bluuebunny/crossref_metadata_2025 huggingface dataset:
Preferential Degradation of Soluble Fibrin Monomers in Streptokinase-activated Plasma
Authors: Konttinen Y. P , Lalla M.L.T , Turunen O
Published: June 2018

Degradation of 125I-labelled fibrin monomers and 125I-fibrinogen in streptokinase-activated plasma was investigated. Release of the label proceeded at a much higher rate from soluble fibrin monomer complexes than from fibrinogen until ca. 70% of the label was recovered in the serum. It is concluded that the fibrinolytic system has an essentially higher affinity to monomers than to fibrinogen, but that the specificity of the monomers as substrate may be limited to the early steps of degradation. It is suggested that the preferential splitting of fibrin monomer complexes may be important in the body’s defence against pathological fibrin formations derived from paracoagulable fibrin.
Leucocytes and Thrombosis
Authors: Banks D.C , Mitchell J.R.A
Published: June 2018

White cell loss from a rotating flask system is enhanced by noradrenaline, adrenaline, isoprenaline, 5HT and tendon extracts. Noradrenaline is the most active agent in this test of white cell behaviour and unlike the other materials its action is not blocked by an ADP-removing system.

Adenosine inhibits both platelet loss and white cell loss, but concentrations can be found which selectively block the former.

The implications of these findings for our understanding of the mechanism of polymorphonuclear leucocyte involvement in thrombus formation are discussed, but the role of white cells once they have become involved in thrombus formation has not yet been determined.
Quantitative Determination of Total Antithrombin III in Plasma
Authors: Yue Robert H. , Starr Toby , Gertler Menard M.
Published: June 2018

A relatively simple method has been developed to evaluate the total amount of antithrombin III in plasma. Since purified and stable bovine thrombin was readily prepared in our laboratory, it was selected to be used in our assay system. Purified α2-macroglobulin was found to inactivate thrombin following a pseudo first-order rate of reaction. However, purified antithrombin III inactivated thrombin following a bimolecular type of reaction. The second-order rate constant for antithrombin III and thrombin interaction was found to be 94 1/M/sec. under these conditions. A least-square straight line was drawn between 15 and 90 minute intervals and the intercept yielded the total amount of antithrombin III expressed NIH thrombin units. This assay system and the graphical analysis yielded the total amount of antithrombin III in plasma quantitatively. The results are reproducible.
Blood Coagulation Inhibitor in a Snake Plasma (Bothrops jararaca)
Authors: Nahas Linda , Betti Ferruccio , Kamiguti Aura S. , Sato Hissae
Published: June 2018

Experiments with plasmas samples from the snake B. jararaca and X. merremii have shown that the two differ markedly with respect to their reaction to bovine thrombin. The plasma of X. merremii appears to react in a manner very similar to that of normal human plasma both using the thrombin clotting test and using the antithrombin III assay of Astrup and Darling (Biggs and Macfariane 1962). The plasma of B. jararaca on the other hand, contains, in addition to antithrombin III, an inhibitor which resembles mammalian heparin. This inhibitor prolongs the thrombin clotting time of human and X. merremii plasmas, is recorded by the antithrombin assay and is neutralized by protamine. The inhibitor is however, not identical with mammalian heparin since it is not adsorbed by BaSO4 and Al(OH)3 (which adsorb heparin) and it is heat labil whereas mammalian heparin is heat stable.
Evaluation of a New Preparation of Urokinase
Authors: Prentice C.R.M , Rogers K.M , McNicol G.P
Published: June 2018

The pharmacological effect of a new preparation of urokinase (Leo) has been studied, both in vitro and in six patients suffering from thrombo-embolic disorders. It was a non-toxic, effective fibrinolytic agent if given in sufficient dosage. A regimen consisting of an initial dose of 7,200 ploug units per kg body weight, followed by hourly maintenance therapy with 3,600 ploug units per kg intravenously, gave satisfactory evidence of whole body fibrinolytic activity. The preparation had minor but insignificant thromboplastic activity both when assayed in the laboratory and when given to patients.
The Filter Loop Technique as a Method of Measuring Platelet Aggregation in the Flowing Blood of the Rat; the Inhibitory Activity of 5-oxo-l-cyclopentene-l-heptanoic Acid (AY-16,804) on Platelet Aggregation
Authors: Muirhead Christopher R.
Published: June 2018

The filter loop technique which measures platelet aggregation in vivo in the flowing-blood of the rat was compared to the optical density technique of Born which is carried out in vitro with platelet rich plasma. Using these two experimental models the effect on platelet aggregation of three known inhibitors sulfinpyrazone, dipyridamole and prostaglandin E1, and a novel compound 5-oxo-l-cyclopentene-l-heptanoic acid (AY-16, 804) was determined.

The effects on platelet aggregation of the known inhibitors were consistent with information in the literature. Prostaglandin E1 was the most potent inhibitor in both techniques; sulfinpyrazone inhibited aggregation in both models but was less potent than prostaglandin E1. AY-16, 804 exhibited activity in vitro and in vivo similar to that of sulfinpyrazone. Dipyridamole did not inhibit platelet aggregation in vivo and did not inhibit aggregation in vitro in concentrations at which it remained soluble.

