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:3.4.21.5 (thrombin)
33,306 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To improve the sensitivity and reliability of the ultrasonic detection of intravascular thrombosis, we employed quantitative ultrasonic techniques for the analysis of thrombi induced in vitro. Citrated human blood or platelet-rich plasma was added to sections of canine thoracic aorta mounted vertically in a saline bath. CaCl2 and thrombin were then added to induce thrombosis. Ultrasonic integrated backscatter measured sequentially with a 10-MHz focused transducer exhibited more than a 10 dB increase in integrated backscatter when either whole blood or platelet-rich plasma was clotted (-61.7 dB to -47.4 dB whole blood; less than -70 dB to -52.8 dB platelet-rich plasma). Thus, large significant changes in integrated backscatter measured from structures within the arterial lumen were readily detectable and indicative of thrombosis in situ.
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PMID:Intravascular thrombosis: definitive detection by quantitative tissue characterization in vitro. 250 83

alpha-Thrombin induces a dose-dependent rapid transient increase in platelet cytosolic Ca2+ levels, coming solely from intracellular stores, since EGTA has no effect. In contrast, the post-stimulation equilibrium [Ca2+]in depends upon an influx from the extracellular milieu, and is lower in the presence of EGTA. We measured the Ca2+ transient (with Indo-1, 1-[2-amino-5-(6-carboxyindol-2-yl)-phenoxy]-2-(2'-amino-5'-methylp henoxy)- ethane-N,N,N',N'-tetraacetic acid), cytosolic alkalinization (with BCECF, 2',7-bis-(2-carboxyethyl)-5(and 6)-carboxyfluorescein), membrane depolarization (with diS-C3-(5), 3,3'-dipropylthiodi-carbocyanide iodide), and degranulation (by beta-glucuronidase release) induced in washed human platelets by 9 nM thrombin in the absence or presence of extracellular or intracellular Ca2+ chelating agents (EGTA and BAPTA, 5,5'-dimethyl-bis-(o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid, respectively). Platelets loaded simultaneously with 2 microM Indo-1 and 15 microM BAPTA (each as the acetoxymethyl ester) before addition of thrombin exhibited no cytoplasmic Ca2+ transient or alkalinization, no depolarization or degranulation. Replenishment of such cells with extracellular CaCl2 restored resting [Ca2+]in. Upon stimulation with 9 nM thrombin these replenished platelets exhibited no Ca2+ transient, and a slow gradual increase in [Ca2+]in from extracellular stores, a slow alkalinization and depolarization, and partial degranulation, all abolished by extracellular EGTA. Thus thrombin-induced platelet activation exhibits a biphasic Ca2+ requirement: the initial transient increase in [Ca2+]in comes from intracellular stores only, while the later steps of depolarization, alkalinization, and degranulation can proceed, albeit more slowly, if only extracellular Ca2+ is available.
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PMID:Cytoplasmic Ca2+ is necessary for thrombin-induced platelet activation. 251 Nov 90

Fibrinogen Haifa is a congenital heterozygous fibrinogen variant (gamma 275 Arg----His) characterized by prolonged thrombin and reptilase times and normal fibrinopeptide (FPA, FPB) release. We compared the polymerization rate (by turbidity measurements at 350 nm) and the ultrastructure of Haifa alpha-, beta-, and alpha, beta-fibrin with that of normal. Haifa alpha, beta-fibrin polymerized less rapidly than did normal and formed a highly branched matrix with a smaller mean fiber diameter; this network closely resembled that of normal alpha, beta-fibrin with EDTA added. In the presence of CaCl2 (1 to 10 mM), Haifa alpha, beta-fibrin polymerized more rapidly than in buffer alone and possessed a matrix structure closely resembling that of normal fibrin. From these observations it appears that the functional defect in Haifa fibrin can be related to the inability of the abnormal molecule to effectively utilize available calcium. The polymerization profile of Haifa alpha-fibrin differed only modestly from that of normal alpha-fibrin, whereas that of Haifa beta-fibrin was markedly impaired. This finding plus similarities in the ultrastructure of Haifa and normal alpha-fibrin specimens suggests that the defective gamma chain structure of Haifa fibrinogen results in greater impairment of the carboxy terminal "b" polymerization domain reacting with the site exposed by cleavage of FPB ("B" site) than it does that of the carboxy terminal "a" domain reacting with the site exposed by cleavage of FPA ("A" site). Whether this effect is due to absolute differences in the degree of impairment of these two types of polymerization sites, or whether proper utilization of the "B" to "b" site is dependent upon participation of the "A" to "a" site remains to be determined.
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PMID:The polymerization of fibrin prepared from fibrinogen Haifa (gamma 275Arg----His). 251 77

