bluuebunny/crossref_metadata_2025 huggingface dataset:
A Randomized Comparative Study on Early Prophylactic Treatment of Thrombosis Using Heparin and High-Molecular Dextran Following Injuries of the Lower-Leg
Authors: Gaudernak T. , Kuderna H. , Olbert F. , Pelinka H. , Schlag G.
Published: June 2019

A randomized study was carried out on 50 patient suffering from lesions in the lower segment of the leg (fractures and ruptures of Achilles tendon). The patients were divided into two groups treated with Heparin (3x5000 i.u./die) and Dextran - 70 500 ml/die. respectively.

The incidence of thrombosis was as high as 44%. In all cases, however, it remained limited to the veins of the tibial region. No significant difference was found between Heparin and Dextran patients.

Control by phlebography carried out after one year in 38 patients indicated rechannelling of the affected veins in all patients. It also showed deep lesions of the veins in the form of a post-thrombotic syndrom (II/2) in three patients.

This leads to the conclusion that immediate administration of Heparin or Dextran — 70 is strongly to be recommended for patients suffering from a recent fracture in the tibial region as a prophylactic measure against thrombosis. It greatly reduces the incidence of pulmonary emboli or of the frequent post-thrombotic syndrome.
Effect of Contraceptive Hormonal Steroids on the Cardiovascular System of Dogs: Scanning Electron Microscopic Studies of Arteries and Veins
Authors: Stanczewski B. , Sawyer P. N. , Kammlott G. W.
Published: June 2019

The effects of contraceptive hormonal steroids on the canine cardiovascular system were evaluated. The changes in morphology of arteries and veins and the nature of thrombi deposited following injury to blood vessels were ascertained by scanning electron microscopy (SEM). Twenty-five female dogs were used for the study. The following estrogenic and progestational agents were used at 10 times the human dose, i) mestranol (M); ii) norethindrone (N); iii) M+N; iv) medroxy progesterone (depo-provera); v) ethynyl estradiol; vi) norgestrel, and vii) controls. The dogs were fed orally for a period of 6 to 18 weeks. During this period the physico-chemical effects of the drugs on platelets, red cells and blood coagulation were determined. On the date of sacrifice the dogs were anaesthetized and femoral and carotid arteries as well as femoral and jugular veins were exposed. In each animal one artery and vein were left intact as internal control and the others were injured by hemostatic crush, and Fogarty balloon catheterization. After injury, the vessels were excised and perfused with 10% formalin for SEM and light microscopic studies. Visible differences in deposition and organization of thrombi could be observed in arteries and veins of several groups of animals. Thus in the animals fed with mestranol one could see several blood cells attached to the surface even under ‘normal’ (no injury) conditions. Vessels from dogs fed norethindrone, under similar conditions, show deposits of fibrin and platelets. The combination of mestranol and norethindrone reveals more well organized fibrin and cellular deposits attached to the vascular endothelium. The cummulative SEM results indicate that these hormonal steroids appear to increase the cell deposition on the intimal surfaces.
Activation of Endotoxin-Induced Intravascular Coagulation in Congenitally C6-Deflcient Rabbits
Authors: Lohmann Elke , Müller-Berghaus G.
Published: June 2019

Activation of endotoxin-induced coagulation is thought to be caused by platelet factor 3. Since the release of platelet factor 3 after mixing platelets and endotoxin is dependent on the presence of complement factors, studies were performed in rabbits congenitally deficient in the sixth component of complement (C6). Endotoxin infusion (40 μg/kg × hr) caused a decrease in platelet counts in C6-deficient as well as in normal rabbits. Furthermore, after endotoxin infusion activation of intravascular coagulation occurred in C6-deficient rabbits to the same degree as in normal rabbits : prothrombin time and partial thromboplastin time were prolonged, factor V- and factor VII-activities were decreased, and ethanol gelation tests revealed positive results. Nine out of 10 C6-deficient rabbits demonstrated fibrin-rich microdots in the glomerular capillaries. Since platelet factor 3 is not released by endotoxin in the C6-deficient rabbits, these experiments make it likely that the release of platelet factor 3 does not represent the trigger mechanism of endotoxin-induced intra-vascular coagulation.

(Supported by the Deutsche Forschungsgemeinschaft, Bad Godesberg, Germany.)
Factor VIII, Small Active Factor VIII Fragments, and von Willebrand Factor
Authors: Wagner R. H.
Published: June 2019

Factor VIII activity of human, canine, and bovine preparations is associated with a molecule of high molecular weight. These preparations dissociate almost completely in 0.25 M Ca2+, yielding a small active fragment with factor VIII activity, and a high molecular weight fraction (carrier protein) with almost no factor VIII activity. With Ca2+ absent, the small active fragment recombines with the carrier protein to form a large protein with factor VIII activity. Mixtures of normal human small active fragment with human hemophilic carrier protein give normal recombination. Comparable experiments with human von Willebrand and canine “carrier protein fractions” show no recombination. The data suggest a working model of a small active fragment(s) linked by noncovalent bonds to a carrier molecule, comprised of repeating subunits, deficient in von Willebrand’s disease.

