Plasma carboxypeptidase U (CPU, CPB2, TAFIa) generation during in vitro clot lysis and its interplay between coagulation and fibrinolysis

2017 ◽  
Vol 117 (08) ◽  
pp. 1498-1508 ◽  
Author(s):  
Dorien Leenaerts ◽  
Jef Aernouts ◽  
Pieter Van Der Veken ◽  
Yani Sim ◽  
Anne-Marie Lambeir ◽  
...  
Keyword(s):  
Time Course ◽  
Active Form ◽  
Clot Lysis ◽  
Specific Substrate ◽  
Composite Effect ◽  
Lysis Time ◽  
Supplementary Material ◽  
Maximum Turbidity ◽  
Coagulation And Fibrinolysis

SummaryCarboxypeptidase U (CPU, CPB2, TAFIa) is a basic carboxypeptidase that is able to attenuate fibrinolysis. The inactive precursor procarboxypeptidase U is converted to its active form by thrombin, the thrombin-thrombomodulin complex or plasmin. The aim of this study was to investigate and characterise the time course of CPU generation in healthy individuals. In plasma of 29 healthy volunteers, CPU generation was monitored during in vitro clot lysis. CPU activity was measured by means of an enzymatic assay that uses the specific substrate Bz-o-cyano-Phe-Arg. An algorithm was written to plot the CPU generation curve and calculate the parameters that define it. In all individuals, CPU generation was biphasic. Marked inter-individual differences were present and a reference range was determined. The endogenous CPU generation potential is the composite effect of multiple factors. With respect to the first CPU activity peak characteristics, we found correlations with baseline proCPU concentration, proCPU Thr325Ile polymorphism, time to clot initiation and the clot lysis time. The second CPU peak related with baseline proCPU levels and with the maximum turbidity of the clot lysis profile. In conclusion, our method offers a technique to determine the endogenous CPU generation potential of an individual. The parameters obtained by the method quantitatively describe the different mechanisms that influence CPU generation during the complex interplay between coagulation and fibrinolysis, which are in line with the threshold hypothesis.Supplementary Material to this article is available online at www.thrombosis-online.com.

ASPIRIN ACETYLATES FIBRINOGEN AND ENHANCES FIBRINOLYSIS IN VIVO. FIBRINOLYTIC EFFECT IS INDEPENDENT OF CHANGES IN PLASMINOGEN ACTIVATOR LEVELS

1987 ◽  
Author(s):  
D Thorir ◽  
M D Bjornsson ◽  
Henry Berger
Keyword(s):  
Plasminogen Activator ◽  
Time Course ◽  
Clot Lysis ◽  
Thrombin Time ◽  
Lysis Time ◽  
Coagulation Tests ◽  
Clot Lysis Time ◽  
Increased Susceptibility

In addition to its antiplatelet effect, aspirin has been reported to have fibrinolytic and hypoprothrombinémie effects. The objective of this study was to investigate possible mechanisms underlying the enhanced fibrinolysis observed after aspirin. Five healthy subjects received 650 mg of aspirin ql2hr for five days. Blood samples were collected before aspirin (control) and immediately before (0 hr) and two hours after (2 hr) the last dose for determinations of clot lysis time, time course of thrombin-induced fibrin aggregation, tissue plasminogen activator (tPA), intrinsic pathway fibrinolytic activity (IPFA), plasminogen, fibrinogen, aspirin and salicylic acid, and the coagulation tests activated partial thromboplastin time, thrombin time and prothrombin time. Clot lysis time was shorter after aspirin, control: 9.1±12.4 min (mean±s.d.), 0 hr: 4.6±4.0 min, 2 hr: 5.7±6.2 min (p:0.04), and the fibrin aggregation curves showed increased turbidity (expressed as AUC over 10 min), control: 72.7±17.8 mnumin, 0 hr: 94.6±1.6 mnumin, 2 hr: 112.8±45.1 mnumin (p:0.02). Control values of tPA (0.11±0.04 IU/ml), IPFA (2.20±0.59 IU/ml), plasminogen (10.9±1.0 mg/dl), fibrinogen (288±37 mg/dl), and the coagulation tests were not differnet from those after aspirin. Plasma aspirin concentrations were below detection limits at 0 hr and averaged 1.63±0.97μg/ml at 2 hr. In vitro studies using fibrinogen-free plasma and added acetylated fibrinogen sfiowed an inverse relationship between the extent of acétylation and clot lysis time. Studies using 14C-acetyl-labeled aspirin and fibrinogen showed that fibrinogen is acetylated to form ε-N-acetyl-lysine on both D and E domains of the molecule (50.6±2.0 and 49.4±2.0%, respectively) and on α β and γchains of the molecule (34.4±1.6, 30.7±3.4 and 34.9±4.7%, respectively), with preferential acétylation on the E domain. On the average, 2.88±1.49 ε-N-acetyl-lysyl residues were formed on each fibrinogen molecule. These results suggest that N-acetylation of lysyl residues of fibrinogen is responsible for the increased susceptibility of fibrin clots to lysis after aspirin.

