bluuebunny/crossref_metadata_2025 huggingface dataset:
Early Platelet Activation During Perfusion of a Hollow Fiber Artificial Kidney
Authors: Kim B. K. , Baldini M.G.
Published: May 2019

Studies of early platelet changes caused by contact with artificial surfaces in a flow-system were done by the use of a bench-model of a hollow fiber (silicone rubber) mini-kidney. The artificial device contained 64 fibers in a polyurethane casing and was inserted in a silicone rubber circuit provided with a roller pump. Human platelet-rich plasma was used for perfusion. Perfusion for 60 m’ at a rate of 6 ml/m' caused no decrease in platelet count and no visible thrombus formation in the fibers. However, after perfusion for 5 m' there was a significant increase in ADP induced platelet aggregation with values of 75 ± 10% (control 56 ± 5.8%). The values rose to 80 ± 8.5% after 30 m' of perfusion. Collagen induced platelet aggregation also increased with values of 81 ± 9.3% (control 67 ± 5.3%) after 5 m' and 82 ± 6.3% after 30 m'. Platelet F3 availability measured by the Kaolin clotting time was 108 ± 5.5 sec before perfusion and became progressively enhanced during perfusion. The Kaolin clotting time became 93 ± 2.9 sec after 5m’, 84 ± 1.7 sec after 15 m', 76 ± 3.3 sec after 30 m' and 71 ± 3.7 sec after 60 m'. Platelet release of 14C-serotonin was only 2% to 3% after 60 m' perfusion. The content of purine nucleoside Phosphorylase (a cytoplasmic enzyme) and of β-glucuronidase (a lysosomal enzyme) detected in the plasma medium after 60m’ perfusion were only 1% to 2% of the respective platelet enzyme activity. It was concluded that contact with silicone rubber surfaces during short-time perfusion of hollow fibers caused significant platelet activation with enhancement of platelet aggregability and platelet F3 availability in the absence of significant degrees of release reaction.
Platelet Dysfunction with Glutathione Reductase Deficiency After 1,3-Bis(2 Chloroethyl)-l-Nitrosourea
Authors: McKenna R. , Ahmad T. , Frischer H.
Published: May 2019

We have previously shown that patients receiving antitumor chemotherapy with 1,3-bis-(2 chloroethyl)-1-nitrosourea (BCNU) rapidly acquire a severe generalized glutathione reductase (GSSG-R) deficiency. We now report that when platelets were exposed in vitro to BCNU (30 minutes, 37°, at 10-3 M, six separate studies), the resulting GSSG-R deficiency was associated with marked impairment of platelet aggregation in response to ADP (1 μM and 3 μM), to epinephrine and to collagen (all p’s< 0. 001). Platelet factor 3 availability was also markedly reduced (p<0.001); prothrombin consumption and glass adhesiveness were unaffected. In additional experiments to evaluate the dose response relationship, epinephrine induced aggregation was abnormal at 5 × 10-6 M BCNU, ADP (1 μM) induced aggregation was abnormal at 10-5 M BCNU, while ADP (3 μM) and collagen aggregation became abnormal at 5 × 10-5 M BCNU. GSSG-R deficiency (less than 11% of control activity), without glucose-6-phosphate dehydrogenase and 6-phosphogluconic dehydrogenase deficiencies, preceded all platelet abnormalities. We have demonstrated that a specific platelet GSSG-R deficiency after BCNU precedes the development of severe platelet dysfunction.
The Source of Hydrogen Peroxide and of Chemiluminescence Observed in Activated Human Platelet Preparations
Authors: Levine P.H. , Scoon K.L. , Hardin J.C. , Krinsky N.I.
Published: May 2019

Human platelet suspensions can be observed to produce small amounts of H2O2 (.04 nmoles H2O2/min/2.5×105 cells mm-3) and measurable chemiluminescence when exposed to target particles for phagocytosis, such as latex spherules. Both H2O2 production and chemiluminescence are characteristic of phagocytosing polymorphonuclear leukocytes (PMN) and analysis of the purified platelets indicates contamination by PMN at the level of 0.2%. The amount of H2O2 produced and the chemiluminescence observed can be duplicated by adding latex spheres to a preparation of PMN at a concentration equivalent to the contaminant in the platelet preparations. We conclude that the H2O2 produced and chemiluminescence observed from activated platelets is due to the presence of small amounts of contaminating PMN. The production of H2O2 and free radicals by the PMN which contaminate platelets may contribute to loss of platelet function during storage. These studies also indicate the importance of controlling for PMN contamination in studies of platelet biochemistry and physiology.
Studies on Anti-Platelet Antibodies. Its Correlation with ACID Phosphatases. Preliminary Results
Authors: Hendler H.N. , Sack E.S.
Published: May 2019

The platelet factor 3 immuno-injury technic (Karpatkin and Siskind, Blood 33:795,1969) has been evaluated as a test for the detection of anti-platelet antibodies in 126 patients with quantitative and qualitative platelet disorders. Plasmatic acid phosphatases in platelet-poor plasma (PAP-PPP) were simultaneously determined in a selected group of patients. Also the effect of serum incubation (from patients and controls) on acid phosphatase availability of platelet-rich plasma (APA-PRP) was investigated.