Thrombin increases the activity of factor VIII preparations or the small active fragment obtained by Ca2+ dissociation. When a factor VIII preparation is treated with thrombin and subjected to gel chromatography or ultracentrifugation, the factor VIII activity appears to be associated with a small molecule. Factor VIII activity of this small molecule is not enhanced by thrombin. These and other data will be discussed.
Mechanism of the Prolonged Bleeding Time in yon Willebrand’s Disease (vWd)
Authors: Mannucci P. M. , Holmberg L. , Ruggeri Z. M. , Nilsson I. M.
Published: June 2019

Factor-VIII antigen was characterized by means of an immunofluorescence technique in biopsy specimens obtained at dental extractions from the buccal mucosa of ten patients with vWd. No activity could be elicited in five patients with severe recessive vWd, whereas venules and arterioles stained positively in five patients with a moderate form characterized by dominant pattern of inheritance. In six patients, immunofluorescence was also carried out in specimens obtained after cryoprecipitate infusion at a time when the plasma defects (VIIIAHF, VIIIAGN and VIIIVWF) were corrected but the long bleeding time was not modified. No activity was detectable on the vessel wall of those patients who were negative before the infusion. It is suggested that the defect of primary hemostasis in patients with vWd is related to the vascular contents of factor-VIII related properties rather than to their plasmatic levels.

Supported by grants from the Fondazione Angelo Bianchi Bonomi, the Swedish Research Council and the Faculty of Medicine, University of Lund.
The Histochemical Assay of Plasminogen Activator in the Venous Wall - An Inter-Ethnic Study
Authors: Franz R. B. , Hugo N. , Jansen C. R.
Published: June 2019

Despite intensive research in the field of blood coagulation and fibrinolysis facilitated by several studies emanating from this continent, the results still afford no satisfactory explanation for the well established observation of the relative rarity of clinically manifest venous thrombo-embolic disease in the Black African.

Pandolfi’s modification of the “fibrinolytic autograph” method of Todd was selected to assess the histochemical localization and the degree of activator activity in the venous wall of the upper and lower limbs in a White control group (18) and in a group of 20 Black patients.

Both ethnic groups showed greater fibrinolytic activity in the veins of the upper limb - a difference which appeared to satisfy the test of actuarial cross examination (P < 0,00001). Of particular interest was the significantly greater fibrinolytic activity in the lower limb of the African patients as compared with the lower limb veins of the White group (P < 0,05).

The increased fibrinolytic activity in the lower limb veins in Black Africans may be an additional anti-thrombogenic component in the haemostatic profile of this particular ethnic group.
Studies on Circulating Plasminogen Activator
Authors: Ogston D. , Bennett B. , Mackie M.
Published: June 2019

A plasminogen activator was prepared in crude form from postvenous occlusion plasma by gel filtration of the plasma on Sephadex G-200 in 0.005 M phosphate buffer (pH 7.5) containing 0.15 M NaCl followed by further gel filtration of the fractions with maximal activator activity on Sephadex G-200 in 0.005 M phosphate buffer containing 1 M NaCl. The preparations were free of plasminogen, α1-antitrypsin, α2-macroglobulin, CI inactivator and antithrombin III as assessed by immunological techniques. The activity of the partially purified plasma activator preparations was markedly more stable at 37° C than the original plasma, was stable at 4° C for several days, and withstood heating at 56° C for 1 hour. The activity was inhibited by exposure to phenylmethyl sulphonyl-fluoride, but not by tosyl-L-lysine chloromethyl ketone or iodoacetamide.
Detection of Carriers of Hemophilia by Hemorrhagometry
Authors: Jesdinsky-Buscher J. , Sutor A. H. , Niederhoff H.
Published: June 2019

Hemorrhagometry, a new in-vivo method, measures bleeding time, intensity and total blood loss from a small standardized wound (Sutor et al.: Blut 22, 27, 1971). In patients with hemophilia these values are within the normal range when the test is performed at room temperature (24° C). However, when the wound is cooled to 17° C (cold tolerance test), bleeding time is abnormally long in hemophiliacs. Therefore, we investigated this test in carriers of hemophilia. The cold tolerance test was performed in 16 ‘proven’ and 6 ‘probable’ carriers (criteria according to Nilsson). 14 proven and 4 probable carriers showed abnormal bleeding times of 15 min and more (2 proven carriers of hemophilia B did not differ markedly from 20 carriers of hemophilia A). When hemorrhagometry was performed at room temperature, the carriers could not be distinguished from normal control persons.

Of 15 sisters and aunts of hemophiliacs 9 had abnormal cold tolerance test findings, in fair agreement with the probability of .5 to be expected theoretically. Thus the hemorrhagometry cold tolerance test is helpful in detecting carriers of hemophilia.
Prevention of Venous Thrombosis in Neurosurgical Patients by Intermittent Calf Compression
Authors: Turpie A. G. G. , Beattie A. S. , Gallus A. S. , Hirsh J.
Published: June 2019