The filter loop technique is a suitable model for measuring platelet aggregation in the flowing blood of the rat. It is a relatively simple method of determining aggregation and easily adapted to other species.
The Migration of Human Platelets in Vitro
Authors: Nathan Paul
Published: June 2018

A simple procedure is described for measuring migration of human platelets in vitro. Migration increased with temperature reaching maximum values at approximately 30 °C. Addition of normal plasma to the incubation chambers increased migration in all tests and the increase reached twice the control values in 3 of 4 tests. The rate of migration in vitro was relatively rapid for the first 12 hrs. of incubation and then continued at a slower rate for at least an additional 12 hours.
Impairment of Human Platelet Aggregation and Serotonin Release Caused in Vitro by Echis Colorata Venom
Authors: Biran Haim , Dvilansky Alexander , Nathan Ilana , Livne Avinoam
Published: June 2018

Aggregation of washed human platelets, induced by either ADP, thrombin or collagen, was decreased by Echis colorata venom (EVC). With ADP as an inducer, the inhibition of aggregation was proportional to the venom concentration, starting from 0.27 μg/ml and attaining full inhibition with venom concentration of 9 μg/ml. Higher concentrations were required for comparable venom effects when collagen or thrombin were used as inducers. Based on serotonin release measurements and platelet counting, it is concluded that the ECV-diminished aggregation is not due to platelet lysis. Thrombin-dependent serotonin release was inhibited by the venom to an extent proportional to the log ECV concentration at a range of 0.27 to 90 μg/ml. ECV effeces on serotonin release are apparently independent on its effects on aggregation, since similar results were obtained either with or without EDTA.

Endothelial damage and defibrination are already known to be associated with the bleeding tendency caused by ECV. The present data disclose a functional impairment of platelets as an additional antihemostatic effect of this venom.
Anticoagulant Factors from Viperidae Venoms
Authors: Boffa Marie Claire , Delori P , Soulier J. P
Published: June 2018

Two proteins presenting anticoagulant activities were isolated from Viperidae venoms : one from Vipera aspis, the other from Vipera berus. They interfere with the formation of prothrombinase. Their inhibitory effect results from a specific action of a complex which blocks the active sites of phospholipids. This complex is easily dissociated by antivenoms allowing normal clot formation.

Berus inhibitor possesses a high phospholipase activity while aspis inhibitor is completely devoid of this activity. The anticoagulant action of these venom factors does not appear to be enzymatic.

The affinity of both inhibitors for phospholipids seems similar though aspis inhibitor shows a more marked anticoagulant action which is neutralized by the addition of a greater amount of phospholipid.

The study of these venom inhibitors confirms the fact that, in the absence of platelet phospholipids, clot formation is not possible.
Clinical Studies Concerning Factor XIII; with Special Reference to Hyperlipemia
Authors: Cucuianu Mircea P. , Vasile Vasile V. , Popescu Titus A. , Opincaru Augustin , Crîsnic Ion , Tapalagå Daniela
Published: June 2018

Factor XIII, estimated according to the method of Heene, was found to be diminished in patients with cirrhosis of the liver, in acute renal failure and, to a lesser extent, postoperatively, owing probably to both a decreased synthesis and an accelerated utilization. When compared to values obtained in healthy, normal weight, normolipemic control subjects, the level of factor XIII was higher in hyperlipemic and especially in hypertriglyceridemic patients, whether clinical atherosclerosis was present or not. Low levels of factor XIII were usually accompanied by a decrease in serum pseudocholinesterase, while high values of this enzyme produced by the liver were noted in hypertriglyceridemic patients. The hypothesis of an enhanced synthesis of factor XIII by the liver, in subjects with endogenous hypertriglyceridemia is considered. It is also suggested that accelerated stabilization of fibrin might contribute to the fibrinolytic insufficiency of hyperlipemic patients and enhance certain atherogenic mechanisms.
Effects of Puff Adder Venom on the Coagulation Mechanism
Authors: Phillips Louise L. , Weiss H. J , Christy N. P
Published: June 2018

Studies of the in vitro effects of puff adder venom (Bitis arietans) on the coagulation system of human plasma have suggested this venom activates factor XI. Incubation with the venom results in a prolongation of the partial thromboplastin times in glass which is associated with a depletion of factors V, VIII, and fibrinogen. The latter may be due to the proteolytic properties of the venom. Soluble fibrin monomer complexes are formed rapidly after addition of the venom to normal plasma. The venom also causes platelet aggregation which is not inhibited by heparin.
Controlled-flow Instruments for Simulating in Vivo Thrombosis
Authors: Clark H. G , Shinoda B. A , Mason R. G
Published: June 2018

A technique has been developed which allows an estimation of the clotting tendency of blood under conditions which simulate in vivo flow in various regions of the circulatory system. Minimum contact between blood and foreign surfaces other than the walls of the tubular test cell is obtained. Early pre-clotting and clotting events which alter the apparent viscosity of blood are resolved within three seconds of their onset. Increases in apparent viscosity slow the oscillations of the test system, and frequency, differential frequency, or amplitude can be monitored continuously. A standard tubing material may be used to evaluate effects of antithrombotic or other agents. For the present studies, the instrument was used with native blood at 28° or 37° C to produce shear rates comparable to those present in venous flow. Gum rubber, vinyl, glass and silicone tubing have been investigated for their effects on activation of the intrinsic coagulation system of human blood. In order of increasing-time to formation of thrombus are glass, vinyl, rubber, silicone. Thrombi were adherent to glass and rubber but not to Tygon or Silastic tubes. Time sequence studies during flow in silicone tube with native blood indicate progressive shortening of Stypven and partial thromboplastin time values, and progressive decrease in leukocyte and platelet counts. Addition of a commercial bovine collagen preparation to native normal blood in Silicone tubing decreased the thrombus time value by 40% while addition of the experimental agents RA 233 or VK 744 decreased this value by 50% and 70% respectively. Aspirin therapy produced a 158% increase in thrombus time values. Light microscopic studies of thrombi formed in this closed system revealed thrombus structure comparable to that of classic venous thrombi.
Effect of Corticosteroids upon Fibrinogen Metabolism in Rabbits
Authors: Seligsohn Uri , Rapaport Samuel I. , Shen Stephen M.-C. , Kuefler Paul R.
Published: June 2018

