The Contribution of Plasma Protease Inhibitors to Antiplasmin Activity in Man

1976 ◽  
Vol 51 (2) ◽  
pp. 215-218
Author(s):  
G. P. M. Crawford ◽  
D. Ogston ◽  
A. S. Douglas
Keyword(s):  
Human Plasma ◽  
Protease Inhibitors ◽  
Trypsin Inhibitor ◽  
Antithrombin Iii ◽  
The Other ◽  
Plasmin Activity ◽  
Neutralizing Activity ◽  
Α2 Macroglobulin

1. Human plasma contains a variety of proteins that are capable of inhibiting plasmin activity. Whole plasma possesses ‘rapid’ and ‘progressive’ plasmin-neutralizing activity: this study assesses the contribution of individual protease inhibitors to this plasmin-neutralizing property of plasma. 2. Rapid and progressive antiplasmin activities of human plasma correlate with α2-macroglobulin and α1-antitrypsin concentrations respectively. 3. Fluctuations in the amounts of the other measured inhibitors (antithrombin III, Cl inactivator and inter-α-trypsin inhibitor) did not influence the measured antiplasmin activity.

The Contribution of Individual Protease Inhibitors to Plasma Antiplasmin Activity

1975 ◽  
Author(s):  
G. P. M. Crawford ◽  
D. Ogston ◽  
A. S. Douglas
Keyword(s):  
Protease Inhibitors ◽  
Trypsin Inhibitor ◽  
Antithrombin Iii ◽  
Correlation Coefficients ◽  
Zero Order ◽  
Plasmin Activity ◽  
Valid Assessment ◽  
Alpha 1 Antitrypsin ◽  
The Individual

The plasma proteins alpha-1-antitrypsin, alpha-2-macroglobulin, antithrombin III, Cl inhibitor and inter-alpha trypsin inhibitor have been shown to be capable of inhibiting plasmin activity. Whole plasma possesses both ‘rapid’ and ‘progressive’ plasminneutralising activity: the study was intended to assess the contribution of individual protease inhibitors to this plasminneutralising activity.The antiplasmin properties, assessed as ‘rapid’ and ‘progressive’ by the neutralisation of plasmin in a caseinolytic assay, and the levels of the individual protease inhibitors, assayed by single radial immunodiffusion, were measured in the plasma of 35 subjects. Statistical analysis using zero order correlation coefficients and zero order partial correlation coefficients indicated that ‘rapid’ and ‘progressive’ antiplasmin activity correlated with the plasma concentration of alpha-2-macroglobulin and alpha-1-antitrypsin respectively, but not with antithrombin III, Cl inhibitor and interalpha trypsin inhibitor.It is concluded that alpha-2-macroglobulin and alpha-1-antitrypsin are the major plasma inhibitors of plasmin and that immunological determination of these proteins provides a valid assessment of plasma antiplasmin activity.

scholarly journals Different molecular forms of plasminogen and plasmin produced by urokinase in human plasma and their relation to protease inhibitors and lysis of fibrinogen and fibrin

1974 ◽  
Vol 143 (2) ◽  
pp. 273-283 ◽  
Author(s):  
Sten Müllertz
Keyword(s):  
Human Plasma ◽  
Protease Inhibitors ◽  
Gel Electrophoresis ◽  
Protease Activity ◽  
Gel Filtration ◽  
Molecular Forms ◽  
Crossed Immunoelectrophoresis ◽  
Low Concentrations ◽  
Α2 Macroglobulin ◽  
Esterase Inhibitor

Urokinase-activated human plasma was studied by gel electrophoresis, gel filtration, crossed immunoelectrophoresis and electroimmunoassay with specific antibodies and by assay of esterase and protease activity of isolated fractions. Urokinase induced the formation of different components with plasminogen+plasmin antigenicity. At low concentrations of urokinase, a component with a KD value of 0.18 by gel filtration and post β1 mobility by gel electrophoresis was detected. The isolated component had no enzyme or plasminogen activity. In this plasma sample fibrinogen was not degraded for 10h, but when fibrin was formed, by addition of thrombin, fibrin was quickly lysed, and simultaneously a component with a KD value of 0 and α2 mobility appeared, which was probably plasmin in a complex with α2 macroglobulin. This complex showed both esterase and protease activity. After gel filtration with lysine buffer of the clotted and lysed plasma another two components were observed with about the same KD value by gel filtration as plasminogen (0.35), but β1 and γ mobilities by gel electrophoresis. They appeared to be modified plasminogen molecules, and possibly plasmin with γ mobility. Similar processes occurred without fibrin at higher urokinase concentrations. Here a relatively slow degradation of fibrinogen was correlated to the appearance of the plasmin–α2 macroglobulin complex. The fibrin surface appeared to catalyse the ultimate production of active plasmin with a subsequent preferential degradation of fibrin and the formation of a plasmin–α2 macroglobulin complex. The gel filtration and electrophoresis of the plasma protease inhibitors, α1 antitrypsin, inter-α-inhibitor, antithrombin III, and C1-esterase inhibitor indicated that any complex between plasmin and these inhibitors was completely dissociated. The β1 and post β1 components appear to lack correlates among components occurring in purified preparations of plasminogen and plasmin.