The risk of intracranial haemorrhage precludes the use of anticoagulant prophylaxis for venous thrombosis in neurosurgical patients. We have studied prospectively in this group, the effect of intermittent calf compression by inflatable plastic boots on the prevention of deep vein thrombosis detected by 125 Iodine-labelled fibrinogen leg scanning. Intermittent calf compression was continued for 5 days in the treatment group. In the control and treatment groups, leg scanning was continued until the patients were fully mobile or for a maximum of 14 days. In all patients during prophylaxis, deep vein thrombosis was detected in 12/62 (19.35%) patients in the untreated group and 1/60 (1.67%) patients in the prophylaxis group (P < 0.002). In patients who had craniotomy for brain tumour, subarachnoid haemorrhage, or subdural haeinatoma, leg scan detected deep vein thrombosis occurred during the 5 days of prophylaxis in 9/43 (20.93%) untreated and 0/43 (0%) treated patients (P < 0.002). At 14 days, deep vein thrombosis was detected in 13/43 (30.23%) untreated and 4/43 (9.3%) treated patients (P < 0,05). In η on-operated patients, leg scan detected deep vein thrombosis occurred in 3/19 (15.79%) untreated and in 1/17 (5.88%) treated patients (P = 0.34). The results indicate that intermittent calf compression is effective prophylaxis against leg scan detected deep vein thrombosis in neurosurgical patients. In post-operative patients the difference is highly significant during intermittent calf compression and the difference is maintained after prophylaxis is discontinued. In non-operated patients, although there is a trend, the difference in incidence of deep vein thrombosis between control and treated groups is not significant.
The Effect of Inhibition of Prostaglandin Synthesis on Microvascular Haemostatic Plug Formation in the Rabbit
Authors: Bergqvist D. , Svensjö E. , Arfors K.-E.
Published: June 2019

Bleeding induced by microvascular transection in the rabbit mesentery stops by the formation of a haemostatic plug. Normal platelets as well as the normal coagulation and fibrinolytic systems are essential for haemostatic plug formation, the initial formation being mainly ADP-dependent and the stability mainly an effect of fibrin formation. The difference in haemostasis between arterioles and venules was abolished by aspirin (Arfors et al. Scand. J. Haematol. 9, 322, 1972). In this study we have investigated the effect of indomethacin. As with aspirin, venular haemostatic plug formation time was shortened and plug stability increased. Local infusion of PGE1 into the cranial mesenteric artery significantly prolonged arteriolar and venular haemostatic plug formation time. Measuring blood flow velocity, vessel contraction and haemostatic plug volume makes it possible to determine the proportion of platelets participating in the formation of an effective plug in individual vessels. Platelet aggregability is significantly higher in plugs formed at injuries on the arteriolar side of the microcirculation than on the venular, but this difference is totally abolished after indomethacin. In conclusion the difference in haemostasis between arterioles and venules in this model can be explained by prostaglandin being formed in the mesenteric preparation.
On the Diagnostic Criteria of DIC in Childhood and the Indication of Streptokinase Therapy in Various Primary Diseases
Authors: Schreinert B.
Published: June 2019

In contrast to the diagnostic parameters of disseminated intravascular coagulation (DIC) in adults the criteria in children must be new defined. The value of the levels of fibrinogen, F V, VIII, XIII, FDP, fibrinmonomers, and the platelet count are quite often different in adults and children as demonstrated. The results were taken from 144 healthy children and 42 paediatric cases of DIC in various primary diseases with special respect to the spontaneous and induced platelet aggregation as an important diagnostic symptom in early DIC. As a result of the investigation new criteria of DIC in childhood as an indication for a thrombolytic therapy are presented and discussed. The main role in this new concept play the platelet count and function, the levels of F V, VIII, and the presence of FDP and fibrinmonomers as a supplement of the global coagulation tests. Finally the approach in DIC in childhood will be represented.
High Fibrinogen Concentrations and a Positive Ethanol Gelation Test
Authors: Slaastad R. A. , Godal H. C.
Published: June 2019

In patients with a positive ethanol gelation test other findings indicating consumption coagulopathy was more frequently demonstrated than in patients with a negative test. Furthermore, high fibrinogen levels were closely associated with a positive test, even, when all the other parameters studied were normal.

To see if high concentrations of fibrinogen alone produced gelation with ethanol fibrinogen was administered intravenously to rabbit. During infusion of highly purified fibrinogen, the test remained negative even at fibrinogen levels above 1500mg%. If the fibrinogen was treated with traces of thrombin prior to infusion, the ethanol test became positive at fibrinogen values about 500 mg%. Thus, soluble fibrin is necessary to give a positive ethanol test even at high fibrinogen levels. This was also supported by in vitro studies.
The Value of a Prothrombin Complex Concentrate in the Coagulation Defect of Liver Disease
Authors: Thomson Jean M. , Poller L. , Green G. , Dymock I. W.
Published: June 2019

A prothrombin complex concentrate rich in factor VII has been used in the management of the clotting defect in thirteen patients with liver disease. Adequate correction of coagulation was achieved immediately after the infusion in all cases. Within four hours there was some deterioration and by twenty-four hours the results approximated to pre-infusion values. Liver biopsy procedures were performed without haemorrhagic complication in the immediate post-infusion period. There was no evidence of induced intravascular coagulation.

Other prothrombin complex concentrates have proved disappointing both in their failure to correct the clotting defect and their production of DIC. From our results the use of this factor “VII rich concentrate appears to be the treatment of choice in patients with liver disease requiring temporary correction of the coagulation defect.
Thromboplastin and Platelet Factor 3: Protein and Phospholipid Components Required for Procoagulant Activity
Authors: McCoy L. E. , Liu D. T. H. , Wu V. Y.
Published: June 2019

Characteristics of bovine brain and lung thromboplastins and platelet factor 3 are compared, emphasizing protein and phospholipid moieties involved in procoagulant activity. Isolated tissue and platelet procoagulant lipoproteins were separated into lipid and protein fractions by repeated ethanolic extraction. Lipid composition and quantitation was ascertained by thin layer chromatography. The protein-free lipid mixture functioned as a partial thromboplastin. Protein fractions were purified by deoxycholate solubilization, gel filtration, and chromatography on diethylaminoethyl cellulose. Comparative biochemical properties were determined by gel filtration, Polyacrylamide gel electrophoresis, sedimentation velocity and by carbohydrate and amino acid composition. Purified protein components were devoid of enzymatic activity. They were also free of procoagulant activity on prothrombin or au top roth rombin III (Factor X) unless they were combined with one or more phosphatides of ethanolamine (PE), choline (PC), serine (PS), or inositol (PI). The proteins of thromboplastin appear to have homologous biochemical characteristics and phospholipid requirements for expression of proeoagulant activity, while differences in platelet factor 3 activity relate to differences in both protein and phospholipid requirements.

