bluuebunny/crossref_metadata_2025 huggingface dataset:
Increased Tissue Thromboplastin Activity in Monocytes of Patients with Meningococcal Infection: Related to an Unfavourable Prognosis
Authors: Østerud B , Flægstad T
Published: July 2018

In 16 patients, 13 with meningococcal infection and 3 suspected to have this infection, 8 patients were found to possess significant higher level of tissue thromboplastin activity of their monocytes isolated from the blood at the admission to the hospital than normal. Five of those 8 patients had an extremely high concentration, > 60-300 fold increase, and all these patients died. The exposed tissue thromboplastin activity on the surface of the endotoxin stimulated monocytes is probably the direct inducer of disseminated intravascular coagulation (DIC) in meningococcal infection.
Increased Urinary β-Thromboglobulin Excretion in Diabetes Assayed with a Modified RIA Kit-Technique
Authors: van Oost Bernard A , Veldhuyzen Bernard , Timmermans Ans P M , Sixma Jan J
Published: July 2018

A modification of the commercial radioimmunoassay (RIA) kit adapting to the assay of β-thromboglobulin (β-TG) in the urine, is described. The urine was concentrated by dialysis against dry Sepharose G 200. l31I-albumin was used as marker. Experiments in which l25I-β-TG and 131I-albumin were added simultaneously showed a parallel recovery of both markers in the final sample. The sensitivity of the RIA was enhanced by incubation overnight of the urine sample with the antiserum. Increased urinary pTG values were found in 20 patients with diabetes mellitus as compared to 20 young healthy normals. Increased urinary β-TG values correlated with decreased creatinine clearance, but the difference between diabetics and normals was still observed when patients with a creatinine clearance of less than 100 ml/min were excluded. The urine βTG level was correlated with the presence of neovascularization within the diabetic group.
High Molecular Weight Forms of Antithrombin III Complexes in Blood
Authors: Marciniak E , Gora-Maslak G
Published: July 2018

A double antibody competition radioimmunoassay was developed that allowed to detect specifically as little as 15 ng antithrombin III (AT III) per ml of the assayed material. In normal plasma examined by this assay, AT III concentration averaged 199 ± 21 μg/ml. Complexes of AT III with thrombin or factor X a crossreacted with free AT III in 87% and 95%, respectively. Molecular forms of AT III produced in plasma treated with coagulation enzymes, or in serum, were assessed by measuring immunoreactive AT III in fractions obtained by gel filtration chromatography on Sephadex G-200. AT III bound by thrombin in fibrinogen free-plasma ranged in molecular size from 160,000 to above 250,000. Similar aggregation occurred when monomeric complex of purified AT III and thrombin, of 90,000 Mr, was added to plasma. Presence of heparin intensified the degree of aggregation. In factor Xa-treated plasma AT III was converted into components with 160,000 Mr, or less. No complexes below 200,000 Mr were present in serum. They decreased in size to 160,000 Mr after affinity chromatography on heparin-Sepharose. These results indicated that blood represents a unique milieu conducive to aggregation of bound AT III. It appears, however, that AT III complexes present in blood may not only aggregate, but also associate with other serum proteins through unstable binding most likely caused by the enzyme component of the complex.
Fibrin Network Structure: Modification by Platelets
Authors: Dhall T Z , Shah G A , Ferguson I A , Dhall D P
Published: July 2018

Platelets play a fundamental role in haemostasis and thrombosis. They are known to undergo characteristic changes including release of subcellular material during clotting. The effect of subcellular platelet material on fibrin network structure, however, has not previously been investigated. Using opacity ratio, syneresis, permeation and electron microscopy it was found that subcellular platelet material extracted into NaCl is able to influence fibrin network structure of clots made from purified fibrinogen as well as platelet-poor plasma. Such clots had higher opacity ratio, reduced syneresis and lower permeability than control clots. Further, the responsible platelet material is heat labile and is released from the platelets during their aggregation with several common aggregating agents. Morphometric analysis of transmission electron micrographs has shown that fibrin fibres in plasma clots made in the presence of platelet subcellular material are thinner than those in control clots. In addition, plasma clots made in the presence of platelet extract had a higher resistance to fibrinolytic digestion than control clots. Thus, platelets play a hitherto undescribed role in regulating fibrin network structure.
The Basis for the Increase in Factor VIII Procoagulant Activity During Exercise
Authors: Kopitsky Robert G , Switzer Mary Ellen P , Williams R Sanders , McKee Patrick A
Published: July 2018

We studied the effect of acute exercise on the ability of thrombin to activate plasma factor VIII (FVIII) activity in 20 healthy males. The subject showed an average exercise-related increase in FVIII activity of 54.5±8.2% over pre-exercise FVIII activity (p<0.001). When exposed to the same concentration of thrombin, post-exercise FVIII activity showed greater enhancement than pre-exercise FVIII activity: 157.1±12.8% increase in activity versus 117.3±9.9%, respectively (p<0.01). The degree of the potentiated thrombin effect in post-exercise samples relative to pre-exercise samples was linearly correlated with the degree of the exercise-related increase in FVIII activity. Taken together with our previous observations that the extent of thrombin enhancement of FVIII activity varies inversely with the mole ratio of FVIII/von Willebrand factor subunits to thrombin, these findings imply that release of FVIII does not occur during exercise, and that the exercise-related increase in FVIII activity results primarily, if not completely, from activation of already circulating but inactive FVIII.
Heparin Counteracts the Antiaggregating Effect of Prostacyclin by Potentiating Platelet Aggregation
Authors: Bertelé Vittorio , Roncaglioni Maria Carla , Donati Maria Benedetta , Gaetano Giovanni de
Published: July 2018

