Antithrombin Activity of Intact Human Platelets

1975 ◽  
Vol 34 (01) ◽  
pp. 115-126 ◽  
Author(s):  
Kiyoake Watanabe ◽  
Francis C Chao ◽  
James L Tullis
Keyword(s):  
Antithrombin Iii ◽  
Human Platelets ◽  
Significant Loss ◽  
Red Cells ◽  
Antithrombin Activity ◽  
Α2 Macroglobulin ◽  
Rapid Reaction ◽  
Pre Treatment ◽  
Different Characteristics

SummaryAntithrombin activity has been identified in intact washed human platelets. An apparent activity was demonstrated at platelet concentrations above 0.31 × 109/ml, when platelet suspensions were incubated with 2.0 NIH units/ml of thrombin. Neither red cells nor white cells revealed antithrombin activity. No significant loss of the platelet antithrombin activity was observed after ten successive washings or after treatment of platelets with antibodies to antithrombin III or α2-macroglobulin. Almost the same amount of antithrombin activity as normal platelets was demonstrated in the platelets from an afibrinogenemic patient. Pre-treatment of platelets with trypsin, papain, and neuroaminidase reduced the activity significantly, whereas lipase was without effect. The platelet antithrombin reacted with thrombin in less than 3 seconds, and this rapid reaction of platelet antithrombin was different from that of plasma antithrombin III or fibrinogen. The thrombin-like clotting activity of ancrod was inhibited by fibrinogen but not platelets. Also, unlike plasma antithrombin III or fibrinogen, brief exposure to heat (56° C or 60° C) reduced considerable amounts of platelet antithrombin activity. These results suggest that platelets possess a specific antithrombin with different characteristics from other known antithrombins.

Antithrombin Activity of Intact Human Platelets

1975 ◽  
Author(s):  
K. Watanabe ◽  
F. C. Chao ◽  
J. L. Tullis
Keyword(s):  
Antithrombin Iii ◽  
Human Platelets ◽  
Significant Loss ◽  
Red Cells ◽  
Antithrombin Activity ◽  
Α2 Macroglobulin ◽  
Rapid Reaction

Antithrombin activity was identified in intact washed human platelets. Red cells and white cells had no antithrombin activity. There was no significant loss of platelet antithrombin activity after ten washings or after treatment with antibodies to antithrombin III or α2-macroglobulin. Platelets from an afibrinogenemic patient showed almost the same antithrombin activity as normal platelets. Platelet antithrombin reacted with thrombin in less than 3 seconds. This rapid reaction was different from that of antithrombin. Ill or fibrinogen. The clotting activity of ancrod was inhibited by fibrinogen but not platelets. Unlike plasma antithrombin III or fibrinogen, brief exposure to 60° C reduced platelet antithrombin activity. These results suggest that platelets possess a specific antithrombin with characteristics different from other known antithrombins.

Comparison of Progressive Antithrombin Activity and the Concentration of Three Thrombin Inhibitors in Nephrotic Syndrome

1981 ◽  
Vol 46 (03) ◽  
pp. 623-625 ◽  
Author(s):  
B Boneu ◽  
F Bouissou ◽  
M Abbal ◽  
P Sie ◽  
C Caranobe ◽  
...  
Keyword(s):  
Nephrotic Syndrome ◽  
Antithrombin Iii ◽  
Heparin Therapy ◽  
Thrombin Inhibitors ◽  
Compensatory Mechanism ◽  
Dramatic Reduction ◽  
Antithrombin Activity ◽  
Α2 Macroglobulin ◽  
At Iii ◽  
A Prospective Study

SummaryIn order to compare the plasmatic progressive antithrombin activity to the concentration of three thrombin inhibitors, antithrombin III (AT III), α2 macroglobulin (α2, M), α1 anti-trypsin (α1 AT) in nephrotic syndrome, a prospective study was carried out on a group of 28 children affected with the disease. A dramatic reduction of the level of AT III and of α1 AT, two inhibitors of molecular weight close to that of albumin, was observed. The decreased level of AT III was counterbalanced by an increase in α2 M. This phenomenon accounts for the increased progressive antithrombin activity observed in all the affected children. It is suggested that the above compensatory mechanism explains the absence of thrombotic accidents in this series and that the benefit of heparin therapy is doubtful in these conditions.

