rat2 cells promoted their proliferation. Accordingly, the mitotic index was increased in the transgenic loaches expressing human TAF-Iα or TAF-Iß. TUNEL assay as well as downregulation of p53 gene and upregulation of bcl-2 gene in these transgenic loaches demonstrated that both isoforms suppressed apoptosis. Thus, TAF-Iα isoform exerts the same oncogenic potential as TAF-Iß, likely by suppressing the apoptosis and promoting cell proliferation.

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.