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Biomedical subjects

D Collen

Publications and source records attributed to D Collen.

At least 667 records · Page 37Linked to original sources
PubMed1977

The interaction in human plasma of antiplasmin, the fast-reacting plasmin inhibitor, with plasmin, thrombin, trypsin and chymotrypsin.

The inhibition of plasmin, (EC 3.4.21.7), thrombin (EC 3.4.21.5), trypsin (EC 3.4.21.4) and chymotrypsin (EC 3.4.21.1) by antiplasmin, the recently described fast-reacting plasmin inhibitor of human plasma, was studied. To determine the quantitative importance of antiplasmin relative to the other plasma protease inhibitors, enzyme inhibition assays were performed on whole plasma and on plasma specifically depleted in antiplasmin, after addition of excess enzyme. Plasmin was the only enzyme for which the inhibitory capacity of antiplasmin-depleted plasma was lower than that of normal plasma. To determine the affinity of the enzymes for antiplasmin, as compared to the other inhibitors, various amounts of enzymes were added to normal plasma and the formation of enzyme-antiplasmin complexes studied by crossed immunoelectrophoresis using specific antisera against antiplasmin. Plasmin and trypsin, but not thrombin or chymotrypsin formed complexes with antiplasmin. It is concluded that antiplasmin is the only fast-reacting plasmin inhibitor of human plasma. It is also a fast-reacting inhibitor of trypsin but only accounts for a very small part of the fast-reacting trypsin-inhibitory activity of plasma. This can be explained by the low concentration of antiplasmin (1 muM) in normal plasma, compared to the other inhibitors (e.g. alpha1-antitrypsin: 40-80 muM).

Chymotrypsin↗
PubMed1977

A latex agglutination test for rapid quantitative estimation of the plasmin-antipalsmin complex in human plasma.

An antiserum was raised in rabbits against human plasmin-antiplasmin complex and rendered specific for neoantigens of this complex by absorption with purified plasminogen and plasma. Polystyrene particles were coated with the specific antibodies and used in an agglutination test for the determination of plasmin-antiplasmin complex in the plasma from various patients. Purified plasmin-antiplasma complex at a concentration of 0.1-0.2 mg/l was found to cause a clear agglutination of the particles. Activation of fresh human plasma with urokinase caused progressive generation of agglutinating activity up to a plasma dilution of 1/480. Intravenous infusion of streptokinase into patients resulted in an increase of the plasmin-antiplasmin titre of at least 1/240. Sera from patients with rheumatoid factor also agglutinated the particles but this activity could be removed by absorbing rheumatoid factor on insolubilized human IgG. Out of 101 male and twenty-three female control subjects, only three men had a plasmin-antiplasmin titre above 1/16. Of 230 hospitalized patients, plasmin-antiplasmin titres of 1/40 or more were detected in twenty-five patients. Most of these patients had diseases which are frequently associated with in vivo coagulation or fibrinolysis, but among them there was only one who showed diffuse intravascular coagulation detectable by classical methods. In the absence of an increased plasmin-antiplasmin titre none of the haemostasis analyses were indicative of in vivo coagulation or fibrinolysis. Seven out of eight patients with diffuse intravascular coagulation of various origin had plasmin-antiplasmin titres of 1/80 or 1/160. Thus, the present latex agglutination test, owing to its simplicity and sensitivity, appears to be a practical routine screening test for detecting fibrinolytic activation in plasma.

Adolescent↗
PubMed1977

Metabolism of antithrombin III (heparin cofactor) in man: effects of venous thrombosis and of heparin administration.

The metabolism of human antithrombin III (heparin cofactor) was studied in four control subjects, in four subjects with peripheral obliterative arterial disease, in six patients with recent venous thrombosis and in one patient with clinically severe haemophilia A. The labelled antithrombin III has a high specific activity (5.75 units/mg) and displayed a single band on SDS-polyacrylamide gel electrophoresis. On Sephadex G-100 gel filtration the labelled material eluted in the same position as the antithrombin III activity in plasma. Crossed immunoelectrophoresis of a mixture of fresh plasma and labelled antithrombin III against a specific antiserum, revealed a single precipitin line in which radioactivity was concentrated. The changes in electrophoretic mobility of both the plasma antithrombin III and the labelled material following the addition of heparin to the mixture or following coagulation were identical. The purified antithrombin III behaved as a homogeneous protein in the turnover experiments. The plasma radioactivity data were approximated by a sum of two exponential terms and the metabolism of antithrombin III represented by a two compartment mammillary model. Results in the control subjects were as follows: plasma antithrombin III concentration 19.6 +/- 2.3 mg/100 ml; intravascular fraction 0.45 +/- 0.05; fractional catabolic rate 0.55 +/- 0.02 of the plasma pool per day; half-life of the plasma radioactivity 2.83 +/- 0.26 days. Circulating large molecular weight degradation products of labelled antithrombin III could not be detected by Sephadex G-100 gel filtration. No significant differences in these parameters were found in the patients with peripheral arterial insufficiency. The turnover rate of antithrombin III was normal in the patient with haemophilia A. In three patients with venous thrombosis not treated with heparin, the turnover of labelled antithrombin III was in the normal range. In three patients with venous thrombosis, treated with heparin, the plasma radioactivity half-life was significantly shortened (2.13 +/- 0.08 days) and the fractional catabolic rate increased (0.75 +/- 0.05) of the plasma pool per day). In one of these patients, the labelled antithrombin III had been incubated with an equimolar amount of heparin prior to injection. In this patient the plasma radioactivity half-life was in the same range as in the other two patients (2.15 days).

Adult↗
PubMed1977

Turnover of fibrinogen, plasminogen, and prothrombin during exercise in man.

