bluuebunny/crossref_metadata_2025 huggingface dataset:
The Uptake of Adenine by Isolated Human Platelet Membranes
Authors: French P. C. , Sixma J. J. , Holmsen H.
Published: June 2019

Adenine uptake into isolated platelet membranes had about the same Km (181±21.9 nM) as uptake into intact platelets (151 ±21 nM) and was also competitively inhibited by papaverine and hypoxanthine. No uptake occured at 0° and accumulated adenine was converted to AMP. AMP was not bound to protein as judged by chromatography of triton X-100 solubilized membranes chromatographed on Sephades G-25. The pH optimum for adenine uptake was atpH 5.5. Exogenous 5-phosphoribosyl-l-pyrophosphate stimulated uptake.

These data may be explained by uptake of adenine by facilitated diffusion followed by conversion to AMP by adenine phosphoribosyltransferase (E.C. 2.4.2.7.) but group translocation cannot be entirely excluded.
Postoperative Deep Vein Thrombosis
Authors: Gordon-Smith I. C.
Published: June 2019

Subcutaneous heparin therapy has been shown in several recent studies to be an effective agent in the prevention of postoperative deep vein thrombosis (DVT). However, DVT has not been abolished by such therapy and an increase in haemorrhagic complications, notably wound haematomata, has been reported after its use.

In an ongoing double blind randomised controlled study the effects of heparin on DVT and postoperative complications have been correlated with blood heparin levels. Thus far some fifty patients have been studied with the following preliminary conclusions :

(I) Subcutaneous heparin administration results in markedly variable blood levels,

being maximal two hours after injection at between 0.01-0.30 I.U./ml. (II) 12-hourly injections of 5000 I.U. heparin did not have a cumulative effect over one week.

(III) Of seven patients with blood heparin levels in excess of 0.2 I.U./ml, four developed a wound haematoma. Of seventeen patients with blood heparin levels of less than 0.2 I.U./ml only two patients developed this complication.

(IV) Thus far 125I-Fibrinogen detected DVT has not occurred in the heparin treated patients.
Heparin Kinetics in venons Thrombosis and Pulmonary Embolism
Authors: Hirsh J. , Aken W. G. van , Gallus A. S. , Dollery C. T. , Cade J.
Published: June 2019

Patients with venous thromboembolism differ considerably in their heparin response and requirements. We have investigated the mechanism for, and practical consequence of this variation in 20 patients with proven venous thromboembolism (VTE). A standard i/v heparin dose 70 U/kg bolus followed 90 min later by 400 U/kg/24 hrs. by continuous infusion was given. Blood was drawn pre heparin and 15, 45 and 90 min post bolus and then during maintenance infusion for the following tests: heparin tolerance, activated partial thromboplastin time (APTT), heparin level. In addition heparin recovery and clearance were calculated. There were large inter-individual variations in the anticoagulant response to heparin which was due in part to differences in rates of heparin inactivation and clearance and in part to marked differences in the APTT response to heparin. For example, the heparin level at 15 min post bolus varied from 0.2 U to 0.9 U and during maintenance from 0.0 U to 0.5 U. The APTT varied from 45 to 200 sees, at 15 min and 38 to 96 sees during maintenance infusion. In addition the APTT prolongation over the pre-treatment APTT varied up to 9 fold between patients for a given heparin concentration. There was a marked and significant difference in heparin half life between patients with venous thrombosis (70±17 min) and pulmonary embolism (PE) (38±1 min) p < 0.005. This rapid clearance in PE may be due to release of antiheparin activity from platelets which interact with thrombin onto the surface of the embolus. The results suggest that the heparin dose should be individualized in VTE and that larger doses should be initially given to patients with PE.
Coagulation Studies in C4-Deficient Guinea Pigs
Authors: Dodds W. J. , Raymond S. L. , Moynihan A. C. , Pickering R. J. , Amiraian K.
Published: June 2019

Coagulation and complement parameters of our colony of NIH-multipurpose strain, C4-deficient guinea pigs were compared to those of our age- and sex-matched control guinea pigs (Hartley strain). In addition to very low (< 1 unit) C4 levels, the deficient pigs had significantly lowered CI and C2 activities. Terminal complement component (C3-C9) activity was similar in both strains of animals. Relative to the control population, C4-deficient pigs had significantly higher leukocyte and platelet counts, and more active clot retraction; lower α1- and β2-globulins; prolonged partial thromboplastin and short prothrombin consumption times; and reduced factor VIII activity. Some sex-related differences, such as increased factor XI levels in C4-deficient females, were also observed. Whole blood-clotting and clot lysis times, fibrinogen and plasminogen levels, and factors II, VII, and X activities were not significantly different.

These findings indicate reduced intrinsic coagulation in the C4-deficient animals relative to the controls. The coexistence of complement and coagulation deficiencies further supports the functional relationship between these systems.