It has recently been reported that heparin neutralizes the inhibitory effect of prostacyclin (PGI2) on human platelet aggregation. The mechanism of this interaction has not yet been unequivocally established. We present here evidence that heparin (Liquemin Roche) does not react directly with PGI2 but counteracts its inhibitory effect by potentiating platelet aggregation. In the absence of heparin, PGI2 was a less effective inhibitor of platelet aggregation induced by the combination of ADP and serotonin than by ADP alone. Moreover, the inhibitory effect of PGI2 was similarly reduced when increasing the concentrations of ADP (in the absence of heparin). The lack of a specific interaction between heparin and PGI2 is supported by the observation that, in the presence of heparin, other prostaglandins such as PGD2 and PGE1, and a non-prostanoid compound such as adenosine also appeared to lose their inhibitory potency. It is concluded that heparin opposes platelet aggregation inhibitory effect of PGI2 by enhancement of platelet aggregation.
Covalent Complexes Between Low Molecular Weight Heparin Fragments and Antithrombin III – Inhibition Kinetics and Turnover Parameters
Authors: Hoylaerts M , Holmer E , Mol M de , Collen D
Published: July 2018

Two high affinity heparin fragments (A/r 4,300 and M, 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 (t'/j) of both low Afr-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 ≃ 7.7 hr), approaching that of free antithrombin III (t1/2 ≃ 11 ± 0.4 hr) and resulting in a 30fold longer life time of factor Xa inhibitory activity in plasma as compared to that of free intact heparin (t1/2 ≃ 0.25 ± 0.04 hr).
A New Dysfibrinogenemia: Fibrinogen Oslo IV
Authors: Stormorken H , Brosstad F , Seim H
Published: July 2018

A family with dysfibrinogenemia is described. The abnormal fibrinogen occurred in three successive generations indicating a dominant hereditary pattern. Thrombin and reptilase times were about twice the normal value. This was shown to be caused by a polymerization defect, fibrinopeptide release being normal. Platelet aggregation was undisturbed, indicating normal platelet-fibrinogen binding. The bleeding time was normal and there was no bleeding tendency. However, an obscure recurrent pulmonary ailment may, or may not, be related to the dysfibrinogenemia. The abnormal fibrinogen was tentatively termed Oslo IV.
Effects of Dipyrone on Prostaglandin Production by Human Platelets and Cultured Bovine Aortic Endothelial Cells
Authors: Eldor A , Polliack G , Vlodavsky I , Levy M
Published: July 2018

Dipyrone and its metabolites 4-methylaminoantipyrine, 4-aminoantipyrine, 4-acetylaminoantipyrine and 4-formylaminoan- tipyrine inhibited the formation of thromboxane A2 (TXA2) during in vitro platelet aggregation induced by ADP, epinephrine, collagen, ionophore A23187 and arachidonic acid. Inhibition occurred after a short incubation (30–40 sec) and depended on the concentration of the drug or its metabolites and the aggregating agents. The minimal inhibitory concentration of dipyrone needed to completely block aggregation varied between individual donors, and related directly to the inherent capacity of their platelets to synthesize TXA2.

Incubation of dipyrone with cultured bovine aortic endothelial cells resulted in a time and dose dependent inhibition of the release of prostacyclin (PGI2) into the culture medium. However, inhibition was abolished when the drug was removed from the culture, or when the cells were stimulated to produce PGI2 with either arachidonic acid or ionophore A23187.

These results indicate that dipyrone exerts its inhibitory effect on prostaglandins synthesis by platelets or endothelial cells through a competitive inhibition of the cyclooxygenase system.
Pre- and Postoperative Levels of Antithrombin III with Special Reference to Thromboembolism After Total Hip Replacement
Authors: Fredin H , Nilsson B , Rosberg B , Tengborn L
Published: July 2018

A prospective study of antithrombin HI, determined by electroimmunochemical assay or an amidolytical method, was carried out with special reference to thromboembolism after total hip replacement. Two hundred and seven patients were randomly allocated to thromboembolic prophylaxis with dextran 70 or low dose heparin combined with dihydroergotamine. Deep vein thrombosis determined by phlebography of the operated leg or pulmonary embolism diagnosed with perfusion/ventilation scintigraphy developed in 51% of the total material and did not differ significantly between the two groups of prophylaxis or between patients with a preoperative At III below normal and those with a normal value. The correlation between the two assay methods for At HI was 0.61. An initial, postoperative decrease in At III was noted with a parallel fall in hematocrit and fibrinogen, later followed by an increase of the plasma proteins. It is concluded that the immediate postoperative decrease of At III is mostly due to hemodilution.
Factors Influencing the Deaggregation of Human and Rabbit Platelets
Authors: Kinlough-Rathbone R L , Mustard J F , Perry D W , Dejana E , Cazenave J-P , Packham M A , Harfenist E J
Published: July 2018