Four mouse hybridomas secreting monoclonal antibodies specific for human protein S (PS) have been generated. The antibodies, all of the IgG1 subclass, were designated S2, S3, S8, and S10. In a fluid phase radioimmunoassay, the binding of monoclonal antibodies to PS was about 30% greater in the presence of EDTA and totally inhibited in presence of Ca2+. Using the same technique, we performed displacement curves of 125I-labeled PS by purified PS, thrombin-cleaved PS, normal plasma, plasma from a patient on warfarin therapy, and plasma from a patient with no free PS and only PS bound to C4b-binding protein. The slopes of the curves show that the monoclonal antibodies reacted equally with all the tested forms of PS indicating that the antigenic site(s) to which the monoclonal antibodies are directed are present and exposed in free and bound PS, in thrombin-cleaved PS, and in the coumarin form of the protein. Each EDTA-dependent antibody, immobilized on Sepharose 4B-CNBr was used to purify PS from the barium citrate-absorbed, ammonium sulphate-soluble fraction of plasma. The fraction eluted from the immunoabsorbent with a buffer containing 4 mmol/l CaCl2 and analysed by SDS-PAGE, contained two bands, one migrating with conventionally purified PS and the other with purified C4b-binding protein. Homogeneous PS was obtained by chromatography of the barium citrate adsorbate on a DEAE-Sephadex column. The protein peak containing the bulk of PS was subsequently applied to the immunoadsorbent and eluted with 4 mmol/l CaCl2.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Monoclonal antibodies directed to the calcium-free conformation of human protein S. 253 Jun 47

The thrombolytic efficacy of recombinant tissue-type plasminogen activator (tPA) in the presence and absence of the selective endoperoxide/thromboxane A2 (TXA2) receptor antagonist, sulotroban (BM 13.177, SK&F 95587) was studied in a model of femoral artery thrombosis in the anesthetized rabbit. The thrombus was formed by injection of thrombin, CaCl2 and whole blood into an isolated segment of the femoral artery. After 30 min of stable thrombotic occlusion of the femoral artery, tPA was infused IV for 90 min at doses of 5.0, 7.5 and 10.0 micrograms/kg/min. In other experiments, sulotroban was administered as a bolus dose of 1 mg/kg/IV, followed by a constant infusion of 1 mg/kg/hr concurrent with tPA infusion. Sulotroban had no effect on the incidence of tPA-induced reperfusion at any dose studied or on residual clot weight. However, at a tPA dose of 10 micrograms/kg/min, IV lysis time was reduced in sulotroban treated animals from 65 min to 29 min (P less than 0.05), and the magnitude of femoral artery blood flow achieved as a result of tPA-induced reperfusion was greater in sulotroban-treated animals. These data suggest that adjunctive therapy with a selective endoperoxide/TXA2 antagonist improves the response to tPA when tPA is administered at a maximal or near maximally effective pharmacological dose.
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PMID:Effect of selective endoperoxide/thromboxane A2 receptor antagonism with sulotroban on tPA-induced thrombolysis in a rabbit model of femoral arterial thrombosis. 253 38

Ca(II) ions are crucial during proteolytic conversion of Factor XIII zymogen into the active enzyme Factor XIIIa. Factor XIII proteolyzed by thrombin or trypsin in the presence of 5 mM-EDTA resulted in rapid inactivation of transglutaminase activity. Factor XIIIa formed by thrombin or trypsin in the presence of 40 microM-Tb(III) ions, however, was indistinguishable from Factor XIIIa formed in the presence of 2-5 mM-Ca(II) ions with respect to molecular mass and transglutaminase activity. Thrombin treatment of Factor XIII in the presence of 1-5 microM-Tb(III) ions resulted in three fragments (76 kDa, 51 kDa and 19 kDa) with simultaneous loss of transglutaminase activity. Tb(III) ions at concentrations greater than 40 microM made platelet Factor XIII resistant to proteolysis by either thrombin or trypsin. Other lanthanide(III) ions [Ln(III) ions] tested [Ce(III), La(III) and Gd(III) ions] functioned similarly to Tb(III) ions during proteolytic activation of Factor XIII. Ln(III) ions (10-100 microM) were unable to replace the Ca(II) ions required for transglutaminase activity of Factor XIIIa. Tb(III) ions also inhibited in a non-competitive manner the transglutaminase activity of Factor XIIIa (Ki 71 microM) even when measured in the presence of 200-fold molar excess of Ca(II) ions. Factor XIII selectively bound to a Tb(III)-chelate affinity column, and could not be eluted by 100 mM-CaCl2. Binding of Tb(III) ions to Factor XIII was demonstrated by fluorescence emission due to Forster energy transfer. A 10(4)-fold molar excess of CaCl2, but not NaCl, partially quenched Tb(III) fluorescence. Low concentrations (5-20 microM) of Tb(III) ions also inhibited the binding of Factor XIII to des-A-fibrinogen by about 43%, whereas higher concentrations (40-100 microM) promoted binding. Conformational changes in Factor XIII consequent to the binding of Tb(III) ions could be responsible for the observed effects on protein structure and function.
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PMID:Tb(III)-ion-binding-induced conformational changes in platelet factor XIII. 256 74