Different mechanisms contribute to the biphasic pattern of carboxypeptidase U (TAFIa) generation during in vitro clot lysis in human plasma

2003 ◽  
Vol 89 (02) ◽  
pp. 264-271 ◽  
Author(s):  
Judith Leurs ◽  
Britt-Marie Wissing ◽  
Viveca Nerme ◽  
Katinka Schatteman ◽  
Petter Björquist ◽  
...  
Keyword(s):  
Human Plasma ◽  
Time Course ◽  
Thrombin Inhibitor ◽  
Clot Lysis ◽  
Time Period ◽  
Lysis Time ◽  
Two Peaks ◽  
Plasmin Inhibitor ◽  
Clot Lysis Time

SummaryCarboxypeptidase U (CPU, TAFIa) recently gained interest as a significant player in dampening the fibrinolytic rate. The aim of this study was to investigate the time course of the generation of CPU activity during coagulation and fibrinolysis using an in vitro clot lysis model in human plasma. A first peak of CPU activity appeared after initiation of the coagulation phase and a second rise in CPU activity was observed during the fibrinolysis. The decrease in the proCPU plasma concentration followed the same trend as the appearance of the CPU activity. The direct thrombin inhibitor inogatran eliminated the CPU generation during coagulation but not during fibrinolysis. Addition of the plasmin inhibitor aprotinin during fibrinolysis resulted in a decrease in CPU activation during the lysis phase. These results demonstrate that proCPU was activated during coagulation by thrombin and during fibrinolysis by plasmin. Addition of a CPU inhibitor before initiation of clotting decreased the clot lysis time as expected. However, addition in the time period between the two peaks of CPU activity had no apparent effect on the clot lysis time.

Identification and characterisation of monoclonal antibodies that impair the activation of human thrombin activatable fibrinolysis inhibitor through different mechanisms

2011 ◽  
Vol 106 (07) ◽  
pp. 90-101 ◽  
Author(s):  
Niraj Mishra ◽  
Ellen Vercauteren ◽  
Jan Develter ◽  
Riet Bammens ◽  
Paul J. Declerck ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Clot Lysis ◽  
Dependent Manner ◽  
Thrombin Activatable Fibrinolysis Inhibitor ◽  
Lysis Time ◽  
Human Thrombin ◽  
Fibrinolysis Inhibitor ◽  
Dose Dependent ◽  
Clot Lysis Time