Anti-platelet antibodies were present in 52% of patients fulfilling the diagnostic criteria for idiopathic thrombocytopenic purpura and in 38% of patients with miscellaneous conditions. The following table express the results of acid phosphatase determina studied:

Platelet destruction is the probable cause of the thrombocytopenia in the group with negative anti-platelet antibodies. Further, their serum lacks membrane-injury action. On the other hand the results in the group with positive antibodies suggest that heterogeneous immunologic mechanisms are operative
Morphologic Platelet Changes in Citrate Blood and PRP
Authors: Wiedemann R. , Breddin K. , Grun H.
Published: May 2019

Using high power Nomarski optics the film presents the shape change of thrombocytes in vitro. Their native shape can be studied by immediate fixation of blood at venepuncture. Platelets appear as flat discs. Less than 25% show pseudopodes. In citrate blood and PRP a pro-qressing shape change occurs depending on time after blood sampling and on incubation temperature. Platelets tend to swell and tentacle formation continues with time. At room temperature 90% of thrombocytes have undergone these changes within 60 min. The addition of ADP (10-6 molar) to PRP induces the formation and rupture of large vesicles in single platelets. Thrombocytes are aggregating, many of the aggregated platelets show the same large vesicles. They are bursting and the release of a granular material can be observed. The remaining platelet material is fusing to an unstructured mass. Bencyclan (2 × 10-4 molar) affects platelet morphology by inducing a spherical transformation, which is paralleled by the inhibition of platelet adhesiveness, spreading and aggregation. Finally a platelet is shown spreading on a glass surface. The mor-nhologic changes of single platelets (primary shape change) demonstrated in the film probably represent basic processes in hemostasis and thrombus formation.
Platelet Water Content is Decreased by Gel-Filtration
Authors: Wiley J.S. , Wray G. , Cooper I.A.
Published: May 2019

One approach to platelet sizing is to measure the intracellular water space of platelets with 3H-H2O since the % water content of platelets remains constant in states with different platelet sizes. Fresh citrated blood was centrifuged for 10 min at 150 'g' to obtain PRP. Aliquots of PRP were briefly incubated with either 3H-H2O or 14C-sucrose then layered over 0.3 ml dibutylphthalate and spun 4 min at 8000 'g'. The cell pellet was solubilized and counted to enable spaces to be calculated. The extracellular (sucrose) space was subtracted from the total water space of the pellet to give a mean intracellular water space of 0.56 ± 0.12 μ1/108 platelets (n =19). Assuming a water content of 7 5% and a density of 1.04, the mean platelet volume for normal subjects is 7.2 fl. Gel-filtration of platelets (GFP) on Sepharose-2B reduced their mean water space by 0.12 μl/108 platelets. However the amount of shrinkage on gel-filtration depended on the initial water space of the platelets in PRP and there was a linear relation between these two variables (r = 0.82). Shrinkage was 40% for an initial platelet water space of 0.70 μl/108 platelets but there was almost no shrinkage below a water space of 0.40 μl/108 platelets. Recovery of platelets from each column averaged 8 0% and showed no relation to the reduction in the mean cell water space. The lower water space of GFP may indicate a reduction in mean cell volume due to gel-filtration.
Circulating Crosslinked Fibrinoligomers. Clinical Observations and Turnover Studies in Rabbits
Authors: Graeff H. , von Hugo R. , Hafter R.
Published: May 2019

Blood samples from patients with coagulation disorders in obstetrics and with advanced carcinoma of the kidney were examined for the presence of cross-linked fibrinoligomers. Quantitative gel filtration of ß-alanine precipitated plasma samples and chain characterization of isolated fibrin derivatives by SDS-PAA gel electrophoresis after reduction with mercaptoethanol were performed. Electrophoresis of immunoadsorbed material was additionally applied. In all cases of intravascular coagulation crosslinked fibrinoligomers in amounts of 8-25 per cent of the total fibrinogen content were observed. Severe cases revealed a molecular weight pattern of derivatives ranging from 5 million and more down to 45 000. X, Y, D, E and D-dimer were found in the ß-alanine supernatant. In 5 patients with advanced renal carcinoma a cross-linked dimeric derivative was observed predominantly.

In vitro produced crosslinked 125 I-fibrinoligomers were injected into rabbits. Radioactivity was measured in gel filtrated fractions from ß-alanine precipitated samples, and a half-life time of approximately 7 hours of the high molecular weight fraction averaging 5 million daltons was found.

It is concluded that circulating crosslinked fibrinoligomers which differ in regard to complex formation and half-life time to soluble fibrin monomer complexes may indicate intravascular coagulation.
Horseshoe Crab Coagulogen: Its Structure and Gelation Mechanism
Authors: Iwanaga S. , Nakamura S. , Takagi T. , Morita T. , Finlayson J.S. , Takahashi K. , Niwa M.
Published: May 2019

A clottable protein, isolated from the amoebocyte lysate of horseshoe crab (Tachypleus tridentatus) was a basic protein with a molecular weight of approximately 17,000. It contained a total of 155(± 5) amino acid residues and had a single NH2-terminal alanine and COOH-terminal phenylalanine. Upon gelation of the coagulogen by an endotoxin-activated clotting enzyme(s), a large peptide, named peptide C, which has 28 amino acid residues, was released, and the resulting gel protein consisted of A and B chains, bridged by disulfide linkages. The chemical analyses on these polypeptide segments indicated that the enzymatic formation of gel involves limited proteolysis of the Arg-46-Gly-47 and Arg-18-Thr-19 bonds located in the N-terminal portion of coagulogen, liberating peptide C. Moreover, the amino acid sequence studies on peptide C and A chain provided that their COOH-terminal octapeptide sequences have a remarkable homology with primate fibrinopeptide B. Up to now, the following NH2-terminal sequence of whole coagulogen was established: NH2-Ala-Asp-Thr-Asn-Ala-Pro-Ile-Cys-Leu-Cys-Asp-Glu-Pro-Gly-Val-Leu-Gly-Arg-Thr-Gln-Ile-Val-Thr-Thr-Glu-Ile-Lys-Asp-Lys-Ile-Glu-Lys-Ala-Val-Glu-Ala-Val-Ala-Gln-Glu-Ser-Gly-Val-Ser-Gly-Arg-Gly-Phe-Ser-Ile-Phe-Thr-His-Arg-Pro-Val-Phe-Arg-Glu-Cys-Gly-Lys-Tyr-Glu-Cys-Arg-Thr-Val-His-Arg-Pro-Glu-Ser-Arg-Cys-Tyr-Asn-Phe-Pro-Pro-Phe-Thr-His-Phe-Lys------------Phe-COOH.
Studies on the Degradation of Human Fibrinogen by the Action of “Caterpillin”
Authors: Arocha-Piñango C. L. , Perales J.
Published: May 2019

