bluuebunny/crossref_metadata_2025_split huggingface dataset:
Selective Staining of Platelet Glycoproteins Using Two-Dimensional O’Farrell Gel Electrophoresis and Avidin-Biotin-Conjugated Lectins
Authors: Kehrel Beate , Kokott Renate , Balleisen Leopold , Stenzinger Werner , Clemetson Kenneth J , van de Loo Jürgen 
Published: June 2018

SummaryA time-saving and sensitive high resolution method lor the analysis and identification of platelet glycoproteins using non-radioactive compounds has been developed. Platelet proteins (50 μg) of normal subjects were separated by isoelectric focusing and SDS-PAGE. Proteins were either stained with silver or electroblotted onto nitrocellulose. Nitrocellulose blots were treated with the following biotinylated lectins : Abrus precatorius lectin, concanavalin A, Lens culinaris lectin, or wheat germ agglutinin. Staining was achieved by avidin-biotin-coupled peroxidase using 4-chloro-l-naphthol as the substrate. A rapid overview of platelet glycoproteins may be obtained by applying all the lectins to a single blot.
Hypothyroidism Leads to More Small-Sized Platelets in Circulation
Authors: van Doormaal J J , van der Meer J , Oosten H R , Halie M R , Doorenbos H 
Published: June 2018

SummaryThe effect of induced hypothyroidism on platelet count and platelet volume distribution was studied in twelve athyreotic patients, After a two weeks withdrawal of triiodothyronine supplementation, platelet count and the ratio between platelet and red cell count were increased in all patients. Furthermore, mean platelet volume was declined and platelet distribution width was risen. Thus, hypothyroidism appears to increase the number of circulating platelets, especially the smaller ones.
Two Genes Homologous with Human Protein S cDNA Are Located on Chromosome 3
Authors: Amstel J K Ploos van , van der Zander A L , Bakker E , Reitsma P H , Bertina R M 
Published: June 2018

SummaryA cDNA coding for the carboxy-terminal region of human protein S and containing a complete 3’-untranslated region, was isolated by a combination of antibody screening of a λgt11 human liver cDNA expression library and in situ hybridization of a pUC9 human liver cDNA library.Hybridization analysis of human total DNA with radiolabelled non-overlapping cDNA restriction fragments revealed the existence of two genes per haploid genome homologous with the protein S cDNA. Both genes were mapped to chromosome 3 using human-rodent cell hybrids. Neither of the genes showed polymorphism for sixteen different enzymes upon hybridization with the protein S cDNA.
Congenital Deficiency of Alpha-2-Adrenoceptors on Human Platelets : Description of Two Cases
Authors: Tamponi Giacomo , Pannocchia Antonella , Arduino Carlo , Bazzan Mario , Dora Nadine Della , Schinco Piercarla , Buraglio Mauro , Eandi Mario 
Published: June 2018

SummaryThe biochemistry and functionality of platelets from two related subjects (mother and son) with alpha-2-adrenoceptor-deficient platelets has been evaluated. Radioligand binding experimentes with the specific alpha-2-adrenergic-receptor antagonist, 3H-yohimbine, showed a drastic reduction of alpha-2-adrenoceptors in platelets from both subjects in comparison with the control values. Electron microscopy studies revealed a normal morphology and a normal number of alpha granules and dense bodies. Levels of adenine nucleotides; 5-hydroxytryptamine; B-thromboglobulin; platelet-factor-4 and thromboxane A2 production were within normal limits.Platelet aggregation and 5-hydroxytryplamine production in response to adrenalin (at concentrations up to 50 μM) were absent, whereas ADP, AA, PAF, collagen and thrombin-induced aggregation, secretion, Ca++ flux and thromboxane A2 production were normal.The inhibitory effect caused by different concentrations of prostacyclin on Ca++ flux, aggregation, secretion and thromboxane A2 production of platelet functionally lacking of alpha-2-adrenoceptor was not distinguishable from control platelets and platelets preincubated with yohimbine.
Immunoaffinity Purification of HTC Rat Hepatoma Cell Plasminogen Activator-Inhibitor-1
Authors: Zeheb Ron , Rafferty Usha M , Rodriguez Miguel A , Andreasen Peter , Gelehrter Thomas D 
Published: June 2018