The mechanisms involved in platelet deaggregation are unclear. Washed platelets from rabbits or humans aggregated by ADP can be deaggregated by EDTA or PGI2 if the release reaction has not occurred; during deaggregation 125I-fibrinogen dissociates from the platelets. Human platelets suspended in a medium without calcium undergo the release reaction during ADP-induced aggregation; EDTA, PGE, or PGI2 do not deaggregate these platelets although EDTA displaces much of the 125I-fibrinogen that associates with them during aggregation. Rabbit platelets aggregated by low concentrations of releaseinducing stimuli (sodium arachidonate, collagen or thrombin) can be deaggregated by EDTA, PGI2 or PGE1 and 125I-fibrinogen dissociates from them; with high concentrations of collagen or thrombin, deaggregation and dissociation of l25I-fibrinogen is slower. Human platelets that have undergone the release reaction in response to thrombin, collagen or a combination of sodium arachidonate and ADP are not readily deaggregated by EDTA or PGE1. Since aggregation and fibrinogen binding involving the glycoprotein IIb/IIIa complex are readily reversed by EDTA, and since Ca2+ is required for thrombospondin binding to activated platelets, there may be a third type of platelet-platelet adherence that is not disrupted by EDTA; this type of binding plays a greater role with human than with rabbit platelets.
Hemarthroses and Warfarin: Joint Destruction with Anticoagulation
Authors: Andes W Abe , Edmunds J Ollie
Published: July 2018

A 56 year old man was seen because of recurrent right shoulder hemarthroses and a superior vena caval syndrome. Intermittent symptoms had occurred for six years and resulted in marked destructive changes in the right shoulder. Arthroscopy added pathological evidence of this unusual mechanism of arthropathy. Patients on anticoagulants deserve close attention to symptoms of occult bleeding.
Lipid Peroxidation and Experimental Disseminated Intravascular Coagulation in Rats Induced by Endotoxin
Authors: Yoshikawa T , Murakami M , Furukawa Y , Kato H , Takemura S , Kondo M
Published: July 2018

The experimental model of disseminated intravascular coagulation (DIC) in rats induced by a 4 hr sustained infusion of 100 mg/kg endotoxin was used to study the relationship between lipid peroxidation and DIC. Serum thiobarbituric acid (TBA) reactive substances, important and damaging products of lipid peroxidation, increased gradually during the infusion of endotoxin. The levels of TBA reactants in serum, abdominal aortic wall and ileum mucosa were significantly increased in rats infused with 100 mg/kg of endotoxin for 4 hr, as compared with the control rats continuously given 11.4 ml of physiological saline for 4 hr. However, the levels in liver and kidney tissues, and in gastric, jejunum and colon mucosa did not increase significantly.
Thrombin Induces Thromboplastin Synthesis in Cultured Vascular Endothelial Cells
Authors: Galdal K S , Lyberg T , Evensen S A , Nilsen E , Prydz H
Published: July 2018

Cultured human umbilical vein endothelial cells responded to thrombin (10−2 – 10 NIH u/ml) with a 2-5 fold increase in thromboplastin activity. The maximum response was reached after 4 hr in serum-free medium. The effect of thrombin was fully inhibited by the presence of 50% (v/v) fetal calf serum or more in the medium, by preincubation of thrombin with hirudin or by treatment of thrombin with N-bromosuccinimide or phenylmethylsulfonyl fluoride. The thrombin-induced thromboplastin activity was inhibited by incubation of the cells with cycloheximide (2 μg/ml) or actinomycin D (2 μg/ml) showing that the response depended on de novo protein and RNA synthesis. It was also suppressed by exposure of the cells to two different phosphodiesterase inhibitors, 3-butyl-l-methyl-xanthine (5 · 10−4 M) and rac-4 (3-butoxy-4-methoxybenzyl)-2-imidazole (5 · 10−4 M), to the transmethylation inhibitors 3-deazaadenosine (10−5 M) and 1-homocysteine thiolactone (2 · 10−5 M) in combination and to the intracellular calcium antagonist 8-(N,N-diethylamino)-octyl 3,4,5,-tri-methoxybenzoate hydrochloride (8 · 10−5 M). Our results suggest that small amounts of thrombin can induce thromboplastin synthesis in endothelial cells in vitro and that this synthesis probably is regulated by the intracellular level of cAMP, by cytoplasmic Ca2+ and possibly also by transmethylation reactions.
A Multivariate Pattern Recognition Study of Risk-Factors Indicating Postoperative Thromboembolism Despite Low-Dose Heparin in Major Abdominal Surgery
Authors: Kjærgaard J , Esbensen K , Wille-Jørgensen P , Jørgensen T , Thorup J , Berning H , Wold S
Published: July 2018

The object of the present investigation was to identify those who, among high-risk patients, would “break through” low-dose heparin prophylaxis and develop thromboembolism after major abdominal surgery.

Twenty-nine variables (clinical characteristics, pre- and postoperative coagulation and fibrinolytic factors) from 19 patients with and 26 patients without thromboembolism were analyzed by means of a multivariate supervised pattern recognition technique (SIMCA).

We found no statistically significant difference between patients with and without thromboembolism. Thus, in the studied group of high-risk patients it was not possible to identify a predictive index for selection of individual patients liable to develop postoperative thromboembolism despite low-dose heparin prophylaxis in major abdominal surgery.
Plasminogen Activation in Diabetes Mellitus: Normalization of Blood Sugar Levels Improves Impaired Enzyme Kinetics In Vitro
Authors: Geiger Margarethe , Binder Bernd R
Published: July 2018

