Factor XIIa Is a Kinetically Favorable Plasminogen Activator

1999 ◽  
Vol 82 (09) ◽  
pp. 1041-1046 ◽  
Author(s):  
Karina Feddersen ◽  
Rasmus Røjkjær ◽  
Inger Schousboe
Keyword(s):  
Competitive Inhibitor ◽  
Assay System ◽  
Clot Lysis ◽  
Factor Xii ◽  
Physiological Concentration ◽  
Lysis Time ◽  
Factor Xiia ◽  
Specificity Constant ◽  
Endpoint Assay

SummaryInitiation of the plasma contact system has been shown to play a significant role in the fibrinolysis, activating both pro-urokinase and plasminogen. The aim of the present study was to further evaluate the functional role of the factor XIIa catalyzed activation of plasminogen. Activation of plasminogen by factor XIIa followed the Michaelis-Menten rate equation. In a continuous assay system the Km was 0.27 μΜ; the kcat 0.078 min−1 and the kcat/Km 0.31 × 106 Μ−1 × min−1. In an endpoint assay system the Km was 0.58 μΜ; the kcat 0.096 min−1 and the kcat/Km 0.16 × 106 Μ−1 × min−1. The discrepancy between the kcat in the two assays is not significant. Theoretically the higher Km in the end-point assay system may be due to the presence or generation of an unidentified competitive inhibitor in this assay system. Comparing the catalytic constants of factor XIIa with those of urokinase activation of plasminogen, the specificity constant, kcat/Km, of factor XIIa activation of plasminogen was 20-fold lower. However, taking the low physiological concentration of urokinase into account, the efficiency of activated factor XII is equivalent to that of urokinase. When monitoring factor XIIa activation of plasminogen in a clot lysis assay, the clot lysis time was 2- to 4-fold as long as that accommodated by urokinase at factor XIIa concentrations equivalent with 5-20% of the zymogen concentration in plasma. The factor XIIa mediated clot lysis was prevented completely by the presence of a polyclonal antibody to factor XII.

Platelet-Bound Prekallikrein Promotes Pro-Urokinase-Induced Clot Lysis: A Mechanism for Targeting the Factor XII Dependent Intrinsic Pathway of Fibrinolysis

1994 ◽  
Vol 71 (03) ◽  
pp. 347-352 ◽  
Author(s):  
Jean-Pierre Loza ◽  
Victor Gurewich ◽  
Michael Johnstone ◽  
Ralph Pannell
Keyword(s):  
Platelet Rich Plasma ◽  
Specific Inhibitor ◽  
Specific Effect ◽  
Clot Lysis ◽  
Factor Xii ◽  
Intrinsic Pathway ◽  
Clot Retraction ◽  
Enzymatic Mechanism ◽  
Low Concentrations ◽  
Factor Xiia

SummaryClots formed from platelet rich plasma were found to be lysed more readily by low concentrations of pro-urokinase (pro-UK) than clots formed from platelet poor plasma. This was not a non-specific effect since the reverse occurred with tissue plasminogen activator. A mechanical explanation due to platelet-mediated clot retraction was excluded by experiments in which retraction was inhibited with cyto-chalasin B. Therefore, a platelet-mediated enzymatic mechanism was postulated to explain the promotion of fibrinolysis. Casein autography of isolated platelets revealed a ≈ 90 kDa band of activity which comigrated with plasma prekallikrein (PK)/kallikrein, a known activator of pro-UK. Furthermore, treatment of platelets with plasma PK activator (PPA), consisting essentially of factor XIIa, induced activation of pro-UK and of chromomgenic substrate for kallikrein (S-2302). This activity corresponded to approximately 40-200 pM kallikrein per 10 8 washed and gel filtered platelets per ml. The activation of pro-UK by PPA-pretreated platelets was dose-dependent and inhibited by soybean trypsin inhibitor but not by bdellin, a specific inhibitor of plasmin, nor by the corn inhibitor of factor XIIa. Kinetic analysis of pro-UK activation by kallikrein showed promotion of the reaction by platelets. The KM of the reaction was reduced by platelets by ≈ 7-fold, while the kcat was essentially unchanged. In conclusion, PK was shown to be tightly associated with platelets where it can be activated by factor XIIa during clotting. The activation of pro-UK by platelet-bound kallikrein provides an explanation for the observed platelet mediated promotion of pro-UK-induced clot lysis. Since pro-UK and plasminogen have also been shown to be associated with platelets, the present findings suggest a mechanism by which the factor Xlla-dependent intrinsic pathway of fibrinolysis can be localized and targeted to a thrombus.