The effect of pharmacologic doses of adrenal corticosteroids upon plasma fibrinogen level, incorporation of 75Se-Methionine (75SeM) into fibrinogen, and disappearance of plasma 125I-fibrinogen was evaluated in rabbits. Single 25 mg doses of hydrocortisone or cortisone transiently increased both 75SeM incorporation into fibrinogen and plasma fibrinogen levels, but 1 to 5 mg doses of hydrocortisone had no significant effect upon plasma fibrinogen levels. Repeated daily injections of 25 mg of hydrocortisone did not increase plasma fibrinogen levels progressively. Two doses of 25 mg of hydrocortisone, given 24 hr apart, failed to alter the rate of disappearance of plasma 125I-fibrinogen radioactivity, i.e., plasma fibrinogen catabolism. Intramuscular injection of 2.5 ml of propylene glycol solvent caused a prolonged rise in plasma fibrinogen level, whereas injection of 25 mg of corticosterone dissolved in propylene glycol resulted in an attenuated and transient rise in plasma fibrinogen level. Apparently, the presence or absence of a coexistent inflammatory stimulus determines the overall effect of large pharmacologic doses of corticosteroids upon fibrinogen metabolism in the rabbit. In the presence of inflammation, corticosteroids decrease synthesis with a resultant fall in plasma fibrinogen level; in the absence of inflammation, corticosteroids may increase synthesis transiently with a resultant slight increase in plasma fibrinogen level.
Effect of Aspirin and Benadryl on Human Platelet Oxidative Phosphorylation and Aggregation
Authors: Yue Kenneth T. N. , Davis James W. , Aldridge E. Gray
Published: June 2018

Tightly coupled oxidative phosphorylation in washed human platelets and in lysates has been demonstrated. Both aspirin (25 μM) and Benadryl (0.15 mM) inhibited phosphorylating oxidation but had no effect on uncoupled respiration. It is likely that they were not involved in the electron transport chain but specifically affected the phosphorylation reactions similar to the action of oligomycin. Preincubation of Benadryl (1.3 mM) with platelet-rich plasma (PRP) at room temperature for 2 min inhibited ADP-induced platelet aggregation by 62% and abolished the second phase of aggregation. Addition of Benadryl to aggregating platelets in PRP during ADP-induced aggregation caused immediate disaggregation whether added during the first or second phase of aggregation. Aspirin had no such effect.
Technical Remarks on the Microscopical Demonstration of Tissue Plasminogen Activator
Authors: Pandolfi M , Bjernstad A , Nilsson Inga Marie
Published: June 2018

The authors examine some variables capable of influencing the results obtained with the fibrin slide technique for microscopical demonstration of fibrinolytic activators in tissues (Todd’s method).

The speed of freezing of the tissue specimen proved to be especially important. Slow freezing of the specimen should be avoided since it results in sections of poor quality and may alter the distribution of fibrinolytic activity. The optimal thickness of the sections lies between 6 and 8 (jim. The concentration of fibrinogen used as substrate may vary between 0.7 and 2.0% and the thickness of the fibrin film between 0.07 and 0.21 mm. Increasing the temperature of incubation from +4° C to +37° C enhanced fibrinolysis, but had no effect on the distribution of fibrinolytic activity in the sections. Storage of the sections at —20° C and at +4° C for 6 days had no apparent effect on their fibrinolytic activity. Storage at 28° C resulted in a considerable loss of the fibrinolytic activity of the sections. The error of a method to assess the strength of fibrinolytic activity of the section is determined.
Gerinnungsphysiologische Analyse der Wirkung synthetischer Thrombininhibitoren
Authors: Maekwaedt F
Published: June 2018

Es wurde der Einfluß von jeweils einem Vertreter aus der Gruppe der kompetitiv, der substratartig und der irreversibel wirkenden synthetischen Inhibitoren für Serinproteinasen auf den Ablauf der Blutgerinnung analysiert. Demnach läßt sich die Gerinnung nur von den kompetitiv wirkenden Inhibitoren aufhalten, deren Reaktionsgeschwindigkeit die der Ferment-Substrat-Reaktion erreicht. Ihre Wirkung kommt durch eine Hemmung des Gerinnungsfermentes Thrombin zustande, die sich nicht nur auf die Fibrinogenumwandlung beschränkt, sondern auch auf andere Wirkungen des Thrombins im Gerinnungssystem, wie die Auslösung der Plättchenaggregation und die Reaktion mit dem Antithrombin des Blutes, erstreckt.
Platelet Aggregation in Diabetes Mellitus and the Effect of Insulin in Vivo on Aggregation
Authors: Hassanein A. A , El-Garf Th. A , El-Baz Z
Published: June 2018

ADP-induced platelet aggregation and calcium-induced platelet aggregation tests were studied in 14 diabetic patients in the fasting state and half an hour after an intravenous injection of 0.1 unit insulin/kg body weight. Platelet disaggregation was significantly diminished as compared to a normal control group, and their results were negatively correlated with the corresponding serum cholesterol levels. Insulin caused significant diminution in the ADP-induced platelet aggregation as a result of rapid onset of aggregation and disaggregation. There was also a significant increase in platelet disaggregation. In the calcium-induced platelet aggregation test, there was a significant shortening of the aggregation time, its duration, and the clotting time. The optical density fall due to platelet aggregation showed a significant increase. Insulin may have a role in correcting platelet disaggregation possibly through improvement in the intracellular enzymatic activity.
Some Effects of Calcium and Strontium Ions on Three Basic Reactions of Blood Coagulation
Authors: Sakuragawa Nobuo , McCoy Lowell E. , Seegers Walter H.
Published: June 2018