SummaryIncubation of HTC rat hepatoma cells with the synthetic glucocorticoid dexamethasone rapidly inhibits tissue-type plasminogen activator activity by inducing a specific plasminogen activator-inhibitor (PAI-1). Using immobilized polyclonal antibodies raised against HT-1080 human fibrosarcoma PAI-1, we have purified HTC PAI-1 from serum-free medium conditioned by dexamethasone-treated HTC hepatoma cells and shown it to be antigenically related to human PAI-1. Greater than 100-fold purification with greater than 75% yield was achieved in a single step. The purified PAI-1 migrates on SDS-polyacrylamide gels as a single major band of 49 kDa with a minor band of 46 kDa. Digestion of PAI-1 with endoglycosidase F causes a shift toward faster migrating species which retain inhibitory activity. The purified PAI-1 was stable at pH 2.5, lost 50% of its activity after 15 min at 45° C, and showed marked activation after treatment with SDS or guanidine-HCl. Purified PAI-1 rapidly inhibited and formed complexes with both tissue-type and urokinase-type plasminogen activators. Polyclonal rabbit antirat PAI-1 antibodies were raised which immunoprecipitate both free and complexed PAI-1.
Fibrinolytic Response and Fibrin Fragment D-Dimer Levels in Patients with Deep Vein Thrombosis
Authors: Declerck P J , Mombaerts P , Holvoet P , De Mol M , Collen D 
Published: June 2018

SummaryAn enzyme-linked immunosorbent assay for fragment D-dimer was developed with the use of two monoclonal antibodies directed against specific non-overlapping antigenic determinants, present in fragment D-dimer of crosslinked fibrin but not in fragment D of non crosslinked fibrin or of fibrinogen, The lower limit of sensitivity of the assay when applied to human plasma, is 25 ng/ml. Concentration of fragment D-dimer in plasma from healthy individuals was 177 ,± 83 ng/ml (mean ± SD). In plasma of 11 out of 12 patients with phlebographically confirmed acute deep vein thrombosis, fragment D-dimer levels were significantly increased. Fragment D-dimer was not increased in 9 out of 10 patients with recurrent idiopathic deep vein thrombosis during clinically silent episodes.Total t-PA antigen and free t-PA antigen concentrations were measured using previously developed ELISAs. Nine of the 12 patients with acute deep vein thrombosis showed a significant increase of total t-PA antigen (from 8.6 ± 6.9 ng/ml to 21 ± 16 ng/ml) after venous occlusion but in 3 of these free t-PA remained undetectable. Five of the 10 patients with recurrent deep vein thrombosis responded to venous occlusion with a significant increase of total t-PA antigen (from 6.7±3.2 ng/ml to 14 ± 7.9 ng/ml) but, in all patients, free t-PA antigen remained undetectable.It is concluded that the combined assays of total and free t-PA antigen and of fragment D-dimer may be useful for the evaluation of the dynamics of the fibrinolytic system in physiological and pathological conditions.
Antithrombin III Binding to Surface Immobilized Heparin and Its Relation to F Xa Inhibition
Authors: Kodama K , Pasche B , Olsson P , Swedenborg J , Adolfsson L , Larm O , Riesenfeld J 
Published: June 2018

SummaryThe mode of F Xa inhibition was investigated on a thromboresistant surface with end-point attached partially depoly-merized heparin of an approximate molecular weight of 8000. Affinity chromatography revealed that one fourth of the heparin used in surface coating had high affinity for antithrombin III (AT). The heparin surface adsorbed AT from both human plasma and solutions of purified AT. By increasing the ionic strength in the AT solution the existence of high and low affinity sites could be shown. The uptake of AT was measured and the density of available high and low affinity sites was found to be in the range of 5 HTid 11 pic.omoles/cmf, respectively Thus the estimated density of biologically active high and low ailmity heparm respectively would be 40 and 90 ng/cm2 The heparin coating did not take up or exert F Xa inhibition by itself. With AT adsorbed on both high and low affinity heparin the surface had the capacity to inhibit several consecutive aliquots of F Xa exposed to the surface. When mainly high affinity sites were saturated with AT the inhibition capacity was considerably lower. Tt was demonstrated that the density of AT on both high and low affinity heparin determines the F Xa inhibition capacity whereas the amount of AT on high affinity sites limits the rate of the reaction. This implies that during the inhibition of F Xa there is a continuous surface-diffusion of AT from sites of a lower class to the high affinity sites where the F Xa/AT complex is formed and leaves the surface. The ability of the immobilized heparin to catalyze inhibition of F Xa is likely to be an important component for the thromboresistant properties of a heparin coating with non-compromized AT binding sequences.
Sources of Variability in Coagulation Factor Assays
Authors: Thompson S G , Martin J C , Meade T W 
Published: June 2018