The Measurement of Human Plasma Antiplasmin Activity by Radial Diffusion Assay

1978 ◽  
Vol 39 (02) ◽  
pp. 437-449 ◽  
Author(s):  
W A Andes
Keyword(s):  
Human Plasma ◽  
Antithrombin Iii ◽  
Normal Subjects ◽  
Radial Diffusion ◽  
Physical Methods ◽  
Α2 Macroglobulin ◽  
Radial Diffusion Assay ◽  
Diffusion Assay

SummaryAn assay of human antiplasmins has been developed utilizing radial diffusion of plasma from wells cut in plasmin-enriched, fibrinogen-agarose plates. After diffusion the fibrinogen is clotted. Zones of fibrin protected from background fibrinolysis develop as the result of plasma antiplasmin activity. A pooled plasma standard was taken to contain 100 % antiplasmin activity. Antiplasmin activity of 52 normal subjects varied from 64 to 132 %. Washed platelets contained 1-5 % antiplasmin activity. Using antisera to precipitate individual inhibitors, physical methods of separation, and electrophoresis of plasma in agarose, several different proteins were found to have antiplasmin activity in this assay. Thus, α2-macroglobulin contributed 56%, α1-antitrypsin 20%, antithrombin III 2%, and other proteins 22% of the total antiplasmin activity. 1 ml of whole plasma neutralized 7.0 CTA units of plasmin.

Relative Importance Of Plasma Protease Inhibitors In The Inactivation Of Kallikrein In Human Plasma

1981 ◽  
Author(s):  
M Schapira ◽  
C F Scott ◽  
R W Colman
Keyword(s):  
Human Plasma ◽  
Antithrombin Iii ◽  
Plasma Concentrations ◽  
The Other ◽  
High Molecular Weight Kininogen ◽  
Physiological Conditions ◽  
First Order ◽  
Complete Inactivation ◽  
Important Regulator ◽  
Normal Human

Human plasma contains several inhibitors of plasma kallikrein (KAL), including C1-inhibitor (C1INH), α2-macro- globulin (α2M), antithrombin III (ATIII), and α1-antitrypsin (αlAT). Studies in purified systems have allowed us to quantitate the kinetic constants of each isolated inhibitor. We also have demonstrated that high molecular weight kininogen (HMWK) is an important regulator since it decreases the inactivation rate of KAL by all inhibitors except α2M. When purified inhibitors and HMWK are present at plasma concentrations, it can be calculated that C1INH, α2M, ATIII and α1AT account respectively for 49%, 49%, 0.8% and 0.2% of the KAL inhibitory activity. To assess if this prediction derived from purified system adequately describes the inhibition of KAL under more physiological conditions, we incubated KAL with normal human plasma (NHP), C1INH-deficient plasma (C1INH-D), and α2M-deficient plasma (α2M-D). KAL activity_was assessed using H-D-Pro-Phe-Arg- Nan as a substrate. C1INH-D was obtained from a patient with hereditary angioedema. C1INH in C1INH-D was 15% of the normal value as assessed by radial immunodiffusion. α2M-D was obtained by selective and complete inactivation of α2M by methylamine. The pseudo-first-order rate constants for the inactivation of_KAL by NHP, C1INH-D, α2M-D, and plasma deficient in both C1INH and α2M were respectively 8.8, 5.0, 5.5, and 1.8 S-1(×l03). Therefore, C1INH accounted for 63% of the observed inhibition, α2M for 35%, and all the other inhibitors for 2%. When these values were corrected for the concentration of HMWK, C1INH accounted for 47% of the inhibition, α2M for 51%, and all the other inhibitors for <2%. Thus, there is an excellent agreement between the results obtained when KAL is inhibited in purified systems and those obtained when it is inhibited in plasma. Moreover, these results indicate that C1INH and α2M are the only important inhibitors of KAL in NHP.

