Behaviour and Quantitation of Extrinsic (Tissue-Type) Plasminogen Activator in Human Blood

1983 ◽  
Vol 50 (02) ◽  
pp. 518-523 ◽  
Author(s):  
C Kluft ◽  
A F H Jie ◽  
R A Allen
Keyword(s):  
Plasminogen Activator ◽  
Venous Occlusion ◽  
Tissue Type ◽  
Functional Assay ◽  
Uterine Tissue ◽  
Tissue Type Plasminogen Activator ◽  
Type Plasminogen Activator ◽  
Baseline Values

SummaryFunctional assay of extrinsic (tissue-type) plasminogen activator (EPA) in plasma on fibrin plates was evaluated. Using specific quenching antibodies, we demonstrated the method to be specific for EPA under all conditions tested. Contributions of urokinases and intrinsic activators were excluded. The quantity of EPA in blood samples, as compared with purified uterine tissue activator, shows 1 blood activator unit (BAU) to be comparable to 0.93 ng.The median values for EPA activity for healthy volunteers were: baseline, 1.9 BAU/ml (n = 123); diurnal, 5.5 BAU/ml (n = 12); DDAVP administration, 11.7 BAU/ml (n = 39); exhaustive exercise, 25 BAU/ml (n = 24); venous occlusion (15 min), 35 BAU/ml (n = 61). A large inter-individual variation in EPA activity was found, while individual baseline values tended to be constant for periods of weeks.In vitro in blood EPA activity shows a disappearance of 50% in about 90 min at 37° C; EPA activity in euglobulin fractions is stable for ≤2 hr at 37° C.A rapid decrease in EPA activity occurs in vivo, as noted after extracorporal circulation and exercise stimulation (t½ decay, 2-5 min).

scholarly journals Inhibition of clot lysis and decreased binding of tissue-type plasminogen activator as a consequence of clot retraction

Blood ◽  
1992 ◽  
Vol 79 (6) ◽  
pp. 1420-1427 ◽  
Author(s):  
S Kunitada ◽  
GA FitzGerald ◽  
DJ Fitzgerald
Keyword(s):  
Plasminogen Activator ◽  
Tissue Type ◽  
Clot Lysis ◽  
Plasminogen Activator Inhibitor 1 ◽  
Clot Retraction ◽  
Tissue Type Plasminogen Activator ◽  
Type Plasminogen Activator ◽  
Pai 1

Tissue-type plasminogen activator (t-PA) is less active in vivo and in vitro against clots that are enriched in platelets, even at therapeutic concentrations. The release of radioactivity from 125I-fibrin-labeled clots was decreased by 47% 6 hours after the addition of t-PA 400 U/mL when formed in platelet-rich versus platelet-poor plasma. This difference was not due to the release of plasminogen activator inhibitor-1 (PAI-1) by platelets. Thus, the fibrinolytic activity of t- PA in the supernatant was similar in the two preparations and fibrin autography demonstrated only a minor degree of t-PA-PAI-1 complex formation. Furthermore, a similar platelet-dependent reduction in clot lysis was seen with a t-PA mutant resistant to inhibition by PAI-1. The reduction in t-PA activity correlated with a decrease in t-PA binding to platelet-enriched clot (60% +/- 3% v platelet-poor clot, n = 5). This reduction in binding was also shown using t-PA treated with the chloromethylketone, D-Phe-Pro-Arg-CH2Cl (PPACK) (36% +/- 13%, n = 3), and with S478A, a mutant t-PA in which the active site serine at position 478 has been substituted by alanine (43% +/- 6%, n = 3). In contrast, fixed platelets and platelet supernatants had no effect on the binding or lytic activity of t-PA. Pretreatment with cytochalasin D 1 mumol/L, which inhibits clot retraction, also abolished the platelet- induced inhibition of lysis and t-PA binding by platelets. These data suggest that platelets inhibit clot lysis at therapeutic concentrations of t-PA as a consequence of clot retraction and decreased access of fibrinolytic proteins.

