bluuebunny/crossref_metadata_2025 huggingface dataset:
A Pharmacologic Approach To The Modification Of The Electrokinetic And Aggregability Characteristics Of Rabbit Platelets In Vitro
Authors: Lucas Thomas R , Firriolo D , Sawyer Philip N
Published: May 2019

An attempt was made to correlate changes in the electrokinetic characteristics of rabbit platelets with alterations in platelet function. In an effort to modify the surface charge characteristics of the rabbit platelet, several pharmacologic agents were used, i.e., heparin, chlorpromazine, and neuraminidase. A Seaman microelectrophoresis apparatus and a chronolog platelet aggregometer were used to measure platelet membrane changes and aggregation. The control electrophoretic mobility (EM) was 0.875±0.005µ for 18 independent blood samples. Heparin increased the EM to 0.99±0.005 in 10 samples. Chlorpromazine decreased the EM to 0.860±0.005 and neuraminidase decreased the EM to 0.73±0.001. The results were significant when compared to baseline (p=0.005). Heparin inhibited thrombin induced platelet aggregation while neuraminidase enhanced primary aggregation and chlorpromazine blocked the secondary wave of platelet aggregation. The results indicate that increased surface negativity reduces platelet aggreg-ability, while a decrease in surface negativity results in altered platelet function affecting primary platelet interaction. The results suggest altered platelet surface charge characteristics as a possible mechanism for platelet aggregation and thrombosis. This study also suggests in part the role of heparin in reducing thrombosis formation by inhibiting platelet-platelet interaction via increased surface negativity.
Malaric Thrombopathy
Authors: Ospina L , Bernal M D E , Santos C
Published: May 2019

The purpose of this investigation was to demonstrate platelet aggregation defects in patients infested with Plasmodium, giant and normal platelets counts.

Laboratory studies were performed by using a “Payton Aggregometer”. ADP, Collagen, Epinephrine and Ristocetin as commercial reagents. Coagulation tests were done by regular methods. Thirty out of 31 patients were found to have marked defects in secondary aggregation wave with ADP. Aggregation with Collagen was constantly abnormal. Fifteen patients showed primary and secondary aggregation abnormality with ADP, Adrenaline and Collagen. Aggregation with Ristocetin was always normal. Aggregation defects were not drug induced since the patients were taking no medicines at least five days prior to studies.

Studies done in volunteers taking antimalarics, Dipirone, Acetaminophen showed no interference with aggregation. Aggregation abnormalities were mostly found in patients infested with chronic Falciparum malaria.
Most of the patients did not have bleeding manifestations but had multiple organ involvement. We think this platelet defect is a manifestation of some defense mechanism to prevent massive thrombosis frequently found in Falciparum Malaria.
HAEMODIALYSIS INDUCED HYPOXIA: A COMPARISON OF THE EFFECT OF PROSTACYCLIN, METHYL PREDNISOLONE, ULTRAFILTRATION AND BICARBONATE DIALYSATE
Authors: Dodd N J , Turney J H , Weston M J
Published: May 2019

In order to test certain hypotheses regarding the cause of arterial hypoxia during haemodialysis(HD),we investigated the effect of:- 1 Heparin bolus-infusion; or heparin plus:- 2.Prostacyclin ( PGI2 ) infusion ( 5ng/Kg/mi n );3. Methyl Prednisolone bolus(30mg/Kg) 30 minutes prior to dialysis; 4. Ultrafiltration for one hour followed by dialysis(UF/HD); 5.40mmo1/l Bicarbonate dialysate instead of standard acetate dialysate . Seven patients on long term HD were examined by all methods. Arterial blood samples were taken before,and at intervals during ,dialysis for blood gases, platelet count,and neutrophil count. All results are expressed as percentage of pre-dialys is value (means).

Results fo r each group were tested fo r significance against the Heparin group(l.)using a paired t tes t . The only significant sparing of hypoxia occurred at 60 minutes during UF/HD.PGI2 spared platelets , as we have previously shown. Methyl Prednisolone failed to prevent dialysis induced neutropenia. The mechanism of haemodialysis induced hypoxia remains obscure
Clinical studies of the use of a fluorogenic substrate assay method for the determination of plasminogen
Authors: Triplett Douglas , Harms Cathy , Hermelin Lori , Huseby Rolf , Mitchell Gary , Pochron Sharon
Published: May 2019

Utilization of a recently reported method for the determination of plasminogen by fluorometric technique (Ref. 3, Pochron, et. al., Thrombosis Research, Vol. 13, pg. 733, 1978) was compared with acceptable methodologies (e.g. the caseinolytic and radial–immunodiffusion assays). Excellent results were seen with the normal range of 2.4–3.8 CTA U/ml of plasminogen by the synthetic substrate assay as compared to 8.7–14.3 mg/dl by the radialimmunodiffusion assay. Normal ranges for the caseinolvtic assay were seen to be 2.2–4.0 CTA U/ml. Patients’ studies were performed on a general medical population but centered primarily on “pre” and “post” operative coronary-pulmonary bypass patients. In all cases studied, a drop in Plasminogen level was noted in this group of patients following surgery (20-50%). Other clinical data will be presented indicating that the determination of plasminogen by the fluorometric assay procedure is sensitive and accurate for clinical situations.
An evaluation of currently available methods for plasma fibrinogen
Authors: Rickard K , Burridge J , Exner T , Power P
Published: May 2019