scholarly journals Rapid Loss of Factor XII- and XI-Activity in Ellagic Acid Activated Normal Plasma: Role of Plasma Inhibitors and Implications for Automated Aptt-Recording

1977 ◽  
Author(s):  
J. H. Joist ◽  
J. F. Cowan ◽  
M. Khan
Keyword(s):  
Ellagic Acid ◽  
Clinical Laboratory ◽  
Coagulation Factors ◽  
Activation Process ◽  
Factor Xii ◽  
Naturally Occurring ◽  
Normal Plasma ◽  
Factor Xiia ◽  
Loss Of Contact

Rapid prolongation of the activated partial thromboplastin time (aPTT) of normal plasma upon incubation with ellagic acid containing aPTT reagents was observed. The aPTT prolongation was not due to time dependent changes in pH in the incubation mixtures or loss of activity of the labile coagulation factors VIII and V, but occurred as a result of rapid progressive inactivation of ellagic acid activated factors XII and XI. Prolongation of the aPTT and loss of contact factor activities was not observed in plasma incubated with particulate activator reagents. This finding seemed to indicate that adsorption of factors XII and XI to larger particles during the activation process may protect these factors from inactivation by naturally occurring plasma inhibitors. Evidence is presented which supports previous observations that Ci-inhibitor, α1-antitrypsin and antithrombin III (in the presence of heparin contribute to factor XIIa- and XIa-inactivation in ellagic acid activated plasma and that plasma albumin may compete with factor XII for ellagic acid binding. The findings indicate that ellagic acid containing aPTT reagents have unfavorable properties which seriously limit their usefulness in the clinical laboratory, particularly in respect to recording of the aPTT with certain fully automated clot timers.

Histidine-rich glycoprotein binds factor XIIa with high affinity and inhibits contact-initiated coagulation

Blood ◽  
2011 ◽  
Vol 117 (15) ◽  
pp. 4134-4141 ◽  
Author(s):  
Jessica L. MacQuarrie ◽  
Alan R. Stafford ◽  
Jonathan W. Yau ◽  
Beverly A. Leslie ◽  
Trang T. Vu ◽  
...  
Keyword(s):  
Partial Thromboplastin Time ◽  
Activated Partial Thromboplastin Time ◽  
Factor Xii ◽  
Intrinsic Pathway ◽  
High Affinity ◽  
Factor Xiia ◽  
Platelet Release ◽  
Contact Pathway ◽  
Physiologic Role

Abstract Histidine-rich glycoprotein (HRG) circulates in plasma at a concentration of 2μM and binds plasminogen, fibrinogen, and thrombospondin. Despite these interactions, the physiologic role of HRG is unknown. Previous studies have shown that mice and humans deficient in HRG have shortened plasma clotting times. To better understand this phenomenon, we examined the effect of HRG on clotting tests. HRG prolongs the activated partial thromboplastin time in a concentration-dependent fashion but has no effect on tissue factor–induced clotting, localizing its effect to the contact pathway. Plasma immunodepleted of HRG exhibits a shortened activated partial thromboplastin time that is restored to baseline with HRG replenishment. To explore how HRG affects the contact pathway, we examined its binding to factors XII, XIIa, XI, and XIa. HRG binds factor XIIa with high affinity, an interaction that is enhanced in the presence of Zn2+, but does not bind factors XII, XI, or XIa. In addition, HRG inhibits autoactivation of factor XII and factor XIIa–mediated activation of factor XI. These results suggest that, by binding to factor XIIa, HRG modulates the intrinsic pathway of coagulation, particularly in the vicinity of a thrombus where platelet release of HRG and Zn2+ will promote this interaction.