scholarly journals SUBSTRATES OF HAGEMAN FACTOR

1974 ◽  
Vol 140 (6) ◽  
pp. 1615-1630 ◽  
Author(s):  
Louis W. Heck ◽  
Allen P. Kaplan
Keyword(s):  
Trypsin Inhibitor ◽  
Gel Electrophoresis ◽  
Antithrombin Iii ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Lima Bean ◽  
Hageman Factor ◽  
Α2 Macroglobulin ◽  
Coagulant Activity ◽  
Bean Trypsin Inhibitor

Unactivated partial thromboplastin antecedent (PTA) has been purified by sequential chromatography of plasma on quaternary aminoethyl Sephadex, sulphoprophyl Sephadex, Sephadex G-150, and passage over an anti-IgG immunoadsorbant. The preparation gave a single band after alkaline disc gel electrophoresis, sodium dodecyl sulfate (SDS) gel electrophoresis and isoelectric focusing in acrylamide gels and was found to have a mol wt of 175,000 by gel filtration, 163,000 by SDS gel electrophoresis, and an isoelectric point of 8.8–9.4 (peak 9.0–9.1). Pre-PTA was activated directly by activated Hageman factor or by Hageman factor prealbumin fragments. Its coagulant activity was inhibited by DFP, soybean trypsin inhibitor and trasylol but not by lima bean trypsin inhibitor or ovomucoid trypsin inhibitor indicating that activated PTA possesses the same inhibition profile utilizing these reagents as does plasma kallikrein. A major plasma inhibitor of activated PTA was found to be a 65,000 mol wt α-globulin which was isolated free of α1-chymotrypsin inhibitor, inter α-trypsin inhibitor, α2-macroglobulin, and the other known inhibitors of activated PTA, the activated first component of complement (C1 INH), and antithrombin III. Its physicochemical properties were identical to α1-antitrypsin, and it was absent in α1-antitrypsin-deficient plasma thereby identifying this PTA inhibitor as α1-antitrypsin.

An Endogenous Protease Activating Plasma Inactive Renin

1978 ◽  
Vol 55 (s4) ◽  
pp. 133s-134s ◽  
Author(s):  
B. J. Leckie
Keyword(s):  
Human Plasma ◽  
Serine Protease ◽  
Protease Inhibitors ◽  
Trypsin Inhibitor ◽  
Soya Bean ◽  
Acid Activation ◽  
Bean Trypsin Inhibitor ◽  
Inactive Renin ◽  
Inhibited Acid ◽  
Endogenous Protease

1. The protease inhibitors Trasylol and soya-bean trypsin inhibitor prevented the activation of plasma inactive renin by acid. 2. N-Ethylmaleimide inhibited acid-activation to some extent but o-phenanthroline had no effect. 3. Acid-activation of the inactive renin in human plasma is mediated by a serine protease.

Blood Coagulation Inhibitor in a Snake Plasma (Bothrops jararaca)

1973 ◽  
Vol 30 (01) ◽  
pp. 106-113 ◽  
Author(s):  
Linda Nahas ◽  
Ferruccio Betti ◽  
Aura S. Kamiguti ◽  
Hissae Sato
Keyword(s):  
Human Plasma ◽  
Blood Coagulation ◽  
Antithrombin Iii ◽  
The Other ◽  
Normal Human Plasma ◽  
Clotting Time ◽  
Bovine Thrombin ◽  
Heat Stable ◽  
Coagulation Inhibitor ◽  
Normal Human

SummaryExperiments with plasmas samples from the snake B. jararaca and X. merremii have shown that the two differ markedly with respect to their reaction to bovine thrombin. The plasma of X. merremii appears to react in a manner very similar to that of normal human plasma both using the thrombin clotting test and using the antithrombin III assay of Astrup and Darling (Biggs and Macfariane 1962). The plasma of B. jararaca on the other hand, contains, in addition to antithrombin III, an inhibitor which resembles mammalian heparin. This inhibitor prolongs the thrombin clotting time of human and X. merremii plasmas, is recorded by the antithrombin assay and is neutralized by protamine. The inhibitor is however, not identical with mammalian heparin since it is not adsorbed by BaSO4 and Al(OH)3 (which adsorb heparin) and it is heat labil whereas mammalian heparin is heat stable.