We have demonstrated previously that fibrin enhanced plasmin formation by the vascular plasminogen activator was significantly impaired, when components isolated from the plasma of three uncontrolled diabetic patients (type I) were used to study plasminogen activation in vitro. In the present study it can be demonstrated that functional properties of the vascular plasminogen activators as well as of the plasminogens from the same three diabetic patients are significantly improved after normalization of blood sugar levels and improvement of HbAlc values. Most pronounced the Km of diabetic vascular plasminogen activator in the presence of fibrin returned to normal values, and for diabetic plasminogen the prolonged lag period until maximal plasmin formation occurred was shortened to almost control values. From these data we conclude that the observed abnormalities of in vitro fibrinolysis are not primarily associated with the diabetic disease, but might be secondary to metabolic disorders caused by diabetes.
Dietary Supplementation with Vitamin E in Hyperlipoproteinemias: Effects on Plasma Lipid Peroxides, Antioxidant Activity, Prostacyclin Generation and Platelet Aggregability
Authors: Szczeklik A , Gryglewski R J , Domagala B , Dworski R , Basista M
Published: July 2018

In a placebo-controlled trial healthy volunteers and patients with hyperlipoproteinemias types II and IV received orally vitamin E at doses of 300 mg and 600 mg daily for 2 weeks. Serum tocopherol levels increased two-fold, but serum concentrations of total lipids, cholesterol, triglycerides, ceruloplasmin and transferrin remained unchanged. Dietary supplementation with vitamin E suppressed elevated concentrations of plasma lipid peroxides and this effect was correlated with an increase in serum antioxidant activity. In patients a mild platelet suppressant effect of vitamin E (600 mg daily) was observed.

Feeding an atherogenic diet to rabbits for a week resulted in elevation of plasma lipid peroxides and a 90% decrease in arterial generation of prostacyclin. Enrichment of the atherogenic diet with 100 mg vitamin E daily prevented the increase in plasma lipid peroxides and protected the prostacyclin generating system in arteries. Thus, in hyperlipoproteinemias vitamin E corrects certain abnormalities of lipid metabolism which might predispose to atherosclerosis.
Protein C Levels in Disseminated Intravascular Coagulation and Thrombotic Thrombocytopenic Purpura: Its Correlation with Other Coagulation Parameters
Authors: Takahashi Hoyu , Takakuwa Etsuko , Yoshino Noriko , Hanano Masaharu , Shibata Akira
Published: July 2018

Protein C was measured by means of enzyme-linked immunosorbent assay (ELISA) in plasmas from 58 normal subjects, 39 patients with disseminated intravascular coagulation (DIC) and 5 patients with thrombotic thrombocytopenic purpura (TTP). Protein C levels ranged from 69.7 to 163.6% (95% confidence limits) in normal subjects. In patients with DIC, protein C concentrations were significantly decreased, with a geometric mean value of 42.1%. Protein C concentration was positively correlated with plasma prothrombin, antithrombin III and serum pseudocholinesterase, and was negatively correlated with von Willebrand factor antigen (vWF: Ag) and vWF: Ag/factor VIII ratio. These findings suggest that low protein C concentrations in DIC mean a consumption of protein C probably due to its activation by thrombin and/or impaired liver synthetic function. In patients with TTP, protein C levels were normal with a geometric mean value of 116.7%, indicating that the pathophysiology of TTP is quite different from that of DIC.
Effects of Adrenaline Infusion on Platelet Number, Volume and Release Reaction
Authors: Lande K , Gjesdal K , Fønstelien E , Kjeldsen S E , Eide I
Published: July 2018

At the end of a diagnostic right heart catheterization ten patients received an intravenous infusion of l-adrenaline which gradually increased the arterial plasma adrenaline concentration from resting physiological values to high values as seen during myocardial infarction, pheochromocytoma and hypoglycemia. Blood was sampled from the brachial artery, femoral vein and hepatic vein. During the adrenaline infusion venous beta-thromboglobulin concentrations increased 23% from 61 ± 5 to 80 ± 7 μg/l (mean ± SE), arterial platelet counts 20% from 212 ± 17 to 253 ± 25 × 109/l and arterial platelet volume 4% from 7.25 ± 0.20 to 7.56 ± 0.21 femtoliter. All changes were significant at the 5% level. Thus, acute increments of arterial plasma adrenaline significantly stimulated the blood platelet parameters studied.
Epidemiological Study on Factor VII, Factor VIII and Fibrinogen in an Industrial Population: I. Baseline Data on the Relation to Age, Gender, Body-Weight, Smoking, Alcohol, Pill-Using, and Menopause
Authors: Balleisen L , Bailey J , Epping P-H , Schulte H , van de Loo J
Published: July 2018

The Münster Arteriosclerosis Study (MAS) is a prospective, longitudinal epidemiological study on an industrial population in Westfalia aimed to establishing clinical and laboratory data with possible relationship to cardiovascular events. The data presented here describe the baseline measurements of fibrinogen, factor VIIc and factor VIIIc from the recruitment of 2880 male and 1306 female persons and their relationship to age, gender, body-weight, smoking, alcohol, pill-using and menopause. The correlations were made by means of a multiple regression analysis. We found an increase of those coagulation factors with age, a correlation of F VII and fibrinogen with body-weight index and of fibrinogen with cigarette smoking. No correlation was found for alcohol consumption. F VIII and F VII were significantly higher after onset of menopause and F VII and fibrinogen in women using the pill.
Evening Primrose Oil (Efamol) in the Treatment of Raynaud’s Phenomenon: A Double Blind Study
Authors: Belch J J F , Shaw B , O’Dowd A , Saniabadi A , Leiberman P , Sturrock R D , Forbes C D
Published: July 2018