Calcium ions accelerated all three basic reactions of blood coagulation at an optimum concentration of 0.01 M. Strontium ions were practically ineffective as substitutes for calcium ions in the formation of autoprothrombin C (factor Xa), but functioned just as effectively as calcium ions in the formation of thrombin. Inhibition was the primary effect of strontium ions in fibrin formation. It is suggested that calcium ions function differently in each reaction. Echis carinatus venom was found to form thrombin directly from its precursor and, likewise, formed autoprothrombin C from its precursor.
Quantitative Monitoring of Intravascular Coagulation and Fibrinolysis in the Lungs of Rats
Authors: Coombey D , Tyler H. M
Published: June 2018

A method is described whereby the arrival and subsequent lysis of thrombin- induced, 125I-labelled, pulmonary emboli can be quantitatively monitored in the rat. A scintillation probe placed over one lung monitors radioactivity in that organ ; serial counts or a continuous recording of radioactivity can be made. Radioactivity counted in serial blood samples also reflect the defibrination and fibrinolytic processes. The influence of the inhibitor EACA, and the lytic agent Brinase are described.
Glutaraldehyde Fixation of Human Blood Platelets
Authors: Sixma J. J , Linssen W. H , Geuze H. J
Published: June 2018

Hypertonic glutaraldehyde solutions produce dilatation of the surface connected system and shrinking of the cytoplasma. Hypertonic solutions produce swelling of the platelets. Good results are obtained with 1% glutaraldehyde M 0.07 M phosphate buffer (275 mosmol) 4 parts to 1 part platelet rich plasma. High glutaraldehyde concentrations do not affect the platelet ultrastructure.
The Effect of Alpha- and Beta-Receptor Blocking Agents on Collagen-Induced Platelet Release Reaction (A Comparison with ADP-Induced Release)
Authors: Kubisz P , Suranova J
Published: June 2018

The mechanism of the platelet release reaction, induced by collagen, is in certain respects different from that of ADP. Besides other facts, the PF 4 release and the availability of PF 3 and APh are not influenced by the concentrations of alpha or beta-receptor blocking agents which clearly interfere with the ADP-induced release.

The possible role of thrombosthenin (or its alpha-receptors in the participation of the release reaction is discussed.
The Dual Sample Aggregometer
Authors: Gerloff J , von Kaulla K. N
Published: June 2018

An improved platelet aggregometer is described which permits the simultaneous recording of two aggregation curves obtained under technically strictly identical conditions which can be recorded with any single pen recorder. The curves are superimposed thus facilitating the visual or mathematical assessment of differences in the aggregation pattern. The instrument is particularly suitable for serial testing of aggregation inhibiting agents. It is equally suitable in the conventional way as any single channel aggregometer for routine testing of platelet aggregation.
Disseminated Intravascular Coagulation Induced by Liquoid in Normal, Thrombocytopenic and Granulocytopenic Rabbits
Authors: Skjørten F , Evensen Stein A.
Published: June 2018

Disseminated intra vascular coagulation was induced by the synthetic acidic polymer Liquoid (sodium polyanethol sulfonate) in normal rabbits, in rabbits with a combined thrombocytopenia and granulocytopenia induced by busulphan, and in rabbits with a selective granulocytopenia induced by nitrogen mustard. One hour after the injection of Liquoid, light microscopy showed deposits with the staining properties of fibrin in glomerular capillaries and hepatic sinusoids of animals from all pretreatment groups. After 2 h, there was an increase in the deposits in all groups, most marked in rabbits with normal platelet and leukocyte counts prior to Liquoid injection.

By electron microscopy, normal animals injected with Liquoid showed predominantly fibrillar deposits in glomerular and hepatic capillaries. The center of some of these deposits was composed of granular masses. Animals with combined thrombocytopenia and granulocytopenia, or with selective granulocytopenia showed purely granular deposits.

We conclude that both types of deposits are derived from fibrinogen. The fibrillar material is assumed to be polymerized fibrin, whereas the granular material may possibly represent complexes formed between fibrin monomers and other plasma proteins, including fibrinogen precipitated by Liquoid.
Studies of the Metabolism and Distribution of Prothrombin in Healthy Men with Homologous 125I-Prothrombin
Authors: Takeda Y
Published: June 2018

A method was described for preparation of purified human prothrombin using repeated barium citrate adsorption and citrate elution, (NH4)2S04 fractionation, Sephadex chromatography and isoelectric precipitation. The produced prothrombins were then physicochemically analyzed. The extinction coefficient was 12.7 ± 0.82 (SD) and the specific activity, 2,563 ± 420 (SD) Iowa units per mg prothrombin. The amino acid composition in g% was as follows: aspartic acid, 8.64; threonine, 5.87; serine, 4.16; glutamic acid, 11.15; proline, 3.91; glycine, 3.96; alanine, 4.65; half cystine, 4.68; valine, 4.46; methionine, 1.14; isoleucine, 2.93; leucine, 6.18; tyrosine, 4.52; phenylalanin, 4.19; lysine, 4.78; histidine, 1.73; ammonia, 1.85; arginine, 6.91. The tryptophan and carbohydrate contents were not determined. The prothrombins were then labeled with 125I in a ratio of about 0.25 atom iodine per molecule of prothrombin and were used for studies of the metabolism and distribution of prothrombin in thirteen healthy men. A model was proposed for the human prothrombin system based on the assumptions that prothrombin is catabolized intravascularly without the formation of thrombin, and was tested against the observed behavior of injected 125I- prothrombin. The model closely predicted the observed behavior of 125I-prothrombin. The analytical results of the tracer data in thirteen healthy men by the methods derived from the model were as follows: The plasma prothrombin, 6.65 ± 1.10 (SD) mg per kg; the interstitial prothrombin, 6.0 ^ 1.9 (SD) mg per kg; the half-life of plasma 125I-prothrombin, 2.29 ± 0.14 (SD) days ; the transcapillary transfer rate of prothrombin, 10.2 ± 3.9 (SD) mg per kg per day; and the catabolic (synthetic) rate of prothrombin, 4.3 J: 1.0 (SD) mg per kg per day. These results support the concept that in healthy men prothrombin is degraded in plasma or spaces in rapid exchange with it generation of appreciable amounts of thrombin.
The Effects of Bile Salts on Some Coagulation Components and Related Enzymes
Authors: Engel A. M , Alexander B
Published: June 2018