Assessment of the Relative Contribution of Different Protease Inhibitors to the Inhibition of Plasmin In Vivo

1993 ◽  
Vol 69 (02) ◽  
pp. 141-146 ◽  
Author(s):  
Marcel Levi ◽  
Dorina Roem ◽  
Angela M Kamp ◽  
Jan Paul de Boer ◽  
C Erik Hack ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Protease Inhibitors ◽  
Proteolytic Enzyme ◽  
Antithrombin Iii ◽  
C1 Inhibitor ◽  
Relative Contribution ◽  
Α2 Macroglobulin ◽  
Diffuse Intravascular Coagulation

SummaryIt has been shown that the most important inhibitor of plasmin is α2-antiplasmin, however, other protease inhibitors are able to inhibit this proteolytic enzyme as well. The contribution of the various protease inhibitors to the inhibition of plasmin in vivo has never been quantitatively assessed.To assess the relative contribution of the different protease inhibitors on the inhibition of plasmin we developed a series of sensitive immunoassays for the detection of complexes between plasmin and the protease inhibitors α2-antiplasmin, α2-macroglobulin, antithrombin III, α1antitrypsin and C1-inhibitor, utilizing monoclonal antibodies that are specifically directed against complexed protease inhibitors and a monoclonal antibody against plasmin.It was confirmed that α2-antiplasmin is the most important inhibitor of plasmin in vivo, however, complexes of plasmin with α2-macroglobulin, antithrombin III, α1antitrypsin- and C1-inhibitor were also detected. Particularly during activation of fibrinolysis complexes between plasmin and inhibitors other than α2-antiplasmin were detected. It was observed that during different situations the inhibition profile of plasmin was not constant e.g. in patients with diffuse intravascular coagulation plasma levels of plasmin-α1-antitrypsin and plasmin-C1-inhibitor were increased whereas in plasma from patients who were treated with thrombolytic agents complexes of plasmin with α2-macroglobulin and with antithrombin III were significantly elevated.In conclusion, we confirmed the important role of α2-antiplasmin in the inhibition of plasmin, however, in situations in which fibrinolysis is activated other protease inhibitors also account for the inhibition of plasmin in vivo. Further investigations to assess the role of the various protease inhibitors in the fibrinolytic system can be assisted by the assays described in this study.

The Inhibition of Tissue Activator and Urokinase by Human Plasma

1982 ◽  
Vol 47 (03) ◽  
pp. 265-268 ◽  
Author(s):  
J E Walker ◽  
D Ogston
Keyword(s):  
Molecular Weight ◽  
Human Plasma ◽  
High Molecular Weight ◽  
Antithrombin Iii ◽  
Inhibitory Effect ◽  
The Other ◽  
Alpha1 Antitrypsin ◽  
Inhibitory Activities ◽  
Fast Acting ◽  
Molecular Weight Fractions

SummaryPlasma reduced the activity of urokinase and, to a lesser extent, tissue activator on unheated fibrin plates. After depletion of plasminogen by lysine-Sepharose the inhibitory effect of plasma was increased.Normal plasma separated on DEAE-Sepharose or, after passage through lysine-Sepharose, on Sephadex G-100 provided fractions in three regions capable of inhibiting tissue activator. The high molecular weight fractions inhibited both activators, but after plasma fractionation on Sephadex 4B the inhibitory activities against tissue activator and urokinase eluted separately. Urokinase inhibition eluted with a2-macroglobulin: the inhibition of tissue activator was of higher molecular weight and of greater lability. Inhibition of tissue activator was found in two lower molecular weight regions: the fractions of one region inhibited tissue activator and coincided with the elution of fast-acting antiplasmin activity. The other region inhibited both tissue activator and urokinase. Alpha1 antitrypsin and antithrombin III eluted after both inhibitory regions.

Application of Crossed Radio Immunoelectrophoresis for the Demonstration of Thrombin Binding Proteins in Human Plasma

1979 ◽  
Author(s):  
L. Grimmer ◽  
L. Aukrust ◽  
P. Andersen
Keyword(s):  
Human Serum ◽  
Human Plasma ◽  
Binding Proteins ◽  
Antithrombin Iii ◽  
Crossed Immunoelectrophoresis ◽  
Α2 Macroglobulin

In order to demonstrate thrombin binding proteins in human plasma in a direct way, crossed Immunoelectrophoresis (CIE) presipitates were prepared by using human plasma and anti human serum. These presipitates were incubated with 125 I-thrombin and subsequently subjected to autoradiography. Binding of thrombin was demonstrated to α2-macroglobulin only By preiricubating heat defibrinated plasma with 125I-thrombin prior to CIE and performing autoradiography without further incubation with 125I-thrombin, binding to antithrombin III was demonstrated as well. No binding of thrombin to α1-antitrypsin was observed in any of the experiments.

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