(Supported by USPHS, NIH Grant HL 14142).
On the Primary Structure of the A Chains
Authors: Hessel B.
Published: June 2019

During plasma digestion of fibrinogen a degradation product with molecular weight (MW) of about 50,000 appear very early in the digest. As revealed by sodium dodecyl sulphate (SDS) gel electrophoresis this product has its origin in the A chain. Upon further hydrolysis the 50,000 MW fragment is degraded to a 25,000 MW fragment. Both the 50,000 and 25,000 MW fragments have been isolated. After treatment with cyanogenbromide (CNBr) of the 50,000 MW fragment and separation of the peptides obtained, a fragment with MW of 30,000 and two fragments of smaller MW were isolated. As revealed by amino acid sequence analysis and peptide mapping the 30,000 MW fragment was identical to a hydrophilic sulfurcontaining peptide (Hi2-DSK), isolated after CNBr-treatment of the whole fibrinogen molecule. The amino acid sequence analysis of the fragments is in progress. Amino acid sequences analysis of the 25,000 MW fragment gave the following sequence: Ala-Len-Thr- Asp-Met-Pro- Gln-Met- Arg-Met-Glu-Leu-Glu-. From residue number 11 it appears to be identical to the Hi2-DSK fragment. However, the 25,000 MW fragment is shorter at the COOH-terminal end.
Soluble Fibrin Complexes in Coronary Artery Disease
Authors: Colman R.W. , Anaya-Galindo R. , Shattil S. J. , Cooper R. A.
Published: June 2019

Patients with myocardial infarction have been found to have elevated plasma concentrations of soluble fibrin complexes (SFC) as revealed by agarose gel chromatography but whether this is a result of the disease or a predisposing factor is not known. Twenty one individuals undergoing coronary arteriography for evaluation of chest pain were evaluated for evidence of intravascular coagulation. Sixteen patients who had abnormal arteriograms with one or more coronary arteries occluded by more than 80% were comparable in age, total and low density lipoprotein cholesterol and triglycerides (p > .4) to five subjects who had no angiographic abnormalities. Evidence for surface activation of plasma coagulation was not found since both groups of patients had normal levels of factor XII, prekallikrein and kallikrein inhibitors and did not differ significantly from each other, Fibrinogen) split products were not elevated significantly in patients with positive coronary angiograms (p > .2) and their levels did not correlate with the degree of angio-graphic abnormality (product of extent of occlusion and number of vessels affected). In contrast, there was a striking increase in the percent of total fibrinogen present as SFC in the patients with abnormal arteriograms (23.3±3.5 SEM%; normal 4.6±1.3%) compared with the patients without angiographic abnormalities (10.9±1.5%). Moreover, the concentration of SFC showed a good correlation (r = .49; p < .05) with the degree of angiographic abnormality. The findings suggest that measurement of SFC may he a useful noninvasive screening test for assessing the extent of coronary artery occlusion.
Therapeutical Effects of Heparin and Aprotinin on Experimental Disseminated Intravascular Coagulation in the Rat
Authors: Heyes H. , Slijepcevic B. , Glück D.
Published: June 2019

A disseminated intravascular coagulation (DIC) was induced in rats by injection of 1 mg endotoxin and subsequent infusion of isotonic saline (500 ml/kg/5 hours) over a period of 5 hours. Animals were treated beginning at the time of initial glomerular fibrin deposition with heparin (1000 IU/kg) or aprotinin (40000 KIU/kg) or heparin-aprotinin in combination (each drug as given as in single dose). The first half-dasc was given as an bolus injection 2 hours after the endotoxin injection, the second dose immediately was added to the saline infusion. The extent of DIC was controlled by several blocdparameters, histological examination of the kidneys, and by the application of 125–I-fibrinogen (given after starting DIC) and 131-I-fibrinogen (injected after beginning the therapy). By the use of two isotopes it was possible to observe the behaviour of the fibrinogen in two different phases.

Heparin had clearly favorable effects stabilizing the situation of hemostasis, although the fibrinogen turnover was not normalised completely, probably due to fibrin monomers which even polymerize in presence of heparin. The beneficious effect of heparin was diminished by the addition of aprotinin which given alone had deleterious effects inhibiting the spontaneous removal of fibrin thrombi. The fibrinogen turnover was clearly enhanced supporting the concept that the plasma fibrinogen level cannot be protected by blocking the fibrinolysis which is very rarely generalized.
Effects of Various Fibrinolytic Agents on the Physico-Chemical Characteristics of the Cardiovascular System
Authors: Sawyer P. N.
Published: June 2019

