Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.73 (urokinase-type plasminogen activator)
 10,685 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Parameters of the fibrinolytic system were studied in a primate model where the generation of thrombin was promoted in vivo. The procoagulant stimulus used was a combination of human factor Xa in combination with phosphatidylcholine/phosphatidylserine lipid vesicles (PCPS) as the source of coagulant active phospholipid. The dosage of each component was formulated to provide a gradation of thrombin generating potential assessed prior to in vivo study in an in vitro clotting assay. These ranged from 25.25-36.60 pMole/kg (factor Xa) and 18.85-56.30 nMole/kg (PCPS). In each case, the ratio of the dose of factor Xa/PCPS was maintained at 0.65 (pMole factor Xa/nMole PCPS). Individual dosage combinations producing recalcification clotting times in vitro of 15, 20, 25 and 30 s were used in detailed in vivo studies. Previous studies in dogs had confirmed the thrombin generating potential of factor Xa/PCPS infusions and demonstrated an associated activation of protein C and increased fibrinolytic activity. This has now been extensively characterized in the chimpanzee as follows: 10 min after the infusion of the highest dose (36.6 pMole factor Xa/56.3 nMole PCPS kg bodyweight), the level of circulating t-PA had risen to 900 ng/ml (antigen), 885 IU/ml (functional). Dosage was observed with the lowest dose of 12.25 pMole factor Xa and 18.85 nMole PCPS being associated with relatively minor increases in circulating t-PA activity. There were no changes in u-PA at any dosage during the full time course of the experimental period (90 min). Plasminogen activation was also apparent with alpha-2 antiplasmin levels falling to 30-40% of pre-infusion levels at the highest dosages.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:The fibrinolytic potential of the normal primate following the generation of thrombin in vivo. 240 50

Thrombotic thrombocytopenic purpura (TTP) is characterized by widespread occluding and persistent microthrombotic lesions. Evidence for both endothelial damage and primary platelet aggregation as possible pathogenetic mechanisms has been produced. Persistence of microthrombi has not been explained satisfactorily. In patients with TTP we studied plasma fibrinolysis and protein C. Tissue plasminogen activator (t-PA) activity levels, measured functionally, were low or unmeasurable in 11 of 12 patients; t-PA antigen levels, measured immunochemically, were normal in all six observed. The level of potent inhibitor of plasminogen activation directed against both t-PA and urokinase was elevated significantly in all 12, whereas the alpha 2-antiplasmin level was elevated in only two. Protein C antigen levels were low in three of six patients observed. Fibrinolysis levels in patients in remission did not differ from those in patients with acute disease. Plasma exchange resulted in temporary reversal of the abnormalities, but achievement of clinical remission was not associated with permanent normalization of fibrinolysis. Inasmuch as all 12 patients had severely depressed fibrinolytic mechanisms it is possible that a defect in the fibrin-clearing system permits thrombus formation to occur and proceed in an unchallenged fashion, thereby contributing to the complex events leading to arterial ischemia in vital organs.
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PMID:Fibrinolysis in health and disease: abnormal levels of plasminogen activator, plasminogen activator inhibitor, and protein C in thrombotic thrombocytopenic purpura. 243 36

Plasminogen activator inhibitor (PAI) was purified in active form from porcine platelets under nondenaturing conditions. The purified inhibitor (Mr 47,000) reacts with tissue-type plasminogen activator (t-PA), urokinase (UK), and activated protein C (APC) to yield both SDS-stable complexes and a modified PAI of slightly reduced molecular weight. The second-order rate constants for the inhibition of t-PA and UK by PAI are 3.5 X 10(7) and 3.4 X 10(7) M-1 s-1, respectively. Activated protein C reacts with PAI with a second-order rate constant of 1.1 X 10(4) M-1 s-1. This rate is not accelerated by protein S, phospholipid, and calcium, or heparin. It is concluded that (1) PAI can function as both inhibitor and substrate of its target proteases, (2) if APC promotes fibrinolysis via inactivation of PAI, then APC must be present in concentrations several orders of magnitude greater than t-PA, or the interaction of APC and PAI must be accelerated by presently unknown mechanisms, and (3) in the absence of heparin, platelet PAI is the most rapid inhibitor of APC yet described.
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PMID:Platelet plasminogen activator inhibitor: purification and characterization of interaction with plasminogen activators and activated protein C. 250 42