A New Assay for the Measurement of Total Progressive Antithrombin

1975 ◽  
Author(s):  
Janet L. Lane ◽  
Prudence Bird ◽  
C. R. Rizza
Keyword(s):  
Rapid Method ◽  
Antithrombin Iii ◽  
Radial Diffusion ◽  
Agarose Gel ◽  
Antithrombin Activity ◽  
Gel Layer ◽  
Α2 Macroglobulin ◽  
Specific Antisera

A new rapid method for assaying total antithrombin activity has been developed based on the inactivation of thrombin incorporated into an agarose gel, during the radial diffusion of plasma in the gel. The area of thrombin inactivation is subsequently observed by the coagulation of fibrinogen in a separate agarose gel layer poured over the thrombin gel. The method is described in detail and its accuracy assessed with respect to other antithrombin assays. Using specific antisera to α2-globulin (antithrombin III), α2-macro-globulin and α1-antitrypsin, total antithrombin activity measured by this assay consisted of 47% α2-globulin, 29% α2-macroglobulin and 26% α1-antitrypsin.

scholarly journals Differential Determination of Antithrombin Activities of Plasma Antithrombin III and α2-Macroglobulin

1977 ◽  
Author(s):  
T. Matsuda ◽  
M. Ogawara ◽  
N. Hirabayashi ◽  
T. Seki ◽  
M. Murakami
Keyword(s):  
Healthy Subjects ◽  
Antithrombin Iii ◽  
Radial Immunodiffusion ◽  
Thrombin Inhibitors ◽  
Healthy Individuals ◽  
Single Radial Immunodiffusion ◽  
Antithrombin Activity ◽  
Α2 Macroglobulin ◽  
At Iii

Among several thrombin inhibitors in normal plasma, antithrombin III (AT-III) and α2-macroglobulin (α2-M) are especially important.However, there has been no appropriate method to determine AT-III and α2-M separately. This study was carried out to differentiate the actions of these two inhibitors of thrombin in plasma. By using single radial immunodiffusion method, we observed that AT-III was adsorbed to bentonite, although α2-M was not. It was necessary to incubate 1 ml of oxalated plasma with 300 mg of bentonite at 37°C for 10 minutes to remove AT-III completely. Supernated plasma contains neither AT-III nor fibrinogen which affects antithrombin activity of plasma. From these results, it is evident that antithrombin activity of plasma adsorbed with bentonite was attributed to that of α2-M. Antithrombin activities of plasma adsorbed with bentonite in 20 healthy subjects were significantly correlated with concentrations of α2-M in plasma, measured immunologically (r=+0.87, p<0.001). It is reasonable to presume that difference between antithrombin activity of defibrinated plasma by heating at 56°C for 2 minutes and that of bentonite treated plasma mainly indicates activity of AT-III. These differences measured in 20 healthy individuals were significantly correlated with plasma AT-TH levels, assayed immunochemically (r=+0.53, p<0.01).

The Relationship between Antithrombin III, Progressive Antithrombin Activity and Anti Xa Activity in Plasma

1975 ◽  
Author(s):  
Katherine Whigham ◽  
P. W. Howie ◽  
C. D. Forbes ◽  
C. R. M. Prentice
Keyword(s):  
Antithrombin Iii ◽  
Factor Xa ◽  
Normal Subjects ◽  
Strong Positive Correlation ◽  
Positive Correlation ◽  
Antithrombin Activity ◽  
Α2 Macroglobulin ◽  
Immunological Assays ◽  
The Relationship

In 30 normal subjects, progressive antithrombin activity, as measured by the rate of thrombin neutralisation in ancrod-defibrinated plasma, was compared with antithrombin III, as measured by radial immunodiffusion. No significant correlation was found between the two methods of antithrombin measurement (r = —0.101). Similarly, no correlation was found between progressive antithrombin activity and immunological measurements of α2 macroglobulin and α1 antitrypsin. These results were not changed by using thrombin purified by Amberlite 1RC50 chromatography in place of commercial thrombin in the clotting test. There was, however, a strong positive correlation between the measurements of progressive antithrombin activity using the commercial and purified forms of thrombin (r = 0.78, p < 0.001). In contrast, there was a positive correlation between antithrombin III and anti-factor Xa activity in plasma (r = 0.48, p < 0.01). There was no correlation between plasma anti Xa activity and α2 macroglobulin or α1 antitrypsin.These results suggest that plasma antithrombin activity is a measure of the activities of several plasma proteins and that antithrombin III may not be the major determinant of antithrombin activity. There is little evidence that immunological assays of antithrombin III reflect total thrombin inhibitory capacity as measured by the biological assay. Caution must be exercised in extrapolating from immunological measurements of antithrombin III to antithrombin activity in-vivo.