Degradation products of human fibrinogen formed by the action of “Caterpillin” (a proteolytic substance from the hemolymph of a saturnid moth caterpillar) was studied byimmuno---electrophoresis and polyacrilamide gel electrophoresis. The pattern of X, Y, D and E degradation products differ from those formed by the action of plasmin and trypsin. Chain characterization during the degradation process indicated a very rapid loss of the α chain.
Platelet Contractile Proteins
Authors: Pollard T.D.
Published: May 2019

Platelets contain a high concentration of the contractile proteins actin and myosin. The properties of these proteins are generally similar to their counterparts in muscle, suggesting that the platelet actin and myosin can generate contractile force. Both proteins are localized in the cytoplasm and little or none of either is present on the outer surface. The distribution of actin in the filopodia and myosin in the body of activated platelets suggests that the force for clot retraction is generated by retraction of the filopodia using a mechanism similar to that in muscle contraction.
Low Affinity Platelet Factor 4 and High Affinity Platelet Factor 4 - Two Antiheparin Proteins Secreted by Human Platelets
Authors: Niewiarowski S. , Rucinski B. , Budzynski A.Z. , Subbarao K.
Published: May 2019

A material with antiheparin activity released by thrombin from human washed platelets was precipitated by 0.04 M ZnSO4 and applied on heparin-agarose column. Fractions eluted at 0.5 M and 1.5 M NaCl contained antiheparin activities and were denoted as low affinity (LA-PF4) and high affinity (HA-PF4) platelet factor 4, respectively. In Factor Xa-heparin inhibition assay 1 mg of each LA-PF4, HA-PF4 and protamine sulfate neutralized 0.86, 7.6 and 11.7 units of heparin, respectively.LA-PF4 and HA-PF4 were homogenous in SDS electrophoresis and had apparent MW of 13,700 and 10,300, respectively. Treatment with reducing agents had no effect on the migration of HA-PF4 but it decreased apparent MW of LA-PF4 by 50%. It has been shown by immunodiffusion and by radioimmunoassay that LA-PF4 and β-thromboglobulin (βTG) share common antigenic determinants. LA-PF4 and HA-PF4 were antigenically different. LA-PF4 and human PF4 prepared previously in our laboratory by DEAE cellulose column chromatography appeared to bè identical proteins. βTG and LA-PF4 had similar patterns of migration on SDS electrophoresis in reduced and non-reduced systems and similar amino acid composition. However, on cellulose acetate electrophoresis, LA-PF4 appeared as one homogenous band both at pH 8.4 and 9.9 which migrated in the γ-globulin region in contrast to βTG migrating with the β-globulin mobility. Migration of LA-PF4 was not affected by Zn ions. Secretion of LA-PF4 from stimulated platelets occurred in parallel with ADP and serotonin and preceded secretion of HA-PF4. In conclusion, thrombin releases from human platelets two distinct proteins with different antiheparin activities.
Physiological and Clinical Significance of Fibrin Polymerization
Authors: Beck E.A.
Published: May 2019

The physiological importance of fibrin polymerization is not well understood in man. In fact, severe disorders of fibrin polymerization (e.g. in congenital dysfibrinogenemia) are mostly “laboratory diagnoses” without clinical implications. Pathophysiologically, fibrin polymerization may be rated either as “favorable” or “unfavorable”: fibrin deposition supports hemostasis, promotes wound healing and confines activated coagulation enzymes mechanically, but participates in thrombosis, favors serous mucosal adhesions and may block phagocytosis. In many diseases, the balance between favorable and unfavorable effects of fibrin polymerization is delicate. Therapeutically, fibrin polymerization should probably be inhibited more often than promoted.
Physiological and Clinical Significance of Disordered Cross-Linking of Fibrin
Authors: Lorand L.
Published: May 2019

Disorders of fibrin stabilization are hemorrhagic conditions in which the patient’s plasma clot is lacking in inter-fibrin γ-glutamyl-ε-lysine isopeptide linkages. The primary defect occurs either because no fibrinoligase (FXIIIa) activity can be generated or because the enzyme cannot act on fibrin in the patient’s plasma. Distinction is made between hereditary disorders (Class I) and those appearing later in life because of an acquired inhibitor (Class II) directed against one of the steps on the pathway of fibrin stabilization:

Of the genetic deficiencies (Class I), Type I is characterized by a lack of zymogen activity in plasma and Type II by the unreactivity of the cross-linking sites of the patient’s fibrin [“dysfibrin(ogen)emias”] towards fibrinoligase.

There are three varieties of Class II abnormalities. In Type I, the acquired inhibitor interferes with zymogen activation. Type II inhibitors affect transamidation by competing against fibrin for the enzyme. The Type III inhibitor combines with fibrin rendering it unreactive towards fibrinoligase. The Type I and III inhibitors appear to be autoimmune antibodies.(Ann. N. Y. Acad. Sei., 202, 6, 1972).

