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Biomedical subjects

D Collen

Publications and source records attributed to D Collen.

At least 577 records · Page 32Linked to original sources
PubMed1984

Dysfibrinogenemia (fibrinogen Dusard) associated with impaired fibrin-enhanced plasminogen activation.

The fibrin-mediated enhancement of the activation of plasminogen by tissue-type plasminogen activator observed with normal fibrin, is strongly decreased with fibrin Dusard, although the binding of tissue-type plasminogen activator to this fibrin is normal. This impaired fibrin-mediated plasminogen activation is most likely related to the history of recurrent thrombosis and pulmonary embolism observed in this family.

Blood Coagulation Disorders↗
PubMed1984

Kinetics of the activation of plasminogen by natural and recombinant tissue-type plasminogen activator.

The kinetics of the activation of plasminogen by tissue-type plasminogen activator were studied in the presence and the absence of CNBr-digested fibrinogen as a soluble cofactor. Michaelis-Menten kinetics applied and the kinetic parameters obtained were very similar to those previously reported for the activation in the presence of solid phase fibrin (Hoylaerts, M., Rijken, D. C., Lijnen, H. R., and Collen, D. (1982) J. Biol. Chem. 257, 2912-2919). The affinity of the enzyme for plasminogen dramatically increases in the presence of the soluble cofactor while the catalytic rate constant does not change significantly (KM drops from 83 to 0.18 microM and kcat increases from 0.07 to 0.28 s-1 for tissue-type plasminogen activator of melanoma origin). Fragments containing the lysine-binding sites of plasminogen compete with plasminogen for interaction with CNBr-digested fibrinogen. The dissociation constant of this interaction was found to be 4.5 microM for the high affinity lysine-binding site. No difference was found in the kinetic parameters for the activation of plasminogen by either tissue-type plasminogen activator of melanoma origin or by glycosylated forms of tissue-type plasminogen activator obtained by recombinant DNA technology. The present findings obtained in a homogenous liquid milieu support the previously proposed mechanism of the activation of plasminogen by tissue-type plasminogen activator in the presence of fibrin. This mechanism involves binding of both tissue-type plasminogen activator and plasminogen to fibrin.

Cyanogen Bromide↗
PubMed1984

Identification of plasminogen activator releasing activity in the neurohypophysis.

Vasopressin (AVP) and some of its synthetic analogues, for example 1 desamino 8-D-AVP (DDAVP), induce plasminogen activator (PA) release in vivo. It has been proposed that this occurs via the release from the central nervous system of a plasminogen activator releasing hormone (PARH). The present study shows that a crude extract of bovine posterior pituitary, but not of the anterior pituitary or of the hypothalamus, induces a marked increase of circulating PA in anaesthetized rats. PA release appears to be caused by AVP because no other PA releasing activities could be identified in or isolated from the extract. In addition, no PA-releasing activity was found in extracts of pituitary glands of rats congenitally deficient in vasopressin. The following experiments did not reveal central nervous system involvement in the PA-release caused by AVP or DDAVP. Perfusion of isolated organs (rat heart and rat liver) with AVP resulted in PA-release. In squirrel monkeys, comparable PA levels were found in the venous return following intrafemoral or intracarotid injection of AVP. Moreover, when plasma obtained 2 min following AVP or DDAVP injection was transfused to a receiving animal no PA release was observed. Thus our present findings do not support the hypothesis that the fibrinolytic response to AVP is mediated by the release of a specific peptide hormone from the central nervous system.

Animals↗
PubMed1984

Multiple proteases in foot-and-mouth disease virus replication.