An assessment of fibrinogen concentration is basic to any investigation of clotting dysfunction and often an estimate of fibrinogen level is needed rapidly as an indication of consumptive coagulopathy or fibrinolysis. Fibrinogen levels in a variety of clinical plasma samples were assessed concurrently by several methods. Results were correlated against a reference method based on Ancrod-clottable fibrinogen and calibrated by U.V. absorbance with alkaline solutions of carefully dried fibrin standard. The best correlations with the reference method were achieved by an immunologic method using the Centrifichem principle and by heat precipitation with quantitation by packing in micro-haematocrit tubes. A modified clot opacity method also gave acceptable results. The turbidimetric ammonium sulphate and sodium sulphite precipitation methods correlated less well with the reference method, and in particular the sodium sulphite technique gave high apparent fibrinogen levels with jaundiced plasmas. Neither of the turbidimetric methods were useful for fibrinogen levels below 50mg/dl. The thrombin time method showed excellent sensitivity to fibrinogen, even at very low fibrinogen levels, but did not correlate well with the reference method. This apparently conflicts with the findings of a recent CAP survey which strongly favoured the thrombin time method. We believe there is a danger that such surveys promote test methods on which there is good interlaboratory agreement, but which may not be specific in function.
Functional Relationship of Polymerization Sites of Vertebrate Fibrinogens
Authors: Cierniewski C , Krajewski T , Janiak E
Published: May 2019

Various studies on the interaction of immobilized mammalian fibrinogen and fibrin monomers with some fibrinogen derivatives demonstrated the presence of two sets of polymerization sites in the mammalian fibrinogen molecule. We obtained the same results while investigating the fibrinogen molecules of other classes of vertebrates /Pisces. Amphibia. Aves/. Despite significant differences among their subunit structures, all of them contain polymerization sites homologous to mammalian counterparts. Moreover, due to great functional similarity, fibrinogen or fibrin monomers of the analyzed species of Pisces. Amphibia. Aves and Mammalia interacted in a specific way with immobilized pig fibrin monomers or fibrinogen, respectively. Using these pig affinity adsorbents, fibrinogen and fibrin monomers of different vertebrates were isolated directly from plasma and analyzed by SDS polyacrylamide gel electrophoresis. Polypeptide compositions of eluted proteins were identical to those obtained for corresponding fibrinogen preparations isolated by cold-ethanol fractionation method. It appears to indicate that the nature of polymerization sites in vertebrate fibrinogens is alike.
Effect of Suloctidil on Shortened Platelet Regeneration Time in Cardiovascular Patients
Authors: Deperon R , Roncucci R , Lambelin G , Lansen J , Van Stalle F
Published: May 2019

Previous investigations based on the measurement of platelet regeneration time (PRT) rather than platelet survival time (PST) have suggested that this parameter is significantly reduced in patients bearing vascular prosthetic material and in a number of patients with occlusive vascular disease (1). Since it is well-known that shortened PST can be normalized by antiplatelet drug’s, it seemed interesting to investigate whether the measurement of PRT could also detect this effect. Suloctidil (S), a new vasoactive agent, has been chosen for this study which was performed in parallel double-blind conditions vs placebo. 32 patient. (24 males and 8 females) with either prosthetic heart valves, aortoiliac grafts or occlusive arterial disease were investigated [mean PRT t/2 value before treatment for the S group (n=16), 2.8±0.1 days]. They were given S at a dose of 600 mg daily during 3 to 4 weeks. The results obtained indicate that S treatment significantly increases shortened PRT t/2 values (mean PRT t/2 value after S treatment : 3.8±0.2 ; p<.001). Placebo treatment had no effect in a similar group of patients (n=16 ; mean PRT t/2 value before treatment : 3.0±0.1 days ; after treatment : 3.2±0.2 days). (1) R. Roncucci et al., Thromb.Res., accepted for publication.
The “Dead Digit, Palpable Pulse” Syndrome: A Preliminary Study of the Value of Antiplatelet Drugs in This Disorder
Authors: Morris-Jones W , Preston F
Published: May 2019

We report here the results of a pilot study to investigate the value of antiplatelet drugs in patients with pregangrene of one or both feet associated with palpable peripheral pulses but without clinical or laboratory evidence of a myeloproliferative disorder in diabetes mellitus. 32 patients were studied. For each patient pregangrene of one or both feet had been present for a minimum period of six weeks. All patients had failed to respond to medical therapy. Platelet aggregation, using ADP, collagen and adrenaline was assessed before therapy. PRP was also tested for spontaneous platelet aggregation (SPA). BTG assays and platelet aggregate ratios were also performed. Aspirin, 325mgs t.d.s. and persantin 100mgs t.d.s. produced significant and maintained clinical improvement in 5 of the 7 patients with SPA- 7 of 12 patients with hypersensitive platelets and 3 of 13 patients with normo-or hyposensitive platelets. Sympathectomy was performed in patients who failed to respond to drug therapy. The mean PAR was 0.94 for the non-responder group and 0.78 for the responder group. The mean BTG for non-responders was 44ng/ml (normal 50). The mean BTG for the responder group was 59.1ng/ml. The results suggest that antiplatelet drugs may be of therapeutic value in patients with the “dead-digit palpable pulse” syndrome arid that controlled clinical trials are indicated.
Action And Interaction Of Sulfinpyrazone In Anticoagulated Patients
Authors: Walter E , Hodok-Laibach P , Zimmermann R , Weber E
Published: May 2019