Certain purified bile salts, individually or in a mixture, profoundly affect - either inhibiting or enhancing - the esterolytic activities of thrombin, trypsin, and plasmin, the clotting activity of thrombin, and caseinolysis by trypsin. They also promote SK-induced fibrinolysis and impair FI clottability. These effects are directly related to bile salt concentration but not to the critical micellar concentration.

A very unusual effect was observed with deoxycholate and FI : besides inhibiting FI clottability, the salt induces spontaneous gelation. In addition, it binds strongly to the protein, as has been already reported for another plasma protein, albumin, and to a lesser degree, to alpha- and beta-globulins.

Noteworthy is the fact that activation of pancreatic juice trypsinogen by thrombin also was increased by prior thrombin exposure to the salts. On the other hand, thrombin-induced platelet aggregation was slightly inhibited by the bile salt mixture, which, when added to PRP moderately inhibited the ADP-induced aggregation. No effect was observed on the conversion of F II in plasma via the thromboplastic mechanism when deoxycholate, or cholate, or glycocholate was added to the system.

It is postulated that the bile salt mixture enhances SK-induced fibrinolysis by direct action, either on SK, or on the SK-activator complex, attributable to the detergent properties of the salts.

The physiologic and pathologic implications of our results with respect to hemostasis and pancreatitis are discussed.
Identification of Fibrinogen Derivatives in Plasma Samples
Authors: Graeff H , von Hugo R
Published: June 2018

The observation of fibrinogen derivatives with a molecular weight higher than the parent molecule in human cases of DIC initiated the present methodological study. These derivatives were identified by the following methods : 2.5 M β-alanine precipitation of the plasma samples, PAA gel electrophoresis, intra gel immunoprecipitation and agarose gel chromatography. In the plasma of a patient with severe eclampsia and laboratory signs of DIC two derivatives with a molecular weight higher than that of fibrinogen were identified according to their relative electrophoretic mobilities: 0.18 and 0.28 × 10−5 cm2/V × sec (fibrinogen: 0.43 × 10−5 cm2/V × sec). Electrophoretic studies in the presence of 5 M urea indicated that the 0.28 derivative is a complex probably formed by fibrinogen and a fibrin monomer.
An Evaluation of Impedance Plethysmography and 125I-Fibrinogen Leg Scanning in Patients following Hip Surgery
Authors: Cruickshank M K , Levine M N , Hirsh J , Turpie A G G , Powers P , Jay R , Gent M
Published: June 2018

Venous thromboembolism is a common post-operative complication in patients following hip surgery. 125I-fibrinogen leg scanning and impedance plethysmography (IPG), are often used in the detection of venous thrombi in such patients. Information on the sensitivity and specificity of these non-invasive tests for the diagnosis of venous thrombosis following hip surgery is relevant for both patient management and for choosing the appropriate outcome measure for clinical trials evaluating new prophylactic regimens.

We determined the sensitivity and specificity of the IPG alone, the 125I-fibrinogen leg scan alone, as well as the combined use of the two tests from a retrospective analysis of 685 hip surgery patients who participated in clinical trials of anti-thrombotic prophylaxis. These patients were followed prospectively with non-invasive tests. Bilateral venography was attempted either when one or both screening tests became positive or on day 10-14 post-operatively if both screening tests remained negative. Adequate venography was obtained in 1,010 (73.7%) legs and thrombi were identified in 198 (19.6%) legs.

The sensitivities of the IPG and leg scanning were 12.9% and 44.6% respectively; the corresponding specificities were 98.1% and 95.0%. The sensitivity of a positive result on one or both screening tests was 49.6% with a specificity of 93.9%.

Therefore, leg scanning and IPG, even in combination, are not sufficiently accurate to be recommended as the only strategy for the diagnosis of venous thrombosis following hip surgery. Venography should be considered in all patients undergoing surveillance testing either when one or both of the screening tests become positive or on day 10-14 if the screening tests remain negative.
The Polymerization of Fibrin Prepared from Fibrinogen Haifa (γ275Arg→His)
Authors: Siebenlist Kevin R , Mosesson Michael W , Di Orio James P , Tavori Shulamith , Tatarsky lllana , Rimon Abraham
Published: June 2018