The effects of fibrinolytic agents Brinase, Brmolase (enzymes isolated from aspergillus oryzae) and Thrombolysin (streptokinase and human plasmin) on the cardiovascular system of dogs and rabbits have been determined by the following methods: (a) electro-phoretic mobility of canine erythrocytes and platelets; (b) alterations of the vascular intimal surface charge as determined by electroosmosis and streaming potentials; (c) alterations in platelet and erythrocytes adhesion characteristics; (d) changes in coagulability of blood as demonstrated by several standard determinations; (e) histology of rabbit blood vessels, and (f) scanning electron microscopy of canine blood vessels. The results indicate that Brinase at 2 mg/kg increases the net negative surface charge of the vascular intima. There is a byphasic response in altering the electrophoretic mobility of erythrocytes and blood platelets, an initial increase in the surface negativity followed by a return to normal after two days. Brinase reduces platelet adhesion to a metal surface as well as to the vascular intima. Histologic examination of hyperlipidemic rabbit vessels indicates that Brinase is effective in reducing fat infiltration with a minimal amount of destruction to the vascular wall. Brinolase has been administered at doses 4.3 mg/kg and 1 mg/kg. The former being a full proteolytic dose results in irreversible incoagulability, total proteloysis and usually death of the animal. At 1 mg/kg the hematologic and physico-chemical effects on the cardiovascular system are similar to those produced by Brinase. The overall effects of Thrombolysin are comparable to those induced by Brinase at 2.0 mg/kg in the long term study.
Formation of Soluble Complexes of Fibrinogen Related Material During Ancrod Infusion
Authors: McKillop Caroline , Edgar W. , Forbes C. D. , Prentice C. R. M.
Published: June 2019

Seven pationts undergoing therapeutic defibrination by ancrod infusion were studied. Blood samples were obtained before treatment and after 6 and 24 hours ancrod infusion. Fibrinogen and its derivatives were precipitated with beta-alanine and separated by ΰ per cent agarose gel filtration. A range of soluble complexes were demonstrated after 6 hours infusion. Polyacrylamide gel electrophoresis in SDS showed that the soluble complexes were largely composed of units with molecular weight similar to a minimally degraded early Fragment X. Polyacrylamide gel electrophoresis in SDS and mercaptoethanol showed a marked loss of intact alphachain in the soluble complexes when compared with the uncomplexed material, suggesting that the soluble complexes had undergone preferential fibrinolytic digestion. It is suggested that, during ancrod therapy, FDP may be produced directly from soluble complexes rather than insoluble micro-thrombi as has been suggested previously.
Cytochalasins: Effects on Platelet Aggregation, Adenylate Cyclase and Membrane Transport Processes
Authors: Mills D. C. B. , Macfarlane D. E.
Published: June 2019

Cytoehalasins A and B (CA and CB) are fungal metabolites that inhibit cellular motility, platelet aggregation and clot retraction. CA differs from CB only in the substitution of a carbonyl in CA for a hydroxy in CB. This confers on CA reactivity towards thiol groups. We have studied the effects of CA and CB on various membrane functions of intact human platelets in citrated plasma. CA inhibited the uptake of adenosine and 5 HT without affecting intracellular nucleotide metabolism. CA (56 uM) but not CB increased the rate of adenine uptake from 11.7±0.2 to 80.1±04.4 pmoles/inin/108 platelets, while the affinity of the uptake mechanism was reduced (Km apparent increased from 0.12 ±0.01 to 1.31 ±0.12 uM). CA inhibited platelet aggregation induced by ADP at concentrations lower than those that blocked the shape change. CA (20–50 uM) abolished the inhibition by ADP of cyclic AMP accumulation in platelets exposed to PGE1 and the phosphodiesterase inhibitor RA 233, whereas CB (170 uM) was inactive. This effect of CA was prevented but not reversed by 1 mM cysteine.

The observed actions of CA are similar to those of other thiol reagents but occurred at concentrations below that of plasma protein thiol, indicating a preferential reactivity of CA towards the platelet membrane.

(Supported by N. I. H. HL 14217.)
Studies on Some Thrombin-Induced Platelet Membrane Alterations
Authors: Hagen I.
Published: June 2019

Membrane alterations in connection with thrombin-induced release reaction have been studied with washed, human platelets. The release reaction was induced by 0.01–1.0 NIH units of thrombin, incubated for 3 min in the presence of 3 mM EDTA. Membrane proteins exposed on the surface were labeled with 125I-by the lactoperoxidase-iodination technique.

Three radioactive peaks (approx. MW 97.000, 120.000 and 150.000) were seen on subsequent SDS Polyacrylamide gel electrophoresis of control platelets. Treatment of prelabeled platelets with thrombin did not reveal any changes in the “specific” radioactivity of the three peaks, indicating that no labeled fragments had been split off.

Treatment with thrombin before iodination under conditions where the release reaction occurred, resulted in a drastic reduction in the incorporation of radioactivity. When the release reaction was inhibited by antimycin and 2-deoxyglucose before thrombin treatment and subsequent iodination, very little or no effect could be observed on the incorporation of radioactivity.