(Supported in part by USPHS Grant HL-09902 from the National Heart and Lung-Institute, National Institutes of Health.)
The Role of Membrane Bound Complement in the Aggregation of Mammalian Platelets by Collagen
Authors: Chater B. V.
Published: June 2019

It has been observed in dogs decomplemented with purified Cobra Venom Factor, that their platelets lose the ability to aggregate in response to collagen stimulation. Further investigation of the effect of CVF in vitro in man, dog and rabbit, and in vivo in dog, reveals that in each case CVF abolishes the collagen response of platelets, and that this effect is dose related. Resuspension of CVF inactivated platelets in plasma containing complement, results in a total return of sensitivity to collagen. Examination of CVF inactivated platelets with the electron microscope, fails to show any marked difference from control platelets. Serotonin granules are still present and the platelets retain a discoid appearance. Incubation of platelets with antibodies to Cl, C3 and C5, results in inhibition of the collagen response, this effect also being dose related. Light microscopy studies indicate that CVF does not affect the adhesion of platelets to collagen but appears to prevent subsequent aggregation. It is suggested that the complement system is involved in the induction of release by collagen, and that inhibition by CVF and anti-complement antibodies is the result of a blocking of the release reaction.
Products of Platelet Aggregation and Release as a Chemotactic Factor for Human Leucocytes
Authors: Clawson C. C. , Repine J. E.
Published: June 2019

Human platelets, doubly washed and resuspended in protein-free balanced saline, were aggregated with bacteria (S. aureus), adenosine diphosphate or collagen. The soluble products of platelet release were separated from the aggregates by centrifugation at 105,000 g for 60 min. Washed human neutrophils in protein-free balanced saline in Boyden chambers were attracted as well by the supernatants of platelet aggregation (297 ± 35) as they were by zymosan-activatedpooled serum (275 ± 12). Ultrasonic disruption of washed, unaggregated platelets failed to release comparable soluble chemotactic components. Control supernatants of unaggregated platelets, bacteria, ADP or collagen and the pellets from aggregated or sonicated platelets all failed to stimulate leucotaxis. Neutrophils incubated with inhibitors of prostaglandin synthesis failed to respond to platelet aggregation products although their random mobility was not impaired. The soluble products of platelet aggregation also stimulated chemotaxis of human eosinophils and monocytes. Neutrophil metabolism of glucose 1-14C was not enhanced by incubation with the supernatants of platelet aggregation. These findings indicate that the leucotactic agent of platelets is a soluble product of physiologic aggregation and release, and that the response of neutrophils to the platelet product is dependent upon prostaglandin synthesis and independent of increased activation of the hexose-monophosphate shunt.

(Aided by grants of the NIH and American Heart Association.)
Endothelial and Smooth Muscle Cells as Antagonists in Vascular Fibrinoysis
Authors: Hegt V. Noordhoek
Published: June 2019

Endothelial plasminogen activator activity in different types of human blood vessels obtained from fifty necropsies and thirty-five biopsies was detected and localized by means of plasminogen-rich fibrin slides. Great differences in endothelial activator activity were found along and across (vasa vasorum) the wall of the human vascular system.

The same blood vessels were simultaneously investigated by a modified fibrin slide technique using plasminogen-free fibrin slides covered by plasmin to detect and localize inhibition of fibrinolysis in the vascular wall. The great variation in plasmin inhibition in different vessels revealed by this “fibrin slide sandwich technique” appeared to be closely associated with the localization and number of smooth muscle cells present in the walls of the vascular system. Strong plasmin inhibition was generally found at sites which showed no activator activity with the regular fibrin slide technique, while areas with a high endothelial fibrinolytic activity mostly revealed no inhibitory capacity.

These results indicate that much of the variation in endothelial fibrinolytic activity on fibrin slides is due to inhibitory effects from the surrounding smooth muscle cells rather than to variability in the plasminogen activator content of the endothelium itself.
Comparative Investigations on Factor VIII Assays
Authors: Pflugshaupt R. , Moser S. , Züger K. , Bütler R.
Published: June 2019

Six one stage methods and one two stage method were tested for precision and reproducibility. With each method twenty calibration curves of normal plasma and two lots of Factor VIII concentrates were established. Statistical evaluation revealed only minor differences. Neither one of the methods was optimal for both the physiological-pathological region and the region of high activity preparations.

Three selected methods were tested in vivo for accuracy: nine patients with hemophilia A were treated with equal amounts of Factor VIII concentrates or kryoprecipitates respectively. The methods showed different activities for preparations as well as for patient’s plasma. The discrepancy between measured and expected recovery differed for each method.
The Factor VIII Response to Synthetic Vasopressin Analogues in Man
Authors: Cash J. D. , Sas G.
Published: June 2019

Biological and immunological assays for factor VIII were performed on 6 healthy male volunteers before, during and after the intravenous infusion (10 μg in 15 minutes) of a variety of synthetic vasopressin analogues, which included arginine vasopressin (AVP), lysine vasopressin (LVP), l-Desamino-8-d-arginine vasopressin (DDAVP), vasotocin (VOT), l-Desamino-carb6-8-d-arginine vasopressin (DCAVP), and 1-Desamino-N-methylar-ginine8 vasopressin (DMAVP). No significant changes occurred during or after the administration of VOT and DMAVP. However, AVP, LVP, DDAVP and DCAVP all produced a significant and parallel increase in the biological and immunological levels of factor VIII. These results point to the possibility that vasopressin may play a role in controlling the release and/or synthesis of factor VIII in man, and, as DDAVP and DCAVP are devoid of unpleasant side-effects, indicate new therapeutic possibilities for the future management of patients with mild or moderate haemophilia A.
Automated Techniques for Estimating Factor VIII Levels in Concentrate Preparations
Authors: Seghatchian M. J. , Miller-Anderson M. , Howarth D. J. , Stirling Y. , Brozovic M.
Published: June 2019

An. automated coagulometer (a modification of Electra 600D) using an optical end-point for clot detection, has been used in both one and twostage methods of assaying factor VIII. Fifity clotting times can be measured in a hour. The methods for which the coagulometer has been used are (1) one-stage assay with kaolin-platelet substitute and haemophilic plasma, (2) one-stage assay with activated cephaloplastin (Dade, Nyegaard) and haemophilic plasma, and (3) two-stage method using combined serum reagent (Diagen). With the first two methods, a shortening of standard and blank clotting times occurred during the hour-long course of the assays (2-5 seconds per hour); this was particularly marked in the presence of kaolin, and due in part to activation of factor VII. With the two-stage method, longer standard and blank clotting times, up to 8 seconds per hour, were often recorded as the assays proceeded. These results emphasize the need to incorporate standards at regular intervals during every run, and to take account of the changes with time that occur.