scholarly journals Human Prekallikrein (Plasminogen Proactivator): Purification, Characterization and Activation by Activated Factor XII

1977 ◽  
Author(s):  
Bonno N. Bouma ◽  
John H. Griffin
Keyword(s):  
Human Plasma ◽  
Plasminogen Activator ◽  
Clot Lysis ◽  
Factor Xii ◽  
Globulin Fraction ◽  
Factor Xi ◽  
Form Analysis ◽  
Plasminogen Activator Activity ◽  
Fibrin Plate ◽  
Factor Xiia

In order to resolve conflicting reports about the possible identity of prekallikrein and Factor XII-dependent plasminogen proactivator (FXII-PPA), the γ-globulin fractions of prekallikrein-deficient (Fletcher trait) and of normal plasma were assayed for FXII-PPA. Based on both fibrin plate and clot lysis tests, FXII-PPA in the γ-globulin fractions of prekallikrein-deficient plasmas from 2 unrelated patients was undetectable, i.e. <1% of the FXII-PPA in the normal γ-globulin fraction. However, PPA independent of FXII was detected in both the Fletcher and the normal γ-globulin fractions at 4% of the FXII-PPA present in the normal γ-globulin fraction.Human plasma prekallikrein was purified 2,000-fold (specific clotting activity 22 units/mg) and was greater than 95% homogeneous on SDS-gels. FXII-PPA was always copurified with prekallikrein and was totally separated from Factor XI. No Factor XII-dependent or Factor XII-independent plasminogen activator activity was detected in purified Factor XI preparations at 40 units/ml. Purified prekallikrein in its precursor form gave 2 protein bands on SDS-gels at 82,000 and 78,000 MW. Upon reduction, a single 85,000 MW band was observed. Kallikrein and plasminogen activator activity were generated upon incubation with purified human Factor XIIa (28,000 MW form). Analysis of this reaction mixture on SDS-gels without reduction showed 2 bands with apparently identical MW’s as the precursor protein bands, whereas reduction showed cleavage of both protein bands.These results suggest that prekallikrein is identical to FXII-PPA in normal human plasma and that activation of this zymogen by Factor XIIa involves limited proteolytic cleavage.

DEMONSTRATION OF Cl-INHIBITOR COMPLEXES IN PLASMA BY HIGHLY SENSITIVE RADIOIMMUNOASSAYS USING A MONOCLONAL ANTIBODY AGAINST A NEODETERMINANT ON COMPLEXED Cl-INHIBITOR

1987 ◽  
Author(s):  
J H Nuijens ◽  
C C M Huijbregets ◽  
L G Thijs ◽  
C E Hack
Keyword(s):  
Monoclonal Antibody ◽  
Contact System ◽  
Factor Xii ◽  
Optimal Conditions ◽  
Spleen Cells ◽  
Highly Sensitive ◽  
Factor Xiia