Aggregation of these platelet suspensions with bovine factor VIII, and ristocetin plus human ristocetin cofactor indicate that platelets which have undergone the release reaction aggregate poorly with these agents. The membrane conformation necessary for iodination of the three membrane polypeptides and the ability of the platelets to aggregate in the factor VIII-dependant systems thus seem to be altered during thrombin-induced release reaction, possibly because of alterations in the membrane structure.
Platelet Membrane Glycoproteins and their Possible Relationship to the ADP Mechanism of Platelet Aggregation
Authors: Phillips D. R. , Jenkins C. S. P. , Meyer D. , Larrieu M.-J. , Luscher E. F.
Published: June 2019

Platelets isolated from patients with Glanzmann’s Thrombasthenia release in the presence of thrombin and other stimuli but fail to respond to ADP. Since the iutia.l interaction, between the platelet and ADP is at the membrane surface, it would appear that this surface lacks the necessary receptor for ADP. The surface structure of normal and thrombasthenia platelets was compared using the lactoperoxidase iodination technique. Iodination of normal platelets results in the labelling of four glycoproteins, I, IIa IIb and III, with relative ratios of 1 : 1 : 1 : 3 plus other non-characterised polypeptides. Thrombasthenic platelets similarly treated revealed a drastically altered expression of the glycoproteins on the membrane. The relative ratios (1.5: 1:0.4:0.5) revealed the decrease of glycoprotein IIb and the marked reduction of glycoprotein III. Arguments and data will be presented which point to the possibility that glycoprotein IIb is involved in ADP-induced aggregation.
Absence of Interaction between Strepto-klnase/Plasmin(ogen) Complexes and α2-Macroglobulin (α2M)
Authors: Steinbuch M. , Friess A. , Drouet J. , Amouch P.
Published: June 2019

Streptokinase, plasmin(ogen), the activator complex and α2M have different, well defined electrophoretie mobilities in agarose/acrylamide in function of their electric charge and molecular size. Highly purified I131-labeled streptokinase has been used to study whether an interaction with α2M can be seen. No radioactivity associated with α2M could be found. Thus, with respect to α M, streptokinase does not behave like urokinase. Furthermore, activator complexes formed in the cold (4°C) to avoid any dissociation did not react with α2M either a,s shown by the same technique whereas I131-labeled plasmin became normally fixed to α2M undoT the same conditions. Several anodic radioactive bands are observed after the interaction between I131-labeled streptokinase and plasminogen) thus showing a striking modification of the streptokinase molecule.

It can be concluded that the activator complex is not inhibited by α2M.
Evidence for a Functionally Important Plasmin in Inhibitor in Human Plasma, Different from α2-Macroblobulin, α1-Antitrypsin and Antithrombin III
Authors: Edy J. , Collen D. , Verstraete M.
Published: June 2019

Gel nitration ou Sephadex G-200 of urokinase-activated fresh human plasma containing a trace amount of radiolabeled plasminogen and of serial plasma samples obtained during streptokinase therapy in patients following injection of labeled plasminogen has been performed. A parallel disappearance of radioactivity and enzymatic activity eluted in the plasminogen position was observed concomitant with the appearance of two radioactive peaks, eluted at the void volume (P-α2M) and just before the globulin peak (P-AP) (D. Collen, Plasminogen and prothrombin metabolism in man, Aggregaatsthesis, Leuven, 1974). The relative distribution of radioactivity over the P-α2M and P-AP peaks depends on the rate of plasminogen activation and may vary from 3 to 1 to 1 to 3, slow activation favoring P-AP formation.

The purified P-α2M complex has been identified as plasmin-α2-macroglobulin complex. The purified P-AP complex, obtained in high yield, does not cross-react with antisera against α1-antitrypsin or antithrombin III, has a mol wt of about 130,000 by SDS-polyacrylamide gel electrophoresis and upon reduction displays a broad band with a molecular weight of about 70,000. Recently Mullertz (Btochem, J., 143, 273, 1974) has probably identified the same P-AP complex which in addition did not react with antisera against inter-α-trypsin inhibitor and C1-esterase inhibitor. All these data suggest that human plasma contains a functionally important, plasmin inhibitor which is different from the five main protease inhibitors in plasma. We have provisionally named this inhibitor α1antiplasmin, in accordance with the nomenclature of Norman (J. Exp. Med.. 108, 639, 1958).
Current Major Trials with Antithrombotic Agents
Authors: Verstraete M.
Published: June 2019

Numerous clinical trials have been recently carried out or are being planned with antithrombotic agents. What is the current knowledge on agents affecting the platelet function: are they clinically useful? The clinical validity of anti-aggregating agents is to be demonstrated in controlled clinical trials having a vascular occlusion as the end point to judge the drug effect. A survey will be presented on the results available with one of the three agents currently available, acetylsalicylic acid, dipyridamole and sulfinpyrazone. Following clinical conditions are to be discussed separately: peripheral arterial disease, coronary artery disease, patients with prosthetic heart valves, certain renal disease, ITP, haemolytic uraemic syndrome and conditions often complicated with deep venous thrombosis.

To what extent and subcutaneous heparin used in low-dosage can decrease the incidence of post-operative deep-vein thrombosis and outweight its possible disavantages is another item to be reviewed.
Low-Dose Heparin Prophylaxis Against Fatal Pulmonary Embolism
Authors: Kakkar V. V. , Corrigan T. P.
Published: June 2019

Several controlled trials have shown that low-dose heparin is effective in reducing the incidence of deep vein thrombosis without increasing the risk of bleeding. However its effectiveness in preventing fatal pulmonary embolism has yet to be determined and, for this, a multicentre tiral was organised in which 31 centres took part; patients over the age of 40, undergoing only major elective abdominal, thoracic or orthopaedic surgery were included. They were randomly allocated to a control or heparin group and, for each patient entered in the trial, essential information was recorded in a proforma designed for computer analysis. The incidence of fatal pulmonary embolism was determined by autopsy examination. 4,121 patients were admitted to the trial – 2,076 in the control group and 2,045 in the heparin group. 16 patients in the control group died due to acute massive pulmonary embolism confirmed at autopsy, but only 2 in the heparin group. The difference is statistically significant (P < 0.01). There was no evidence of excessive blood loss during or after surgery. These results will be presented in detail. Low-dose heparin prophylaxis can now be recommended as the method of choice for preventing postoperative fatal pulmonary embolism.
Prevention of Deep-Venons Thrombosis after acute Myocardial Infarction. Comparison ot Effectiveness of ASA, Dipyridamole + ASA, and Phenprocoumon
Authors: Hart H. Gh. , Vries L. J. de , Smit J.W. Wisse
Published: June 2019