Translation of foot-and-mouth disease virus RNA in a rabbit reticulocyte lysate for short time intervals resulted in the production of the peptides P20a , P16, and P88 (Lab, Lb, and P1) (R. R. Rueckert , Recommendations of the 3rd European Study Group on Molecular Biology of Picornavirus, Urbino , Italy, 1983). If further translation was prevented, the structural protein precursor P88 was not cleaved, even after prolonged incubation. This result indicates that the mechanism of the cleavage between P20a -P16 and P88 and of that between P88 and P52 (P2) differs from the mechanism of the secondary cleavages which produce the structural proteins. Furthermore, treatment of foot-and-mouth disease virus-infected cells with the protease inhibitor D-valyl phenylalanyl lysyl chloromethyl ketone prevented the in vivo cleavage between P20a -P16 and P88 but had no effect on any of the other cleavage events. These results suggest that the cleavage of the foot-and-mouth disease virus polyprotein utilizes two different host proteases.

Animals↗
PubMed1984

Coronary thrombolysis with intravenously administered human tissue-type plasminogen activator produced by recombinant DNA technology.

Coronary thrombolysis was induced by intravenous infusion of human tissue-type plasminogen activator (recombinant human t-PA or rt-PA) obtained by expression of the cloned gene in a mammalian cell system. Thrombolysis was detected by the appearance of reperfusion arrhythmia and confirmed by repeat angiography in anesthetized dogs with 1-hr-old thrombi of the left anterior descending coronary artery that were induced with a copper coil. Infusion of 1000 IU (10 micrograms)/kg/min intravenous rt-PA (n = 9) elicited reperfusion within 13.7 +/- 1.9 min (mean +/- SE) without producing systemic fibrinolysis or distal coronary embolization. Infusion of urokinase at the same rate elicited thrombolysis in seven of 10 dogs within an average of 19.3 +/- 2.2 min. However, distal coronary embolization occurred in two dogs and systemic fibrinolysis was observed in all. In three dogs treated with urokinase thrombolysis was obtained only with subsequent intracoronary infusion. Restoration of myocardial perfusion and metabolism assessed with positron-emission tomography was consistently noted in dogs treated with rt-PA. Thus, rt-PA, a clot-selective thrombolytic agent that does not activate the fibrinolytic system systemically and that is potentially available in large quantities, in view of its synthesis by recombinant DNA technology, offers a promising practical approach for coronary thrombolysis in patients with acute myocardial infarction.

Angiography↗
PubMed1984

Improvement of regional myocardial metabolism after coronary thrombolysis induced with tissue-type plasminogen activator or streptokinase.

To assess the effects on the heart itself of coronary thrombolysis induced with either tissue-type plasminogen activator (t-PA) or streptokinase (SK), we performed positron emission tomography with 11C-palmitate in 19 patients with initial transmural myocardial infarction immediately after admission and again within 48 to 72 hr after intracoronary administration of t-PA (n = 2) or SK (n = 17). Clots were persistent in eight patients treated with SK despite an average dose of 336,000 IU, sufficient to markedly deplete fibrinogen. In the absence of lysis, favorable tomographic changes did not occur. In contrast, in each of the 11 patients in whom lysis was induced (two with t-PA and nine with SK) myocardial accumulation of 11C-palmitate improved by an average of 29% in late compared with early studies (p less than .001). Results were comparable in patients with anterior and those with inferior infarction. Thus clot lysis induced with either t-PA or SK led to improved regional myocardial metabolism.

Adult↗
PubMed1984

Coronary thrombolysis with recombinant human tissue-type plasminogen activator: a prospective, randomized, placebo-controlled trial.

Forty-five patients with acute transmural myocardial infarction and angiographically confirmed complete coronary occlusion were prospectively randomized, two for one, to treatment of acute coronary thrombosis with intravenous recombinant human tissue-type plasminogen activator (rt-PA) or placebo. Each of five additional consecutive patients was treated with a high dose of rt-PA for 2 hr. Twenty-five of 33 patients (75%) receiving 0.5 to 0.75 mg/kg of rt-PA over 30 to 120 min had angiographically proven recanalization within 90 min of initiation of therapy. Only one of 14 patients given placebo had spontaneous recanalization within 45 min (p less than .001). Thirteen placebo-treated patients were crossed over to the intracoronary rt-PA group. Nine (69%) exhibited subsequent recanalization within 45 min. Levels of circulating fibrinogen decreased after treatment with rt-PA by an average of only 8% of baseline values. None of the patients manifested a depletion of fibrinogen level to below 100 mg/dl. Six patients who were completely unresponsive to rt-PA were subsequently treated with intracoronary streptokinase and none responded. Thus, either intravenous or intracoronary rt-PA induced coronary thrombolysis without eliciting clinically significant fibrinogenolysis in patients with evolving myocardial infarction due to thrombotic coronary occlusion.