Differential diagnostic criteria for this family of molecular disorders will be discussed.
Isolation of Platelet Membrane Collagen Receptor
Authors: Livne A. , Swanson A.L. , Scheuenstuhl H. , Michaeli D.
Published: May 2019

A fraction which competes with intact platelets for interaction with collagen was isolated from human platelets by Sepharose 2B-collagen (SC) affinity columns. Binding of platelets to SC columns, followed by lysis with a nonionic detergent and extensive wash with water (treated SC columns), reduced the columns’ capacity to subsequently bind platelets and induce serotonin release. This reduction was proportional to the number of platelets that had been applied to SC columns. Treated SC columns could be partially regenerated with solutions of high ionic strength (1 M NaCI or Tris-HCl) and most effectively with 0.3% sodium dodecylsulfate (SDS), but not with 8 M urea, 15% ethanol or 1% Triton X-100, indicating an ionic interaction. A fraction eluted with SDS from treated SC columns manifested receptor activity: when rebound to collagen it caused a dose-dependent decline in interaction of the collagen with intact platelets, as measured by binding and serotonin release. The receptor activity was sensitive to heat and was absorbed by an anionic exchange resin. When the membrane of intact platelets were labeled with 125I and lactoperoxidase, the derived receptor fraction contained a small proportion (about 1%) of the label.
Effects of Photolysable Azido-Analogues of Adenosine, AMP and ADP on Human Platelets
Authors: Cusack N.J. , Born G.V.R.
Published: May 2019

The aggregating effect of ADP on human platelets and the inhibiting effect of adenosine are apparently mediated by different receptors on the cells’ outer membranes (R.J. Haslam & G.M. Rosson, 1975, Molec. Pharmacol., 11, 528). Photolysable azido-analogues of adenosine & ADP have been prepared so that the receptors for them can be labelled. 2-Azidoadenosine inhibits adenosine deaminase competitively before and irreversibly after irradiation at 365 nm (N.J. Cusack and G.V.R. Born, 1976, Proc. Roy. Soc. B, 193, 307). 2-Azidoadenosine inhibited platelet aggregation by ADP more than adenosine itself and increased platelet cAMP as effectively as did adenosine or 2-chloroadenosine. 2-Azido AMP inhibited aggregation much more effectively than AMP itself or than 2-chloro AMP. 2-Azido ADP was about 5 times more potent than ADP in causing aggregation. All these 2-azido derivatives were photolysable by irradiation at 365 nm which did not affect platelet functions.
The Number and Nature of ADP Receptors on Human Blood Platelets Determined with a Photoaffinity Label
Authors: Macfarlane D.E. , Mills D.C.B.
Published: May 2019

ADP interacts with platelets to cause shape change, aggregation and inhibition of adenylate cyclase. Previous attempts to measure the binding of ADP to its receptor on intact platelets have been frustrated by the low ratio of specifically bound ADP to the ADP trapped in centrifuged pellets. Several 2-substituted derivatives of ADP have higher apparent affinities for the ADP receptor, improving this ratio. We have prepared 2-azido 5'-ADP from 2-chloroadenosine in 20% yield. It was more active than ADP as an antagonist of PGE1-induced elevation of cyclic AMP (Ki = 96 nM, cf ADP Ki = 760 nM) and also as an inducer of shape change and aggregation. 2-Azido [β32P]-5'-ADP (2.5 Ci/mmole) was prepared and used to study binding. Platelets were separated from plasma by centrifugation through silicone oil (1.023 gm/ml) in a modified Eppendorf centrifuge. Scatchard plots of the binding data were resolved into two linear components, one of zero affinity, corresponding to trapped plasma, measured independently with [14C] sucrose, and a high affinity component with apparent KD = 110-160 nM. There were 400-700 of these binding sites per platelet. 2-Azido 5'-ADP is photolysable with light at <310 nm, forming a reactive nitrene potentially suitable for photoaffinity labelling. 2-Azido [β32P]-5'-ADP was photolysed in the presence of washed platelets which were subsequently rewashed, solubilized and electrophoresed on SDS-polyacrylamide gradient slabs. Several peaks of radioactivity were observed in the high molecular weight region of the gel (ca 100-250 kD) one of which was less labelled in the presence of excess ADP or ATP to block access of the label to the receptor.
The Platelet-Leukocyte Aggregates in Occlusive Arterial Disease and the Effect of Hydroxyethylrutosides
Authors: Bielawiec M. , Kiersnowska-Rogowska B. , Łukjan H.
Published: May 2019

The diagnostic value and the effect of the therapy on the number and kind of platelet-leukocyte aggregates in patients with arteriosclerosis obliterans of the lower limbs were investigated. The modified Silbergleit’s method was used. 8 types of aggregates were found. The mean number of aggregates in 20 healthy subjects was 8.2, in 52 patients suffering from obliterative arteriosclerosis of the lower limbs this number was significantly higher, that is 63.9. In healthy subjects most of the aggregates were type 1 to 4 whereas in the patients types 3 and 5 were most frequently found. 30 patients were treated for 3 weeks with hydroxyethylrutosides /HR/ - orally 600 mg three times daily or intravenously 1000 mg twice a day. A significant decrease of the number of platelet - leukocyte aggregates in the groups of patients receiving HR was found. The type of aggregates after HR was similar to that found in the control group. In 22 patients receiving nicotinic acid derivatives /orally 300 mg three times daily and intramuscularly 300 mg twice a day/ for 3 weeks no significant changes in the parameter investigated was observed. The method of differential counting of platelet-leukocyte aggregates may be of value in early diagnosis of occlusive arterial disease and in evaluation of their therapy.
Intraperitoneal Activity of Heparin During Peritoneal Dialysis
Authors: Gomperts E.D. , Furman K.I. , Hockley J.
Published: May 2019