The turnover of biologically intact high-purity 125I- and 131I-labeled fibrinogen, plasminogen, and/or prothrombin was studied in 16 untrained healthy subjects before and during strenuous physical exercise on a bicycle ergometer (repeated 4 times/day for 2 days). The exertion resulted in the well-known changes in the coagulation and fibrinolytic tests but did not induce changes in the concentration of fibrinogen, plasminogen, or prothrombin in the plasma. A significantly increased catabolism of fibrinogen and plasminogen but not of prothrombin was observed. The extent of Aalpha-chain degradation of fibrinogen in the plasma was quantitated before and 2 h after exercise, and a significant increase in degraded Aalpha chains was found in the postexercise samples. Immunochemical estimation of plasmin-antiplasmin complex in four subjects revealed a small increase after exercise in three of them. All these data support the concept that plasminogen activation and plasmin-induced fibrinogen degradation occur to some extent in man following strenuous physical exercise.

Adult↗
PubMed1977

Comparison of the reactions of neutral granulocyte proteases with the major plasma protease inhibitors and with antiplasmin.

Reaction mixtures of human serum and increasing amounts of granulocyte collagenase, elastase and chymotrypsin-like enzyme were studied by crossed immunoelectrophoresis utilizing antibodies against alpha1-antitrypsin, alpha1-antichymotrypsin, and antiplasmin. The increasing complex formation of alpha1-antitrypsin and alpha 1-antichymotrypsin with the different granulocyte proteases was not accompanied by any changes in the electrophoretic mobility or precipitate pattern of antiplasmin until the protease binding capacity of serum was saturated. The antiplasmin component in the reaction mixtures of human serum and granulocyte collagenase or elastase was not precipitated by antibodies against the proteases. The results indicate that none of the granulocyte proteases are bound by antiplasmin and that these enzymes do not activate plasminogen in serum.

Antibodies↗
PubMed1976

Identification and some properties of a new fast-reacting plasmin inhibitor in human plasma.

Fresh plasma was seeded with trace amounts of highly purified biologically intact iodine-labelled plasminogen and the plasmin-inhibitor complexes formed after activation with streptokinase or urokinase separated by gel filtration. Two radioactive peaks were observed, the first one eluted in the void volume and the second one just before the 7-S globulin peak. In incompletely activated samples, the second peak was always predominant over the first one. Both components were purified with high yield by a combination of affinity chromatography on lysine-agarose and gel filtration, and investigated by dodecylsulphate-polyacrylamide gel electrophoresis and immunoelectrophoresis. Neither component reacted with antisera against alpha1-antitrypsin, antithrombin III, C1-esterase inhibitor, inter-alpha-trypsin inhibitor or alpha1-antichymotrypsin. The component of the first peak appeared to be a complex between plasmin and alpha2-macroglobulin which reacted with antisera against human plasminogen and against alpha2-macroglobulin. The component of the second peak had a molecular weight (Mr) of 120000-140000 by dodecyl-sulphate-polyacrylamide gel electrophoresis and lpon reduction displayed a doublet band with an Mr of 65000-70000 and a band with Mr 11000. It reacted with antisera against plasminogen and with antisera raised against this complex and absorbed with purified plasminogen. The latter antisera reacted with a single component in plasma which is different from the above-mentioned plasma protease inhibitors. Specific removal of this component from plasma by immuno-absorption resulted in disappearance of the fast-reacting antiplasmin activity whereas alpha2-macroglobulin was found to represent the slower-reacting plasmin-neutralizing activity. In the presence of normal plasma levels of these proteins, the specific removal or absence of alpha1-antitrypsin, antithrombin III or C1-esterase inhibitor did not alter the inactivation rate of plasmin when added to plasma in quimolar amounts to that of plasminogen. It is concluded that only two plasma proteins are important in the binding of plasmin generated by activation of the plasma plasminogen, namely a fast-reacting inhibitor which is different from the known plasma protease inhibitors and which we have provisionally named antiplasmin, and alpha2-macroglobulin, which reacts more slowly.

Blood Proteins↗
PubMed1975

Primary structure of human fibrinogen and fibrin. Isolation and partial characterization of chains of fragment D.

Fragment D has been isolated as an apparently single molecular weight species (molecular weight about 100,000) from plasmin digests of humman fibrinogen, using a combination of affinity chromatography on insolubilized "fibrin monomer" and gel filtration. This fragment consists of three chains with molecular weights of 15,000 (Dbeta), 42,500 (Dgamma1) or 39,500 (Dgamma2), and 14,000 (Dalpha) held together by disulfide bonds. The S-carboxymethyl derivatives of the chains have been separated by gel filtration and ion exchange chromatography, and their identity has been confirmed by peptide mapping and immunological analysis. The chain with a molecular weight of 45,000 is a fragment of the Bbeta chain of fibrinogen. The chain derived from the gamma chain of fibrinogen occurred in two molecular forms having molecular weight 42,500 and 39,500. The chain derivative with molecular weight 14,000 is most likely derived from the Aalpha chain of fibrinogen. The chains were characterized by NH2-terminal sequence analysis, amino acid composition, and carbohydrate staining. The two molecular analysis, amino acid composition, and carbohydrate staining. The two molecular forms of the gamma chain appeared to be identical except for an NH2-terminal peptide extension of 23 amino acid residues in the longer chain. The latter has sequences in common with the COOH-terminal part of the gamma chain of the NH2-terminal disulfide knot (BROMBACK, B., BRONDAHL, N. J., HESSEL, B., IWANAGA, S., and WALLEN, P. (1973) J. Biol. Chem. 248, 5806-5820); its NH2-terminal residue being Ala-63 of the gamma chain of fibrinogen.

Amino Acid Sequence↗