Ilexonin A is an effective compound isolated from Ilex pubescens Hook. et Arn. It has been used in the clinic to treat cardiovascular diseases such as cerebral embolism and myocardial infarction. Experimental studies have shown that it inhibited thrombosis, platelet adhesion, and platelet aggregation and 5-HT release induced by collagen, ADP or A23187 in vitro and in vivo. The inhibitory effect of platelet aggregation induced by A23187 was reduced by adding CaCl2 (1mM) to the medium. It suggests that the effects of llexonin A on platelet function are possibly related to calcium. The present experiments were designed to investigate the effects of Ilexonin A on thrombin-induced Ca2+ fluxes of platelets, in which quin-2 was used to measure the cytoplasmic Ca2+ concentration. The results indicated that the free calcium concentrations of resting platelets and the platelets activated by thrombin were 78.65 +/- 7.74 nM and 871.10 +/- 123.63 nM respectively. After adding EGTA (lmM) to the medium, the calcium concentrations reduced to 34.75 +/- 7.77 nM and 50. 86 +/- 7.44 nM. Ilexonin A and verapamil markedly inhibited the thrombin induced Ca2+ influx. The IC50 was 76.8 microM and 67.5 microM respectively. But both had no effect on thrombin-induced Ca2+ release from dense tubular system. It suggests that Ilexonin A acts most likely as a calcium slow channel blocker.
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PMID:[Ilexonin A may be a slow calcium channel blocker]. 261 56

Chemical pleurodesis induced with fibrin glue (fibrinogen 1.0 g, thrombin 500 mu, 2% CaCl2 10 ml, tranexamic acid 10 ml) was performed in 6 cases with spontaneous pneumothorax in whom surgery was not indicated because of various reasons, such as low pulmonary function or old age. These cases were complicated with left pneumonectomy, bilateral emphysema, pulmonary tuberculosis and interstitial pneumonia. In all cases, favorable results were obtained and there was no recurrence. As side effects, only transient low grade fever and slight chest pain were observed with no liver damage or pleural thickening. These results suggest that chemical pleurodesis induced with fibrin glue is very useful in the treatment of inoperable spontaneous pneumothorax.
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PMID:[Chemical pleurodesis induced with fibrin glue in the treatment of inoperable spontaneous pneumothorax]. 261 97

We report a study on the importance of factor IX activation in thromboplastin-dependent coagulation in plasma. Diluted, CaCl2-containing thromboplastin solutions at constant phospholipid concentration were used to trigger the coagulation in plasma from patients with congenital factor IX and factor VIII deficiency in the presence and absence of added factors IX and VIII, and the generation of thrombin activity was monitored. When coagulation is triggered with the high thromboplastin concentrations normally used in clinical routine tests, the generation of thrombin activity in plasma of patients with congenital factor IX deficiency before and after reconstitution with purified factor IX appears identical. When, however, coagulation is triggered with low thromboplastin concentrations, a clear dependency of the generation of thrombin activity on the concentration of factor IX becomes evident at factor IX concentrations lower than 30 nM (about 40% clotting factor activity). Factor VIII is a compulsory cofactor for this factor IX activity because the prothrombinase activity at optimal factor IX concentration is still critically dependent upon the amount of factor VIII present. The lower the amount of thromboplastin, the higher the importance of factor IX and factor VIII activation in thromboplastin-dependent coagulation. This suggests a role of this pathway in pathophysiological thrombin formation.
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PMID:Importance of factor-IX-dependent prothrombinase formation--the Josso pathway--in clotting plasma. 262 Aug 66

Methylene blue (MB) has been suggested as a therapeutic alternative for heparin reversal in patients sensitive to protamine. We investigated the impact of MB on the assembly and structure of thrombin induced fibrin and plasma gels. MB (1,600 micrograms/ml) reduced the thrombin clotting time (TCT) of plasma by 30% and of purified fibrinogen by 46%. Above 1,600 micrograms/ml, TCTs were prolonged due to MB mediated fibrinogen precipitation. The presence of 5 mM CaCl2 masked the effect of MB in both purified and plasma systems and lowered the threshold for MB-induced purified fibrinogen precipitation to 800 micrograms/ml. MB shortened the lag phase prior to thrombin-induced turbidity increase, and enhanced final gel turbidity. The fibrin fiber mass/length ratio increased from 5.2 to 13.1 X 10(13) dalton/cm in purified fibrin gels and from 3.2 to 10.4 X 10(13) dalton/cm in plasma gels as the MB concentration increased from 0 to 200 micrograms/ml. Due to the photooxidant effect of MB on fibrinogen, rapid time-dependent loss of fibrinogen clottability was obvious at low MB concentrations (50 to 400 micrograms/ml). At high MB concentrations, intense MB light absorption partially protected fibrinogen within the sample. Accurate measurements could only be made, however, when MB was added just prior to thrombin and the assays were performed in the dark. While erythrocytes may reduce the impact of MB photooxidation in whole blood, plasma samples must be shielded from light if reproducible results are to be obtained.
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PMID:Methylene blue enhances lateral association of fibrin resulting in rapid gelation and thick fiber formation. 274 96


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