Monoclonal Antibodies to Tissue-Type Plasminogen Activator which Prolong its Clearance In Vivo

1989 ◽  
Vol 61 (02) ◽  
pp. 259-261
Author(s):  
Thomas M Reilly ◽  
Robert M Knabb ◽  
Andrew T Chiu ◽  
David L Bradfute ◽  
Pieter B M W M Timmermans
Keyword(s):  
Monoclonal Antibodies ◽  
Plasminogen Activator ◽  
Half Life ◽  
Cell Receptor ◽  
Tissue Type ◽  
Human Hepatoma Cells ◽  
Tissue Type Plasminogen Activator ◽  
Type Plasminogen Activator

SummaryThree murine monoclonal antibodies to tissue-type plasminogen activator (t-PA) were evaluated for their effects on the binding of iodinated t-PA to cultured human hepatoma cells (Hep G2), and on extending the half-life of t-PA injected into rabbits. Two of the antibodies, AE5 and EG2, significantly inhibited t-PA binding in vitro, and extended the in vivo half-life of t-PA four to five-fold. A third antibody, BA10, which had a much smaller inhibitory effect on t-PA binding, had no influence in extending t-PA’s half-life. MOPC-21, a control antibody not directed to t-PA, had no effect on either test. Our results are the first to correlate different compounds’ effects on t-PA binding with their ability to retard t-PA clearance in vivo, and provide additional evidence for the importance of a liver cell receptor in the t-PA clearance process.

EVIDENCE FOR SYNERGISM BETWEEN TISSUE-TYPE PLASMINOGEN ACTIVATOR AND SINGLE CHAIN UROKINASE ON IN VITRO PLASMA CLOT LYSIS

1987 ◽  
Author(s):  
R S Rappaport ◽  
M R Blume ◽  
R L Vogel ◽  
M H Levner ◽  
P P Hung
Keyword(s):  
Plasminogen Activator ◽  
Tissue Type ◽  
Clot Lysis ◽  
Single Chain ◽  
Plasma Clot ◽  
Tissue Type Plasminogen Activator ◽  
Molar Ratios ◽  
Type Plasminogen Activator

There is mounting evidence from animal models and the clinic that combination thrombolytic therapy with tissue-type plasminogen activator (tPA) and single chain urokinase (scuPA) is synergistic. Yet, efforts to demonstrate synergism between these two plasminogen activators in vitro have met with discordant results. Collen et al (Thromb. Haemostasis, 56:35, 1986) reported an absence of synergism between these two agents on clot lysis in an in vitro plasma milieu when they were evaluated at molar ratios of 1:4 (tPA:scuPA and vice versa). Gurewich and Pannell (Thromb. Res., 44:217, 1986), however, reported a synergistic effect on fibrin-specific clot lysis in vitro when the agents were combined in concentrations exceeding molar ratios of 1:4 (tPA:scuPA). Here, we present evidence that synergism between tPA and scuPA may be demonstrated in vitro provided that the molar ratio of tPA to scuPA exceeds 1:4 and that the concentration of clot bound or unbound tPA is minimized. In order to achieve this experimental condition, the standard in vitro plasma clot lysis assay was modified. Human plasma clots were incubated first for a short time in plasma containing varying amounts of tPA. After incubation, the clots were washed thoroughly and reimmersed in plasma alone or in plasma containing varying amounts of scuPA or tPA. Under these conditions, lysis proceeded at a greater rate and to a greater extent when tPA clots were immersed in plasma containing an appropriate amount of scuPA than when they were immersed in plasma alone or in plasma containing appropriate amounts of tPA. Lysis of untreated clots or clots exposed first to scuPA and then to plasma containing varying amounts of scuPA proceeded far less efficiently with a characteristic lag. The enhanced lysis produced by tPA and scuPA obeyed the classical definition of synergy: the same biological effect can be obtained with two drugs together at algebraic fractional combinations of less than 1 (Berenbaum, M.C., Clin. Exp. Immunol., 28:1-18, 1977). Thus, conditions that more closely mimic the in vivo situation resulting from a bolus injection of tPA followed by infusion with scuPA, may provide a system for duplication of in vivo synergism in. vi tro and investigation of the mechanism thereof.