Prostaglandin E1 (PGE1) and prostacyclin have been used in Raynaud’s phenomenon (RP) but are unstable and require intravenous administration. An alternative approach is to stimulate the body’s own PGE1 production via administration of the precursor essential fatty acid. We studied the effect of 12 capsules/day of evening primrose oil (EPO) on the manifestations of RP. 21 patients received a two week course of placebo, thereafter 11 received EPO for 8 weeks and 10 patients received placebo. As the weather worsened the placebo group experienced significantly more attacks than the EPO group. Visual analogue scales assessing the severity of attacks and coldness of hands improved in the EPO group. No changes were seen in either group in hand temperatures and cold challenge plethysmography. Blood tests showed some antiplatelet effects of the drug. In conclusion patients receiving EPO benefited symptomatically. This was not matched however by any change in objective assessment of blood flow, although changes in platelet behaviour and blood prostanoids were observed.
Immunodetection of Human Fibrin Using Monoclonal Antibody-64C5 in an Extracorporeal Chicken Model
Authors: Hui Kwan Y , Haber Edgar , Matsueda Gary R
Published: July 2018

The potential use of a fibrin-specific monoclonal antibody-64C5 as a thrombus detecting agent was examined in a quantifiable extracorporeal circulatory model using the chicken. This species was selected because antibody-64C5 did not cross-react with chicken fibrin. Human fibrinogen was clotted with human thrombin and factor XIII on filter paper disks, which were then inserted into an ex-vivo chamber that interrupted an extracorporeal loop between a carotid artery and a jugular vein. Antibody-64C5 was injected intravenously and after an equilibration period, disks bearing human fibrin were placed in the extracorporeal chamber and exposed to circulating antibody. Uptake which was quantified by treating the disks with 125I-goat antimouse-(Fab’)2 was rapid, with 70 percent of maximal uptake occurring within 10 min. Fibrinolysis, although measurable, had no adverse effect upon uptake of the monoclonal antifibrin-64C5.
Plasma Concentrations of Platelet-Specific Proteins in Different Stages of Essential Hypertension: Interactions between Platelet Aggregation, Blood Lipids and Age
Authors: Yamanishi J , Sano H , Saito K , Furuta Y , Fukuzaki H
Published: July 2018

Plasma β-thromboglobulin (βTG) and platelet factor 4 (PF4) were significantly higher in a group of 116 hypertensive men than in a normotensive group of 142 men. They increased with the stage of hypertension but the level did not correlate with the age of the subjects. Platelet aggregation was similar in the two groups and positively correlated with the age of the subjects in the normotensive group but not in the hypertensive group. A strong positive correlation was observed between the levels of plasma βTG and PF4 and between platelet aggregation to ADP and that to epinephrine in both the hypertensive and normotensive groups. However, there was no correlation between the level of plasma βTG or PF4 and platelet aggregation. Plasma antithrombin III was lower in the hypertensive group than in the normotensive group.

These studies suggest that plasma levels of PTG and PF4 are closely related to the stage of hypertension and are better indicators than aggregation of in vivo platelet activation in hypertensive subjects. Enhanced platelet activation may be involved in the acceleration of hypertensive arteriovascular damage and atherosclerosis.
Deaggregation of Human Platelets Aggregated by Thrombin
Authors: Kinlough-Rathbone R L , Perry D W , Packham M A , Mustard J F
Published: July 2018

Human platelets that have undergone the release reaction do not deaggregate readily. We examined conditions under which washed human platelets can be deaggregated after they have undergone an extensive release reaction induced by thrombin (1 or 5 U/ml). To make fibrinogen receptors unavailable, either CP/ CPK (or apyrase) was used to remove released ADP, or PGEi was used to increase cAMP. Chymotrypsin was used to digest proteins that might link platelets, and heparin to interact with released proteins and interfere with their binding to platelets and to each other. Individually, none of these caused deaggregation; heparin did not inhibit the effect of thrombin because no antithrombin III was present. Platelets exposed to thrombin (1 U/ ml) which was neutralized at 90 sec by hirudin, could be deaggregated by combinations of CP/CPK (or apyrase) and chymotrypsin, or PGE1 and chymotrypsin. When a higher concentration of thrombin was used (5 U/ml) these combinations caused platelets to deaggregate only when heparin was added before thrombin induced the release reaction. Thus, when extensive release occurs three mechanisms may come into play to link human platelets: one that requires the fibrinogen receptor; a heparin-sensitive reaction that may involve the binding of released proteins; and a linkage that can be disrupted only by proteolysis, providing the other two mechanisms are also inhibited.
Standardization of β-Thromboglobulin (β-TG) and Platelet Factor 4 (PF4): A Collaborative Study to Establish International Standards for β-TG and PF4
Authors: Kerry P J , Curtis A D
Published: July 2018

An international collaborative study was carried out to determine the suitability of freeze-dried preparations of β-TG and PF4 to serve as international standards, and to compare these materials with other purified preparations and with plasma samples. Although problems remain with the accurate measurement of these proteins, it has been demonstrated that common standards improve the precision of measurement by RIA and provide an essential foundation for future work into the effects of assay system differences.