Heparin is frequently added to peritoneal dialysate to prevent the formation of thrombi with the resulting obstruction of peritoneal catheters. As a guide for such therapy the pharmacokinetics of intraperitoneal heparin was studied in 11 patients undergoing maintenance peritoneal dialysis. The heparin activity was assessed by adding dialysate to control plasma and measuring the prolongation effect on the activated partial thromboplastin time (A-PTT). It was observed that the A-PTT was prolonged in proportion to the amount of heparin in the peritoneal fluid. The decay of this activity was relatively slow with the mean T½ in the peritoneal cavity being 10.78 ± 0.93h. Systemic blood coagulation was unaffected by single 10,000 U intraperitoneal doses of heparin in that plasma A-PTT's were not lengthened over the ensuing 6 hours. Antithrombin III assessed by immunochemical and functional procedures was present in low concentrations in residual peritoneal fluid aspirated prior to commencing dialysis. Generally this was less than 1/3 of normal plasma values, and with the repeated dilution and outflow sequences of dialysis, the cofactor concentrations fell to negligible levels, usually below 1% by the end of the second cycle. These results indicate therefore that despite the persistence and slow decay of heparin within the peritoneal activity, therapeutic efficacy is unlikely to be achieved except in those cases where adequate cofactor might be introduced as a result of massive intraperitoneal haemorrhage.
Functional Properties and in Vivo Survival of Normal and Asialo Human Factor VIII/VON Willebrand Protein
Authors: Sodetz J.M. , Pizzo S.V. , McKee P.A.
Published: May 2019

Recent evidence suggests that the molecular defect in von Willebrand’s disease resides in the carbohydrate moiety of Factor VIII/von Willebrand factor (FVIII/vWF). In light of this, we have examined and compared certain properties of normal and asialo FVIII/vWF. Purified human FVIII/ vWF was desialylated using a protease-free neuraminidase. At various incubation times, sialic acid released, ristocetin-induced platelet aggregating activity(RPA) and procoagulant activity (PCA) were measured. RPA decreased with increasing amounts of sialic acid released. When completely desialylated (>95%), only 35±10% of the initial RPA was retained. In contrast, PCA remained constant with up to 80-85% sialic acid removed. Normal and asialo FVIII/vWF displayed similar immunological cross-reactivity to normal FVIII/vWF rabbit anti-sera. However, immuno-electrophoretic results clearly indicated a reduced anodic mobility for asialo FVIII/vWF relative to normal. Normal and asialo FVIII/vWF were then radiolabeled with 125I, infused in rabbits and the circulatory survival times measured. Asialo FVIII/vWF was cleared from circulation at a rate 50-fold greater than that observed for normal FVIII/vWF. This clearance was accompanied by quantitative appearance of radioactivity in the liver. In addition, simultaneous infusion of human asialo α1-acid glycoprotein, a protein known to bind to hepatic asialoglycoprotein receptors, competitively inhibited asialo FVIII/vWF clearance. These results indicate that desialylation of FVIII/vWF does not alter PCA, decreases RPA and, as observed with other plasma asialoglycoproteins, facilitates its rapid hepatic clearance. Understanding these properties of asialo FVIII/vWF represents an important initial step towards defining a possible carbohydrate defect as a cause for von Willebrand’s disease.
Immunoelectrocataphoresis of Fibrinogen: A New Method Applied to the Detection of Fibrinogen Electrophoretic Abnormalities
Authors: Ruíz-Reyes G. , Ruíz N. Landero de , Armenta-Olvera T.
Published: May 2019

Migration of fibrinogen in an electric field is useful to separate abnormal fibrinogens in those which migrate normally, anodally of cathodally. Using the conventional Immunoelectrophoresis technic of plasma, the distinction between normal mobilities and the slightly abnormal ones is not easy to establish in some cases. After electrophoresis of plasma in agar gel, a very clear visualization of the fibrinogen motility is obtained by precipitation of the protein with a monoespecific antiserum applied to the gel surface in cellulose acetate strips. The name of “immunoelectrocataphoresis” is proposed to call the method (from the greek cata=downward) considering that this denomination describes clearly the basic principle involved in the procedure and the descending direction of the antibody to meet and precipitate with the antigen. It is faster and cheaper than the conventional immunoelectrophoretic technique and can be easily incorporated into routine laboratory work. The method can be applied to study the fibrinogen motility of hereditary or acquired cases of dysfibrinogenemi as.
Primary Shape Change of Thrombocytes and Platelet Aggregation after Administration of Acetyl Salicylic Acid (ASA)
Authors: Krzywanek H.J. , Breddin K.
Published: May 2019

Studies using interference contrast microscopy (Nomarski optics) with immediate glutaraldehyde fixation of blood at venepuncture revealed the disc like shape of native thrombocytes. Soon after blood drawing platelets in citrate blood or PRP are swelling and the formation of pseudopodes can be observed. If citrate blood is incubated at 37°C, 38% of the thrombocytes have formed pseudopodes after 30 min. The form variation is inhibited in some platelet defects (storage pool disease, aspirin like defect) and is enhanced in certain malignancies (Hodgkin’s disease, bronchial carcinoma). After oral intake or intravenous administration of 1.0 g ASA the primary shape change is partly inhibited for a period of six hrs. with the greatest effect at two hrs. In contrast to this short lasting effect on primary shape change the inhibition of ADP or collagen-induced aggregation by a single dose of ASA can be demonstrated for at least four days.