SummaryEstimates of the components of variation of a number of coagulation factor assays are presented and are compared with those for other haematological and biochemical variables. They have been derived from a study in which fourteen volunteers gave blood samples approximately bi-monthly for three years. The systematic between-batch variation w7as generally only a small component of the total variation. However, the proportion of the total variation due to differences between people ranged from about 15% in the case of two measures of antithrombin III to about 85% for cholesterol and haemoglobin. For factor VII activity and fibrinogen, the figure was just over 70%. These estimates of the components of variation are not generally available in the literature. In view of the increasing interest in the role of coagulation factors in the pathogenesis of arterial disease, estimates of this kind are needed both for the interpretation of observed relationships as well as for planning sample sizes for future studies.
Risks to the Fetus of Anticoagulant Therapy During Pregnancy
Authors: Ginsberg Jeffrey S , Hirsh Jack , Turner D Christoper , Levine Mark N , Burrows Robert 
Published: June 2018

SummaryThe use of anticoagulants during pregnancy is problematic because of the potential adverse effects to the mother and the fetus. Heparin does not cross the placenta, and thus, it was surprising that a recent report concluded that heparin therapy during pregnancy was as risky as oral anticoagulant therapy. Therefore, we performed a literature review of fetal/infant outcomes following anticoagulant therapy during pregnancy. We examined 186 reports which described fetal/infant outcomes in 1,325 pregnancies associated with anticoagulant therapy. The rates of adverse fetal/infant outcomes including death, prematurity and congenital malformations following treatment with heparin, oral anticoagulants, or both were calculated. The previously described high rate of adverse feta/infant outcomes with heparin- treated patients, could be accounted for by the frequent use of heparin in pregnancies with comorbid conditions independently associated with adverse outcomes and by reports of uncomplicated prematurity. After excluding such pregnancies, outcomes in heparin-treated patients are similar to the normal population.
The Relative Antithrombotic Effectiveness of Heparin, a Low Molecular Weight Heparin, and a Pentasaccharide Fragment in an Animal Model
Authors: Thomas D P , Merton R E , Gray E , Barrowcliffe T W 
Published: June 2018

SummaryThe antithrombotic efficacy of unfractionated heparin (UFH), a low molecular weight heparin (LMWH) and a synthetic pentasaccharide (PENTA) has been compared in an animal model for stasis thrombosis. We have also compared the relative ability of these three agents to impair thrombin generation in vitro and in vivo, and measured their effects on anti-Xa activity and thrombin clotting times. UFH, LMWH and PENTA all had the capacity to impair thrombogenesis, although there were marked differences in their relative effectiveness. Reduction of thrombin generation to 20% of control values was closely correlated with the prevention of thrombosis after 20 minutes’ stasis, but this was only achieved with UFH. The same dry weight dose of LMWH or PENTA reduced thrombin generation to about half control values, and neither significantly impaired thrombus formation after 20 minutes’ stasis. Impaired thrombin generation correlated better than anti-Xa activity with prevention of stasis thrombosis. In this model, UFH was clearly superior to LMWH and PENTA as an antithrombotic agent.
A Rapid and Simple Method for the Separation of Four Molecular Forms of Human Plasminogen
Authors: Nieuwenhuizen W , Traas D W 
Published: June 2018

SummaryAt least four molecular forms of plasminogen are known. Two of those forms have glutamic acid at their amino-terminal end, and are designated as glu-plasminogen. The other two have lysine, methionine and/or valine as amino-terminal amino acid and are collectively designated as lys-plasminogen. Two subforms (I and II) each of glu- and lys-plasminogen exist. The I-forms are glycosylated at asn-288 and thr-345, whereas the II-forms are only glycosylated at thr-345. In a previous publication (Thromb Haemostas 1984; 52: 347-349) we have described the separation of the I- and II-forms of plasminogen in lysine-Sepharose in phosphate buffers. Now we have combined those findings with the differential affinity of glu- and lys-plasminogen for aminohexyl-Sepharose through their aminohexyl-sites, recently described by Christensen (Biochem J 1984; 223: 431-421). Acid/urea electrophoresis, end-group determination and carbohydrate analysis show that the combination of affinity chromatography on lysine-Sepharose in phosphate buffers, and on aminohexyl-Sepharose provides an efficient procedure to separate the four molecular forms of plasminogen.
Affinity Purification of Plasma Proteins: Characterization of Six Affinity Matrices and their Application for the Isolation of Human Factor VIII
Authors: te Booy M P W M , Riethors W , Faber A , Over J , König B W 
Published: June 2018