SummaryThrombin activatable fibrinolysis inhibitor (TAFI) forms a molecular link between coagulation and fibrinolysis and is a putative target to develop profibrinolytic drugs. Out of a panel of monoclonal antibodies (MA) raised against TAFI-ACIIYQ, we selected MA-TCK11A9, MA-TCK22G2 and MA-TCK27A4, which revealed high affinity towards human TAFITI- wt. MA-TCK11A9 was able to inhibit mainly plasmin-mediated TAFI activation, MA-TCK22G2 inhibited plasmin- and thrombin-mediated TAFI activation and MA-TCK27A4 inhibited TAFI activation by plasmin, thrombin and thrombin/thrombomodulin (T/TM) in a dose-dependent manner. These MA did not interfere with TAFIa activity. Using an eightfold molar excess of MA over TAFI, all three MA were able to reduce clot lysis time significantly, i.e. in the presence of exogenous TM, MATCK11A9, MA-TCK22G2 and MA-TCK27A4 reduced clot lysis time by 47 ± 9.1%, 80 ± 8.6% and 92 ± 14%, respectively, compared to PTCI. This effect was even more pronounced in the absence of TM i.e. MATCK11A9, MA-TCK22G2 and MA-TCK27A4 reduced clot lysis time by 90 ± 14%, 140 ± 12% and 147 ± 29%, respectively, compared to PTCI. Mutagenesis analysis revealed that residues at position 268, 272 and 276 are involved in the binding of MA-TCK11A9, residues 147 and 148 in the binding of MA-TCK22G2 and residue 113 in the binding of MATCK27A4. The present study identified three MA, with distinct epitopes, that impair the activation of human TAFI and demonstrated that MATCK11A9 which mainly impairs plasmin-mediated TAFI activation can also reduce significantly clot lysis time in vitro.

The Effect of Erythropoietin on Platelet Function and Fibrinolysis in Chronic Renal Failure

1994 ◽  
Vol 17 (3) ◽  
pp. 141-145 ◽  
Author(s):  
D. Stenver ◽  
L. Jeppesen ◽  
B. Nielsen ◽  
J. Dalsgaard Nielsen ◽  
C. Hædersdal ◽  
...  
Keyword(s):  
Platelet Function ◽  
Clot Lysis ◽  
Platelet Activity ◽  
Platelet Factor ◽  
Human Erythropoietin ◽  
Lysis Time ◽  
Euglobulin Clot Lysis Time ◽  
Erythropoietin Treatment

The influence of erythropoietin therapy on platelet function and fibrinolysis was evaluated in 12 anemic hemodialysis patients. Six months of therapy with human erythropoietin (50 to 80 IU/kg initially) raised the hemoglobin level to 10.8 g/dl but did not increase platelet activity in vivo as measured by beta-thromboglobulin or platelet factor 4. There was no change in the platelet aggregation thresholds in vitro for ADP, adrenaline, thrombin or collagen during treatment. Platelet number and volume were also unaffected. Fibrinolytic activity intensified as erythropoietin treatment proceeded, with a fall of euglobulin clot lysis time and rise in the activity of t-PA. PAI-1 levels also showed a downward trend, without reaching significance. Thus erythropoietin treatment in modest doses does not seem to adversely influence the hemostatic system in patients on hemodialysis.

Inhibition of fibrinolysis by recombinant factor VIIa in plasma from patients with severe hemophilia A

Blood ◽  
2002 ◽  
Vol 99 (1) ◽  
pp. 175-179 ◽  
Author(s):  
Ton Lisman ◽  
Laurent O. Mosnier ◽  
Thierry Lambert ◽  
Evelien P. Mauser-Bunschoten ◽  
Joost C. M. Meijers ◽  
...  
Keyword(s):  
Hemophilia A ◽  
Recombinant Factor Viia ◽  
Factor Viia ◽  
In Vitro Study ◽  
Clot Lysis ◽  
Severe Hemophilia ◽  
Large Variability ◽  
Lysis Time ◽  
Clot Lysis Time