The World Health Organization established in 1984 the purified preparation of β-TG (83/501) and the purified preparation of PF4 (83/505) as International Standards, with assigned potencies of 500 International Units per ampoule and 400 International Units per ampoule, respectively.
Protamine-Induced Thrombocytopenia and Leukopenia
Authors: Al-Mondhiry Hamid , Pierce Williams S , Basarab Robert M
Published: July 2018

Protamine has been reported to cause thrombocytopenia and granulocytopenia. In this article, we report studies examining the relative contribution of protamine and heparin in the pathogenesis of this phenomenon, the dose-effect relationship, and the possible mechanism of cell loss. Protamine alone infused into experimental animals causes mild, transient granulocytopenia and thrombocytopenia. The sequential administration of heparin and protamine results in a more severe cytopenia lasting 30 to over 60 min. Organ scanning with 111In-labeled platelets shows a striking though transient accumulation of radioactivity in the lungs following heparin-protamine infusion. Platelet survival, however, is not shortened. Incubation of 125I-labeled protamine with blood cells in the presence of heparin results in tight binding of the drug to platelets and granulocytes. These observations suggest that protamine and heparin form a complex that binds to blood cells. The sequestration of coated cells in the lungs results in transient granulocytopenia and thrombocytopenia.
Neutralisation of Heparan Sulphate and Low Molecular Weight Heparin by Protamine
Authors: Hubbard A R , Jennings C A
Published: July 2018

The neutralisation by protamine sulphate (PS) of heparan sulphate (HS), a low molecular weight heparin (LMWH), and a reference preparation of unfractionated heparin (UH), was studied by activated partial thromboplastin time (APTT) and anti-Xa clotting assays. UH was most easily neutralised in the APTT assay by PS (on a weight for weight basis), followed by LMWH and HS. The neutralisation of APTT activity by PS closely followed the loss of activity in the anti-Xa clotting assay, when plasma was used as the source of At III. When the anti-Xa clotting assay was carried out using purified At III in place of plasma, HS and LMWH were neutralised by much lower amounts of PS and resembled UH neutralisation more closely. Resistance of HS anti-Xa activity to PS neutralisation decreased with increasing plasma dilution. The presence of bovine albumin with purified At III concentrate increased the resistance of HS to PS neutralisation. It is concluded that PS binding to UH, HS and LMWH is probably related more to their degree of sulphation than molecular weight and that non-specific interactions between PS and plasma proteins inhibit the binding of PS to HS and LMWH.
PAF-Acether May Not Mediate the Third Pathway of Platelet Aggregation since Self-Desensitization Reduces the Effects of Low Thrombin but Enhances Those of Convulxin
Authors: Wal Françoise , Joseph Danielle , Touqui Lhousseine , Vargaftig B Boris
Published: July 2018

PAF-acether (platelet-activating factor) was hypothesized as the mediator of the ADP and thromboxane-independent activation of platelets induced by thrombin (Thr) and by the snake venom glycoprotein convulxin (Cx). Aspirinized rabbit platelets self-desensitized to PAF-acether were less responsive to low amounts of Thr, as expected if PAF-acether would be formed, but were hyper-reactive to Cx, in contradiction with its hypothesized mediating role. Aggregation by higher concentrations of Thr overcame inhibition. Experiments with ADP-depleted platelets showed that secretion is neither involved with desensitization to PAF-acether nor with hyper-reactivity to Cx. Those effects required the presence of PAF-acether in the platelet suspension and persisted when transformation of PAF-acether into its recognized metabolite alkyl-acyl-glycerophosphorylcholine was inhibited. The ADP and thromboxane-independent activation of rabbit platelets by low and medium concentrations of Thr may be accounted for by platelet formation of PAF-acether, but overall the contrasting effects of platelet desensitization to PAF-acether on responsiveness to Thr and to Cx suggest that the third pathway of aggregation requires other explanations.
Enhanced Platelet Release Reaction, Shortened Platelet Survival Time and Increased Platelet Aggregation and Plasma Thromboxane B2 in Chronic Obstructive Arterial Disease
Authors: Zahavi J , Zahavi M
Published: July 2018

Plasma β-thromboglobulin (βTG) and thromboxane B2 (TXB2) level, platelet aggregation (PA) in platelet rich plasma, platelet survival time using luIndium radiolabelled platelets and platelet sensitivity to prostacyclin (PGI2) were measured in chronic obstructive arterial disease (COAD) patients. Severity of the disease was assessed by the ankle pressure index using Doppler auscultation.

Platelet survival time was shorter, plasma βTG and TXB2 and the rate and extent of PA induced by ADP or 1-epinephrine (but not collagen) were greater in the patients than in controls. βTG was inversly correlated with the pressure index and positively with TXB2 indicating increased platelet TXA2 synthesis. Platelet sensitivity to PGI2 was similar in the patients and controls.

These results indicate increased platelet consumption and enhanced in vivo platelet activation and PA in COAD. The enhanced activation correlates with the severity of the disease and the activated platelets presumably synthesize increased amounts of TXA2. It is therefore concluded that platelets might be involved in the pathogenesis of COAD. The normal platelet sensitivity to PGI2 suggests that the administration of the compound may improve the abnormal platelet function in COAD patients and may attenuate the progression of the disease.
Release of Plasminogen Activator from Isolated Perfused Dog Heart
Authors: Matsubara Fujio , Sueishi Katsuo , Ishii Yutaka , Tanaka Kenzo
Published: July 2018

The release and the local activity of plasminogen activator (PA) were studied in isolated perfused dog hearts, without or with intimal injury induced by means of a balloon catheter inserted into the left anterior descending coronary artery (LAD). Thrombin but not DFP-thrombin induced a dose-dependent PA release in doses of 8 to 32 units. ADP 20 or 200 μmol but not ergonovine 20 or 200 μg induced a weak PA release. The local PA activity was much lower in the LAD at 1 or 4 weeks after this injury than in the intact LAD. However, the release of PA from the hearts after intimal injury was similar to findings in the intact hearts.