scholarly journals A monoclonal antibody preventing binding of tissue-type plasminogen activator to fibrin: useful to monitor fibrinogen breakdown during t-PA infusion

Blood ◽  
1986 ◽  
Vol 67 (5) ◽  
pp. 1482-1487 ◽  
Author(s):  
P Holvoet ◽  
HR Lijnen ◽  
D Collen
Keyword(s):  
Plasminogen Activator ◽  
Tissue Type ◽  
Clot Lysis ◽  
Plasminogen Activation ◽  
Plasma Clot ◽  
Tissue Type Plasminogen Activator ◽  
Type Plasminogen Activator ◽  
Additional Support

Abstract One (MA-1C8) of 36 monoclonal antibodies obtained by fusion of P3X63- Ag8–6.5.3 myeloma cells with spleen cells of mice immunized with purified human tissue-type plasminogen activator (t-PA) blocked the activity of t-PA on fibrin plates but not on chromogenic substrates. MA- 1C8 at a concentration of 200 micrograms/mL inhibited plasma clot lysis and binding of t-PA to the clot. MA-1C8 had no influence on the activation of plasminogen by t-PA, which obeys Michaelis-Menten kinetics with Km = 105 mumol/L and kcat = 0.05 s-1; however, it abolished the influence of CNBr-digested fibrinogen on Km. These findings confirm that the stimulatory effect of fibrin on the activation of plasminogen by t-PA is mediated by binding of t-PA to fibrin and provide additional support for the kinetic model. Addition of t-PA to pooled fresh human plasma to a concentration of 5 micrograms/mL resulted in extensive fibrinogen breakdown after incubation for one hour at 37 degrees C or during storage at -20 degrees C for one day. In both instances, fibrinogen degradation was completely prevented by addition of MA-1C8 to a concentration of 200 micrograms/mL of plasma. MA-1C8 also effectively prevented in vitro fibrinogen degradation and in vitro plasminogen activation in plasma samples obtained during infusion of recombinant t-PA in patients with thromboembolic disease. Thus, MA-1C8 is a useful tool for discriminating between in vivo and in vitro fibrinolysis during thrombolytic therapy with t-PA.

scholarly journals Retroviral gene transfer of tissue-type plasminogen activator targets thrombolysis in vitro and in vivo

Gene Therapy ◽  
2007 ◽  
Vol 14 (21) ◽  
pp. 1537-1542 ◽  
Author(s):  
Y S Gong ◽  
K L Zhang ◽  
X G Jiang ◽  
Z W Wang ◽  
Z Q Sun ◽  
...  
Keyword(s):  
Gene Transfer ◽  
Plasminogen Activator ◽  
Tissue Type ◽  
Tissue Type Plasminogen Activator ◽  
Retroviral Gene Transfer ◽  
Type Plasminogen Activator

scholarly journals Lack of interference by heparin with thrombolysis or binding of tissue- type plasminogen activator to thrombi

Blood ◽  
1988 ◽  
Vol 71 (5) ◽  
pp. 1347-1352 ◽  
Author(s):  
ET Fry ◽  
BE Sobel
Keyword(s):  
Plasminogen Activator ◽  
Plasma Concentrations ◽  
Concomitant Administration ◽  
Tissue Type ◽  
Clot Lysis ◽  
Chloromethyl Ketone ◽  
Tissue Type Plasminogen Activator ◽  
Type Plasminogen Activator