Four different types of concentrate preparations commonly used in the treatment of haemophilia were assayed in two laboratories, each using conventional and automated (Electra 600D) methods. There was good agreement between methods and between laboratories.
A More Uniform Measurement of Factor VIII Inhibitors
Authors: Kasper C. K. , Aledort L. , Aronson D. , Counts R. , Edson J. R. , Eys J. v. , Fratantoni J. , Green D. , Hampton J. , Hilgartner M. , Levine P. , Lazerson J. , McMillan C. , Penner J. , Shapiro S. , Shulman N. R.
Published: June 2019

A group of hematologists, involved with hemophilia research and care in the U.S.A., met under the sponsorship of the Division of Blood Diseases and Resources of the National Heart and Lung Institute. In order to improve future communication among ourselves, we decided to alter our individual methods of measurement of inhibitors to the extent necessary to permit a uniform, although arbitrary, description of inhibitor units. We agreed to the following standards: (1) The incubation mixture consists of one part citratecl patient plasma, undiluted or diluted, plus an equal part of citrated pooled normal human plasma. (2) A control incubation mixture consists of equal parts of normal pooled plasma and imidazole buffer, as formulated by Dr. Biggs. (3) The mixtures are incubated at 37° C for two hours. (4) Assays specific for Factor VIII are then performed and the Factor VIII activity in the patient mixture is divided by the Factor VIII activity in the control mixture to determine the percent residual Factor VIII activity. (5) A patient plasma giving a residual Factor VIII activity of 50 percent in this test is said to contain one “Bethesda unit” of inhibitor per ml. (6) On a graph, the log percent residual Factor VIII activity is plotted against inhibitor units. If the residual Factor VIII activity of the incubation mixture is between 75 and 25 percent, the inhibitor units are read from the graph. Plasmas containing strong inhibitors are diluted with imidazole buffer before being placed in the incubation mixture. A dilution is sought which will result in a residual Factor VIII activity between 75 and 25 percent. The units of inhibitor read from the graph are then multiplied by the dilution factor to determine the number of Bethesda units of inhibitor per ml of undiluted patient plasma.

We invite interested colleagues to join us in the use of this method, and we invite discussion of better methods of describing inhibitor potency.
Dissociation of “Factor VIII Complex” in Various Animal Species.
Authors: Lavergne J. M. , Meyer Dominique , Jenkins C. S. P. , Larrieu Marie-José
Published: June 2019

Under conditions of high salt concentration, “Factor VIII complex” (Factor VHI activity, Willebrand Factor activity – measured using a washed platelet system and ristocetin – and Willebrand antigen) may be dissociated into a high (M. W. > 106) and a low molecular weight fragment. The dissociation of “Factor VIII complex” was studied by a two step procedure. Human or animal plasma or cryoprecipitate was submitted to gel filtration on Sepharose 4B, using 0.15 M NaCl, Imidazole or Tris-HCl Buffer as eluant. The void volume fraction, containing the three entities of “Factor VIII complex” was concentrated and submitted to a second gel filtration using a dissociating buffer as eluant (1 M NaCl or 0.25 M CaCl2). The three entities of “Factor VIII complex” were measured in the eluted fractions. “Factor VIII complex” was found to dissociate using the high salt buffer in some but not all animal species. When dissociation occurred, Willebrand Factor activity and antigen eluted in the void volume, and Factor VIII activity in later fractions. Heterologous antisera were raised against the different fractions and the reactivity towards human and animal plasma was studied.
Prophylaxis of Deep Vein Thrombosis after Total Hip Replacement
Authors: Rosengarten David S. , McNeur J. Clarke
Published: June 2019

In a prospective randomized study designed to compare the effectiveness of intraoperative electrical calf stimulation, perioperative low dosage heparin, and aspirin, either alone or in combination, as prophylactic measures for deep vein thrombosis following total hip replacement, the incidence of thrombosis, as diagnosed by the radioactive fibrinogen uptake test in the study groups was: Control – 9 out of 20 (45%); Calf Stimulation – 9 out of 22 (41%); Aspirin – 9 out of 20 (45%); Heparin – 7 out of 21 (33%); Aspirin plus Calf Stimulation – 10 out of 20 (50%); Heparin plus Calf Stimulation – 0 out of 25.

Following this study only 2 patients developed thrombosis during the period of heparin administration out of a further 104 consecutive patients given low dosage heparin and calf stimulation. Of these patients, thrombosis occurred within 48 hours of cessation of heparin in 11 out of 52 (23%) given heparin until the tenth postoperative day, and in 7 out of 75 (9% ) given heparin until the 14th postoperative day.

