Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.73 (urokinase-type plasminogen activator)
10,685 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Adenosine 3',5'-cyclic monophosphate (cAMP) elevation in cultured rat mesangial cells causes urokinase-dependent adhesion loss, stress-fiber fragmentation, and shape change. Thrombin cleaves single-chain urokinase (scu-PA), causing its inactivation, but not two-chain u-PA [tcu-plasminogen activator (PA)] or tissue-type PA. We tested the ability of thrombin to inhibit the effects of cAMP elevation in mesangial cells and inactivate cell-associated scu-PA. In an assay of trypsin-sensitive adhesion, 65.9% of control cells and 5.5% of cells treated with isoproterenol + methylisobutylxanthine (IM) remained adherent. In the presence of 0.01, 0.1, 1.0, and 10.0 unit/ml thrombin, 20.9, 46.6, 50.4, and 53.3%, respectively, of IM-treated cells remained attached. Thrombin also inhibited stress-fiber fragmentation and shape change. The effects of thrombin were blocked by hirudin or antithrombin III plus heparin. Direct zymography in gels containing gelatin and plasminogen revealed loss of a closely spaced pair of PA bands with thrombin treatment (1.0 unit/ml). Hirudin blocked the loss. alpha-Thrombin inactivated by diisopropyl fluorophosphate neither inhibited shape change nor caused loss of the PA bands; however, gamma-thrombin was nearly as active as native alpha-thrombin in both regards. Pretreatment of the cells with as little as 1.0 unit/ml thrombin for 1.0 min caused marked inhibition of shape change and near total loss of the slower migrating u-PA band (of the doublet). The faster migrating band was inhibited less. The results indicate that the slower migrating band represents scu-PA; the nature of the faster migrating band is less certain. Thrombin reversed the adhesion loss and shape change caused by 8-(4-chlorophenylthio)-cAMP and MIX. Thus physiological concentrations of thrombin rapidly inactivate mesangial cell scu-PA and inhibit and reverse cAMP-stimulated adhesion loss and shape change.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Regulation of mesangial cell adhesion and shape by thrombin. 165 8

Interaction of vitronectin with glia-derived nexin (GDN), thrombin, and the complex GDN-thrombin was demonstrated in direct binding assays that indicated the formation of binary and ternary complexes. The concentration of vitronectin necessary to obtain 50% saturation of the immobilized GDN-thrombin complex binding sites (EC50) was about 1 nM. Under similar experimental conditions, the EC50 of vitronectin for the immobilized antithrombin-III-thrombin complex was about fivefold higher. A tight complex was also formed between vitronectin and immobilized GDN (EC50 approximately 1.5 nM) but when vitronectin was immobilized, GDN displayed a reduced affinity for vitronectin (EC50 approximately 10 nM). These results suggest differences between the immobilized and free conformations of GDN and/or vitronectin. In contrast, vitronectin displayed negligible affinity for antithrombin III. Biotinylated GDN was used to characterize further the binding of GDN or the GDN-thrombin complex to vitronectin. The interaction of the biotinylated GDN-thrombin complex with immobilized vitronectin (EC50 approximately 2 nM) was completely blocked by nonbiotinylated complexes of thrombin with either GDN or antithrombin III, whereas free GDN, free thrombin and the GDN-trypsin complex were only weak competitors. Active-site-blocked urokinase and the complex GDN-urokinase also strongly competed for binding of the biotinylated GDN-thrombin complex to vitronectin. Binding of biotinylated GDN to immobilized vitronectin was specific, saturable and was competed with decreasing efficiency by the GDN-thrombin complex, free GDN and free antithrombin III. These interactions between the adhesive component vitronectin and the serine protease inhibitor GDN may relate to localized control of thrombin and/or urokinase action at certain extravascular sites. These results are discussed in terms of binding sites for vitronectin on GDN, thrombin, and the GDN-thrombin complex.
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PMID:Specific interaction of vitronectin with the cell-secreted protease inhibitor glia-derived nexin and its thrombin complex. 169 27

Three chimeric mutants of plasminogen activator inhibitor 1 (PAI-1) have been constructed where the strained loop of wild type PAI-1 (wtPAI-1) has been replaced with a 19-amino acid region from either plasminogen activator inhibitor 2 (PAI-2), antithrombin III, or with an artificial serine protease inhibitor superfamily consensus strained loop. The inhibitors were expressed in Escherichia coli, and the purified proteins had specific activities toward urokinase-type plasminogen activator (uPA) or the single- and two-chain forms of tissue type plasminogen activator (tPA) that were similar to wtPAI-1. Experiments suggest that the strained loop of PAI-1 is not responsible for the transition between the latent and the active conformations or for binding to vitronectin. Second-order rate constants for the interactions with uPA and single- or two-chain tPA were similar to those of wtPAI-1. Values range from a low of 1.8 x 10(5) M-1 s-1 for the interaction of the PAI-2 chimera with single-chain tPA to a high value of 1.6 x 10(7) M-1 s-1 for the consensus mutant with two-chain tPA. This former value is 200 times higher than the reported rate constant for the interaction between PAI-2 and single-chain tPA, suggesting that structures outside of the strained loop are responsible for the major differences in specificity between PAI-1 and PAI-2.
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PMID:Structure-function studies of the SERPIN plasminogen activator inhibitor type 1. Analysis of chimeric strained loop mutants. 170 Jul 86