Levels of factor XIIa- and kallikrein-Cl inhibitor (Cl-Inh) complexes in plasma reflect activation of the contact system in vivo. Here, we report the development of radioimmunoassays (RIAs) for these complexes using a monoclonal antibody (mAb K0K12) that reacts with a neodeterminant exposed on Cl-Inh after interaction with proteases. mAb K0K12 was obtained by a fusion experiment with spleen cells of a mouse hyperimmunized with Cl-Inh complexes.Experiments with purified Cl-Inh incubated with either Cls or elastase revealed that the determinant for mAb KOK12 is exposed on complexed as well as proteolytically inactivated (modified) Cl-Inh.Radioimmunoassays (RIAs) for the detection of factor Xlla-Cl-Inh and kallikrein-Cl-Inh complexes were performed as follows: mAb K0K12 was coupled to Sepharose and incubated with the sample to be tested. Binding of Cl-Inh complexes was detected by a subsequent incubation with 125I-antibodies against factor XII or (pre)kallikrein.With these RIAs, activation of 0.1% of factor XII or prekal-likrein in plasma is easily detected.Optimal conditions for blood sampling and processing were established, i.e. conditions that prevented any in vitro activation of factor XII and prekallikrein. Levels of factor XIIa-Cl-Inh and kallikrein-Cl-Inh complexes in plasma samples from normal donors were less than 0.1 U/ml (100 U/ml is the maximal amount of Cl-Inh complexes generated in pooled plasma by DXS). Considerably higher, and fluctuating levels were observed in patients with diseases such as septicaemia. These highly sensitive RIAs will facilitate studies concerning the role of the contact system in human pathophysiology.

PLASMA-CLOT LYSIS INDUCED BY MONOCLONAL ANTIBODY AGAINST α2-PLASMIN INHIBITOR

1987 ◽  
Author(s):  
Y Sakata ◽  
J Mimuro ◽  
Y koike
Keyword(s):  
Clot Lysis ◽  
Blood Collection ◽  
Plasma Clot ◽  
Specific Patient ◽  
Lysis Time ◽  
Pi Deficiency ◽  
Dose Dependent ◽  
Plasmin Inhibitor