The Reversible Antithrombin Activity of Incubated Plasma

1977 ◽  
Vol 37 (01) ◽  
pp. 123-135 ◽  
Author(s):  
E. D Gomperts ◽  
J. C Stavridis ◽  
R. S Mibashan
Keyword(s):  
Calcium Ions ◽  
Antithrombin Iii ◽  
Thrombin Time ◽  
Platelet Poor Plasma ◽  
Antithrombin Activity ◽  
Physicochemical Changes ◽  
Enhanced Activity ◽  
Α2 Macroglobulin ◽  
Glass Tubes ◽  
Fibrin Monomers

SummaryThe thrombin time of normal citrated plasma is progressively prolonged on incubation in open glass tubes at 37° C. The phenomenon is dependent on the temperature and duration of incubation, on the pH, and on the concentration of calcium ions present. Platelet-rich citrated plasma fails to exhibit augmented antithrombin activity when similarly incubated, and the addition of washed platelets to platelet-poor plasma inhibits this activity. The clot retarding action of incubated plasma against thrombin is also manifested against Arvin (Ancrod), but not against Reptilase-R. This thrombin time lenghening may be inhibited by incubation with anti-antithrombin III antiserum thus indicating that the phenomenon of thrombin time lengthening is consistent with enhanced activity of antithrombin III. It is unlikely that alterations in the activity of α2-macroglobulin, is important in the reduced thrombin-coagulability of incubated plasma. Interference with the polymerisation of fibrin monomers by the physicochemical changes may contribute to the observed phenomenon.

Effect Of Heparin And Protamine On The Antithrombin Activity Of α2-Macroglobulin

1981 ◽  
Author(s):  
J N Shanberge ◽  
J Love ◽  
N Swanborg
Keyword(s):  
Inhibitory Action ◽  
Antithrombin Iii ◽  
Protamine Sulfate ◽  
Radial Immunodiffusion ◽  
Clotting Time ◽  
Antithrombin Activity ◽  
Α2 Macroglobulin ◽  
Heparin Complexes ◽  
Heparin Fractions

It has been shown that certain fractions of porcine mucosal heparin may form complexes with α2-macroglobulin but still retain their capacity to activate antithrombin III. These heparin fractions are not neutralized by protamine. The effect of heparin and protamine on the antithrombin activity of α2-macroglobulin has now been studied.Plasma to which a tritium-labelied heparin was added was chromatographed on Sephadex G-200. The macromolecular fractions, in which there is heparin (indicated by radioactivity) and α2-macroglobulin (demonstrated by radial immunodiffusion), have no immediate inhibitory action on the thrombin clotting time (TCT) of fibrinogen but do inhibit the TCT of whole plasma. In addition, these fractions have progressive antithrombin activity (PATA) but less than that in fractions containing α2-macroglobulin obtained with non-heparinized plasma. Fischer et al stated that heparin complexes with thrombin to prevent its inactivation by α2-macroglobulin. However, when the heparinized plasma is “neutralized” with protamine sulfate prior to chromatography, the PATA of the macromolecular fractions is reduced further even through these fractions still have immediate inhibitory action on the TCT of plasma. When the same concentration of protamine is added to non-heparinized plasma, the PATA in the macromolecular fractions is not altered. Treatment of the macromolecular fractions from heparinized plasma with triethylaminoethyl cellulose removes the immediate inhibitory action on the TCT of plasma but does not affect the PATA of the fractions. How the heparin and protamine interact to decrease the PATA of α2-macroglobulin containing fractions is not as yet determined. Recently it was found that a similar complexing of heparin fractions with α1-antitrypsin may occur in the same way as with α2-macroglobulin.