Abstract Coronary thrombolysis with t-PA is generally implemented with concomitant administration of heparin. However, results of studies in vitro suggest that heparin competes with fibrin for binding of tissue- type plasminogen activator (t-PA), augments activation of free plasminogen, decreases fibrin specificity, and impairs thrombolysis. To define the biological implications of these observations, we characterized effects of therapeutic concentrations of heparin on the binding of t-PA to thrombi formed in whole blood, effects of heparin on activation of plasminogen by t-PA in plasma, and effects of heparin on thrombolysis induced by t-PA in a clot lysis system designed to simulate conditions in vivo. The amount of t-PA bound to thrombi was not affected by heparin (0, 0.5, 1.0, and 5.0 U/mL). When t-PA activity was selectively and irreversibly inhibited by D-Phe-Pro-Arg- chloromethyl ketone (PPACK) the amount of t-PA-PPACK bound was similarly unaffected by heparin. Thrombolysis measured by 125I- fibrin(ogen) release and by reduction of mass of thrombi were not altered by heparin. Heparin did not affect plasminogen consumption induced by t-PA. Plasma concentrations of alpha-2-antiplasmin after exposure of blood to t-PA were less depressed with increasing concentrations of heparin. Thus, heparin in therapeutic concentrations does not interfere with binding of t-PA to thrombi, augment activation of free plasminogen, or inhibit thrombolysis. Accordingly, it appears likely that concomitant administration of heparin will not impair thrombolysis with t-PA implemented clinically.

The brain-specific tissue-type plasminogen activator inhibitor, neuroserpin, protects neurons against excitotoxicity both in vitro and in vivo

2005 ◽  
Vol 30 (4) ◽  
pp. 552-558 ◽  
Author(s):  
Nathalie Lebeurrier ◽  
Géraldine Liot ◽  
José P. Lopez-Atalaya ◽  
Cyrille Orset ◽  
Monica Fernandez-Monreal ◽  
...  
Keyword(s):  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
Tissue Type ◽  
Tissue Type Plasminogen Activator ◽  
Type Plasminogen Activator ◽  
Specific Tissue ◽  
The Brain ◽  
Activator Inhibitor

Inhibitor-Resistant Tissue-Type Plasminogen Activator: An Improved Thrombolytic Agent In Vitro

1994 ◽  
Vol 71 (01) ◽  
pp. 124-128 ◽  
Author(s):  
R V Shohet ◽  
S Spitzer ◽  
E L Madison ◽  
R Bassel-Duby ◽  
M-J Gething ◽  
...  
Keyword(s):  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
Tissue Type ◽  
Wild Type ◽  
Plasminogen Activator Inhibitor 1 ◽  
Tissue Type Plasminogen Activator ◽  
Type Plasminogen Activator ◽  
Pai 1

SummaryPlatelet-rich clots are inefficiently lysed by current fibrinolytic agents. Platelets contain a great deal of plasminogen activator inhibitor 1 (PAI-1), the principal endogenous inhibitor of tissue-type plasminogen activator (t-PA). We have tested whether PAI-1 resistant t-PAs would be more effective thrombolytic agents in an in vitro model of platelet rich clots. Clots were formed with recalcified human plasma without or with the addition of platelets. The lysis of these clots was followed by the release of incorporated 125I-fibrinogen. Mutant and wild-type t-PA were almost equally effective against clots lacking platelets but the mutant was twice as effective at lysing platelet-rich clots. A mechanism for this effect is suggested by the demonstration that a complex between wild-type t-PA and extruded platelet contents resembles that between purified t-PA and PAI-1 and that the PAI-1 resistant t-PA does not interfere with formation of this adduct. Because of its enhanced ability to lyse platelet-rich clots in vitro, further in vivo work may find that PAI-1 resistant t-PA is a more efficacious therapeutic agent than wild-type t-PA in situations where platelets contribute to the failure of thrombolysis.

scholarly journals Tissue-type plasminogen activator regulates macrophage activation and innate immunity

Blood ◽  
2017 ◽  
Vol 130 (11) ◽  
pp. 1364-1374 ◽  
Author(s):  
Elisabetta Mantuano ◽  
Pardis Azmoon ◽  
Coralie Brifault ◽  
Michael A. Banki ◽  
Andrew S. Gilder ◽  
...  
Keyword(s):  
Plasminogen Activator ◽  
Macrophage Activation ◽  
Innate Immune ◽  
Tissue Type ◽  
Tissue Type Plasminogen Activator ◽  
Type Plasminogen Activator ◽  
Key Points ◽  
Proinflammatory Responses

Key Points Tissue-type plasminogen activator inhibits the activity of the innate immune system in macrophages in vitro and in vivo in mice. Suppression of macrophage proinflammatory responses by tPA requires the NMDA receptor.

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