Data will be presented to support the following conclusions : 1. The incidence of thrombosis is (i) high (about 50%), (ii) over 50% of thromboses occur either during or soon after operation, (iii) no more frequent in either the operated or non-operated legs. 2. Clinical Diagnosis of thrombosis is grossly inadequate. 3. Prophylaxis is: (i) ineffective with aspirin and calf stimulation, either alone or in combination, or with low dosage heparin alone, (ii) significant with a combination of intraoperative electrical calf stimulation and perioperative low dosage heparin in that the incidence is markedly reduced and the onset delayed the longer the heparin is administered. 4. Low dosage heparin or aspirin does not increase the risk of haemorrhage.
A Patient with Acquired von Willebrand’s Syndrome and Platelet Aggregating Activity in the IgC Fraction
Authors: ten Cate J. W. , Wanders R. , Schneider-Trip M.
Published: June 2019

A new case is described of acquired von Willebrand’s syndrome: bleeding time 20’; platelet-adhesion 20%; F. VIII 9% (procoag. act.); F. VIII R. A. not demonstrable; von Willebrand factor activity (Weiss assay) not demonstrable; absent Ristocetin aggregation. No anti-F. VIII activity could be demonstrated in vitro. Cryoprecipitate infusion resulted in low recovery of f. VIII activity and no f. VIII-synthesis. The serum contained a platelet aggregating activity when tested on normal PHP or washed platelets of 30 random selected donors, and also in PRP of hemophilia A or severe von Willebrand patients. The aggregating activity was present in serum, heated serum (30’ 56° C), BaSO4 absorbed serum, in the IgG fraction isolated both by column chromatography and by ultracentrifugation. Absorption of serum with Vo fraction (sepharose 6B) of normal cryoprecipitate did not remove the aggregating activity. Aggregation was not inhibited by aspirin, adenosine or heparin and was inhibited by EDTA and Cocain and independant of complement. The aggregation was characterized by serotonin and nucleotide release in comparable amounts as thrombin. E. M. studies revealed degranulation and viscous metamorphosis.

The platelet aggregating activity is probably due to platelet antibodies.
Von Willebrand Disease (vWd) Characterized by Increased Ristocetin Aggregation: A Study of Eleven Cases
Authors: Ciavarella N. , Pareti F. I. , Ruggeri Z. M. , Mannucci P. M.
Published: June 2019

The defective ristocetin aggregation occurring in patients with vWd is thought to depend on the decrease of a plasmatic factor related to factor VIII (Willebrand factor, VIIIVWF) 10 patients from 3 families had a mild to moderate bleeding tendency with autosomal dominant pattern of inheritance and laboratory findings suggestive for vWd (decreased levels of antihemophilic factor, and factor- VIII related antigen reduced platelet retention to glass beads columns and prolonged bleeding time). However, ristocetin aggregation in PRP was markedly increased, although VIIIVWF plasma levels were decreased. Aggregation induced in PRP by other agents (such as ADP, adrenaline and collagen), and the release of 14C serotonin was normal; the addition of purified bovine factor VIII was followed by normal aggregation in patients’ PRP and washed platelets. Patients’ washed platelets added to normal plasma aggregated to ristocetin more than normal washed platelets with patients’ plasma; the most marked aggregation response, however, was obtained when patients’ washed platelets were mixed with their own plasma. These findings suggest that a, platelet component is involved in the hyperaggregation response to ristocetin of these patients; however, the interaction of platelets with patients’ plasma is needed to produce the maximum response.

Supported by a grant of the Fondazione Angelo Bianchi Bonomi.
Clinical and Genetic Aspects in 22 Patients with Thrombasthenia
Authors: Seligsohn U. , Reichert N. , Ramot B.
Published: June 2019

Glanzmann’s thrombasthenia has been observed in Israel relatively frequently especially among Iraqi Jews. This paper presents a countrywide survey. Twenty two patients (12 males and 10 females) fulfilled the minimal criteria for establishing the diagnosis of thrombasthenia which were: A lifelong history of a bleeding tendency, a normal platelet count, absent clot retraction and at least 2 of the following: Prolonged bleeding time, isolated platelets on a blood smear and markedly impaired ADP aggregation. The frequencies of bleeding manifestations were: Purpura 100%, monorrhagia 100%, following circumcision 100%, epistaxis and gum 86%, gastrointestinal 68%, hematuria 32% and hemarthrosis 9%. Platelet transfusions seemed beneficial in preventing excessive bleeding in 4 patients who underwent a Caeserian section, 2 circumcisions and extensive abdominal surgery, respectively.

Genetic analysis was possible for 21 patients who belong to 12 unrelated kindreds: 11 Jewish (of whom 10 are Iraqi) and one Arab. Parental consanguinity was present in 9 of sibships. The corrected segregation ratio was 0.20, which is compatible with autosomal recessive inheritance. Twelve obligatory carriers were available for studies. In none of them was it possible to detect impaired platelet functions.
Platelet Factor XIII in Hyperlipidemias
Authors: Scrobohaci M. L. , Stäncioiu N. , Stejänescu N. , Idu S.
Published: June 2019

The total platelet factor XIII–T. P. F. XIII (after destroying by triton) and the released one – R. P. F. XIII (during thrombin induced platelet aggregation) were quantificated in IIa, IIb, IV types of hyperlipemia and myeloproliferative disorders subjects compared to normal control subjects.

T. P, F. XIII was increased in IIb and IV, and normal in IIa types of hyperlipemia while it was decreased in myeloproliferative disorders. R. P. F. XIII was increased only in IIb, “nul” in IIa, IV types of hyperlipemia, myeloproliferative disorders and control subjects too.