For 8 weeks 29 anticoagulated patients with elevated serum uric acid levels were controlled with 14 days intervals. From week 3 till the end of 6 they received 600 mg sulfinpyrazone p. d. Because of excellent compliance the uric acid decreased from 7.0±0.2 mg% to 3.7±0.2 and raised to 7.1±0.3 (x±sx) after omitting the drug. In contrary to this collagen-induced aggregation (5ug/ml, Vmax = Δ E/min.) decreased during the first l4 days of therapy from 0.388±0.048 to 0.228±0.045 (p < 0.02) and reached. 0.166±0.031 (p < 0.01) at the end of the therapy, but with 0.269±0.040 did not return to the starting point two weeks after stopping the drug. The increasing effect of therapy and the long lasting effect point to an active metabolite of sulfinpyrazone, which is slowly eliminated. The interaction of sulfinpyrazone with phenprocoumon consisted in a displacement of the anticoagulant from protein-binding sites, resulting in a fall of the Thrombotest value from 11.2 % to 9.0 % (p<0,0l). During sulfinpyrazone therapy the GGT increased significantly, indicating an induction of drug metabolizing enzymes in the liver, which had to be counter- acted with a higher phenprocoumon dosis for therapeutic anticoagulation.
Clot Promoting Effect of Platelet-Vessel Wall Interaction
Authors: Hornstra G
Published: May 2019

When a piece of vascular tissue (rat, rabbit) is incubated in plasma, clotting occurs.
The presence of platelets accellerates this clotting process.Inhibition of vascular prostacyclin (PGI2) production, either by drugs or by inducing arachidonic acid deficiency, causes the platelets to become activated by the vessel wall, resulting in platelet shape change and -aggregation and in a further enhancement of the coagulation response. Hirudin very actively inhibits both aggregation and clotting whereas ADP scavenging has only a weak effect. This indicates that the aggregation response, occurring upon platelet-vessel wall interaction is for the greater part mediated by thrombin.

Saturated fat feeding increases the clot promoting effect of the platelet-vessel wall interaction. This appeared not to be due to differences in the production of vascular PGI2or platelet endoperoxides and thromboxanes (measured as malondialdehyde).

These findings indicate that:
1. Thrombin, locally generated upon the interaction between damaged vessel wall and blood constituents, contribute to arterial thrombus formation;

2. Prostacyclin regulates local thrombin formation via its inhibiting effect on platelet activation;

3. The existence of another, platelet bound affector is postulated.
Histochemical And Ultrastructural Study Of The Platelet-Arterial Subendothelium Interaction
Authors: Legrand Y , Bariety J , Birembaut P , Michel H , Fauvel F , Caen J
Published: May 2019

In order to characterize the subendothelial macromolecules able to interact with platelets rabbit aortas were de-endothelialized and incubated either with chymotrypsin (24 h at 37° C), with a highly purified bacterial collagenase (2 h at 37° C), or with chymotrypsin
followed by collagenase or the reverse. Histochemical ultrastructural studies were performed before and after blood perfusion, using three different staining procedures : tannic acid (for the microfibrillar structures and elastin), ruthenium red (for the polyanions) and two peroxidase-labelled lectins (concanavalin A and ricinus communis (for the detection of saccharidic determinants)). After chymotrypsin, the tannic acid +ve, ruthenium red +ve and lectin +ve microfibrils had been digested ; platelet adherence to the remaining sparse collagen fibrils persisted. After collagenase, these microfibrils are preserved, collagen has been digested, but the platelets can still adhere. After combined incubation with both enzymes, elastin is the only remaining constituent and no platelet adhesion can be observed.

These results show that in the rabbit aorta subendothelium besides collagen, microfibrillar glycoproteins are able to interact with human platelets.
In Vitro Synthesis of Bovine Prothrombin in a Purified System
Authors: de Metz M , Vermeer C , Soute B , Hemker H
Published: May 2019

Coumarin treatment of the cow results in the appearance of a prothrombin precursor (de-carboxyprothrombin) in the blood. We have isolated from bovine liver the enzyme (prothrombin synthase) that is able to convert purified decarboxyprothrombin into prothrombin. Prothrombin synthase is localized in the microsomal fraction and extracted therefrom with a buffer containing 2% Triton X-l00. Further purification of the enzyme was accomplished by column chromatography on Sepharose 4B, DEAE Sephacell and Sephadex G 100. The molecular weight of prothrombin synthase was found to be about 60,000 D. The enzyme activity is defined as the ability to increase the prothrombin concentration at standard conditions and it is measured as a decrease of the clotting time in the one-stage coagulation assay. In the presence of NaH14CO3, radioactive label is incorporated into decarboxyprothrombin, parallel to the generation of prothrombin.

After extraction with a suitable organic solvent prothrombin synthase was separated from endogenously bound vitamin K. After extraction the water phase contained the enzyme, the activity of which was completely restored by supplementing the reaction mixtures with vitamin KH2. The organic phase contained the vitamin K as was deduced from U.V.spectra. Prothrombin synthase is dependent on Mn2+and molecular oxygen and is inhibited by warfarin, Cl-K and Cu2+.
The Inhibition of the Vitamin K Dependent Carboxylation by Free Radical Scavengers
Authors: Walter S , Burgess A , Esnouf M , Green M , Hill H , Okolow-Zubkowska M
Published: May 2019