Fibrinogen Haifa is a congenital heterozygous fibinogen variant (γ275 Arg→His) characterized by prolonged thrombin and reptilase times and normal fibrinopeptide (FPA, FPB) release. We compared the polymerization rate (by turbidity measurements at 350 nm) and the ultrastructure of Haifa α-, β-, and α, β-fibrin with that of normal. Haifa α, β-fibrin polymerized less rapidly than did normal and formed a highly branched matrix with a smaller mean fiber diameter; this network closely resembled that of normal α, β-fibrin with EDTA added. In the presence of CaCl2 (1 to 10 mM), Haifa α, β-fibrin polymerized more rapidly than in buffer alone and possessed a matrix structure closely resembling that of normal fibrin. From these observations it appears that the functional defect in Haifa fibrin can be related to the inability of the abnormal molecule to effectively utilize available calcium. The polymerization profile of Haifa α-fibrin differed only modestly from that of normal α-fibrin, whereas that of Haifa β-fibrin was markedly impaired. This finding plus similarities in the ultrastructure of Haifa and normal α-fibrin specimens suggests that the defective γ chain structure of Haifa fibrinogen results in greater impairment of the carboxy terminal “b” polymerization domain reacting with the site exposed by cleavage of FPB (“B” site) than it does that of the carboxy terminal “a” domain reacting with the site exposed by cleavage of FPA (“A” site). Whether this effect is due to absolute differences in the degree of impairment of these two types of polymerization sites, or whether proper utilization of the “B” to “b” site is dependent upon participation of the “A” to “a” site remains to be determined.
Procoagulant Activity of T Lymphoblastoid Cells in Extrinsic and Intrinsic Coagulation Systems
Authors: Barrowcliffe T W , Marshall Dianne E , Trickett Lynne P , Hubbard A R , Gray Elaine , Thorpe R
Published: June 2018

We have measured the procoagulant activity (PCA) of four T lymphoblastoid cell lines (Jurkat, CEM, HSB-2 and Molt 4) as well as normal peripheral blood T lymphocytes, before and after stimulation with phytohaemagglutinin (PHA), using clotting and amidolytic methods.

Of the four cell lines only one, Jurkat, gave enhanced PCA after stimulation with PH A. This activity was shown to be tissue factor-like by its dependence on factor VII in plasma and in an amidolytic assay with purified factors VII and X. Jurkat was also the only one of the four cell lines to secrete interleukin-2. All four cell lines promoted the generation of large amounts of thrombin in platelet-free plasma in glass tubes. This activity was dependent on the presence of plasma factor VIII, and was probably due to phospholipids in the cell membranes. Normal T lymphocytes gave intrinsic PCA in the thrombin generation test which was only 15% of that of the lymphoma cells.

These results show that some T lymphocytes can develop PCA in both intrinsic and extrinsic systems and this should be taken into account in studies of the PCA of mixed leukocyte populations.
Distinguishing Plasma Lupus Anticoagulants from Anti-Factor Antibodies Using Hexagonal (II) Phase Phospholipids
Authors: Rauch Joyce , Tannenbaum Marion , Janoff Andrew S
Published: June 2018

The association of lupus anticoagulant antibodies with thrombosis, thrombocytopenia, and multiple spontaneous abortions underlines the importance of diagnostic assays which are able to distinguish these antibodies from anti-factor antibodies and factor deficiencies, as all three prolong in vitro coagulation assays measuring activated partial thromboplastin time (APTT). Heparin also prolongs the APTT assay and often interferes with the detection of lupus anticoagulant activity. The present study describes a direct and simple dilute APTT assay in which plasma is preincubated with hexagonal (II) phase phosphatidyl ethanolamine (PE). Using this system, the lupus anticoagulant antibody activity of 10 randomly selected plasmas from SLE patients was inhibited by 81.2-99.5%, while prolongation of the APTT assay by 6 different anti-factor antibody-containing plasmas, 5 factor deficient plasmas, and 6 heparin-containing plasmas remained unaffected. Inhibition was dependent on epitopes exposed when PE was presented in the hexagonal (II) phase. This data suggests that hexagonal (II) PE is specifically recognized by lupus anticoagulant antibodies in SLE patients and may play a role in the etiology of the disease.
Molecular Forms of Human Protein C: Comparison and Distribution in Human Adult Plasma
Authors: Greffe Brian S , Manco-Johnson Marilyn J , Marlar Richard A
Published: June 2018

Protein C (PC) is a vitamin K-dependent protein which functions as both an anticoagulant and profibrinolytic. It is synthesized as a single chain protein (SC-PC) and post-transla-tionally modified into a two chain form (2C-PC). Two chain PC consists of a light chain (LC) and a heavy chain (HC). The present study was undertaken to determine the composition of the molecular forms of PC in plasma. PC was immunoprecipitated, subjected to SDS-PAGE and Western blotting. The blots were scanned by densitometry to determine the distribution of the various forms. The percentage of SC-PC and 2C-PC was found to be 10% and 90% respectively. This is in agreement with previous work. SC-PC and the heavy chain of 2C-PC consisted of three molecular forms (“alpha”, “beta”, and “gamma”). The “alpha” form of HC is the standard 2C form with a MW of 40 Kd. The “beta” form of HC has also been described and has MW which is 4 Kd less than the “alpha” form. The “gamma” species of the SC and 2C-PC has not been previously described. However, its 3 Kd difference from the “beta” form could be due to modification of the “beta” species or to a separate modification of the alpha-HC. The LC of PC was shown to exist in two forms (termed form 1 and form 2). The difference between these two forms is unknown. The molecular forms of PC are most likely due to a post-translational modification (either loss of a carbohydrate or a peptide) rather than from plasma derived degradation.
The Relationship between Anti-Factor Xa Level and Clinical Outcome in Patients Receiving Enoxaparine Low Molecular Weight Heparin to Prevent Deep Vein Thrombosis after Hip Replacement
Authors: Levine M N , Planes A , Hirsh J , Goodyear M , Vochelle N , Gent M
Published: June 2018