SummaryFor the purification of coagulation factor VIII, (1,1’-carbonyldiimida zole [CDI] -activated) Sepharose CL-48 was functionalized with two aminoalkyl and four aminoalkyl-carbamylalkyl ligand- spacer combinations. The affinity matrices were contacted with human plasma. All affinity matrices showed complete adsorption of factor VIII (>90%) and three aminoalkyl-carbamylalkyl Sepharoses gave factor-VIII recoveries of 50-65% and a factor-VIII preparation with a specific activity of 1–2 U factor VIII/mg of protein. Furthermore, no fibrinogen, immunoglobulin G and albumin could be detected in the isolated factor VIII. Optimal results were obtained using the di-methyl-aminopropyl-carbamylpentyl-Sepharose affinity matrix.
In Vitro Carboxylation of a Blood Coagulation Factor IX Precursor Produced by Recombinant-DNA Technology
Authors: Soute Berry A M , Balland Alain , Faure Thérèse , de la Salle Henri , Vermeer Cees 
Published: June 2018

SummaryBlood coagulation factor IX (Christmas factor) is a plasma protein which is required for normal haemostasis. A functional deficiency of factor IX results in haemophilia B, a bleeding disorder which is generally treated by infusions of factor IX concentrates prepared from pooled human plasma. The use of human blood products is connected with the risk of transmitting viral agents responsible for diseases such as hepatitis B and AIDS. Recombinant DNA techniques may provide the means to produce the required proteins without exposing the patients to these risks and at lower costs. One of the problems which has to be overcome before recombinant factor IX can be used for therapeutical purposes is related to the vitamin K-dependent carboxylation of its 12 NH2-terminal glutamate residues. In cell cultures this carboxylation, which is required to render the protein its procoagulant activity, is far from complete, especially at high expression levels. In this paper we describe the in vitro carboxylation of non and/or partly carboxylated recombinant factor IX produced by transformed Chinese hamster ovary cells. The identity of the newly formed Gla residues was verified and it could be demonstrated that all carboxyl groups had been incorporated into the recombinant factor IX.
A Prospective Study of Haemostatic Tests at 28 Weeks Gestation as Predictors of Pre-Eclampsia and Growth Retardation
Authors: Thornton J G , Molloy B J , Vinall P S , Philips P R , Hughes R , Davies J A 
Published: June 2018

SummaryA panel of haemostatic tests was perfomed on 400 primiparous women at 28 weeks to test whether one or more could predict the development of pregnancy complications. Fifteen women subsequently developed pre-eclampsia with significant proteinuria and 13 delivered growth retarded infants. There were no significant differences between mothers in the pre-eclampsia group and 22 randomly selected controls. A stepwise logistic discriminant analysis of the data did not produce a significant model. In the growth retarded group only beta thromboglobulin levels were significantly lower than in the controls (p <0.05), although in the logistic discriminant analysis the inclusion of both beta thromboglobulin and fibrin degradation products led to a borderline significant improvement in fit of the model. We conclude that the haemostatic variables studied are not significantly changed at 28 weeks nor clinically useful predictors of either pre-eclampsia or fetal growth retardation.
Immunogold Electron-Microscopic Localization of Calpain I in Human Erythrocytes
Authors: Samis John A , Elce John S 
Published: June 2018