Prothrombin contains γ-carboxyglutamic acid residues, synthesised at a post-translational step requiring vitamin K quinol, carbon dioxide, molecular oxygen, and a carboxylating system solubilised from liver microsomal fractions of rats (normal and vit.K deficient) and calves by non-fonic detergents. The reaction is assayed by the incorporation of 14C into prothrombin precursor or the pentapeptide, Phe-Leu-Glu-Glu-Val. Superoxide dismutase reduces the incorporation (50% inhibition at 300μM) and so too do copper chelates with known dismutase activity:50% inhibition at 60μM copper (II) aspirinate, 127μM copper (II) penicillamine and 152μM copper (II) tyrosine. The superoxide anion is postulated as a primary product of reduced Vitamin K and dioxygen. Catalase also inhibits at high concentrations (> .5mM), possibly by shifting the equilibrium to favour dismutation or by preventing production of an ’active carbon’ species via hydrogen peroxide. So far it has not been possible to replace Vitamin K with active intermediates, such as t-butyl hydroperoxide, which also inhibits the action of the vitamin, possibly due to the formation of Vitamin K 2,3-epoxide. These results point to the involvement of radical species in the Vitamin K dependent carboxylation of prothrombin.
m-[o-(2-Chloro-5-Fluorosulfonylphenylureido)Phenoxybutoxy]Benzamidine: Affinity Labeling Reagent Which Can Identify Secondary Binding Sites of Coagulation Complement and Fibrinolytic Serine Proteases
Authors: Bing D , Robison D , Andrews J , Laura R
Published: May 2019

We have determined that m-[o-(2-chloro-5-fluorosulfonylphenylureido)phenoxybutoxy]benza-midine [mCP(PBA)-F] is an affinity labeling reagent which labels both polypeptide chains of thrombin, factor Xa, complement component CIS and plasmin. As this means it is reacting outside of the catalytic center, we have called this reagent an exo-site affinity labeling reagent. Progressive irreversible inhibition of these enzymes by this reagent is rapid (k1st 2.5-4.6 x 10-3sec-1), the kinetics of inactivation are consistent with inhibition proceding via formation of a specific enzyme-inhibitor complex analogous to a Michaelis-Menton complex (KL - 115-26 μM), and diisopropylfluorophosphate or p-amidino-phenylmethanesulfonyfluoride Prevent labeling by [3H]mCP(PBA)-F. A molecular model of mCP(PBA)-F shows that the reactive SO2F group can be 17 A from the cationic amidine. The data are consistent with the hypothesis that both peptide chains are required for the specific proteolytic activity exhibited by these proteases and that the peptide chain which does not contain the active site serine is close to the catalytic center. (Supported by NIH and AHA grants
The Effect of Sulfinpyrazone, Aspirin and Their Metabolites on Prostacyclin Production by Rabbit Aortic Rings
Authors: Westwick J , Webb H , Lewis G
Published: May 2019

Since sulfinpyrazone (SU) and aspirin (ASA) are used as anti-thrombotic compounds, it is important to know the effects of these agents and their metabolites on prostacyclin (PGI2) generation. Rabbit aortic rings (RAR) incubated with arachidonic acid (AA) generate a product that inhibits platelet aggregation. That the ring product was PGI2 was shown by 1) a similar temperature-dependent decay curve when assayed against authentic PGI2, 2) the formation of the product was blocked by 15-00H AA (l0µg/ml), 3) the action of the product on platelets was antagonised by antisera to 6-keto-PGF1α which antagonised authentic PGI2 but not PGD2 or PGE1. The optimal conditions for PGI2 generation by RAR were determined by varying storage and incubation media, time and temperature of incubation and substrate concentration. RAR (l0-20mg) were incubated in 300µl buffer with each concentration of drug in quintuplicate for 5 min at 25°C. AA was added, 5 min later a 200µl aliquot was withdrawn and its ability to inhibit ADP-induced platelet aggregation was compared with that produced by vehicle-treated RAR and to authentic PGI2·IC50(µg/ml) of PGI2 production were: SU (210), sulfoxide M of SU (60), p-hydroxy M of SU(inactive), ASA(140), salicylic acid (inactive). Therefore the concentrations of sulfinpyrazone and aspirin necessary to inhibit PGI2 production are higher than those which produced their anti-thrombotic effect.
Clinical Trials in the Secondary Prevention of Myocardial Infarction and Stroke
Authors: Armitage P
Published: May 2019

In the 30 years or so since adequate clinical trials were first conducted the basic principles have scarcely changed. Randomization remains an essential ingredient. Trials have increased in size and duration, and standards of design and execution are now high. Particular attention should be paid to the effect of baseline variables, for correcting imbalance and improving precision, and also to reveal interactions between these variables and the treatments.

Trials in the secondary prevention of stroke and myocardial infarction, as in other fields, frequently appear to engender controversy. This is partly due to dissenting views about aspects of the protocol on which choices had to be made (perhaps an argument for flexibility in the protocol); and partly due to termination at an inconclusive stage. The ethical dilemma prevents any easy solution, but it is suggested that early stopping should only be allowed when evidence for a difference is very strong.

Different approaches to the handling of protocol departures are discussed; the pragmatic approach, with inclusion of all cases after randomization, is advocated.

Factorial designs are likely to be increasingly adopted, in view of the pressure of new drugs and the need to economise in the large numbers of patients required. Some interpretative problems are discussed.
Decreased Reactivity of Plasmin-Streptokinase Towards α-2-Antiplasmin
Authors: Cederholm-Williams S , Lijnen H , De Cock F , Collen D
Published: May 2019

Plasmin (P) is very rapidly (rate constant 2 x 107M-1s-1) inactivated by α-2-antiplasmin (A), and blocking of the lysine binding sites (LSB) of plasmin reduses this rate 50 fold (Eur.J.Biochem.84, 573-578, 1978). Using purified reactants and the plasmin substrate D-Val-Leu-Lys-pNA (S 2251) the influence of streptokinase (SK) on the reaction between P and A was examined. Addition of SK to P reduces the reaction rate with A in a sigmoidal fashion. Assuming the formation of a reversible bimolecular P:SK complex which is unreactive towards A then the dissociation constant Kd of this complex is less than 10-9M. The LSB of P does not play a role in this interaction since similar Kd values are found with modified P, lacking LSB and in the presence of 0.3 mM 6-aminohexanoic acid which blocks the LSB. The P:SK complex is only very slowly inhibited by A with a rate constant < 5 x 102M-1s-1.