Studies in experimental animals have demonstrated that there is a relationship between levels of low molecular weight (LMW) heparin and both bleeding and inhibition of thrombosis. The relationship between these outcomes and ex vivo anti-factor Xa levels has been examined in 163 patients undergoing total hip replacement who were given prophylaxis once daily with a LMW heparin (enoxaparine). Fifty patients received 60 mg of enoxaparine and 113 received 40 mg, both regimens being administered subcutaneously once daily. Blood samples for anti-factor Xa levels were collected 12 hours after the injection on the day of surgery and on days 1, 3 and 6, postoperatively. The incidence of wound hematoma was 5.3% when the maximum anti-factor Xa level was ≤0.2 units per ml, but increased to 24.5% when the anti-factor Xa level exceeded 0.2 units per ml, P = 0.0008. The incidence of postoperative thrombosis was low (6.3%) if the minimum anti-factor Xa level exceeded 0.1 units per ml, but increased to 14.6% when ≤0.1 units per ml, and to 18.8% if the anti-factor Xa level was ≤0.05 units per ml. Regression analysis revealed that there was a statistically significant relationship between anti-factor Xa level and wound hematoma, P = 0.002 and anti-factor Xa level and thrombosis, P = 0.03. These findings suggest that when enoxaparine is administered as a once daily subcutaneous injection, the 12 hour anti-factor Xa level should not exceed 0.2 units per ml to minimize bleeding and levels >0.05 units per ml should be obtained to optimize efficacy.
The Effect of Calcium Chloride on Anti-Xa Activity of Heparin and Its Molecular Weight Fractions
Authors: Barrowcliffe T W , Shirley Yvonne Le
Published: June 2018

The anti-Xa activities of unfractionated heparin (UFH) and nine low molecular weight heparins (LMWH) have been measured in the presence and absence of 3 mM CaCl2, using bovine Factor Xa, purified human AT III and an amidolytic assay. The addition of CaCl2 increased the activity of UFH by 93%, but the effect on LMWH was less, ranging from −20% to +55%. Studies of gel filtration fractions of UFH showed marked Mr dependence of the CaCl2 effect in the range 4,000-12,000. The differences among the various LMW heparins with respect to the effect of CaCl2 were closely correlated with the amount of polysaccharide above an Mr of 6,500. Kinetic studies confirmed the potentiation of the activity of UFH with bovine Xa and showed an even more marked effect using human Xa.
Differential Ability of Agonists to Express Distinct Pools of Fibrinogen (Gpllb/llla) Receptors which Can Mediate the Aggregation of Human Platelets
Authors: Watts Ian S , Keery Rebecca J , Lumley Philip
Published: June 2018

We have investigated the effect of two procedures that modify human platelet surface membrane glycoprotein (Gp) IIb and IIIa complexes upon whole blood platelet aggregation to a range of agonists. (A) Irreversible disruption of complexes by temporary (30 min) Ca2+-deprivation with EGTA at 37° C. (B) Binding of a monoclonal antibody M148 to the complex. EGTA exposure abolished aggregation to ADP, adrenaline and PAF. In contrast, full aggregation curves to collagen and U-46619 could still be established. EGTA exposure reduced M148 binding to platelets by 80%. Excess M148 abolished aggregation to ADP, PAF, collagen and U-46619. However, upon removal of unbound antibody from platelets full aggregation curves to collagen and U-46619 but not to ADP and PAF could be re-established. Thus human platelet aggregation to ADP, PAF and adrenaline appears absolutely dependent upon surface membrane GpIIb/IIIa complexes. In contrast, collagen and U-46619 cause expression of an additional distinct pool of Gp complexes inaccessible to EGTA and M148 in unstimulated platelets which is intimately involved in aggregation to these agonists.
A Variant of Glanzmann's Thrombasthenia Characterized by Abnormal Glycoprotein IIb/IIIa Complex Formation
Authors: Fournier Dominique J , Kabral Arnold , Castaldi Peter A , Berndt Michael C
Published: June 2018

Glanzmann's thrombasthenia is a congenital bleeding abnormality characterized by absent platelet aggregation due to the failure of fibrinogen to bind to activated thrombasthenic platelets. In the majority of cases, this defect is caused by the absence or marked reduction of a specific fibrinogen-binding aggregation receptor, the GP IIb/IIIa complex. E.T., an 18-year-old female with a life-long history of bleeding and easy bruising, had the normal clinical features of Glanzmann's thrombasthenia. Surprisingly, sodium dodecyl sulphate-polyacrylamide gel electrophoresis of her platelets showed no apparent abnormality of the GP IIb/IIIa complex. Control platelets washed in the presence of 2 mM EDTA and control and patient platelets washed in the presence of 2 mM calcium ions showed normal reactivity with anti-GP IIb, anti-GP Ilia, and anti-GP IIb/IIIa complex specific monoclonal antibodies as evaluated by flow cytometry. In contrast, patient's platelets washed in the presence of 2 mM EDTA reacted with anti-GP IIb, anti-GP Ilia, but not with the complex-specific monoclonal antibodies. The increased susceptibility of the patient's GP IIb/IIIa complex to EDTA dissociation was confirmed by crossed immunoelectrophoresis (CIE). CIE analysis further indicated that the patient's GP IIb/IIIa complex did not bind fibrinogen. The combined results suggest that this patient has Glanzmann's thrombasthenia due to an abnormal association of the GP IIb/IIIa complex which results in the failure of the complex to bind fibrinogen.
Mechanism of Potentiation by Manganese Ion of Aggregation of Porcine Pancreatic Elastase-Treated Human Platelets
Authors: Azuma Hiroyuki , Shigekiyo Toshio , Miura Shinji , Uno Yuka , Saito Shiro
Published: June 2018