Both T. P. F. and R. P. F. XIII increased values in IIb parallel the increased platelet aggregation in these subjects. On the contrary the lack of R. P. F. XIII in IIa and IV types is concordant with normal platelet aggregation revealed in our subjects too. In myeloproliferative disorders when the platelets are “pool-vide” the T, P. F. and R. P. F. XIII is much decreased. These data supported the enhance thrombogenic tendency in IIb type hyperlipemia by platelet increased activity and decreased fibrinolysis due partially to enhanced fibrin stabilization. Other factors incriminated in the decrease of fibrinolysis and increased platelet aggregation, not discussed here, may be responsible for the type IV as thrombogenic as IIb and may counter balance our disordant data in these more prevalent types. The myeloproliferative disorders data are consensual with the tendency to bleeding disorders.
Assay of Antiplatelet Antibodies in Immune Thrombocytopenic Patients by Sedimentation Pattern Test
Authors: Pogliani E. , Cofrancesco E. , Volpe A. Della , Cortellaro M. , Masera G. , Polli E. E.
Published: June 2019

15 sera from never transfused thrombocytopenic children (idiopathic thrombocytopenic purpura, drug induced and post-viral thrombocytopenia) were performed in a microtechnique according to the modified sedimentation pattern test of Myllyla (1). Eighter percent of these sera, tested against a pannel of at least five healthy donors, gave a positive result at different dilutions (from ¼ to ⅙). This quantitative platelet agglutination assay, compared with platelet aggregometry and 14 C-serotonin release, seems to be a sensitive, simple and reliable method to detect antiplatelet antibodies. Therefore it could have clinical applicability to the study of patients affected by immune thrombocytopenia.

(1) Myllyla, G.: Aggregation of human blood platelet by immune complexes in the sedimentation pattern test.

Scand, J. Haemat. suppl. 19, 1973.
Serum Fibrin/ Fibrinogen Degradation Products (FDP) after Experimental Pulmonary Embolism in Dogs
Authors: Jones D. R. B. , Cash J. D. , Owens Rosemary , Dalton R. G. , Ruckley C. V.
Published: June 2019

In previous clinical studies we reported elevated serum FDP values after pulmonary embolism. We detected high values in only 50% of patients and the elevation was transitory. In order to determine the exact time and magnitude of the FDP elevation, and whether the increase was consistent, experimental pulmonary emboli were produced in dogs.

Under pentobarbitone anaesthesia emboli of autologous clot were injected via the femoral vein. FDP, platelets and fibrinogen were estimated on frequent samples taken via a jugular vein catheter. The FDP were measured by the Tanned Red Cell Haemagglutination Inhibition Immunoassay with reagents specific for dog fibrinogen.

It 5 control animals FDP values rose gradually during 8-12 hours of anaesthesia. In 11 dogs receiving 3 ml/ kg of clot the mean FDP rose from 9 ug/ml at rest to 142 ug/ml (Range 9–16100 ug/ml). The peak of FDP was reached in 30-60 minutes and remained significantly above control values for up to 7 hours. These data support our view thai’FDP measurement is valuable in the diagnosis of venous thrombo-embolic disease.
Fibrinogen α-Chain Resistance to Arvin (Ancrod) in DIC
Authors: Lane R. S. , Baugh O. H. , Flude P. T.
Published: June 2019

In high concentrations, Arvin (Ancrod) splits fibrinogen α-chains into fragments of 39,000 and 31,000 molecular weight. In a patient with mild chronic DIC, secondary to a dissecting aneurysm, plasma fibrinogen exhibited marked resistance to normal α-chain proteolysis by Arvin. The patient showed a moderate haemostatic defect acquired from the time of aortic dissection: bleeding was stopped with EACA. Tests were consistent with the presence in plasma of high and low molecular weight FDP, thus deficient haemostasis could be explained by defective fibrin polymerization and X-linking.

The α-chain resistance to Arvin was a coincidental finding. Does this behaviour represent a defect and, if so, is it congenital, or acquired representing a previously unencountered phenomenon associated with DIG? The data show Arvin to be a valuable agent for the rapid isolation and investigation of fibrinogen in normal and pathological plasmas.
Intravascular Acoustic Microstreaming as an Initiator of Mural Thrombi in Vivo
Authors: Williams A. R.
Published: June 2019

Acoustic microstreaming (a rapidly rotating small-scale eddy of blood) is developed inside blood vessels when a discrete portion of the vessel wall is driven to oscillate at an ultrasonic frequency. The wall of the intact vessel was set into motion by the external application of a steel probe (a 115 μm radius wire oscillating transversely at 20 kHz or the 52.5 μm diameter tip of an exponentially tapered velocity transformer oscillating in its longitudinal mode at 80 kHz). The intravascular microstreaming field subjects the vascular endothelium immediately adjacent to the probe tip to large hydrodynamic shear stresses which renders that surface thrombogenic. Simultaneously, platelets are traumatized by the same microstreaming field and are bomarded against the vessel wall where they appear to participate in the formation of mural thrombi. Small blood vessels are completely occluded by platelet plugs which may be stabilized by the formation of fibrin.

This technique produces discrete localized intravascular thrombi by the non-invasive application of hydrodynamic shear stress in the absence of any other thermal, chemical or electrical trauma. It therefore provides a novel in vivo model to investigate the effect of biological and pharmacological variables on thrombus growth and development, and the nature of the repair processes.
On Physiological Regulation of the Liquid State and Coagulation of Blood
Authors: Kudrjashov B. A.
Published: June 2019

The fluidity of blood in a healthy organism is kept by means of reflex reaction initiated by thrombin appearing in blood stream in a threshold concentration. Heparin and activator of plasminogen are excreting in blood as a result of the reflex act. Heparin forms the complexes with fibrinogen, factors XIII, plasminogen, adrenalin and other factors. These compounds act as antiaggregating and antistabilizing agents on the fibrin monomer. They dissolve non-stabilized fibrin, even in the presence of EACA or antiplasmin. Complex plasmin-heparin carries out the enzymatic lysis of stabilized fibrin in the presence of the same inhibitors of fibrinolysis. The clearance of I131-thiombin in blood stream depends from the eomplexing of enzyme with heparin. Complex Thrombin-heparin is absorbed in liver. The accumulation of other heparin complexes take place in the lungs, what is important for quick lysis of fibrin monomer aggregates in the vessels of this organ.
Phospholipase A2 and Competitive Inhibition of Procoagulant Phospholipids
Authors: Boffa M. C. , Boffa G. A.
Published: June 2019

A protein with anticoagulant and phospholipase Aa activities was isolated by chromatography, from Vipera berus venom. The phospholipase activity was demonstrated on isolated platelet phospholipids and derivatives were analysed on thin layer chromatography. The strong anticoagulant activity on recalcified platelet poor plasma, is not dependent upon any proteolytic effect. It is due to a competitive inhibition of the phospholipid procoagulant activity. The inhibition constant has been determined.