Recombinant factor VIIa (rFVIIa) is a novel prohemostatic drug for patients with hemophilia who have developed inhibitory antibodies. The postulation has been made that hemophilia is not only a disorder of coagulation, but that hyperfibrinolysis due to a defective activation of thrombin activatable fibrinolysis inhibitor (TAFI) might also play a role. In this in vitro study, the potential of rFVIIa to down-regulate fibrinolysis via activation of TAFI was investigated. rFVIIa was able to prolong clot lysis time in plasmas from 17 patients with severe hemophilia A. The prolongation of clot lysis time by rFVIIa was completely abolished by addition of an inhibitor of activated TAFI. The concentration of rFVIIa required for half maximal prolongation of clot lysis time (Clys½-VIIa) varied widely between patients (median, 73.0 U/mL; range, 10.8-250 U/mL). The concentration of rFVIIa required for half maximal reduction of clotting time (Cclot½-VIIa) was approximately 10-fold lower than the Clys½-VIIa value (median, 8.4 U/mL; range, 1.7-22.5 U/mL). Inhibition of TFPI with a polyclonal antibody significantly decreased Clys½-VIIa values (median, 2.6 U/mL; range, 0-86.9 U/mL), whereas Cclot½-VIIa values did not change (median, 7.2 U/mL; range, 2.2-22.5 U/mL). On addition of 100 ng/mL recombinant full-length TFPI, a nonsignificant increase of Clys½-VIIa values was observed (median, 119.2 U/mL; range, 12.3-375.0 U/mL), whereas Cclot½-VIIa values did not change (median, 8.8 U/mL; range, 2.6-34.6 U/mL). In conclusion, this study shows that rFVIIa both accelerates clot formation and inhibits fibrinolysis by activation of TAFI in factor VIII-deficient plasma. However, a large variability in antifibrinolytic potential of rFVIIa exists between patients.

Changes in Platelet Function, Blood Coagulation and Fibrinolysis During Insulin-Induced Hypoglycaemia in Juvenile Diabetics and Normal Subjects

1982 ◽  
Vol 47 (03) ◽  
pp. 254-258 ◽  
Author(s):  
J Dalsgaard-Nielsen ◽  
S Madsbad ◽  
J Hilsted
Keyword(s):  
Insulin Infusion ◽  
Adenosine Diphosphate ◽  
Normal Subjects ◽  
Clot Lysis ◽  
Control Group ◽  
Lysis Time ◽  
Euglobulin Clot Lysis Time ◽  
Juvenile Diabetics ◽  
Clot Lysis Time

SummaryHaemostatic parameters were assessed before insulin induced hypoglycaemia and 0, 1 and 2 fu after discontinuation of insulin infusion in 7 non-diabetics, aged 28 (22-31) years (mean and range), and 8 juvenile diabetics, aged 3L (27-35) years, with a mean duration of diabetes of 4 years. The patients were normoglycaemic for at least L0 hr before the study.Platelet aggregation in vitro was induced by lower adenosine diphosphate (ADP) concentrations in the diabetics than in the controls before hypoglycaemia and 0 and 60 min after insulin infusion. Platelet counts decreased significantly in the diabetics after hypoglycaemia, whereas no changes were seen in the control group. The activated partial thromboplastin time (APTT) was reduced in both groups and significantly lower in the diabetics than in the controls 120 min after insulin infusion.Fibrinogen and factor VIII R: Ag increased after insulin infusion; highest values were seen in the diabetics. The euglobulin clot lysis time (ELT) was reduced in both groups during insulin infusion; L20 min after end of insulin infusion ELT was significantly longer in the diabetics than in the control group.

Correlation of in Vivo Clot Lysis with other Thrombolytic Variables Following Administration of abbokinase ®(Tissue Culture Urokinase), to Anesthetized Dogs

1979 ◽  
Author(s):  
D. Martin ◽  
J. Cain ◽  
J. Chmiel ◽  
S.E. El Masry
Keyword(s):  
Mathematical Model ◽  
Tissue Culture ◽  
Plasma Levels ◽  
Half Life ◽  
Clot Lysis ◽  
Lysis Time ◽  
Euglobulin Lysis Time ◽  
Anesthetized Dogs