In a controlled prospective trial of anti-aggregating drugs versus phenprocoumon, the incidence of deep-venous thrombosis (D. V. T.) was measured in 150 patients admitted with acute myocardial infarction in a coronary care unit. Patients moribund on admission, with a history of active peptic ulcer or hiatus hernia, a recent cerebrovascular accident, and those who used oral anticoagulants, aspirin, or anti-aggregating drugs regularly were excluded from participation. At admission the patients were randomly allocated to one of three treatment groups: 1.5 gram of acetylsalicylic acid (ASA) in 3 divided doses, a combination of 300 mg dipyridamole (in 4 divided doses) + 1.5 gram of ASA, or phenprocoumon. All patients were confined to bed for 12 days. 135 Patients completed the trial. The 3 groups were well matched statistically with regard to sex, age, weight, and coronary prognostic index. The l25I-fibrinogen test was used for detection of D. V. T.

The difference in frequency of D. V. T. between the ASA-group and the two other groups is highly significant (p < 0.01). The combination dipyridamole + ASA was as effective as phenprocoumon in the prevention of D. V. T. after acute myocardial infarction.
Fibrinogen Metabolism and Disseminated Intravascular Coagulation in Dengue Hemarrhagic Fever
Authors: Srichaikul T. , Nimmanitaya S. , Artchararit N.
Published: June 2019

Fibrinogen metabolism was studied using 125I fibrinogen in 29 Dengue Hemorrhagic Fever (DHF) patients (twelve with grade 2 and 17 with grade 3 & 4 DHF), Hemostatic studies were also performed to search for evidence of disseminated intravascular coagulation (DIC). The findings indicated that increased fibrinogen consumption as judged by rapid T1 and increased fractional catabolic rate (FCR) of radioactive fibrinogen occur in 58% of grade 2-DHF patients. Moreover 2 patients in this group had hemostatic changes suggesting DIC. In grade 3 & 4 patients, the majority of whom were studied during shock, increased fibrinogen consumption was observed as well as increased FCR in 89% of cases. Evidence of DTC was observed in 13 of 17 cases with latter group. A correlation between the increased fibrinogen consumption, DIC and clinical severity and shock was demonstrated. The role of DIC on the pathogenesis of shock in DHF was discussed. The use of heparin was suggested for patients with prolonged shock and severe acidosis when DIC becomes clinically apparent.
The Risk of Deep Venous Thrombosis in Surgical Patients Using Multivariate Analysis
Authors: Nicolaides A. N. , Irving Doreen
Published: June 2019

The purpose of the present study was to develop a method of determining the patients who are at risk and require prophylaxis.

In a prospective study of 535 patients various clinical factors believed to predispose to deep venous thrombosis have been studied using a linear logistic analysis. The patients had been screened with the 125I-fibrinogen test. A standard computer program was used to obtain the maximum likelihood estimates of the coefficients for the various factors studied. The most important factor was age; premedication with omnopon, presence of varicose veins, infection, history of previous DVT, severity and type of operation were also significant.

For any patient the logit y can be estimated and expressed as a percentage risk of thrombosis using the equation y = −6.00 + (Age × 0.0617) + (History of previous DVT × 1.38) + (V. veins × 1.26) + (Premedication with omnopon × 0.97) + (Infection × 0.84) -(Urological operation × 1.94) - (Thoracic operation × 1.15) derived from the significant factors and their coefficients. (The values of factors other than age are either 0 = absent, or 1 = present.)
Electronmicroscopic Studies on Human Thrombocytogenesis
Authors: Kosaki Goro
Published: June 2019

The mechanism of platelet production and platelet release in human bone marrow tissues was studied electronmicroscopically. Promegakaiyocytes and megakaryocytes were classified morphologically into 4 stages. Out of the 126 cells studied, 3.2% were in stage I, which was featured by active synthesis of protein and other cellular constituents; 8.2% were in stage II, in which specific granules were produced; 57.9% were in stage III, in which the demarcation membrane was formed; 21.5% were in stage IV, in which platelets were released. Eleven (8.7%) were unclassified. Seventy-three or 75% of the colls in each stage I, II and III existed in the parenchyma. But almost all megakaryocytes in stage IV existed as free cells presumably in a sinusoid. It is assumed that, after formation of the demarcation membrane in the whole cytoplasm, megakaryocytes in the parenchyma move into the sinusoids, and then the whole cytoplasm of megakaryocytes is broken explosively into small pieces to be designated as platelets.
Thrombocytopathy Induced by Acquired Circulating Factor VIII Inhibitor
Authors: Cortellaro M. , Pogliani E. , Cofrancesco E. , Polli E. E.
Published: June 2019

A reduced ADP and ristocetin platelet aggregation by acquired circulating factor VIII inhibitor (2 U/ml) was found in a young woman with S. L. E. Bleeding time was prolonged, factor VIII activity decreased (25%), Willebrand antigen and Willebrand factor were normal. PF3 assay and PF4 release were normal. Platelet (14C) - serotonin uptake, but not release, was reduced. Antiplatelet antibody was not detected.