We conclude from this study that thrombin plays an important role in regulating the coagulation-fibrinolysis system in endothelial cells and that changes in the properties of the endothelial cells may lead to initiation and enhancement of atherosclerosis.
A Collaborative Study of a Proposed International Standard for Tissue Plasminogen Activator (t-PA)
Authors: Gaffney P J , Curtis A D
Published: July 2018

An international collaborative study involving seven laboratories was undertaken to assess which of three lyophilised preparations might serve as an International Standard (I.S.) for tissue plasminogen activator (t-PA). Two of the preparations were isolates from human melanoma cell cultures while one was of pig heart origin. A clot lysis assay was used by all participants in the study.

The data suggested that both preparations of human cell origin were comparable, in that their log dose-response lines were parallel, while that of the porcine preparation was not. Accelerated degradation studies indicated that one melanoma extract (denoted 83/517) was more stable than the other and it was decided to recommend preparation 83/517 as the standard for t-PA. The International Committee for Thrombosis and Haemostasis (Stockholm 1983) has recommended the use of this material as a standard and it has been established by the Expert Committee on Biological Standardization of the World Health Organization as the International, Standard for tissue plasminogen activator, with an assigned potency of 1000 International Units per ampoule.
Relationship Between Thromboembolic Complications and Intensity of Treatment During Long-Term Prophylaxis with Oral Anticoagulants Following DVT
Authors: Schulman Sam , Lockner Dieter
Published: July 2018

The frequency of thromboembolic recurrencies during secondary prophylaxis after DVT was retrospectively studied and related to the intensity of the oral anticoagulation. All patients receiving oral anticoagulation after DVT at our hospital during April 1972 - May 1980 were studied. Treatment was given to 596 patients for 724 thrombotic events for a total of 4450 months. Thirty-six thromboembolic complications, all objectively verified, occurred. Patients with cancer had complications throughout the entire range of anticoagulation. Patients without neoplastic disease (15 events) never had complications below a prothrombin complex level of 27% as assessed with Simplastin A, corresponding to a BCT-ratio of 1.9. This study confirms, that the lower limit of the therapeutic range, determined by the risk of thromboembolic complications, should be set at a Simplastin A-level of approx. 25% corresponding to BCT 2.0.
Vascular Smooth Muscle Cells Inhibit the Plasminogen Activators Secreted by Endothelial Cells
Authors: Laug Walter E
Published: July 2018

TPure cultures of bovine endothelial cells (EC) produce and secrete large amounts of plasminogen activators (PA). Cocultivation of EC with vascular smooth muscle cells (SMC) resulted in a significant decrease of PA activities secreted by the EC, whereas the cellular PA activities remained unaffected. Secreted PA activities were absent in the growth medium as long as the SMC to EC ratio was 2:1 or higher. The PA inhibitory activity of the SMC was rapid and cell-to-cell contact was not necessary.

The PA inhibitory activity was present in homogenates of SMC as well as in the medium conditioned by them but not in the extracellular matrix elaborated by these cells. Serum free medium conditioned by SMC neutralized both tissue type (t-PA) and urokinase like (u-PA) plasminogen activators. Gel electrophoretic analysis of SMC conditioned medium followed by reverse fibrin autography demonstrated PA inhibitory activities in the molecular weight (Mr) range of 50,000 to 52,000 similar to those present in media conditioned by bovine endothelial cells or fibroblasts. Regular fibrin zymography of SMC conditioned medium incubated with u-PA or t-PA revealed the presence of a component with a calculated approximate Mr of 45,000 to 50,000 which formed SDS resistant complexes with both types of PA.

These data demonstrate that vascular SMC produce and secrete (a) inhibitor(s) of PAs which may influence the fibrinolytic potential of EC.
Characterization of Two Monoclonal Antibodies Against Human Tissue-Type Plasminogen Activator
Authors: Schleef Raymond R , Sinha Manjula , Loskutoff David J
Published: July 2018

A series of hybridoma clones, each producing monoclonal antibodies to human tissue-type plasminogen activator (t-PA), were prepared from mice by standard procedures. Two of these clones were selected for further study. One HI72A1, produced antibodies that bound to t-PA and strongly inhibited its activity, whereas another, LI72D1, produced antibodies that bound to t-PA but did not affect its activity. The specificity of these antibodies was assessed in immunoabsorption experiments. Both immunoprecipitated 125I-labeled t-PA, and both were specific since only t-PA was recognized in conditioned media collected from Bowes melanoma cells cultured in the presence of 3H-leucine. Neither antibody recognized urokinase. t-PA was desorbed from antibody HI72A1-Sepharose columns with 0.5 M NaCl, consistent with its relatively low association constant (Ka = 9.37 × 107 M-1). In contrast, a strong chaotropic agent (i.e., 2 M KI) was required to elute t-PA from antibody LI72D1 columns (Ka = 2.08 × 109 M-1). This latter high affinity antibody was employed to develop an immunoradiometric assay for t-PA having a sensitivity of 0.5 ng/ml.
HTLV-I Antibody Status in Hemophilia Patients Treated with Factor Concentrates Prepared from U.S. Plasma Sources and in Hemophilia Patients with AIDS
Authors: Chorba T L , Jason J M , Ramsey R B , Lechner K , Pabinger-Fasching I , Kalyanaraman V S , McDougal J S , Cabradilla C D , Tregillus L C , Lawrence D N , Evatt B L
Published: July 2018