Effect of Anabolic Steroids on Plasma Antithrombin III

1975 ◽  
Vol 34 (01) ◽  
pp. 106-114 ◽  
Author(s):  
I. D Walker ◽  
J. F Davidson ◽  
P Young ◽  
J. A Conkie
Keyword(s):  
Heart Disease ◽  
Ischaemic Heart Disease ◽  
Oral Contraceptives ◽  
Disseminated Intravascular Coagulation ◽  
Fibrinolytic Activity ◽  
Antithrombin Iii ◽  
Anabolic Steroids ◽  
Intravascular Coagulation ◽  
Clear Cut ◽  
Α2 Macroglobulin

SummaryThe effect of seven different anabolic steroids (Ethyloestrenol, Methenolone acetate, Norethandrolone, Methylandrostenediol, Oxymetholone, Methandienone, and Stanozolol) on three α-globulin antiprotease inhibitors of thrombin and plasmin was studied in men with ischaemic heart disease. In distinct contrast to the oral contraceptives, five of the six 17-α-alkylated anabolic steroids studied produced increased plasma Antithrombin III levels and five produced decreased levels of plasma α2-macroglobulin. The effect on plasma α1antitrypsin levels was less clear-cut but three of the steroids examined produced significantly elevated levels. The increased plasma fibrinolytic activity which the 17-α-alkylated anabolic steroids induce is therefore unlikely to be secondary to disseminated intravascular coagulation.

Isolation of Antithrombin III from Normal and α1-Antitrypsin-Deficient Human Plasma

1977 ◽  
Vol 38 (02) ◽  
pp. 0475-0485 ◽  
Author(s):  
Anna D. Borsodi ◽  
Ralph A. Bradshaw
Keyword(s):  
Enzymatic Activity ◽  
Isoelectric Focusing ◽  
Antithrombin Iii ◽  
Factor Xa ◽  
Antithrombin Activity ◽  
Bovine Trypsin ◽  
Normal Plasma ◽  
Antitrypsin Deficiency ◽  
Simplified Procedure ◽  
Stimulation Of

SummaryThe plasma of individuals, hetero- or homozygous for α1-antitrypsin deficiency, contains greatly decreased amounts of antithrombin activity as assayed against factor Xa. However, heparin stimulation of the residual antithrombin activity is observed, which is comparable to that of normal plasma. Antithrombins isolated from both normal and α1-antitrypsin deficient plasma by a simplified procedure are indistinguishable in both properties and yields. The microheterogeneity observed on isoelectric focusing of both preparations can be eliminated by treatment with neuraminidase. Neither purified human antithrombin nor α1-antitrypsin, when assayed against bovine trypsin, is stimulated by heparin. These results clearly establish the unique natures of antithrombin and α1-antitrypsin and show that about 75% of the antithrombin activity measured in normal plasma is due to α1-antitrypsin. Estimates of anti thrombin III activity in normal plasma by assays dependent on enzymatic activity can probably be obtained only in the presence of heparin.

Antithrombin III Antigen in Human Platelets

1985 ◽  
Vol 54 (03) ◽  
pp. 599-602 ◽  
Author(s):  
M Léon Alhenc-Gelas ◽  
M Aiach ◽  
A Gorenflot ◽  
J P Andreux
Keyword(s):  
Arachidonic Acid ◽  
Enzyme Immunoassay ◽  
Antithrombin Iii ◽  
Human Platelets ◽  
Mean Value ◽  
Normal Blood ◽  
Freezing And Thawing ◽  
Healthy Donors ◽  
At Iii ◽  
Storage Granules

SummaryImmunoreactive AT III was found in human platelets. AT III antigen was quantified in platelets taken from each of 17 healthy donors by a specific competitive enzyme immunoassay using purified AT III and AT III antibodies. AT III antigen levels in extracts of washed platelets disrupted by freezing and thawing ranged from 32 to 140 ng per 109 platelets with a mean value of 70.3 ± 27.3. When stimulated by arachidonic acid, the platelets released AT III antigen together with immunoreactive fibrinogen. These results show that AT III is present in platelets at a level corresponding to approximately 0.01% of total antithrombin in normal blood, and suggest that platelet AT III, like fibrinogen, is contained in the storage granules.

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