It is concluded that the α-2-antiplasmin is less reactive towards the plasmin streptokinase complex due to a modification of the active center of plasmin rather than an effect upon the binding sites.
Viscosity and Red Cell Deformity in Arterial Disease
Authors: Cella G , de Haas H , Rampling M , Kakkar V
Published: May 2019

Haemorrheological factors have been shown to be affected in many kings of vascular disease. The present study was undertaken to correlate these factors in normal subjects and patients suffering from peripheral arterial disease. Twenty-two patients were investigated; they had moderate or severe intermittent claudication, extent of disease being confirmed by aorto-arteriography and ankle-systolic pressure studies. Twenty-five controls with no symptoms or signs of arterial disease were selected with comparable age and sex distribution. Whole blood viscosity was measured at shear rates of 230 secs-1 and 23 secs-lat 37°c using a Wells Brookfield cone plate microvisco meter. Plasma viscosity was also measured in an identical manner. Erythrocyte flexibility was measured by centrifuge technique and fibrinogen concentration as well as haematocrit by standard techniques. The fibrinogen concentration appeared to be the only significant parameter; the mean concentration in patients with peripheral vascular disease of 463 ± 73mg/l00ml in the control group ( < 0.05). Although whole blood viscosity was high in patients, when corrected to a common haematocrit, there was no significant difference between patients and controls. The same megative correlation was found for plasma viscosity. The red cell flexibility was found to be increased in patients as compared to the control group, but this effect appeared to be simply proportional to the fibrinogen concentration.
Increased Leukocyte Thromboplastin Activity by Sensitized Lymphocytes: A Link Between Immunology and Blood Coagulation
Authors: van Aken W , Zeylemaker W , Stricker E , Oh J , van Ginkel C
Published: May 2019

Fibrin deposition and infiltration of mononuclear leukocytes (MN) frequently occurs concomitantly at the site of an immune reaction. As human MN can produce thromboplastin (TP) we have investigated whether immune stimulation of MN can trigger the production of TP by these cells. Both specific antigen (tuberculin PPD) and allogeneic lymphocytes were used for stimulation. MN from 14 donors with a high response to PPD and from 11 donors with a low response to PPD (based on the incorporation of 3H-thymidine) were cultured for 7 days in the presence or absence of PPD and the TP activity of the whole cell suspension was determined. When PPD was absent, TP levels in MN from low and high responders were not different (113±47 and 197±111 u/ml resp. p ˃ 0.10). With PPD, however, MN from high responders produced 1470±413 u TP/ml compared to 126±49 u TP/ml produced by MN from low responders (p < 0.005). Similar results were obtained when MN were kept in culture for either 4 or 11 days. However, after 24 hours in culture, TP synthesis was not different in the MN of the 2 groups. When MN were cultured in a mixed leukocyte reaction, allogeneic MN produced more TP (405±151 u/ml) than control cultures (95±21 u/ml) (p<0.025;n=22). These data indicate that stimulated lymphocytes can enhance TP activity of leukocytes. It is postulated that fibrin formation in immune reactions such as delayed hypersensitivity and rejection of kidney allografts is mediated by immunologically stimulated MN.
Doppler Angiography for the Non-Invasive Investigation of the Carotid Artery in the Neck
Authors: Warlow Charles , Fish Peter
Published: May 2019

The accuracy, and possible clinical application of non-invasive ultra sound imaging of the cervical carotid artery in patients with cerebrovascular disease has been investigated by comparing a pulsed multi-channel directional Doppler imaging devise (“MAVIS” - GEC Medical) with conventional carotid angiography. of 23 normal or minimally diseased internal carotid arteries only 4 appeared as stenosed on Doppler angiography, the other 19 were normal; of 14 stenosed arteries only 1 appeared normal on imaging, 1 appeared occluded and 12 showed clear images of stenosis; all 6 occluded arteries appeared occluded on ultra sound imaging. It was difficult to obtain satisfactory images of the external carotid arteries in all cases. The additional facility to display a velocity flow profile at preselected points along the arterial image should increase the usefulness of this technique, particularly when there is only minor atheromatous irregularity or ulceration, and such a device will be briefly discussed.
The Pathogenesis of Thrombotic Thrombocytopenic Purpura
Authors: Kwaan H. C.
Published: May 2019

The vascular lesions with microthrombi were studied in 12 patients with thrombotic thrombocytopenic purpura (TTP), diagnosed by the characteristic clinical and laboratory findings and confirmed histologically in each case. While defibrination was not observed, and with only minimal changes in the circulating levels of fibrinogen, fibrin degradation products and plasminogen activator, the microthrombotic lesion was invariably present. Immunofluorescent and histochemical studies indicated that both platelet and fibrin were present in the microthrombi with the platelet components dominant in many cases. Using the fibrin slide method, plasminogen activator was demonstrated in the uninvolved blood vessels but totally absent in the vessels occluded by microthrombi. in contrast, fibrinolysis is always present in the vessels afflicted with other types of thrombosis, such as the microthrombi in disseminated intravascular coagulation. Since circulating fibrinolytic activity was normal in TTP, the absence of vascular fibrinolysis is a local defect due to either inhibition by the platelet deposits or by local vascular damage. The inability of thrombolysis may explain the absence of systemic defibrination and the severity of the disease.
Evidence for A Coagulopathy Related to Endotoxaemia in Patients with Obstructive Jaundice
Authors: Hunt D. R. , Allison M. E. M. , Forrester A. , Prentice C. R. M. , Blumgart L. H.
Published: May 2019