Patient’s plasma inhibited the ristocetin and ADP induced platelet aggregation of normal PRP, but not of normal and patient GFP.

After steroid treatment F. VIII inhibitor and thrombocytopathy disappeared.

It is suggested that the circulating inhibitor is able to coat autologous and isologous unwashed platelets, interfering with platelet function.
Purification of Prothrombin by Means of Adsorption Onto Al(OH)3 in the Presence of Cd++ Ions
Authors: Devilee P. P. , Hemker H. C.
Published: June 2019

A simple method for the purification of prothrombin is based on the phenomenon that in the presence of 1 mM CdSO4, Al(OH)3 (1.3% w/v) completely adsorbs the coagulation factors VII, IX, and X from normal plasma but adsorbs factor II for about 50% only. Column chromatography is necessary only once. A 750-fold purification is obtained at a 24% yield. The product is indistinguishable from that obtained by the method of Kisiel and Hanahan (Biochim. Biophys. Acta 304, 103-113, 1973) as judged by amino acid composition, N-terminal amino acid (alanin) molecular weight (73,000) or immunological properties. Comparison of the two preparations in a thrombin generating system shows that although the final yield of thrombin from a given amount of prothrombin in both preparations is the same, the initial velocity of thrombin formation from our preparation is comparable to that of native prothrombin, whereas the other preparation is converted significantly slower.

This phenomenon may be related to the loss of γ-carboxyl glutamic acid residues during the purification procedure as described by Kisiel and Hanahan.
Purification and Properties of Staphylocoagulase
Authors: Muller A.D. , Bas B. M. , Hemker H. C.
Published: June 2019

Staphylocoagulase, an exoprotein of coagulase positive staphylocoagulase, has been purified to a state in which only trace amounts of contaminating proteins are detectable.

Purification was more than 35,000 fold, which is 7 times more than the highest value reported in the literature. The yield was about 15%.

Aspartic acid was found as a single N-terminal amino acid in this preparation. The molecular weight is 61,000 and the isoelectric point lies at pH 4.53.

The amino acid composition was determined.
Interaction Cultured Cells and Blood Platelets
Authors: Holme R. , Oftebro R. , Hovig T.
Published: June 2019

The effects of washed cultured cells (human lymphoma cells and three different lines of Chang liver cells) on blood platelets in heparinized plasma were studied. Whereas suspensions of two of the lines with Chang cells induced platelet aggregation, the third line did not. Suspensions of the lymphoma cells induced marked platelet aggregation. Ultrastructurally the platelet aggregates were densely packed with pronounced alterations of the platelet morphology and presence of fibrin strands. The aggregation reaction was not blocked by hirudin, but could be almost completely abolished by incubation with apyrase. Presence of ADP could be demonstrated in the washing fluids from the cultured cells, except for the one line of Chang cells which did not induce aggregation.

It is concluded that ADP may leak out from damaged cells, especially tumor cells in amounts sufficient to induce microthrombosis.
Partial Purification and Characterization of Factor VIII-Activity Present in the Supernatant of Cryoprecipitates
Authors: Over J. , Bakker-Woudenberg I. , Bouma B.N. , Mourik J. A. van , Sixma J.J.
Published: June 2019

A relative higher percentage of the initial factor VIII activity is found in the supernatant of plasma from patients with von Willebrand’s Disease than in that from normals after cryoprecipitation. Moreover no factor VIII related antigen can be demonstrated in the supernatant after cryoprecipitate. A further study of the characteristics of this cryo-supernatant-factor VIII seemed therefore of interest.

Cryosupernatant-factor VIII was partially purified by poly ethyl enegly col and ammoniumsulfate precipitation, followed by Sepharose 6B gelfiltration. Cryosupernatant-factor VIII was eluted at the void volume of Sepharose 6B columns, but in contrast to cryoprecipitated factor VIII it was retarded on Sepharose 2B columns.

Cryosupernatant-factor VIII has antigenic determiniant(s) in common with normal factor VIII.

Von Willebrand plasma’s obtained 6 to 40 hours after transfusion with normal cryoprecipitate were applied to gelchromatography on Sepharose 6B columns. The factor VIII activity was eluted at the void volume. On Sepharose 2B the elution pattern of the factor VIII activity was comparable to that of normal plasma.
Coagulation in an Agarose Gel and its Application to Detecting Factor VIII Clotting Activity Associated with Factor VIII-Related Antigen
Authors: Bird Prudence , Rizza C. R.
Published: June 2019

The coagulation of citrated plasma in an agarose gel treated with thrombin or calcium chloride may be detected by a change in the gel opacity. Experimental evidence suggests that this opacity represents fibrin formed within the gel matrix. The coagulation of citrated 0% factor VIII haemophilic plasma in an agarose gel treated with calcium chloride is delayed compared with normal plasma. A method for detecting factor VIII clotting activity in agarose has been developed based on factor VIII promoting the coagulation of haemophilic plasma in agarose.

Using this test to detect factor VIII clotting activity in a one dimensional Laurell electroimmunoassay for factor VTII-related antigen, all factor VIII clotting activity detected is found in the same position as the factor VIII-related antigen immunoprecipitate. This result suggests that the molecule containing factor VIII clotting activity carries factor VIII-rolated antigen determinants.