We suggest that the short lasting inhibition of the primary shape change of platelets may be responsible for the efficacy of ASA in postoperative thrombosis prophylaxis.

Regular intake of ASA in doses of 0.5 g each at 6 - 8 hrs. intervals may be necessary to achieve a continuous antithrombotic effect. Our results may explain why studies on thrombosis prophylaxis with aspirin using 0.6 g (BMRC, 1973) or 0.3 g (Elwood, 1974) per day were inconclusive whereas 3 × 0.5 g reduced the thrombosis frequency significantly (Zekert, 1973, Loew, 1973).
The Effects of Cigarette Smoking on the Haemostatic System
Authors: Meade T.W. , Chakrabarti R. , Haines A.P. , North W.R.S. , Stirling Yvonne
Published: May 2019

The main purposes of the Northwick Park Heart Study (NPHS) are to improve the prediction of ischaemic heart disease (IHD) and to elucidate its pathogenesis. Measures of haemostatic function are made along with those of variables already known to be associated with IHD; one such variable is cigarette smoking. It is clearly of interest to know whether the smoking effect may be mediated through changes in haemostatic function; data at recruitment in 1392 men and 580 women have therefore been examined in order to study such changes. Since certain characteristics in NPHS, particularly age and social class, are correlated with both cigarette smoking and with levels of some haemostatic variables, multiple regression analysis has been used to identify smoking effects independent of associations between smoking and these other characteristics. The haemostatic variables included in this, analysis are factors V, VII and VIII, fibrinogen, antithrombin III, fibrinolytic activity, platelet count and platelet adhesiveness. In men, factor VIII levels and fibrinolytic activity are significantly lower in smokers than non-smokers; fibrinogen levels are significantly higher. In women, factor VIII levels are also significantly lower in smokers, but there is no difference in fibrinogen; for fibrinolytic activity there is an interaction between smoking and the use of oral contraceptives (OC). Activity in non-smokers on OC is substantially higher than in those not on OC, but activity in smokers on OC is the same as in women not on OC. It therefore seems that smoking abolishes the protective rise in fibrinolytic activity caused by OC. In men, cigarette smoking may promote thrombosis through effects on fibrinogen and fibrinolytic activity.
Artificial Substrate Plasma for the Specific One-Stage Factor X Assay
Authors: Beeser H. , Kulzer R.
Published: May 2019

As patients who are congenitally deficient in factor X are extremely rare natural substrate plasma for the specific one-stage factor X assay is only poorly available. Asbestos-filtered or charcoal-filtered ox plasma deficient both in factor X and VII is instead widely used as artificial factor X substrate plasma with Russel’s viper venom (BW) and cephalin or lecithin as the thromboplastic agent.

We studied the practicability of removing separately factor X from the plasma of different species (man, horse, ox, sheep, swine) by several chromatographic procedures. Chromatography of swine plasma on DEAE-Sephadex A 25 yielded a substrate virtually free of factor X while containing adequate concentrations of fibrinogen and factor II, V and VII, Using different brands of commercial tissue thromboplastins the so prepared swine plasma has proved to be a specific artificial substrate plasma for the one-stage factor X assay. Calibration curves obtained by plotting the clotting times of various dilutions of pooled normal human plasma determined with this factor X assay system against the percentages showed a linear relationship from 100% to about 2% on log/log paper. Factor X determinations with our method on various patients’ plasmas (i.e. coumarin treatment, congenital factor X deficiency, liver disease) performed in parallel to the RVV-cephalin and the congenital factor X deficient plasma methods gave comparable results.

As our specific artificial factor X deficient swine plasma is fairly simple prepared with a good yield, it has to be considered as an appropriate reagent to overcome the shortness of the rare congenital factor X deficient substrate plasma.
Fibrinogen-Fibrin Monomer Complexes in Cirrhosis of The Liver
Authors: Coccheri S. , Palareti G. , Poggi M. , Oca G.
Published: May 2019

Positivity of paracoagulation tests in a previously studied group of 80 patients with chronic active liver disease did not exceed 5-10% of the cases. In the present study, plasma samples from 20 cases of decompensated liver cirrhosis, assessed by liver biopsy, were investigated by means of agarose cromatography. Fibrinogen related materials were measured immunologically and by Staphylococcal Clumping Test.

First appearance of fibrinogen-like materials occurred at earlier fractions in cirrhotic patients in comparison with normal controls. The relative amount of soluble fibrin monomer complexes (SFMC) as referred to total fibrinogen was significantly increased. No correlation was found between the amount of SFMC and the severity of fibrinogen polymerisation defect.

Circulating SFMC are therefore present in severe liver cirrhosis. However, DIC may not be the only proposed explanation for this finding.
Sensitivity of Thromboplastins
Authors: Altman R. , Rouvier J. , Mosquera G.
Published: May 2019

Using dilutions 1/1-1/16 of different thromboplastins and undiluted plasma, the prothrombin times were longer with the increase of dilutions. When plasma was diluted in saline or buffer (1/5/1/10), a proportional shortness of the clotting times were observed. With adsorbed plasma as diluent a progressive lengthening of the prothrombin times were obtained, related with the clotting times of the undiluted plasmas. Express as ratios, all thromboplastins showed shortness when plasma was diluted with saline or buffer but rather constant values with adsorbed plasma dilutions. Then, thromboplastins were concentrated and, although times of undiluted plasmas were rather similar, the ratios of the different concentrations of tissue extract, considering the 1/10 dilution of plasma, were different. In terms of the so called “sensitivity” this means a better sensitivity when thromboplastin concentration was increased. Similar results were achieved when thromboplastin was additionated with fenol. We concluded that many tissue extracts may achieve a “good sensitivity” by increasing it concentration in the test tube Why plasmas diluted with adsorbed plasma gives constant ratios, remains unknown for us. We feel that standardization of thromboplastin should be made on more biochemical basis.
Inactivation of Trypsin By Antithrombin III
Authors: Feinman Richard D. , Chang Tien-ling
Published: May 2019