A Monoclonal antibody (MCA) against α2-plasmin inhibitor α2-Pl) designated as JTP-1 inhibited antiplasmin act ivity and complex formation of a2-PI with plasmin. By using this MCA we tried to observe plasma-clot lysis (CL) in vitro and to estimate the level of total fibrinolytic capacity in plasma. As reported previously (Blood 55: U83, 1980) spontaneous CL is a striking feature of the plasma derived from a patient with α2-PI deficiency. We showed that this abnormally increased fibrinolysis was solely due to the deficiency of α2-PI. However, the contribution of plasminogen activator (PA) and its inhibitor to this specific patient' plasma -CL has been under discussion. Therefore, to test whether similar CL can be found in normal plasma without an addition of PA, plasma clots were made after incubation of plasma from normal volunteers containing 125i fibrinogen with various concentration of JTP-1, and fibrinolysis was measured by counting the soluble radioactivity. The addition of JTP-1 to plasma led to a dose-dependent enhancement of the soluble 125i fragment-release from the clot. However, JTP-1 had no effect on a2-PI-deficient plasma-CL. Other anti-a2-PI MCAs whose epitopes were not involved in the reactive site of a2-PI had no effect on CL and rabbit anti-mouse immunoglobulin IgG neutralized this JTP-l-inducing CL completely. Immunodepletion of tissue PA (tPA) or plasminogen from plasma decreased the rate of CL but that of prourokinase did not. To determine the role of PA inhibitor (PAl) released from platelets (pits) in the regulation of CL in vitro, plasma clots were made from pits poor plasma (PPP) and pits rich plasma (PRP), and CL was observed in the presence of JTP-1. There was almost no difference of the lysis time between PPP clot and PRP clot, although plasma clot from pregnant woman was lysed slowly. These results strongly suggest that endogenous t-PA in plasma is still functionally active after blood collection and CL is mainly prevented by a2-PI in vitro.-plasmin inhibitor (a2-Pl) designated as JTP-1 inhibited antiplasmin act ivity and complex formation of a2-PI with plasmin. By using this MCA we tried to observe plasma-clot lysis (CL) in vitro and to estimate the level of total fibrinolytic capacity in plasma. As reported previously (Blood 55: U83, 1980) spontaneous CL is a striking feature of the plasma derived from a patient with a2-PI deficiency. We showed that this abnormally increased fibrinolysis was solely due to the deficiency of a2-PI. However, the contribution of plasminogen activator (PA) and its inhibitor to this specific patient' plasma -CL has been under discussion. Therefore, to test whether similar CL can be found in normal plasma without an addition of PA, plasma clots were made after incubation of plasma from normal volunteers containing 125i fibrinogen with various concentration of JTP-1, and fibrinolysis was measured by counting the soluble radioactivity. The addition of JTP-1 to plasma led to a dose-dependent enhancement of the soluble 125i fragment-release from the clot. However, JTP-1 had no effect on a2-PI-deficient plasma-CL. Other anti-a2-PI MCAs whose epitopes were not involved in the reactive site of a2-PI had no effect on CL and rabbit anti-mouse immunoglobulin IgG neutralized this JTP-l-inducing CL completely. Immunodepletion of tissue PA (tPA) or plasminogen from plasma decreased the rate of CL but that of prourokinase did not. To determine the role of PA inhibitor (PAl) released from platelets (pits) in the regulation of CL in vitro, plasma clots were made from pits poor plasma (PPP) and pits rich plasma (PRP), and CL was observed in the presence of JTP-1. There was almost no difference of the lysis time between PPP clot and PRP clot, although plasma clot from pregnant woman was lysed slowly. These results strongly suggest that endogenous t-PA in plasma is still functionally active after blood collection and CL is mainly prevented by a2-PI in vitro.-plasmin inhibitor (a2-Pl) designated as JTP-1 inhibited antiplasmin act ivity and complex formation of a2-PI with plasmin. By using this MCA we tried to observe plasma-clot lysis (CL) in vitro and to estimate the level of total fibrinolytic capacity in plasma. As reported previously (Blood 55: U83, 1980) spontaneous CL is a striking feature of the plasma derived from a patient with α2-PI deficiency. We showed that this abnormally increased fibrinolysis was solely due to the deficiency of a2-PI. However, the contribution of plasminogen activator (PA) and its inhibitor to this specific patient' plasma -CL has been under discussion. Therefore, to test whether similar CL can be found in normal plasma without an addition of PA, plasma clots were made after incubation of plasma from normal volunteers containing 125i fibrinogen with various concentration of JTP-1, and fibrinolysis was measured by counting the soluble radioactivity. The addition of JTP-1 to plasma led to a dose-dependent enhancement of the soluble 125i fragment-release from the clot. However, JTP-1 had no effect on a2-PI-deficient plasma-CL. Other anti-a2-PI MCAs whose epitopes were not involved in the reactive site of a2-PI had no effect on CL and rabbit anti-mouse immunoglobulin IgG neutralized this JTP-l-inducing CL completely. Immunodepletion of tissue PA (tPA) or plasminogen from plasma decreased the rate of CL but that of prourokinase did not. To determine the role of PA inhibitor (PAl) released from platelets (pits) in the regulation of CL in vitro, plasma clots were made from pits poor plasma (PPP) and pits rich plasma (PRP), and CL was observed in the presence of JTP-1. There was almost no difference of the lysis time between PPP clot and PRP clot, although plasma clot from pregnant woman was lysed slowly. These results strongly suggest that endogenous t-PA in plasma is still functionally active after blood collection and CL is mainly prevented by α2-PI in vitro.