Animal studies suggest that endotoxaemia may contribute to the disturbances of coagulation in obstructive jaundice. Two groups of patients, 14 controls (C) and 28 jaundiced (J) having pancreatic or biliary surgery were studied prospectively. Endotoxaemia, soluble fibrin (s. f.) by tanned red cell agglutination, FDP and DVT were measured for comparison with outcome of surgery. in group J more complications occurred with 7 deaths and 10 DVT; by comparison, only one death occurred in group C and no DVT, fever or haemorrhage. S. f. was found in none of group C but in 9 of 24 in group J pre-operatively. Presence of s. f. did not influence outcome of surgery. Endotoxaemia and raised FDP were also seen more frequently pre-operatively in group J. of the group J patients 11 had endotoxaemia or FDP before operation and 7 died. An association between endotoxaemia and FDP was shown in both groups and in group J endotoxaemia appears related to s. f. although s. f. and FDP are independent. A coagulopathy, associated with endotoxaemia is present in some jaundiced patients and they fair badly after surgery.
The Effects of PGI2 on Thrombin, Factor Xa, Plasmin and on Their Inactivation By Antithrombin-III and Heparin
Authors: Blaskó Gy
Published: May 2019

The data presented by the author demonstrate the effect of PGI2 on the activity and the inactivation of enzymes participating in blood clotting /thrombin and Factor Xa/ and fibrinolysis /plasmin/. According to his results PGI2 has no effects on the activity of Factor Xa and plasmin and on theinactivation of these enzymes by antithrombin-III and/or heparin An acceleration of the inactivation of thrombin by antithrombin-III was found in the presence of PGI2 within a concentration range of 50-250 μg/ml. This heparin-like effect of PGI2 is demonstrable only at a concentration of 5 magnitude higher than that of the antiaggregation effect on platelets /5-10 ng per ml/. Factors influencing the inactivation of thrombin and Xa by antithrombin-III and heparin /proteins, ions/ are under discussion.
Fragment D Immunoreactivity: The Influence of Iodination and Calcium Environment
Authors: Kudryfc B. , Blombäck N.
Published: May 2019

Human fragment D (Fg-Ds) has been iodinated using both the Chloramine-T and lactoperoxidase methods. The specific activity was similar regardless of the method used. However, binding to a specific antibody was different for each preparation. The antigen labeled by the Chloramine-T method bound to a maximum of 40%, the other labeled product bound up to 85%. A correlation between the degree of immunoreactivity and avidity for a fibrinmonomer conjugate was found also. Fibrinmonomer bound about twice the amount of lactoperoxidase iodinated Fg-Ds as it did the Chloramine-T product. The use of these conjugates in the purification of immunoreactive Fg-Ds of high specific activity will be discussed. affinity binding sites for calcium have recently been demonstrated in fibrinogen. The presence of bound calcium is also believed to protect Fg-Ds from further digestion by plasmin. This is probably due to the formation of a more compact structure. However, conformational changes for calcium bound fibrinogen or Fg-Ds have not been observed. We tested the immunoreactivity of the lactoperoxidase iodinated Fg-Ds in presence and absence of calcium. Differences were found and this data suggests that some modification of antigenic determinants takes place as a consequence of calcium in the environment.
Determination of α1-Antitrypsin with A Chromogenic Peptide Substrate Methodology and Clinical Application
Authors: Witt I. , Lill H. , Karitzky D. , Flad H.
Published: May 2019

α-Antitrypsin (α-AT) is a relatively unspecific inhibitor of proteinases. The co-dominant hereditary defect of the protein can be associated with obstructive pulmonary emphysema in adulthood or with juvenile hepatocirrhosis.

A rapid and reliable method for the determination of α1-AT activity with the chromogenic substrate Z-Val-Gly-Arg-pNA has been developed: a standardized trypsin solution of known activity is allowed to react with the plasma or serum sample and the residual trypsin activity determined after incubation. The difference between the initial and residual trypsin activities corresponds to the α1-AT activity. The assay is carried out in tris/HCl-buffer at pH 8.0 (ionic strength=0.15) and 25°C; measurements are taken at 405 nm. Both kinetic and 2-point measurement is possible. The coefficient of variation for day-to-day precision is <5%.

The following guide values were determined on 15 healthy probands: 141 ± 15 inhibitor U/ml for plasma, and 130 ± 25 inhibitor U/ml for serum.

The minute volume of plasma or serum used precludes non-specific reactions of the substrate with other proteases. Homozygous carriers of this protein defect can be accurately identified by means of this test.
Congenital Antithrombin III Deficiency in 3 Families (7 Affected Members)
Authors: Conard J. , Samama M. , Horellou M. H. , Cazenave B. , Griguer P. , Barsotti J. , Herreman G. , Godeau P.
Published: May 2019

A congenital Antithrombin III (AT III) deficiency affecting 7 members of 3 families is reported.

The first throrabo-embolic accidents were observed between the age of 22 and 35 : they were spontaneous or occured after delivery or oral contraception. in one patient, a deep vein thrombosis was observed during heparin treatment. in 2 cases, recurrent pulmonary embolic episodes required vena cava ligation. No thromboembolic accident was observed during oral anticoagulation.