We have studied the reaction of antithrombin III (AT) with trypsin as a model of the interaction of the inhibitor with the serine proteases of the coagulation scheme. From equilibrium binding studies, the reaction is reversible and binding is weaker than for the AT-thrombin reaction. Stop-flow kinetic studies show that trypsin reacts much faster than thrombin with AT; the trypsin reaction in the absence of heparin is almost as fast as the reaction of thrombin with the heparin-AT complex. Trypsin reactions, on the other hand, are relatively insensitive to heparin.
Thrombotic Manifestations of Essential Thrombocythemia
Authors: Hussain Shafkat , Schwartz Joel , Friedman Saafor , Chua Segundina
Published: May 2019

The course of essential thrombocythemia (ET) has been observed in ten patients, aged 46-83. Eight have been followed for 1 to 8.5 years. In contrast to previous recorded experience with ET, arterial thrombosis was much more common than hemorrhage, with peripheral arterial or cerebrovascular disease dominating the clinical presentation. Six patients had incipient gangrene of toes, and three patients developed occlusion of tibial and larger arteries. Hemorrhage was seen only in two patients in surgical wounds. Absent or markedly decreased epinephrine-induced platelet aggregation was found in all patients studied at initial diagnosis. Epinephrine-induced aggregation improved with platelet count lowering and was impaired again with increase in platelet count. Collagen-induced aggregation was normal in all but one and ADP-induced aggregation was variable. Leukocyte alkaline phosphatase, serum vitamin B12 and unbound B12 binding capacity determinations were not of differential diagnostic significance. A prolonged remission was easily achieved with busulfan in all patients except one who died of a cerebrovascular thrombosis. The six patients with incipient gangrene showed marked improvement during therapy. Relapses in five patients after 2-87 months responded well to another course of busulfan therapy. No patient has shown evolution to another myeloproliferative disorder. Essential ET should be considered in the differential diagnosis of occlusive arterial disease even in the presence of atherosclerosis.
Factor VIII Response to Venous Congestion in Patients with von Willebrand’s Disease
Authors: Morfini M. , Longo G. , Rossi Ferrini P. , Seri A.
Published: May 2019

Venous congestion(v.c.) is known to increase plasma levels of antihemophilic factor activity (VIII AHF) and fibrinolytic activity in man. In 20 normal subjects and 15 patients with VonWillebrand’s disease(VWD), before and after v.c. on forearm the VIII AHF and related antigen(VIII AGN), the euglobulin lysis time, the haematocrit and platelet’s count were performed. Ethanol-precipitate (v/v 3% at -3°C) is prepared from each sample. Two-dimensional Immunoelectrophoresis and gel filtration through siliconized glass column(K15/30 Pharmacia) packed with Sepharose 6B, was performed by applyng ethanol-precipitate. In normal subjects a significant increase of VIII AHF has been observed after v.c. (F test p<0.001) and of VIII AGN (F test p<0.05). In VWD patients only VIII AHF increase was found (F test p<0.01) but in patients with very low VIII AHF level, the increase did not occur. The anodal migration of AGN on crossed electrophoresis and elution pattern of AHF and AGN did not show any significant variations after v.c. in normal subjects as in VWD patients. A likely mechanism of VIII AHF and VIII AGN increase may be a release from endothelial cells of vascular wall during venous congestion.
Endothelotropic Drugs - Stabilizers of the Endothelial Lining Against Desquamation
Authors: Hladovec J.
Published: May 2019

On the basis of a new test for counting circulating endothelial cells in blood, a group of drugs was redefined which prevent endothelial desquamation after a standard intravenous injection of citrate into rats. The group comprised practically all drugs designated as venotonics or vitamin P-active agents, including some haemostatics, analgesic-antipyretics (acetylsalicylic acid) and some antiatherosclerotics (pyridinolcarbamate, Clofibrate). A marked activity was observed also with several calcium-antagonists (nifedipine, prenylamine, chromonar, dipyridamole and verapamil). Many endothelotropic drugs are simultaneously effective in blocking platelet functions. The suggested common mechanism of action is the influence on the consistency of the cellular surface coat (endothelia cement).
Hemostasis Studies in Patients with Colon Cancer
Authors: Scialla S.J. , Kimball D.B.
Published: May 2019

Coagulation and platelet function studies were evaluated in eighteen patients with colon cancer (all clinical stages) prior to treatment with chemotherapy. All patients were asymptomatic from bleeding or thrombosis and were not taking aspirin. The tests included general coagulation screening, coagulation factor analyses, platelet aggregation, fibrin split product concentration and a serial dilution protamine sulfate test for the presence of fibrin monomer. Average values compared to normal controls showed an elevated fibrinogen 400.5 mg.% (normal 300.0 mg.%), shorter activated partial thromboplastin time 29.4 seconds (normal 36.0), increased factor VIII coagulant activity 127-2% (normal 100%), and elevated fibrin split products 28.8 ug/ml.(normal less than l6 ug/ml.). Seven patients showed the presence of fibrin monomer. Eight patients showed enhanced aggregation reacting at or below 0.16 micromolar epinephrine. Six patients in the study showed more advanced cancer (five requiring intrahepatic artery infusion for extensive liver metastasis). Of this subgroup four patients demonstrated fibrin monomer and three patients showed spontaneous platelet aggregation. In this group, the fibrinogen was 484.22 mg.%, fibrin split products were 28.8 ug/ml., and factor VIII was 176%. A state of hypercoagulation with signs of a chronic process of intravascular coagulation was demonstrated in a group of colon cancer patients which corresponded to their extent of disease.
Stimulation of Thrombasthenic Platelets by Collagen. A Scanning Microscopic Study
Authors: Balleisen L. , Schramm W. , Marx R.
Published: May 2019