Plasminogen activator inhibitor type 1 (PAI-1), the fast-acting inhibitor of tissue-type plasminogen activator (t-PA) and urokinase (u-PA), is a member of the serpin superfamily of proteins. Both in plasma and in the growth substratum of cultured endothelial cells, PAI-1 is associated with its binding protein vitronectin, resulting in a stabilization of active PAI-1. Recently, it has been demonstrated that the PAI-1-binding site on vitronectin is adjacent to a heparin-binding site (Preissner et al., 1990). Furthermore, it can be deduced that the amino acid residues, proposed to mediate heparin binding in the serpins antithrombin III and heparin cofactor II, are conserved in PAI-1. Consequently, here we have investigated whether PAI-1 also interacts with heparin. At pH 7.4, PAI-1 quantitatively binds to heparin-Sepharose and can be eluted with increasing [NaCl]. Binding of PAI-1 to heparin-Sepharose can be efficiently competed with heparin in solution (IC50, 7 microM). In the presence of heparin, the protease specificity of PAI-1 toward thrombin is substantially increased. This is shown by (i) quenching of thrombin activity of PAI-1 in the presence of heparin and (ii) induction of the formation of SDS-stable complexes between thrombin and PAI-1 by heparin. In a dose response curve, both effects reached a maximum at approximately 1 unit/mL and then diminished again upon further increasing the heparin concentration, strongly suggesting a template mechanism as an explanation for the observed effect. In contrast to vitronectin, heparin does not stabilize the active conformation of PAI-1.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Functional interaction of plasminogen activator inhibitor type 1 (PAI-1) and heparin. 170 36

The changes in relevant haemostatic parameters during the course of ten orthotopic liver transplantation were studied when aprotinin was given intra-operatively. Increases of tissue-type (P = 0.008) and urokinase-type (P = 0.009) plasminogen activators during the anhepatic phase could be correlated with hyperfibrinolysis. Thrombin-antithrombin III complexes (TAT) increased after revascularization of the liver graft (P = 0.003). Parallel studies in the perfusate showed that TAT concentrations were 350% and protease inhibitor activities (antithrombin III, protein C) only 52% of the systemic circulation before reperfusion, suggesting that thrombin activation together with protease inhibitor consumption occurs during graft liver reperfusion. The relatively smaller increases in profibrinolytic parameters and a lower blood loss when compared with other groups may be explained by aprotinin administration in our patients.
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PMID:Coagulation changes and the influence of the early perfusate in the course of orthotopic liver transplantation (OLT) when aprotinin is used intra-operatively. 172 11

In order to evaluate precisely the fibrinolytic states in clinical disorders, plasma levels of D dimer (cross-linked fibrin degradation products) were measured by a newly developed, rapid quantitative method based on the latex photometric immunoassay in patients with hematological malignancies, diabetes mellitus, collagen disease, liver disease, thrombotic disease and disseminated intravascular coagulation (DIC). Plasma levels of D dimer were elevated in a variety of diseases, especially in DIC. Patients with hematological malignancies, liver disease and thrombotic disease also had relatively high levels of D dimer. On the whole, D dimer values were positively correlated with plasmin-alpha 2-plasmin inhibitor complex and thrombin-antithrombin III complex. In addition, plasma D dimer was measured during fibrinolytic therapy with urokinase or tissue-type plasminogen activator; its elevation was detected in some patients. These findings indicate that accelerated fibrinolysis is frequently observed in a variety of diseases, and that a rapid quantitative measurement of D dimer would be valuable for the precise assessment of fibrinolysis in these disease states.
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PMID:[Evaluation of clinical usefulness of a rapid quantitative measurement of D dimer (cross-linked fibrin degradation products)]. 177 52

6 patients with deep vein thrombosis triggered by drug therapy, that is oral contraceptives in 5 and the anticonvulsant tranexamic acid in 1, are described. These cases were among 40 symptomatic patients out of a total group of 81 with congenital coagulation inhibitor defects studied over 10 years at the Institute of Medical Semiotics, Padua, Italy. The 5 women with deep vein thrombosis ranged in age from 20-34, and had typically taken oral contraceptives containing 35 mcg ethinyl estradiol in combined or phasic preparations, for 1 to 8 cycles. One women, however, had been prescribed sequential pills containing 50 mcg mestranol. Another had taken oral contraceptives with impunity for 3 years, but developed deep vein thrombosis after taking tranexamic acid for 10 days. All recovered after heparin or oral anticoagulant therapy, except a 21 year old whose condition evolved into complete ileo-caval obstruction up to the renal veins, and was treated with urokinase. the congenital defects involved were 3 probable heterozygous true deficiencies of antithrombin III (low ATIII antigen and activity); a decreased protein C antigen to factor X antigen ratio; a heparin cofactor II deficiency; and a type I protein S deficiency (low free protein S, with normal total protein S and normal levels of C4B-bp.) While 5 of these 6 women had family histories of thromboembolic disease, the drug was prescribed without knowing that they were heterozygous for a coagulation inhibitor deficiency. The incidence of drug-induced thromboembolism was low in this series overall, where most of the events were triggered by surgery or trauma.
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PMID:The role of drugs, particularly oral contraceptives, in triggering thrombosis in congenital defects of coagulation inhibitors: a study of six patients. 178 39