AT III was measured by an amidolytic method and by the Mancini method on plasma and serum ; the antithrombin activity was determined on serum by the von Kaulla method. in 7 patients, a decreased AT III was found by all the methods performed. The AT III level was around 50 % in patients treated or not by oral anticoagulants One patient was studied during heparin treatment and then under oral anticoagulants : AT III levels were lower under heparin.
Reduced Glutathione and Platelet Functions
Authors: Matsuda S. , Ikeda Y. , Kikuchi M. , Ando Y. , Aoki M. , Ogawa T. , Hasegawa M.
Published: May 2019

The role of reduced glutathione(GSH) on platelet functions was investigated utilizing diamide[(CH3)2NCON=NCON(CH3)2], which was shown to be a rather specific agent for the intracellular oxidation of GSH to oxidized glutathione(GSSG). Human normal citrated PRP was incubated with various concentration of diamide for 30 min at 22°C and platelet aggregation studies were performed using ADP, epinephrine, or collagen as aggregating agents. Clot retraction was examined in PRP treated with diamide by adding 100mM CaCl2 and 1.0u/ml thrombin. Simultaneously, GSH in platelets were measured by Beutler’s method. The maximum aggregability of platelets treated with 25μM diamide was significantly decreased with 2.5μM ADP, 5μM epinephrine, or 64μg/ml collagen as aggregating agent. When platelets were incubated with 50μM diamide, no secondary aggregation with 2.5μM ADP or 5μM epinephrine was observed. At a concentration of 100μM, diamide completely abolished primary aggregation with 2.5μM ADP. Clot retraction was grossly inhibited by diamide at concentrations above 50μM. GSH levels of platelets which were incubated with 100μM and 1mM diamide at 22°C for 30 min was 11.41×l0-9and 1.16×10-9moles/109 plts, respectively, while that without diamide treatment was 15.28×l0-9moles/109 plts. Decrease of GSH levels seemed to be dependent on diamide concentration. Membranes were isolated from diamide treated platelets according to the method of Jamieson et al, and were electrophoreses on SDS Polyacrylamide gels. There was no difference in Coomassie blue or PAS stained bands between control and diamide treated platelets. Marked inhibition of platelet aggregation and clot retraction accompanied by intracellular oxidation of GSH to GSSG may imply an important role of GSH on platelet aggregation and clot retraction.
Microheterogeneity of Human Factor VIII
Authors: Mourik J.A. van , Hellings J. A. , Hoorweg E. M.
Published: May 2019

Human factor VIII is a glycoprotein and tends to form a variety of large aggregates, the presence of which was demonstrated by a number of techniques including the use of large pore gel chromatography and electrophoresis using a polyacrylamide gel system designed for high molecular weight aggregates. Factor VIII aggregates can be fractionated in part according to molecular size. Reduction of these polymers by 2-mercaptoethanol results in presumably identical fragments suggesting that all polymers, though differing in size, are composed of identical subunits.

Factor VIII aggregates are particularly sensitive to proteolytic breakdown by trypsin and plasmin as judged by large pore polyacrylamide gel electrophoresis. Short-term incubation of factor VIII with trace quantities (equivalent to plasmin- or trypsin-like activities present in normal plasma) of these enzymes respectively results in substantial fragmentation with concurrent loss of both factor VIII procoagulant and von Willebrand factor activity. However, factor VIII activity is lost prior to discernible protein fragmentation whereas von Willebrand factor inactivation is associated with advanced protein degradation. When the ionic strength of the medium is lowered the susceptibility of factor VIII to proteolytic breakdown by trypsin is increased dramatically. Interestingly, degradation of factor VIII by plasmin is not affected by the ionic strength. These data, then, provide conclusive evidence of microheterogeneity of normal human factor VIII and may account for the observed heterogeneity of factor VIII on crossed immunoelectrophoresis.
Hemophilia a in a Girl with Deletion of a Part of the long ARM of one X Chromosome
Authors: Samama M. , Perrotez Ch. , Houissa R. , Hafsia A. , Seger J.
Published: May 2019

A severe hemorragic diathesis has been discovered in a 10 years old girl whose brother has severe hemophilia with classical haemarthrosis. The factor VIII biological activity is less than 1% in both of them with an almost normal factor VIII related antigen. The bleeding time was normal. There was no consanguinity and paternity was not disproved by extensive blood grouping tests. Cytogenetics studies showed a deletion of a part of the long arm of one X chromosome. The formula of the caryotype is 46, X, del X (q.212). The sex chromatin showed the presence of a phenotypic female. The X chromatin was smaller than normal. This is the first case to our knowledge of true female haemophilia A due to a deletion of one X chromosome.
Interaction of Factor XI and Kallikrein with High Molecular weight Kininogen
Authors: Thompson Russell E. , Mandle Robert J. , Kaplan Allen P.
Published: May 2019

The molecular weight (mol. wt.) of factor XI in normal plasma fractionated on Sephadex G-200 was over 400,000, while the mol. wt. of factor XI in HMW kininogen deficient plasma was 175,000. When HMW kininogen deficient plasma was made 70 μg/ml in HMW kininogen and fractionated on Sephadex G-200, factor XI was again found at mol. wt.~ 400,000. Prekallikrein was found complexed to HMW kininogen (Mandle and Kaplan PNAS 73: 4179, 1976) however, no complex containing both prekallikrein and factor XI was identified and neither Hageman factor nor plasminogen were complexed to HMW kininogen. Gel filtration of normal plasma had a major peak of HMW kininogen at mol. wt. of 200,000 while isolated HMW kininogen had a mol. wt. of 130,000 on reduced SDS gels (SDS-PAGE). Alkaline disc gel electrophoresis of HMW kininogen revealed two bands of equal intensity and each possessed the ability to correct the coagulation defect of HMW kininogen deficient plasma. Digestion of HMW kininogen with kallikrein generated bradykinin and a peptide of mol. wt. 13,000, while SDS-PAGE of the residual HMW kininogen revealed bands of mol. wt.110,000 and 120,000. Upon reduction, bands at mol. wts. 77,000, 66,000, and 37,000 were obtained. Antibody specific for HMW kininogen reacted with either native or kallikrein-treated HMW kininogen but not reduced, kinin-free kininogen. Kinin-free kininogen, reduced kinin-free kininogen, and the major fragment eluted from alkaline disc gels after electrophoresis of reduced kinin-free kininogen all retained coagulant activity. Factor XI as well as prekallikrein therefore circulate complexed to HMW kininogen; these complexes may then interact with surface-bound Hageman factor. Kallikrein digests human HMW kininogen to yield bradykinin, a peptide, and disulfide-linked fragments which retain functional activity.
Molecular Weight Distribution Studies on Heparin
Authors: Barlow Grant H.
Published: May 2019