Arteriosclerosis↗
PubMed1984

Influence of protein C activation on blood coagulation and fibrinolysis in squirrel monkeys.

Protein C is a circulating proenzyme which, upon activation, exerts a potent anticoagulant activity. Infusion of activated bovine protein C into dogs is accompanied by an increase of circulating tissue plasminogen activator (PA) activity. However, the evidence that human protein C shares a similar profibrinolytic capacity is still lacking. Therefore, we investigated the profibrinolytic properties of human protein C in squirrel monkeys (Samiri sciureus). Injection of activated human protein C resulted in prolongation of the activated partial thromboplastin time but was not associated with increased fibrinolytic activity of blood. Similarly, activation of endogenous protein C (up to 20-30%) by infusion of thrombin-thrombomodulin complex markedly reduced blood coagulability without being accompanied by an increase of circulating PA activity. The in vivo-generated anticoagulant activity was identified as activated protein C by the following observations. It was neutralized by rabbit anti-human protein C-IgG, was slowly inhibited by plasma but not by anti-thrombin III, was adsorbable on barium citrate, and expressed amidolytic activity. Activation of protein C appeared to be selective since other parameters such as thrombin time, platelet count, fibrinogen, and factor V levels were unaffected by thrombin-thrombomodulin infusion. Infusion of human plasma derived from whole blood incubated in vitro with human activated protein C also did not induce a fibrinolytic response, suggesting that no second messengers with PA-releasing activity were being generated in blood. It is concluded that in a primate, neither the administration of activated human protein C nor the activation of endogenous protein C are associated with an increase of fibrinolytic activity. These findings question the role of this enzyme in the regulation of PA release in man.

Animals↗
PubMed1984

A functional assay of protein C in human plasma.

A three-step spectrophotometric assay was developed for measuring functional protein C (PC) in human plasma. The assay is based on: (1) adsorption of citrated platelet-poor plasma on barium citrate and elution of the vitamin K-dependent factors with EDTA; (2) activation of PC by incubation of the mixture of vitamin K-dependent factors with a complex of thrombin and its endothelial cell cofactor, thrombomodulin; (3) addition of antithrombin III and heparin to the system to inhibit thrombin and other coagulation enzymes generated during incubation and measurement of the activated PC with a synthetic (chromogenic) substrate. The assay appears to be specific for PC because: (a) PC-depleted plasma (by immunoadsorption) is inactive; (b) addition of purified PC to PC-depleted plasma reconstitutes its activity; and (c) no enzymatic activity is generated in the absence of the thrombin-thrombomodulin complex. Mixtures of a normal plasma pool with PC-depleted plasma yielded an amount of enzymatic activity proportional to the fraction of normal plasma. Using this as a standard curve, the amount of PC in the plasma of 23 normal subjects was 97% +/- 15%. The within-assay coefficient of variation was 3.5% and the between-assay coefficient 6.5%. A linear correlation (r = 0.86) was found between PC as measured with the functional assay and with a radioimmunoassay. In 3 patients with congenital PC deficiency, the functional PC level was 37% +/- 9% and the antigen level 64% +/- 11%. It is concluded that the present assay may be used for reliable and accurate estimation of activatable PC in human plasma.

Adsorption↗
PubMed1984

Biological properties of human tissue-type plasminogen activator obtained by expression of recombinant DNA in mammalian cells.