Serum samples from 85 Austrian hemophilia patients treated with lyophilized factor concentrates prepared from U.S. plasma sources, 24 hemophilia patients from Georgia on a home therapy program with factor concentrates, and 10 U.S. hemophilia patients with acquired immunodeficiency syndrome (AIDS) were analyzed by two different methods for the presence of antibodies to the major internal antigen of human T-cell leukemia virus I (HTLV-I) p24. All but one, a Georgia sample, were negative. The absence of antibody to HTLV-I p24 in the serum of European hemophilia patients, of U.S. hemophilia patients with no symptoms of AIDS, and of U.S. hemophilia patients with AIDS is interpreted as an indication of the lack of ready transmissibility of HTLV-I in lyophilized factor concentrates.
Agglutination of Platelet-Type von Willebrand’s Disease Platelets by Bovine von Willebrand Factor
Authors: Takahashi Hoyu , Shibata Akira
Published: July 2018

It has been shown that platelets from patients with platelet- type von Willebrand’s disease (vWD) agglutinate upon the addition of human von Willebrand factor (vWF) in the absence of ristocetin or botrocetin, suggesting that platelet membrane receptors for human vWF is abnormal. The present work reports the platelet agglutinability on stimulation with bovine vWF in platelet-type vWD. Platelets in patient platelet-rich plasma or washed platelet suspensions and patient platelets treated with formalin agglutinated in the presence of markedly lower concentrations of bovine vWF than those required for normal platelets. This finding provides additional evidence that platelet-type vWD platelets have abnormal expression of binding sites for vWF on their surface, and supports that platelet receptors for bovine vWF are identical or very close to those for human vWF.
Sources of Variability in Dose Response Platelet Aggregometry
Authors: Vickers M V , Thompson S G
Published: July 2018

An experiment designed to assess the components of variability of a number of measures of platelet aggregability showed that the ADP ED50 (the estimated dose of adenosine diphosphate at which primary aggregation occurs at half its maximum velocity) had the least method error of any of the parameters measured, but that none had a very high between-person component of variability. A simultaneous comparison of a syringe technique and a free-flowing technique for venepuncture revealed no differential effects on the aggregation parameters measured. An enforced increase in the stirring speed in the aggregometer led to an experiment which showed that such a change did not apparently affect the ED50s.
Plasma Beta-Thromboglobulin, Fibrinopeptide A and Bβ15–42 Antigen in Relation to Postoperative DVT, Malignancy and Stanozolol Treatment
Authors: Douglas Jessie T , Blamey S L , Lowe G D O , Carter D C , Forbes C D
Published: July 2018

Plasma levels of betathromboglobulin (BTG), fibrinopeptide A (FPA) and Bβ15–42 fragment, indices of platelet release, thrombin generation and plasmin activity respectively, were measured in 32 high risk patients during a double blind study of a single dose of the anabolic steroid stanozolol (50 mg IM) in the prevention of DVT after major gastro-intestinal surgery. The prevalence of malignancy and the incidence of DVT (125I fibrinogen scan) were similar in the two treatment groups. On the first postoperative day, BTG, FPA and Bβ15–42 levels were increased in most patients. Plasma BTG levels were significantly increased on the first post-operative day in patients who developed a DVT (n = 14) compared to those patients who did not (n = 18). A significant increase in FPA levels was found in the DVT group, 7 days after surgery. On the morning before surgery, plasma Bβ15–42 levels were significantly increased in patients who developed a DVT. In patients undergoing surgery for early malignancy (n = 17), we observed a pre-operative increase in FPA levels when compared to patients without malignancy. At post-operative day 7, Bβ15–42 levels were significantly increased in patients who received stanozolol (n = 15), when compared to the placebo group, suggesting that intramuscular stanozolol increases fibrinolysis in vivo.
Severe Protein C Deficiency in Congenital Thrombotic Disease – Description of an Immunoenzymological Assay for Protein C Determination
Authors: Soria J , Soria C , Samama M , Nicolas G , Kisiel W
Published: July 2018

An immunoenzymatic assay (ELISA) is described for the quantitative assay of protein C in plasma. This technique allows a safe, reliable, and sensitive assay of protein C, and is easily used for routine investigation. Using this technique, a protein C deficiency (0.16 U/ml) was discovered in a 16 year old patient with a history of very severe thrombotic disease. Protein C deficiency was also discovered in his mother (0.62 U/ml) and father (0.50 U/ml). We therefore suggest that this case could represent a homozygous deficiency of protein C.
Unreliability of Current Serum Fibrin Degradation Product (FDP) Assays
Authors: Gaffney P J , Perry M J
Published: July 2018

Previously, assays of fibrin-fibrinogen degradation products (FDP) had to be performed on serum samples. However, monoclonal antibodies (Mabs) are now available which permit the measurement of FDP directly in plasma. We have employed two Mabs, one monospecific for FDP originating from crosslinked fibrin and another panspecific for the FDP fraction, to determine normal FDP levels in plasma and serum. The monospecific Mab gave a value of 40 ng FDP/ml in plasma and 10 ng/ml in serum, while the serum level of FDP recorded using the panspecific Mab was >1000 ng/ml, at all the concentrations of thrombin employed. Similarly, when a solution of purified fibrinogen was treated with thrombin, the concentration of FDP present in the clot supernatant was >1000 ng/ml when assayed using the panspecific Mab. Thus during serum preparation as much as 75% of the native FDP is incorporated into the clot while in excess of 1000 ng/ml of laboratory generated FDP, probably incompletely polymerized fibrin, is measured using panspecific antisera. These data indicate that current FDP assays using polyclonal antibodies are not a reliable reflection of the FDP level generated in vivo. The use of FDP-specific Mabs which do not react with fibrinogen is recommended for future FDP assays performed directly on plasma.