The same molecule appeared to be responsible for both activities as demonstrated by continuous gradient chromatography, polyacrylamide gel electrophoresis and electro-focusing.

Inactivation studies permitted to consider that each of both activities was related to a different site on the same molecule.
Complex Role of Fibrinogen A α-Chain in Fibrin Crosslinking and Gel Formation
Authors: McDonagh R. P. , McDonagh Jan , Shen L. L. , Hermans J.
Published: June 2019

The Aα-chain of fibrinogen has several important functions. Its NH2-terminus is the primary proteolytic site for thrombin and most coagulant enzymes. The fragile COOH-terminus, highly susceptible to plasmin proteolysis, renders fibrinogen heterogeneous in plasma. Another important role of the COOH-terminal region is in providing the primary sites of crosslinking of the α-chains of fibrin by factor XIHa, which makes the covalently crosslinked network resistant to proteolysis and increases the elastic modulus of the gel. Degradation of isolated α-polymer and α-chain monomer with cyanogen bromide (CNBr) lias revealed α- single peptide of molecular weight 2,400 which is involved in crosslinking. A core CNBr fragment of approximate m.w. 30,000 is not involved. No other peptide fragments have been detected which participate in α-chain crosslinking. Treatment of fibrinogen at low concentrations with thrombin and small amounts of plasmin results in gels which transiently develop much higher elasticity than in the absence of plasmin. Fibrinogenolysis proceeds to first give proteolysis near the CCOH-termmus of Aα-chains. Assuming that this is true also of fibrinolysis, we see that either splitting of a peptide bond near the COOH-terminus of α-chain or formation of additional isopeptide bonds in this region, may strengthen the structure of fibrin gel.

Supported by Grants HL 14228 and HL 06350 from the National Institutes of Health, U.S.A.
Dextran Induced Changes of the Factor VIII Molecule
Authors: Aberg M. , Hedner U. , Holmberg L. , Bergentz S. E.
Published: June 2019

500 ml of dextran –70 (6% in saline) was given during 30 min to 10 healthy young men. This had no effect on factor VIII-activity (one-stage clotting assay), but resulted in an immediate decrease of factor VIII-related protein (imm. chem. assay). The decrease was maximal 2-4 hours after infusion (initial mean value 111.1%; mean after 4 hours 85.9%; p < 0.001). There was a parallel decrease in platelet adhesiveness (method of Salzman) in two analysed subjects (16% and 14%; initial values 50% and 60% resp.), and a significantly decreased ristocetin aggregation of plasma from all analysed subjects (5) was seen. Factor VIII-activity after gel filtration of plasma showed a different elution pattern after dextran infusion. These findings are in accordance with those seen in a variant fo v. Willebrand’s disease in which an abnormal factor VIII-related protein was demonstra of v. Willebrand’s disease in which an abnormal factor VIII-related protein was demonstrated (Holmberg et al, 1975). Dextran is therefore suggested to interfere with the factor VIII molecule resulting in a defect primary haemostasis.
Inaccurancies of Clotting Tests in Early Diagnosis of Post-Operative Deep Vein Thrombosis (D. V. T.)
Authors: Plante J. , Boneu B. , Gaillard J.
Published: June 2019

Two groups of pneumonectomized patients with diagnosis of pulmonary carcimona were successively studied to assess the diagnostic and predictive values of some clotting test in post-operative D. V. T. The incidence of D. V. T. was determined by the 125I fibrinogen method. Simultaneously blood clotting tests including thromboelastography (T.E.G.), P.T.T. platelet count, fibrinogen level, F. D. P. assay, ethanol test were performed both before and after the surgical procedure on the 2d, 4th, a.nd 7th post-operative days. The first group (32 patients) received subcutaneous Heparin beginning on the third post-operative day. The dose (5000 to 10000 u T. I. D.) was adapted according to the results of the T. E. G. in an efford to obtain an isocoagulable state. The second group (45 patients) received 5000 u of heparin subcutaneously (B. I. D.) with the first injection two hours before the surgical procedure: the dose was adapted on the third day according to the T. E. G.

The results of this study show: (1) The frequency of blood clotting disturbances is similar in both groups in spite of pre-operative heparinotherapy. (2) The first group presented five D. V. T. and two fatal pulmonary embolisms whereas the second group presented only three D. V. T. (P < 0,1). (3) There is no correlation between a positive scanning of the lower limbs and blood clotting disturbances.

In conclusion, clotting disturbances seem to be related to either to surgical procedure or to the carcimona. D. V. T., at early stage, appears to be a local phenomenon which in itself is unable to perturb peripheral blood clotting tests.