Protein C inhibitor was purified from human plasma by a modification of a published procedure (Suzuki, K., Nishioka, J., and Hashimoto, S. J. Biol. Chem. 258, 163-168, 1983). Approximately 1 mg of pure protein was obtained from 1 L plasma, a yield of about 17%. The protein C inhibitor preparation did not lose activity over 4 weeks at 4 degrees C. Second order rate constants were measured for the inhibition of activated protein C, thrombin, and urokinase, and bimolecular complexes of protein C inhibitor with activated protein C and thrombin were visualized by denaturing polyacrylamide gel electrophoresis. Heparin accelerated the inhibition of the three proteinases in a manner consistent with a template mechanism. Plasma or pure protein C inhibitor (at the same concentration) showed the same effect of heparin on activated protein C inhibition, indicating that protein C inhibitor accounts for all the heparin-dependent inhibition of activated protein C in vivo.
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PMID:Protein C inhibitor: purification and proteinase reactivity. 254 38

An assay system for protein C (PC) activity and PC-inhibitor in plasma was developed. The assay was based on: (1) binding of PC to wells of a microtiter plate coated with a murine monoclonal anti-PC antibody (C3) that did not interfere with the activity or activation of PC; (2) activation of immobilized PC with Protac C; (3) incubation with or without a source of activated PC inhibitor; and (4) measurement of amidolytic activity using the substrate S-2366. The activity assay was specific for PC and sensitive to less than 1 microliter of plasma or 4 ng PC. Inhibition of activated PC by plasma followed pseudo first order kinetics. Heparin caused a dose dependent increase in the inhibition rate with half maximal stimulation at approximately 3 U/ml and maximal stimulation at heparin concentrations greater than or equal to 10 U/ml. This assay is suitable not only for determination of functional plasma levels of PC and PC inhibitor activities but also for kinetic studies of inhibition of activated PC in complex systems, such as plasma. Studies showed that urokinase interfered with the inhibition of APC by plasma inhibitor(s).
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PMID:Functional assays for protein C activity and protein C inhibitor activity in plasma. 254 80

Plasma protein C inhibitor (PCI) was purified to homogeneity (greater than 95%) with good recovery (greater than 25%) and reproducibility, and the inhibition of a number of blood clotting and fibrinolytic enzymes by purified PCI was studied. PCI inhibited activated protein C (APC), two-chain urokinase (2c-uPA), two-chain tissue plasminogen activator (2c-tPA), thrombin, factor Xa, plasma kallikrein and factor XIa, and this inhibition was accelerated by heparin. The inhibition of each enzyme was accompanied by formation of enzyme inhibitor complexes and by degradation of the inhibitor to lower molecular weight derivatives. Plasma kallikrein and factor XIa cleaved PCI of native Mr = 57,000 into two products with Mr = 54,000 and 52,000 whereas the other enzymes converted the PCI to a product with Mr = 54,000. PCI did not detectably inhibit alpha-factor XIIa or plasmin. Kinetic studies using PCI yielded the following second-order rate constants for inhibition of human APC, 2c-uPA, 2c-tPA, thrombin, factor Xa, kallikrein and factor XIa respectively: 0.65 x 10(4), 0.22 x 10(4), 0.08 x 10(4), 0.61 x 10(4), 2.01 x 10(4), 6.50 x 10(4), and 9.03 x 10(4) M-1s-1 in the absence of heparin and 1.58 x 10(6), 0.43 x 10(6), 0.03 x 10(6), 0.52 x 10(6), 0.09 x 10(6), 0.18 x 10(6) and 0.74 x 10(6) M-1s-1 in the presence of optimal concentrations of heparin. The rate constants for the inhibition of factor XIa and 2c-uPA by PCI suggest a possible role of PCI in the physiologic regulation of these enzymes. The second order rate constants for inhibition of bovine APC and Gla-domainless bovine APC by human PCI were 0.61 x 10(4) and 0.26 x 10(4) M-1s-1 in the absence of heparin and 0.54 x 10(6) and 0.71 x 10(6) M-1s-1 in the presence of heparin, respectively. Calcium ions (0.05 to 4 mM) did not affect these rate constants. The results obtained with normal and Gla-domainless APC indicate that the Gla domain of APC is not required for inactivation by PGI and is not essential for the heparin stimulation of this reaction.
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PMID:Purification and characterization of plasma protein C inhibitor. 255 Oct 64

Protein C inhibitor (PCI) was purified from human plasma using immunoaffinity chromatography and heparin Sepharose chromatography, a method that allowed the purification of active and inactive inhibitor. PCI purified from outdated plasma was inactive and either in complex with plasma kallikrein or proteolytically degraded. Sequence analysis of cleaved PCI and of complexes between PCI and activated protein C or urokinase identified the previously recognized inhibitor cleavage site Arg354-Ser355. Two additional cleavage sites were observed in the modified inhibitor i.e. Arg357-Leu358 and Arg362-Leu363 which probably represent secondary cleavage of the inhibitor. Furthermore the sequence analysis of the inhibitor, whether purified from fresh or outdated plasma, revealed that it was microheterogeneous in the NH2-terminus as a result of cleavage by a trypsin like enzyme(s).
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PMID:Protein C inhibitor from human plasma: characterization of native and cleaved inhibitor and demonstration of inhibitor complexes with plasma kallikrein. 255 11