The functional operation of the cell surface pro-u-PA and plasminogen activating system has previously been shown to depend on the assembly of u-PA receptors, plasminogen binding sites, and their respective ligands at the focal adhesions of cell extensions. We now show that additional factors operate that affect the persistence of functional activity and that evidently involve charge interactions mediated by polyanions, such as those found in the cell surface proteoglycans. Heparin-like compounds and protamine were identified as fast-acting stimulators of cell surface plasminogen activation. Heparin stabilized surface u-PA activity during plasminogen activation, and we propose that a heparin binding site exists in the kringle structure of u-PA. Heparin at 40 micrograms/ml could reduce u-PA loss to only 20% compared with 60% on control cells activating plasminogen. Protamine (25 micrograms/ml) exerted a strong stimulatory effect on the level of generated bound plasmin and notably prolonged the persistence of this activity, so that 100 minutes after addition of plasminogen the level of plasmin on protamine-treated cells was five times higher than on control-treated cells. The effect of protamine on plasmin clearance suggests that an unknown plasmin inhibitor may be produced by rhabdomyosarcoma cells, whose action is accelerated by endogenous polyanions, in an analogous manner to thrombin inactivation by antithrombin III and protease nexin on endothelial cells and fibroblasts, respectively. The stimulatory effects of heparin and protamine do not affect the inactivation of cell surface u-PA by recombinant PAI-2.
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PMID:Stimulation of cell surface plasminogen activation by heparin and related polyionic substances. 183 80

The potential importance of pleural fibrin deposition in the pathogenesis of pleural injury is supported by both clinical and experimental observations. We hypothesized that the local equilibrium between procoagulant and fibrinolytic activities is disrupted to favor fibrin deposition in exudative pleuritis. To test this hypothesis, we characterized procoagulant and fibrinolytic activities in pleural exudates from patients with pneumonia, lung cancer, or empyema and transudates from patients with congestive heart failure. Procoagulant activity was generally increased in exudative processes and was due mainly to tissue factor. All effusions contained antithrombin III and inhibited factor Xa and thrombin, but endogenous prothrombinase or thrombin activities were variably detected. Pleural fluid fibrinolytic activity was increased in congestive heart failure and was due to both tissue plasminogen activator and urokinase. Depressed fibrinolytic activity was found in pleural exudates despite increased concentrations of plasminogen, mainly glu-1-plasminogen, and was due to inhibition of plasminogen activation by plasminogen activator inhibitors 1 and 2 and of plasmin, in part by alpha 2-antiplasmin. Concentrations of PAI-1 in exudative pleural fluids were increased up to 913-fold, compared with normal pooled plasma. Exudative pleural effusions are characterized by increased procoagulant and depressed fibrinolytic activity, favoring fibrin deposition in the pleural space. The balance of these activities is reversed and favors fibrin clearance in congestive heart failure.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Abnormalities of pathways of fibrin turnover in the human pleural space. 206 28

Eleven patients with systemic sclerosis (SSc) were studied for plasma and cutaneous fibrinolytic activity, residual (potential fibrinolysis) fibrinolytic activity (FA) fo the dermal vessels that is related to the endothelial storage of plasminogen activators that become available due to particular stimuli such as intradermic injection of histamine, and the serum levels of circulating von Willebrand antigen, antithrombin III, plasminogen, beta-thromboglobulin, and platelet aggregate ratio (PAR). Cutaneous FA (autohistographic fibrin film method) appeared normal or increased in non-affected skin, normal in lesional skin, and increased after intradermal (i.d.) injection of 0.1 ml of 0.01% histamine. Monoclonal antibodies directed against the catalytic site of tissue type plasminogen activator completely blocked the fibrinolytic activity, while anti-urokinase antibodies did not abolish the lysis areas. Plasmatic FA, euglobulin lysis time test, (ELT) and the levels of beta-thromboglobulin resulted similar to the controls. A significant increase in von Willebrand Factor VIII antigen (but not of Factor VIII coagulant) was observed in the patients (p less than 0.01). Platelet aggregate ratio, levels of plasma plasminogen and Antithrombin III showed a significant difference (p less than 0.01) when compared with the control subjects. Data suggest that primary injured microvessels in SSc are likely to be arteriolae while venulae could be affected by secondary hypoxia due to the arteriolar damage with consequent release of tissue type plasminogen activator. Therefore, the authors suggest that the fibrinolytic potential is maintained in SSc and that the fibrinolytic therapy should not be used in all patients with SSc but only in those cases with documented exhaustion of plasmatic and/or cutaneous FA.
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PMID:Cutaneous and plasma fibrinolytic activity in systemic sclerosis. Evidence of normal plasminogen activation. 212 82


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