(Supported in part by a Grant of Inserm: ATP-12-74-33.)
Deep Venous Thrombosis of the Legs after Strokes: Incidence, Natural History and Predisposing Factors
Authors: Warlow Charles , Ogston D. , Douglas A. S.
Published: June 2019

Forty of 76 patients who had sustained a cerebro-vascular’ accident developed a deep venous thrombosis of the paralysed leg as detected by the 125I-fibrinogen technique. A further 5 also had a thrombosis in the non-paralysed leg. ‘Definite’ pulmonary embolism, diagnosed at necropsy or by unequivocal clinical criteria, occurred in 9.2% of patients and ‘probable’ embolism, diagnosed on clinical examination and chest x-ray only, occurred in another 6.6% of patients Within the limitations of the 125I-fibrinogen technique it appeared that, in patients after cerebrovascular accidents, thrombosis developed in several independent sites in the venous system of the legs and propagation of the thrombi took place in both a distal and proximal direction. Pulmonary embolism occurred, with one exception, only in patients with detectable leg vein thrombosis.

None of the risk factors studied including age, sex, obesity, side of paralysis, past history of venous thromboembolism, neurological and cardiac states, and interval between onset of stroke and mobilisation was helpful in predicting the likelihood of development of venous thrombosis in the paralysed limb.
A Growth Factor – Another Product of the Platelet Release Reaction?
Authors: Busch C. , Westermark B.
Published: June 2019

Platelets have been shown to be a source of a growth factor promoting growth of mouse fibroblasts (Kohlor and Lipton, 1974), monkey arterial smooth muscle cells (Ross et al., 1974) and human glial cells (Westermark and Wasteson, 1974). This study investigates whether the factor is a product of the platelet release reaction or if it is just a component of the cell liberated when it is completely destroyed. Thus human platelet rich plasma was incubated with 14C-serotonin, divided into fractions which were subjected to different releasing substances in different concentrations (thrombin, collagen, ADP and adrenaline). Supernatants of the spun fractions were added to human glial cell cultures and the growth promoting activity measured by 3H-thymidine incorporation.

Results: Growth stimulating activity was found to be released by all inducing substances, most efficiently by thrombin and collagen. The release of this activity showed a linear relationship with the of 14C-serotonin.

Conclusions: Release of a cell growth stimulating factor seems to be a part of the platelet release reaction. Such a mechanism if operating also in vivo could be of importance e.g. in repair of endothelial damage, regulation of endothelial cell turnover as well as in inflammation and repair in a broader sense.
Soluble Fibrin Monomer Complexes Demonstrated by Agarose Gel Filtration and by Adsorption on Insolubilized Fibrinogen. A Comparative Study
Authors: Hugo von R. , Hafter R. , Stemberger A. , Graeft H.
Published: June 2019

Incubation of fibrinogen with small amounts of thrombin resulted in the occurrence of soluble fibrin monomer complexes. These complexes consisted predominantly of a derivative with a higher molecular weight than that of fibrinogen. It was characterized by its relative electrophoretic mobility in 5% PAA gel (0.28 × 10-5 cm2/V×sec) and its elution position prior to the fibrinogen peak following gel filtration. Using adsorption chromatography on insolubilized fibrinogen the derivative dissociated at a ratio of almost 1 : 1 into one part which was adsorbed and into fibrinogen which was not adsorbed. The part which was adsorbed seemed to be the thrombin mediated fibrin monomer. This study confirms the concept that dissociable dimeric fibrinogonfibrin monomer complexes occur after limited action of thrombin on fibrinogen.

(Supported by DFG ; SFB 51, grant no. 2/Gra – B/6.)
Studies on Soluble Fibrin Complexes as a Function of pH
Authors: Benabid Y. , Concord E. , Suscillon M.
Published: June 2019

Purified fibrinogen solutions, incubated with thrombin. CNBr. Sepharose, were subjected to agarose gel chromatography and eluted at different pH (6.5; 7.5; 8.5). Among high molecular weight derivatives formed by thrombin, the major component was a dimer. Gel chromatography at pH 8.5 showed a complexes peak distinct of that from fibrinogen, whereas at pH 6.5, only the fibrinogen peak appeared: fibrin monomer was eluted with fibrinogen as demonstrated by polyacrylamid gel electrophoresis 3.75% pH 8.9. SDS urea electrophoresis after reduction indicated that complexes peak contained two α-chains (α and α′). When fibrinogen was incubated with thrombin in the presence of FSF and calcium, several derivatives with higher and higher molecular weights were formed besides the dimer, and elution profiles of chromatography were identical at pH 6.5 and 8.5, thus indicating stable complexes formation. If fibrinogen-fibrin monomer mixture was subjected to FSF action at different pH, no complexes were formed at pH 6.5. These results confirm that at pH 6.5, any association was prevented.
Specific Patterns of Fibrinogen Proteolysis in Clinical Syndromes Revealed by Fibrinopeptide Assays
Authors: Nossel H. L. , Ti M. , Soland T. , Spanondis K. S. , Canfield R. E. , Butler V. P.
Published: June 2019

Specific radioimmunoassays were used to study fibrinopeptide cleavage from fibrinogen. Thrombin cleaves fibrinopeptide A (FPA) more rapidly than fibrinopeptide B (FPB) whereas plasmin cleaves a peptide with FPB immunoreactivity much more rapidly than FPA. Clinical blood samples manifested several distinct patterns of fibrinopeptide release (mean values in table).