The effect of manganese ion (Mn2+) on the aggregation of porcine pancreatic elastase-treated platelets (ETP) induced by fibrinogen (Fbg) was studied. Mn2+ enhanced the aggregation of ETP on addition of Fbg specifically and dose-dependently. This effect of Mn2+ was not associated with the formation of thromboxane A2, and was not affected by pretreatment of ETP with acetylsalicylic acid in the presence of Mn2+. Moreover, it was not dependent on extracellular adenosine diphosphate, as shown by removal of extracellular adenosine diphosphate by pretreatment of ETP with creatine phosphate/creatine phosphokinase. Studies on the binding of 125I-Fbg to ETP showed that Mn2+ increased the Kd value of binding but did not affect the number of Fbg binding sites. These results indicate that Mn2+ specifically and dose-dependently potentiated Fbg-induced aggregation of ETP and that this effect of Mn2+ may be due to an increase in the affinity of binding of Fbg to the glycoprotein IIb-IIIa complex on the membranes of ETP.
Thrombin Stimulated Platelet Accumulation on Protein Coated Glass Capillaries: Role of Adhesive Platelet α-Granule Proteins
Authors: Mulvihill Juliette N , Davies J Andrew , Toti Florence , Freyssinet Jean-Marie , Cazenave Jean-Pierre
Published: June 2018

The generation of trace amounts of thrombin at artificial surfaces in contact with blood is likely to be a contributing factor in thrombosis on biomaterials. Using an in vitro capillary perfusion system, platelet accumulation on glass surfaces, uncoated or precoated with purified bovine collagen or human plasma proteins, was determined in the presence or absence of preadsorbed purified human thrombin. Static adsorption for 15 min at 22° C from solutions of thrombin 100 NIH units (33 μg)/ml gave surface concentrations in the range 0.019-0.101 μg/cm2. Protein coated capillaries, thrombin treated or untreated, were perfused for 2 min at 37° C with suspensions of washed 111In-labeled human platelets in Tyrode's-albumin buffer containing 40% washed red blood cells, under conditions of controlled, non pulsatile laminar flow (50 s−1 or 2,000 s−1). Platelet accumulation was increased in the presence of surface adsorbed thrombin on uncoated and albumin or fibrinogen coated glass but little affected on fibronectin or collagen coated glass. On von Willebrand factor (vWF) coated glass, thrombin enhancement was observed only at high shear forces. In experiments using antibodies against human platelet α-granule proteins, thrombin stimulated platelet deposition in uncoated glass capillaries was inhibited at 2,000 s−1 by anti-vWF and to a lesser extent by anti-fibrinogen but not by antithrombospondin antibodies.
Inhibition of Platelet Thromboxane Formation and Phosphoinositides Breakdown by Osthole from Angelica pubescens
Authors: Ko Feng-Nien , Wu Tian-Shung , Liou Meei-Jen , Huang Tur-Fu , Teng Che-Ming
Published: June 2018

Osthole, isolated from Chinese herb Angelica pubescens, inhibited platelet aggregation and ATP release induced by ADP, arachidonic acid, PAF, collagen, ionophore A23187 and thrombin in washed rabbit platelets. It showed a weak activity in platelet-rich plasma. Osthole inhibited the thromboxane B2 formation caused by arachidonic acid, collagen, ionophore A23187 and thrombin in washed platelets, and also the thromboxane B2 formation caused by the incubation of lysed platelet homogenate with arachidonic acid. The generation of inositol phosphates in washed platelets caused by collagen, PAF and thrombin was suppressed by osthole. These data indicate that the inhibitory effect of osthole on platelet aggregation and release reaction was due to the inhibition of thromboxane formation and phosphoinositides breakdown.
Ouabain Enhances Basal and Stimulus-Induced Cytoplasmic Calcium Concentrations in Platelets
Authors: Lees A D , Wilson J , Orchard C H , Orchard M A
Published: June 2018

Incubation of platelet-rich plasma (PRP) with ouabain, an inhibitor of sodium/potassium ATPase (Na+/K+ ATPase), induced a significant rise in basal platelet intracellular calcium concentration ([Ca2+]i) when measured using fura 2. Ouabain induced an enhanced aggregation response to low doses of collagen in both PRP and washed platelets loaded with aequorin. In aequorin loaded platelets this enhanced aggregation response was associated with an enhanced rise in [Ca2+]j such that the relationship between [Ca2+]i and aggregation was unchanged. As inhibition of plasma membrane Na+/K+ ATPase would lead to a raised intracellular sodium ion concentration ([Na+]i) the results suggest that in the platelet, [Na+]i can modulate [Ca2+]i and hence influence the response of platelets to stimuli such as collagen.
Rat Platelets Adhere to Human Thrombin-Treated Rat Lungs under Flow Conditions
Authors: Venturini Catherine M , Fenton John W , Minnear Fred L , Kaplan John E
Published: June 2018

Platelet attachment to thrombin-treated endothelium was examined under flow conditions in the vascular bed of isolated perfused rat lungs. The infused thrombin bound to the pulmonary endothelium in a specific, competitive manner as shown by displacement of 125I-α-thrombin with an excess of cold diiso-propylfluoro-phosphate-α-thrombin. α-Thrombin significantly increased the attachment of isolated 51Cr-labelled rat platelets to the perfused lungs. Scanning and high voltage electron microscopy showed single platelets in various stages of activation attached to pulmonary endothelium. The receptor binding site and catalytic activity of thrombin were essential to the enhancement of platelet attachment. Aspirin given to rats before the lung isolation had no effect on thrombin-induced platelet attachment. Thus, endothelial bound thrombin initiates platelet activation and enhances subsequent platelet attachment.