Systemic activation of the coagulation mechanism is known to exist in patients with colon cancer. The mechanism of such activation was investigated using immunohistochemical techniques applied to fresh frozen sections of resected primary colon cancer specimens. Tumor cells stained for tissue factor, factor V, and urokinase-type plasminogen activator. Perivascular and intercellular areas stained for fibrinogen and the "a" subunit of factor XIII. Staining was minimal or absent for protein C, protein S, plasminogen activator inhibitors 1-3, factor VII, factor X, and fibrin (the antigenic site on the amino-terminal portion of B beta chain that is exposed following thrombin cleavage of fibrinopeptide B was not detected). The lack of an intact thrombin-generating pathway in situ associated with viable colon cancer cells is consistent with the findings of others that coagulation activation in colon cancer may be triggered by a soluble tumor product that exerts its effect at sites distant from the tumor. These results may explain the absence of clinical responsiveness of colon cancer to antithrombotic drug therapy and may clarify therapeutic strategies for this common tumor.
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PMID:Indirect activation of blood coagulation in colon cancer. 269 22

Between August 1983 and December 1987, 23 patients received a 30-minute intraoperative, intraarterial infusion of streptokinase (seven patients) or urokinase (16 patients) because of residual thrombus or persistent ischemia or both after thromboembolectomy. Ages ranged from 21 to 77 years (mean, 58 years). In 15 patients intraoperative lytic therapy was part of the initial operation, whereas in eight patients intraoperative lytic therapy was performed during a secondary operation to treat thrombosis of a recently placed graft. Seven patients in the latter group had hypercoagulable conditions (five had heparin-induced thrombosis; one had protein C deficiency; one had polycythemia with thrombocytosis). Improvement after intraoperative lytic therapy was seen on angiography performed after infusion in 13 of 17 (76%) patients in whom angiography was performed both before and after intraoperative lytic therapy. Grafts in 12 of these patients remained patent without additional intervention, and in one graft thrombus formed again. In contrast, among four patients without angiographic evidence of improvement, thrombus formed again in four grafts (p less than 0.004). Intraoperative lytic therapy was considered successful in 74% of instances (17/23), including four of seven patients with hypercoagulable states. Three of six patients whose grafts failed had major amputations, whereas there were no amputations after successful infusions. Twelve patients were heparinized after intraoperative lytic therapy. Ten patients in this group were considered treatment successes, and two were considered treatment failures. Three of 11 patients not heparinized after intraoperative lytic therapy were considered treatment failures. Four hematomas occurred in the former group and none in the latter (p less than 0.03). No hematomas occurred in the heparin-induced thrombosis group in spite of anticoagulation with sodium warfarin (Coumadin). Only one hematoma occurred within 6 hours of intraoperative lytic therapy, and thus it was attributable to the infusion. We conclude that intraoperative lytic therapy is an effective adjunct to manage residual thrombus or persistent ischemia or both after lower extremity revascularization. Postinfusion angiography is of prognostic value. Heparinization after intraoperative lytic therapy seems beneficial but significantly increases the risk of bleeding complications.
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PMID:Intraoperative infusion of lytic drugs for thrombotic complications of revascularization. 279 66

Purified plasma and urinary protein C inhibitors (PCI) formed heparin-dependent complexes with activated protein C (APC) which were detected by immunoblotting after nondenaturing gel electrophoresis. Bands representing APC.PCI complexes were also seen on immunoblots after incubation of plasma with APC and heparin. The same immunoblot pattern of complexes was detected by three different methods: method A, monoclonal antibody to plasminogen activator inhibitor-3 (PAI-3, urinary urokinase inhibitor) + 125I-labeled anti-mouse IgG; method B, polyclonal antibodies to PCI + 125I-labeled purified plasma PCI; and method C, monoclonal antibody to protein C + 125I-protein C. Plasma depleted of PAI-3 by immunoadsorption with insolubilized monoclonal antibody to PAI-3 showed no detectable antigen or complexes with APC as visualized by methods A or B. This PAI-3-depleted plasma had less than 10% of the heparin-dependent inhibitory activity of normal plasma toward APC. Purified plasma PCI was fully reactive in an enzyme-linked immunoabsorbent assay for PAI-3, and plasma and urinary PCI inhibited urokinase activity in a heparin-dependent manner. These data indicate that heparin-dependent plasma and urinary PCI and PAI-3 are immunologically and functionally very similar if not identical. This observation identifies a new interrelation between the protein C anticoagulant and the fibrinolytic systems. In addition, plasma contains a heparin-independent inhibitor of APC which is not immunologically related to plasma PCI or to PAI-3.
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PMID:Immunological identity of heparin-dependent plasma and urinary protein C inhibitor and plasminogen activator inhibitor-3. 282 68


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