Urokinase-Related Fibrinolytic Activity In Human Plasma

1981 ◽  
Author(s):  
G Wijngaards ◽  
C Kluft
Keyword(s):  
Fibrinolytic Activity ◽  
Double Diffusion ◽  
Clot Lysis ◽  
Factor Xii ◽  
High Concentration ◽  
Related Activity ◽  
Precipitation Line ◽  
Lysis Time ◽  
Different Types ◽  
Antigenic Material

Blood represents a complex system containing plasminogenn activators. At present, three different types of plasminogen activator activity are distinguished. It has been assumed that urokinase is not one of the blood activators, but recently an improbably high concentration of urokinase antigenic material has been claimed to be present in plasma.The present study was performed to establish if urokinase activity participates in blood fibrinolysis. This was investigated by measuring quenching by antibodies. The dextran sulphate euglobulin fraction (DEF) of plasma was prepared and its fibrinolytic activity was measured in a clot lysis time method with and without addition of various anti-urokinase immunoglobulin fractions (AUK’s).Considerable quenching of the DEF activity by the AUK’s was found. The quenchings reached maximum values at high antibody concentrations. The quenching curves of the fibrinolytic activity of both urokinase and DEF could be superimposed for each AUK. This quenchable activity was also very sensitive to inhibition by a specific urokinase inhibitor from human placenta. Addition of kaolin to DEF, to include factor XII-dependent fibrinolysis, showed an increase of the fibrinolytic activity which was not quenched by AUK. Double diffusion analyses did not result in the demonstration of urokinase antigen in plasma. This was in agreement with the amount of activity quenched (comparable with about 1 ng urokinase/ml plasma) and the detection limit for a visible precipitation line.These observations indicate that plasma contains urokinase-related activity. This activity is assumed to belong to the factor XII-independent proactivator system.

scholarly journals Haemostasis and High Altitude

1977 ◽  
Author(s):  
I. Singh ◽  
I.S. Chohan
Keyword(s):  
Factor Viii ◽  
High Altitude ◽  
Bleeding Time ◽  
Clot Lysis ◽  
Factor Xii ◽  
Factor V ◽  
Platelet Adhesiveness ◽  
Clot Retraction ◽  
Platelet Factor ◽  
Lysis Time

On arrival at high altitude there is a tendency towards hypercoaguiation associated with an increase in platelet count, factors X and XII, and thrombo-test activity (TA) which is reflected by decrease in prothrombin time (PT), bleeding time (BT), clotting time in glass (CT-gl) and in silicone (CT-sl), and stypven time (ST). Clot retraction is impaired. This hypercoaguiation state is countered by a compensatory rise in fibrinolytic activity reflected by reduction of clot lysis time (CLT), plasma fibrinogen, and factor VIII.This hypercoaguiation state persists throughout the early fortnight after arrival at high altitude and then starts regressing. On day 3, factor V decreases, BT, and ST are further reduced, and factor VIII shows a rise. On day 7, a progressive rise occurs in factors V, VIII, X and XII, TA, platelet counts and platelet factor 3 (PF-3).CT-gl, CT-sl, PT and ST are further shortened. On day 14, haematocrit rises and of all the parameters, factors V and X and clot retraction return to normal. Throughout the fortnight, factor XII remains high, CLT is short, and platelet adhesiveness remains within normal range.After 2 years’ stay at high altitude, a regression in hypercoagulability occurs and is indicated by persistent short CLT, prolonged BT, CT-sl, PT, ST and reduced TA. Platelet adhesiveness, PF-3, factors V, VIII and XII and clot retraction are restored to normal.

scholarly journals Mechanism of transient adsorption of fibrinogen from plasma to solid surfaces: role of the contact and fibrinolytic systems

Blood ◽  
1988 ◽  
Vol 71 (4) ◽  
pp. 932-939 ◽  
Author(s):  
JL Brash ◽  
CF Scott ◽  
P ten Hove ◽  
P Wojciechowski ◽  
RW Colman
Keyword(s):  
Solid Surfaces ◽  
Factor Xii ◽  
Contact Activation ◽  
Transient Detection ◽  
Normal Plasma ◽  
Plasma Factor ◽  
Factor Xiia ◽  
Factor Xia ◽  
Specific Proteins