Human tissue-type plasminogen activator (t-PA), obtained by expression in mammalian cells of recombinant DNA coding for the entire sequence of t-PA (rt-PA), was compared with natural activator from melanoma cell culture (mt-PA). In an in vitro system, composed of [125I]fibrinogen-labeled plasma clot suspended in circulating human plasma, rt-PA and mt-PA caused a very similar dose-related degree of fibrinolysis without causing extensive fibrinolytic activation and fibrinogen breakdown in the surrounding plasma. Urokinase only induced fibrinolysis at a 5- to 10-fold higher concentration and in association with extensive fibrinogenolysis. Intravenous injection of mixtures of labeled (0.4 microCi/kg) and unlabeled (2000 I.U./kg) mt-PA or rt-PA resulted in a rapid but similar disappearance of activity from plasma (T1/2 of 3 min) and specific accumulation of tracer in the liver. In rabbits with experimental jugular vein thrombosis, rt-PA and mt-PA caused a very similar dose-dependent thrombolysis without causing substantial systemic activation of the fibrinolytic system and fibrinogenolysis. Urokinase induced significant thrombolysis only at a 10-fold higher dose and this was associated with systemic fibrinolytic activation. Infusion of 96,000 I.U./kg (approximately equal to 1 mg/kg) of mt-PA or rt-PA over 4 hr induced approximately 70% lysis, whereas a 10-fold higher dose of urokinase yielded 35 to 40% lysis. Two subfractions of rt-PA differing in the extent of glycosylation had very similar thrombolytic properties. It is concluded that the potentially more readily available rt-PA could constitute a specific, fibrin-selective thrombolytic agent.

Animals↗
PubMed1983

Interaction of heparin with histidine-rich glycoprotein and with antithrombin III.

The interaction between heparin, histidine-rich glycoprotein and antithrombin III was studied in purified systems. Histidine-rich glycoprotein binds heparin and thereby interferes with its interaction with antithrombin III, resulting in neutralization of the anticoagulant activity. This interaction occurs with clinical grade heparin as well as with high affinity (for antithrombin III) heparin and with a high affinity heparin fragment with Mr 4,300. Low affinity heparin competes with high affinity heparin for the binding to histidine-rich glycoprotein which results in an apparent increase of the anticoagulant activity of high affinity heparin. The interaction between heparin and histidine-rich glycoprotein is counteracted by Ca2+-binding anticoagulants, indicating that it is dependent on the presence of divalent metal ions. Ethylenediaminetetraacetate is a much more potent inhibitor of the interaction between heparin and histidine-rich glycoprotein than citrate.

Antithrombin III↗
PubMed1983

Clot-selective coronary thrombolysis with tissue-type plasminogen activator.

Coronary thrombolysis, an intervention that can abort the sequelae of acute myocardial infarction, was accomplished within 10 minutes in dogs by intravenous administration of clot-selective, tissue-type plasminogen activator. In addition to inducing clot lysis, this promising fibrinolytic agent restored intermediary metabolism and nutritional myocardial blood flow, detectable noninvasively with positron tomography, without inducing a systemic fibrinolytic state.

Animals↗
PubMed1983

Covalent complexes between low molecular weight heparin fragments and antithrombin III - inhibition kinetics and turnover parameters.

Two high affinity heparin fragments (Mr 4,300 and Mr 3,200) were covalently coupled to antithrombin III (J. Biol. Chem. 1982; 257: 3401--3408) with an apparent 1:1 stoichiometry and a 30--35% yield. The purified covalent complexes inhibited factor Xa with second order rate constants very similar to those obtained for antithrombin III saturated with these heparin fragments and to that obtained for the covalent complex between antithrombin III and native high affinity heparin. The disappearance rates from plasma in rabbits of both low molecular weight heparin fragments and their complexes could adequately be represented by two-compartment mammillary models. The plasma half-life (t1/2) of both low Mr-heparin fragments was approximately 2.4 hr. Covalent coupling of the fragments to antithrombin III increased this half-life about 3.5 fold (t1/2 congruent to 7.7 hr), approaching that of free antithrombin III (t1/2 congruent to 11 +/- 0.4 hr) and resulting in a 30 fold longer life time of factor Xa inhibitory activity in plasma as compared to that of free intact heparin (t1/2 congruent to 0.25 +/- 0.04 hr).

Antithrombin III↗