The elevated FPA level is interpreted as signifying increased thrombin action in vivo. A rapid FPA generation rate is thought to signify a uniformly elevated blood thrombin concentration whereas a low FPA generation rate in the presence of an elevated FPA level is thought to signify localized thrombin action. Elevated FPB immunoreactivity is postulated to reflect excessive plasmin action on fibrinogen.
The Effect of 6-Substituted Imidazo(4,5-b)Pyridines on Vitamin K-Dependent Procoagulants in Dogs, Warfarin-Sensitive and Resistant Rats
Authors: Bang N. U. , O’Doherty G. O. P. , Barton R. D.
Published: June 2019

Dogs and warfarin-sensitive rats chronically fed a series of 6-substituted imidazo (4,5-b)-pyridines (6-I[4,5-b]P)

General structure:

died with a picture of generalized hemorrhage. The mechanism of action of these compounds was a selective suppression of the activities of vitamin K-dependent clotting factors II, VII, IX and X. Like warfarin, 6-I(4,5-b)P were significantly more effective in suppressing vitamin K -dependent clotting factor activities in male than in female rats. Rats proven resistant to warfarin also exhibited resistance to 6-I(4,5-b)P. Menadione sodium bisulphite did not antagonize the suppression of vitamin K-dependent clotting factor activities by 6-I(4,5-b)P. 6-I(4,5 -b)P structurally far removed from either vitamin K or the conventional vitamin K antagonists, the coumarins and indandiones may represent important tools to further elucidate the chain of events leading to vitamin K-dependent procoagulant synthesis and in pinpointing the biochemical defect responsible for warfarin resistance.
Heparin and Factor XaI Levels in Patients Undergoing Hip Surgery
Authors: Thomas Duncan P. , Sagar S. , Kakkar V. V.
Published: June 2019

Plasma heparin and activated Factor X inhibitor (Xal) levels were measured in 25 patients undergoing total hip replacement. Blood samples were taken before, during and for 5 days after operation. In patients receiving heparin prophylaxis, over 50% of samples taken four hours after a subcutaneous injection of 5,000 units had no detectable plasma heparin; in none of the remaining samples did the level exceed 0.04 units per ml. In patients who developed deep vein thrombosis (DVT) postoperatively, as detected by 125-I-labelled fibrinogen and confirmed by venography, the mean preoperative level of Factor XaI was 73.5% (S. E. M.±6.9). In those patients who did not develop DVT, the mean preoperative level was 101.5% (S. E.M.±4.3) (P > 0.01).

It is concluded that a regimen of 5,000 units 8-hour’ly does not give sustained plasma heparin levels after total hip replacement, which may in part explain the reduced effectiveness of low-dose heparin in preventing DVT in patients undergoing hip surgery. Low levels of Factor XaI in the immediate preoperative period correlated well with the subsequent development of thrombosis in these patients.
C-W Ultrasound Evaluation of Chronic Peripheral Arterial Disease Treated by i. v. Brinase
Authors: FitzGerald D. E. , Frisch E. P.
Published: June 2019

Fortyfour patients with chronically obstructed peripheral arteries have been treated intravenously with brinase. Twentyfour of these patients had rest pain or gangrene (Stage III or IV) and 20 were claudicants (Stage II).

Assessment before and after treatment was by clinical evaluation, blood pressure measurement in the limbs, and C-W ultrasound scanning of the peripheral arteries (Three arterial segments in each limb). Individual dosage requirement was decided by the level of brinase inhibitors in plasma. The more severely diseased patients were anti coagulated with Warfarin. The follow-up period from treatment extends from six months to three years.

At the end of brinase treatment, good clinical improvement was obtained in 11/20 Stage IT, 10/12 Stage III, and 10/12 Stage IV. The arm-leg blood pressure gradient before treatment in Stage II was 88±41 mmHg, in Stage III 108±43 mmHg, and in Stage IV 129 ±45 mmHg. In patients responding to brinase treatment (27/44) a post-treatment gradient of 44±27 mmHg was recorded. In patients who failed to improve the higher gradient of 81 ±49 mmHg was retained.

C-W ultrasound scanning of 228 arterial segments before treatment showed that 67 segments were occluded, 101 stenosed and 60 segments without significant lesions. Following treatment 19 segments remained occluded, 122 showed stenosis and 87 were without significant lesions. The post-treatment figures indicate a reduction in severity of arterial lesions.
The Role of Calcium Ions in the Activation of Blood Platelets
Authors: Massini P.
Published: June 2019

The plasma membrane of the resting platelet is only slightly permeable to Ca2+-ions. Stimulation of platelets with thrombin or other activators induces an increased influx of 45Ca. The influx occurs simultaneously with the release of serotonin. The “Ca influx is inhibited when the energy supply of the platelets has been interrupted. Stimulation with thrombin increases the efflux of 46Ca in platelets which have been labelled with 45Ca for 24 hours.

Ionophores for divalent cations (X-537 A, A 23187) induce the release reaction, aggregation, clot retraction and rapid shape change. The release reaction does not require external Ca2+-ions whereas clot retraction depends on a Ca2+-eontaining medium.

These results strongly suggest that the reactivity of platelets to external stimuli is primarily mediated by an increase of the cytoplasmic concentration of Ca2+-ions.
Isolation and Properties of Elastase-Like Enzymes from Human Platelets and Pig Aorta
Authors: Hornebeck W. , Legrand Y. , Caen J. P. , Robert L.
Published: June 2019

An elastase-like enzyme has been isolated from human platelets. Its purification using precipitations with ammonium sulphate, gel filtration and affinity chromatography on Agarose-elastin, is described. The acrylamide gel of the affinity peak reveals only one band corresponding to a molecular weight of about 25,000 daltons. The amino acid composition is similar to pancreatic elastase. Using the same kind of purification procedure an aortic elastase-like enzyme has also been isolated and characterized. These two enzymes possess comparable proteolytic activity on various synthetic and natural substrates considered as specific for elastases. The ratio of their activity on these substrates differs however from that of pancreatic elastase. The inhibitory effect of α1, antitrypsine and α2 macroglobuline were also studied and shown to differ quantitatively from those on pancreatic elastase. These elastase like enzymes may be responsible for the degradation of elastin occuring in ageing and arteriosclerosis.