The spreading of platelets on a silicone-coated surface has been evaluated as a sensitive test for the stimulation of thrombasthenic platelets by different collagen types. The platelets from six patients with Thrombasthenia were incubated with soluble type I, type III, methylated type I collagen and collagen type I fibrils and allowed to settle for one hour on a silicone surface. The platelets from two patients scarcely reacted, only a few pseudopodes were seen, no platelets had spread and no adhesion on collagen fibrils occurred, whereas the platelets from the four other patients showed many long pseudopodes, some of the platelets had spread and the adhesion on collagen fibrils seemed to be normal. Without collagen the platelets of all patients showed no pseudopodes and no spreading The three collagen types under investigation were qualitatively effective in the same manner, quantitatively was a decreasing activity from the methylated type I collagen to type III to type I collagen. The platelet reactivity correlated with the clinical severity of the disease. The results indicate that there are two groups of patients affected with thrombasthenia and that the reactivity of the thrombasthenic platelets with collagen may contribute to the variable clinical severity of the disease.
Haemostasis Disorders after an Extra-Corporeal Circulation in Cardiac Surgery, Based on a Study of 850 Patients
Authors: Doutremepuich Ch. , Chauve A. , Deckmyn M. , Boisseau M. , Fontan F.
Published: May 2019

The purpose of the study was to determine the influence of Extra-Corporeal Circulation (E.C.C.) on the principal factors of haemostasis, then to define biological troubles with a view their prevention. Tests performed 25 minutes after the conclusion of E.C.C., neutralization of heparin by protamin, Kunitz inhibitor injection, produced the following results: platelet count 75795 + 22045/mm3, prothrombin value 60.81±10.97%, fibrinogen 2.06 ± 0.85 g/1; the euglobulin lysis time was positive within 30 minutes for 1.16% of the patients;an heparinaemia lower than 0.2 Ul/ml was found in 69.41 % of the cases when the E.C.C. was performed in conditions of moderate hypothermia (28°c) and in 41.81% in deep hypothermia. A further injection of protamin was given in all cases where the heparinaemia was found to be above 0.2 UI/ml. A new control reading was made 3 hours later and showed an heparin rebound in 34.05% of the cases. These results allowed us to fomulate a standard monitoring program in case of post-operative bleeding.
The Classification of The Platelet Aggregation Pattern by Two Adp Solution (Double-Adp Method) and its Clinical Evaluation
Authors: Matsuo Takefumi , Ohoki Yasuo , Okinaga Youichi , Matsuo Osamu
Published: May 2019

A new method to describe platelet aggregability was devised. Two concentration of 0.4 µmol and 2 µmol ADP were added to two platelet-rich plasma obtained from the same patient at the same time, and the recording aggregation patterns by use of two channel aggregometer were classified into four types according to the simple criteria of visual impression. The distribution type in healthy adult belonged to either type I (76%) or II (20%), which represents normoaggregability whereas that in diabetic patients in comparison with aged matched control was a majority in type III and IV, which represents hyperaggregability. And the degree and severity of retinopathy in diabetics paralleled the increase of aggravated type. Although platelet aggregation seems to be related to the abnormal concentrations of alpha 2-macroglobulin, alpha 1-antitrypsin, fibrinogen, antithrombin III and blood lipids, the type classification was slightly affected by each of the increase of alpha 2-macroglobulin and fibrinogen, and the decrease of antithrombin III. The effect of aspirin and diet cure on hyperaggregability could be evaluated simply and clearly by the type classification of platelet aggregation based on our double-ADP method. This method is thus of considerable and suitability for clinical use.
Properties and Origin of C1-Inactivator Resistant Fibrinolytic Activity
Authors: Kluft C.
Published: May 2019

Intrinsic and extrinsic activators contribute to the total fibrinolytic activity in plasma. It was found that a percentage of the activity in euglobulin fractions on fibrin plates is not inhibited by C1-inactivator. Studies were carried out to determine the origin and properties of this CI-inactivator resistant fibrinolytic activity (C1-RFA). The Cl-RFA is specifically increased in situations of stimulated fibrinolysis, e.g., in plasma after 15 min venous occlusion. Extrinsic activators (urokinase, human tissue activator and activator from perfused cadaver limbs) are resistant to C1-inactivator. This suggests that C1-RFA could be attributed to extrinsic activators.

Study on the properties of C1-RFA in plasma after venous occlusion showed that the activity could be preserved in vitro by freezing the plasma. After coagulation in this plasma, C1-RFA was low suggesting strong adsorption to fibrin. The C1-RFA diminished gradually (50% in 90 min) at 37°C in vitro in plasma and blood, but was stable in euglobulin fractions. This indicates that its “lability” is due to inhibitors not found in euglobulin fractions but which are present in blood (also postmortem) and plasma after venous occlusion. In vivo disappearance of elevated activities is much more rapid than in vitro, which suggests a different clearing mechanism in vivo under conditions of stimulated fibrinolysis.