An anesthetized dog model, using an extracorporeal loop containing an autologous radioactive clot, was utilized to test the effects of various doses of ABBOKINASE® on the rate of clot lysis and on plasma levels of urokinase, plasmin, antiplasmin, plasminogen and fibrinogen. The effects of ABBOKINASE® on hematocrit, euglobulin lysis time and125 I-clot lysis, in vitro, were also determined. Correlations were sought between plasma urokinase, plasmin, antiplasmin and the rate of clot lysis. Kinetic evaluations of half-lives of urokinase and plasmin and of the rate of regeneration of antiplasmin were made. Some of the conclusions reached were: 1) plasma fibrinogen does not decrease until antiplasmin is depleted and free plasmin appears in blood. 2) plasma urokinase levels are related to the dose infused and decrease with a half-life of about 8 minutes following infusion. 3) the rate of clot lysis in the loop is proportional to the dose of ABBOKINASE® over a defined range of doses and can be fitted to a mathematical model. 4) at lower doses, clot lysis occurs in the absence of measurable free plasmin.

Lysis Timer: a new sensitive tool to diagnose hyperfibrinolysis in liver transplantation

2018 ◽  
Vol 72 (1) ◽  
pp. 58-65 ◽  
Author(s):  
Stéphanie Roullet ◽  
Sylvie Labrouche ◽  
Christine Mouton ◽  
Alice Quinart ◽  
Karine Nouette-Gaulain ◽  
...  
Keyword(s):  
Liver Transplantation ◽  
Laboratory Tests ◽  
Characteristic Curve ◽  
Area Under The Curve ◽  
Clot Lysis ◽  
Signal Acquisition ◽  
Standard Laboratory ◽  
Link Type ◽  
Lysis Time

AimsDiagnosis of hyperfibrinolysis in orthotopic liver transplantation (OLT) remains challenging. Euglobulin clot lysis time (ECLT) is not adapted to clinical situations. ROTEM is specific but seldom sensitive to hyperfibrinolysis. The Lysis Timer assesses ‘Global Fibrinolytic Capacity’ in citrated plasma (GFC/LT). GFC/LT associates reagents for in vitro triggering of the clot (thrombin and calcium) and its lysis (tissue-plasminogenactivator (t-PA)), turbidity signal acquisition by the Lysis Timer, and dedicated software converting the digital signal into an optical curve. A visual check of the curves was systematic to ascertain the lysis time values calculated by the software. The primary aim of this prospective observational study was to evaluate the ability of GFC/LT to recognise hyperfibrinolysis during OLT. The secondary aim was to compare its results with ROTEM maximum lysis (EXTEM ML) and with standard laboratory tests.MethodsThirty consecutive adult patients undergoing OLT were included (NCT03012633). Standard laboratory tests, ROTEM, GFC/LT, ECLT and fibrinolysis parameters were assayed at five sample times.ResultsGFC/LT was correlated with ECLT, plasmin activator inhibitor 1 antigen and activity and t-PA activity (r=0.490, 0.681, 0.643 and –0.359, respectively). Hyperfibrinolysis was defined as ECLT ≤60 min. Receiver operating characteristic curve analysis showed that GFC/LT with a threshold of 31 min detected hyperfibrinolysis with a sensitivity of 0.88 (95% CI 0.73 to 0.96), a specificity of 0.68 (95% CI 0.56 to 0.78) and an area under the curve (AUC) of 0.85 (95% CI 0.74 to 0.94). EXTEM ML >12% did not detect hyperfibrinolysis (sensitivity 0.38 (95% CI 0.24 to 0.55), specificity 0.95 (95% CI 0.86 to 0.99) and AUC 0.60 (95% CI 0.46 to 0.75)).ConclusionsGFC/LT recognised hyperfibrinolysis during OLT with a significant agreement with the other tests of fibrinolysis.Trial registration numberNCT03012633.