The determination of molecular weight distribution using the sedimentation equilibrium analysis developed for polymers by T. Scholte (J. Polymer Sei, 6, 111, 1968) has been adapted for heparin analysis. Pork mucosal heparin separated into molecular weight subfractions by gel filtration on Ultrogel AcA44 (L.K.B.) was used to test the validity and resolving power of the method. Results indicate that the method is able to differentiate molecular weight distribution satisfactorily. Comparisons have been made of molecular weight distribution of samples from different species, organs and manufacturers. Average molecular weights for most samples center around 15,000 Dal tons, but samples show considerable variation in their distribution data. Results suggest that variations between manufacturers is more pronounced than the specie and organ difference indicating the importance of the purification procedure.
Interactions Among Heparin, Cold-Insoluble Globulin, and Fibrinogen in Formation of the Heparin Precipitable Fraction of Plasma
Authors: Stathakis N. E. , Mosesson M. W.
Published: May 2019

Fibrinogen and the cold-insoluble globulin of plasma (CIg) are the main protein components of the heparin cryoprecipi table fraction (HPF) of normal plasma. The interactions between these proteins and heparin were examined. Heparin formed a cold precipitable complex with purified CIg or with mixtures of CIg and fibrinogen (T/2, 0.2; pH 7.2) but not with fibrinogen alone. Cryoprecipitation could be augmented by addition of Ca++ or by selection of optimal heparin levels; it could be reduced or even abolished by raising the ionic strength or pH or both, or by raising the heparin level above that needed for maximum precipitation of CIg. Fibrinogen reduced the threshold level of CIg at which heparin-induced cryoprecipitation occurred and, by co-precipitating with heparin and CIg, increased the total precipitate that formed. In contrast to the HPF from normal plasma which contained both fibrinogen and CIg, that from afibrinogenemic plasma contained CIg but lacked fibrinogen. Normal plasma depleted of CIg failed to form a heparin-induced cryoprecipitate. Thus, CIg is essential for heparin-induced cryoprecipitation to occur. Fibrinogen, as assessed by chromatographic experiments with heparin-Sepharose columns, has a considerably lower heparin-binding affinity than does CIg, indicating that it participates in formation of the HPF mainly, if not entirely, by virtue of its affinity for CIg.
Amino Acid Sequence Studies on the β-Chain of Human Fibrinogen
Authors: Watt K. , Goldbaum D. , Cottrell B. A. , Takagi T. , Doolittle R. F.
Published: May 2019

The β-chain of human fibrinogen contains 480 ± 15 residues, sixteen of which are methionines. In this vein, we have isolated and characterized all seventeen cyanogen bromide peptides. The arrangement of most of these fragments has been achieved by the identification of key overlap peptides derived from enzymatic digestion of β-chains, on the one hand, and a characterization of β-chain fragments isolated from fragments D and E, on the other. In some cases the alignment is based only on homologies with the α- and/or γ-chains, and in a few instances some definite ambiguities still exist. For the most part, however, the general arrangement is in hand. Of particular interest is the plasmin-sensitive segment which is a part of the inter-domainal connection, which in turn we believe is a three-stranded coiled-coil punctuated by an unusual arrangement of disulfide bonds. The positioning of the other cysteine residues in the β-chain is also of considerable interest, since it sheds light on the extent of connections between portions of the three non-identical chains in the parent molecule.
Complement Dependent and Independent Effects of Bacterial Products on Rabbit and Human Platelets
Authors: Semeraro N. , Vermylen J.
Published: May 2019

The effect of 13 lipopolysaccharides (LPS) from aerobic and two LPS from anaerobic bacteria, two mucopeptides and two meningococcal polysaccharides, on rabbit platelets in citrated and heparinized plasma was studied by aggregometry. The LPS from aerobic bacteria showed variable aggregating activity. LPS from S. Minnesota Re 595 (essentially lipid A) was one of the most active. The two mucopeptides were very active whereas the LPS from anaerobic bacteria and the meningococcal polysaccharides were inactive. These effects were abolished by inactivation of complement and by ethyleneglycoltetraacetic acid (EGTA) indicating that they are dependent on activation of the classical pathway. None of these effects was observed using human platelets.

When washed rabbit platelets are mixed with complement-depleted rabbit serum and calcium chloride, generation of thrombin occurs. All the LPS preparations but not the mucopeptides or meningococcal polysaccharides enhanced the rate of thrombin formation to a variable extent. Identical effects have also been found using human platelets (Brit. J . Haemat. in press).

It is concluded that LPS affect rabbit platelets by separate complement dependent and independent mechanisms. However with human platelets only the complement independent phenomenon (enhancement of thrombin generation) is observed.