The transient detection of fibrinogen on surfaces has been described (Vroman effect) and high-mol-wt kininogen (HK) has been shown to play a role in this reaction. In this study, we attempted to identify the form of HK responsible for preventing detection of the fibrinogen initially adsorbed from plasma to various artificial surfaces and to determine if other plasma components were involved. We compared 125I-fibrinogen adsorption in the presence of normal plasma to plasma deficient in specific proteins. On all surfaces tested, we found that fibrinogen was displaced from the surface. The extent of displacement was greatly reduced, however, but not eliminated in HK-deficient plasma. Factor XII- deficient plasma also showed reduced fibrinogen displacement. These data indicate that HK can actually displace fibrinogen; however, factor XII, or a factor XII-mediated reaction also appears to be necessary for this displacement to occur. Furthermore, when normal plasma was first subjected to extensive contact activation by dextran sulfate, during which the HK was extensively degraded to components smaller than the light chain (as assessed by Western blotting), we observed greatly reduced displacement of fibrinogen. Extensive contact activation of Factor XI-deficient plasma failed to show low-mol-wt derivatives, however, and displacement of fibrinogen was similar to normal plasma that had not undergone extensive activation. These data indicate that HKa (active cofactor produced during contact activation by factor XIIa or kallikrein) is primarily responsible for displacing fibrinogen, and that HKi (inactive cofactor generated by factor XIa) cannot displace fibrinogen. The fibrinogen from all plasma samples looked similar by Western blot analysis, suggesting that fibrinogenolysis was not a component of the Vroman effect. In addition, experiments performed with plasma prechromatographed on lysine agarose showed that a lysine- agarose adsorbable protein may be minimally involved in fibrinogen desorption and a synergism may exist between HK and that protein.

Thrombomodulin in Human Plasma Contributes to Inhibit Fibrinolysis through Acceleration of Thrombin-dependent Activation of Plasma Procarboxypeptidase B

1998 ◽  
Vol 79 (02) ◽  
pp. 371-377 ◽  
Author(s):  
Yoshitaka Hosaka ◽  
Yasuo Takahashi ◽  
Hidemi Ishii
Keyword(s):  
Human Plasma ◽  
Clot Lysis ◽  
Direct Inhibition ◽  
Carboxypeptidase B ◽  
Lysis Time ◽  
Tissue Plasminogen ◽  
Dependent Inhibition ◽  
Normal Human ◽  
Clot Lysis Time

SummaryThrombomodulin (TM) expressed on endothelial cells binds thrombin and initiates anticoagulant pathways. Soluble functional proteolytic fragments of TM are also present in circulating plasma. Recently, it was reported that TM accelerated thrombin-dependent plasma procarboxypeptidase B (pro-pCPB) activation in a purified system and suggested that TM may inhibit fibrinolysis in crude plasma. The aim of present study was to evaluate any functional role of soluble TM fragments in plasma or purified TM added into plasma to the regulation of coagulation and fibrinolysis. Addition of rabbit TM (1-200 ng/ml) to plasma resulted in a concentration-dependent prolongation of urokinase (UK)- or tissue plasminogen activator (t-PA)-induced clot lysis time. The concentration of TM required for the inhibition of fibrinolysis was lower than that required for the inhibition of coagulation. Addition of anti-rabbit TM IgG or anti-human TM IgG into plasma reduced UK- or t-PA-induced clot lysis time without affecting clotting times, indicating that exogenous TM or soluble TM fragments in normal human plasma participated in regulation of fibrinolysis. Moreover, the TM-dependent inhibition of fibrinolysis was observed only in the presence of thrombin and blocked by addition of carboxypeptidase B inhibitors, but not mediated by protein C activation or direct inhibition of UK, t-PA or plasmin. Analysis of various substrates and inhibitors indicated that TM accelerated thrombin-dependent pro-pCPB activation in plasma. The present results indicate that TM, including soluble TM fragments in plasma, inhibit fibrinolysis via activation of pro-pCPB in plasma.

Sign in / Sign up

Export Citation Format

Share Document