Thrombin activatable fibrinolysis inhibitor and its fibrinolytic effect in normal pregnancy

2004 ◽  
Vol 92 (11) ◽  
pp. 1025-1031 ◽  
Author(s):  
Hatem Mousa ◽  
Colin Downey ◽  
Zarko Alfirevic ◽  
Cheng-Hock Toh
Keyword(s):  
Protein S ◽  
Significant Negative Correlation ◽  
Normal Pregnancy ◽  
Clot Lysis ◽  
Thrombin Activatable Fibrinolysis Inhibitor ◽  
Significant Drop ◽  
Soluble Thrombomodulin ◽  
Lysis Time ◽  
Fibrinolysis Inhibitor

SummaryWe investigated changes in both thrombin activatable fibrinolysis inhibitor (TAFI) antigen levels and its functional effect on in vitro fibrinolysis in normal pregnancy. 152 pregnant women and 31 women in the immediate postpartum period were studied, with pregnancy divided into 6 windows at 4 weekly intervals. As TAFI influences and is in turn influenced by components of the protein C (PC) pathway, its measurements were correlated with levels of soluble thrombomodulin, PC, protein S (PS) and the overall phenotype of activated PC resistance (APCR). Compared with mean TAFI levels at booking gestation (6.6 +1.2 μg/ml), levels peaked at 35-39 weeks gestation (9.6 +2 μg/ml, p = 0.001), followed by a significant drop within 24 hours of delivery (7.2 + 1.1 μg/ml). In functional terms, the mean clot lysis time (CLT) (101 + 13 min at booking) also peaked at 35-39 weeks gestation (141 + 42 min, p = 0.007) and dropped after delivery (99 + 33 min), and was significantly correlated with gestational age (r = 0.410, p = 0.001) and could be abrogated in the presence of an inhibitor to TAFI activation. A significant negative correlation was found between TAFI levels and APCR (r = −0.478, p <0.001), APCRV (r = −0.598; p <0.001), PS (r = −0.490, P <0.001) and PC (r = −0.198, p = 0.02). In summary, there is a significant increase in TAFI levels, which translates into increased CLT during pregnancy. Furthermore, changes in TAFI contribute to the increasing APCR of pregnancy.

scholarly journals Impaired Fibrinolysis Predicts Adverse Outcome in Acute Coronary Syndrome Patients with Diabetes: A PLATO Sub-Study

2020 ◽  
Vol 120 (03) ◽  
pp. 412-422 ◽  
Author(s):  
Wael Sumaya ◽  
Lars Wallentin ◽  
Stefan K. James ◽  
Agneta Siegbahn ◽  
Katja Gabrysch ◽  
...  
Keyword(s):  
Acute Coronary Syndrome ◽  
Fibrin Clot ◽  
Clot Lysis ◽  
Bleeding Events ◽  
Coronary Syndrome ◽  
Lysis Time ◽  
Increased Risk ◽  
Patients With Diabetes ◽  
The Relationship ◽  
Maximum Turbidity

AbstractHypofibrinolysis is a key abnormality in diabetes but the role of impaired clot lysis in predicting vascular events and mortality in this population is yet to be determined. We aimed to investigate the relationship between fibrin clot properties and clinical outcomes in patients with diabetes and recent acute coronary syndrome (ACS). Plasma samples were collected at hospital discharge from 974 ACS patients with diabetes randomised to clopidogrel or ticagrelor in the PLATO trial. A validated turbidimetric assay was employed to study fibrin clot lysis and maximum turbidity. One-year rates of cardiovascular (CV) death, spontaneous myocardial infarction (MI) and PLATO-defined major bleeding events were assessed after sample collection. Hazard ratios (HRs) were determined using Cox proportional analysis. After adjusting for CV risk factors, each 50% increase in lysis time was associated with increased risk of CV death/MI (HR 1.21; 95% confidence interval [CI] 1.02–1.44; p = 0.026) and CV death alone (HR 1.38; 1.08–1.76; p = 0.01). Similarly, each 50% increase in maximum turbidity was associated with increased risk of CV death/MI (HR 1.25; 1.02–1.53; p = 0.031) and CV death alone (HR 1.49; 1.08–2.04; p = 0.014). The relationship between lysis time and the combined outcome of CV death and MI remained significant after adjusting for multiple prognostic vascular biomarkers (p = 0.034). Neither lysis time nor maximum turbidity was associated with major bleeding events. Impaired fibrin clot lysis predicts 1-year CV death and MI in diabetes patients following ACS. Clinical Trial Registration URL: http://www.clinicaltrials.gov. Unique identifier NCT00391872.

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