SummaryWith a view to understanding the function of calpain I (EC 3.4.22.17) in vivo, the localization of the enzyme was studied by immunoelectron microscopy in human erythrocytes. Thin sections of the cells embedded in Spurr’s resin were exposed to solutions of monoclonal anti-calpain I or control antibodies, biotin antimouse IgG antibodies, and streptavidin-gold. Most of the calpain I (93%) was found to be distributed throughout the cytoplasm, and only 7% of the gold particles were associated with the erythrocyte membrane. Erythrocytes were Ca2+-loaded by means of the calcium ionophore A23187, and the rise in intracellular [Ca2+] was demonstrated both by crenation of the cells, and by activation of calpain which was detected by immunoblotting. The proportions of cytosolic and membrane-bound gold labelling were, however not altered by Ca2+-loading. These results are not consistent with the hypothesis that activation of calpain requires membrane-binding.
Monoclonal Antibodies to Tissue-Type Plasminogen Activator which Prolong its Clearance In Vivo
Authors: Reilly Thomas M , Knabb Robert M , Chiu Andrew T , Bradfute David L , Timmermans Pieter B M W M 
Published: June 2018

SummaryThree murine monoclonal antibodies to tissue-type plasminogen activator (t-PA) were evaluated for their effects on the binding of iodinated t-PA to cultured human hepatoma cells (Hep G2), and on extending the half-life of t-PA injected into rabbits. Two of the antibodies, AE5 and EG2, significantly inhibited t-PA binding in vitro, and extended the in vivo half-life of t-PA four to five-fold. A third antibody, BA10, which had a much smaller inhibitory effect on t-PA binding, had no influence in extending t-PA’s half-life. MOPC-21, a control antibody not directed to t-PA, had no effect on either test. Our results are the first to correlate different compounds’ effects on t-PA binding with their ability to retard t-PA clearance in vivo, and provide additional evidence for the importance of a liver cell receptor in the t-PA clearance process.
Tissue-Type Plasminogen Activator (t-PA) and Dilute Blood Clot Lysis Time in Nephrotic Patients
Authors: Cucuianu Mircea P , Rus Horea G , Roman Stefan , Mărcuşu Codruţa , Spînu Constantin , Manasia Mihai , Niculescu Florin 
Published: June 2018

SummaryWhen compared to normal weight normolipidemic control subjects, dilute blood clot lysis time was found to be obviously (p <0.001) prolonged in hypertriglyceridemic patients without proteinuria and slightly (p <0.05) accelerated in hyperlipidemic nephrotic patients in spite of their very high levels of plasma fibrinogen. As a result the ratio plasma fibrinogen (mg/dl) per clot lysis time (minutes) was 1.241 ± 0.08 (X ± SEM) in control subjects, 0.574 ± 0.07 in hypertriglyceridemic patients and 2.69 ± 0.172 in nephrotic patients. This finding suggesting that a larger amount of fibrin is rather readily dispersed from dilute blood clots of nephrotic patients was associated with higher levels of plasma t-PA: Ag (9.45 ng/ml ± 1.18 in nephrotic patients versus 5.8 ng/ml ± 1.23 in controls before venous occlusion and respectively 33.1 ng/ml ± 3.83 versus 20.3 ± 3.40 in controls after venous occlusion). Plasminogen activator activity of the euglobulins as assessed by the bovine fibrin-agarose plate was significantly higher in nephrotic patients only after venous occlusion. Plasma samples of nephrotic patients exerted a more potent inhibition of fibrinolysis in a urokinase activated system. This effect was, however, mainly due to the high levels of α2 macroglobulin in nephrotic plasma which apparently have little influence on dilute blood clot lysis time.
Fibrinolytic Profiles in Local Low-Dose Thrombolysis with Streptokinase and Recombinant Tissue Plasminogen Activator
Authors: Berridge D C , Earnshawl J J , Westby J C , Makin G S , Hopkinson B R 
Published: June 2018

SummaryFibrinolytic parameters have been monitored in 44 patients undergoing local low-dose intra-arterial thrombolysis for acute peripheral arterial ischaemia. Streptokinase (Sk), at a dose of 5,000 units/hr with 250 units/hr heparin, was used in 23 patients and recombinant tissue plasminogen activator (r-tPA) at a dose of 0.5 mg/hr was used in 21 patients. Successful lysis was seen in 18 (86%) patients following r-tPA and in 15 (65%) patients following streptokinase. There were 4 minor haematomas in each group usually at the catheter entry site. Both agents produced a systemic effect, which was still seen 12 hours post-infusion. However, that produced by r-tPA was delayed and significantly reduced compared to that produced by Sk.These results confirm the relative fibrin specificity of r-tPA. When used as a continuous low-dose intra-arterial infusion, r-tPA offers a significantly lower, potentially safer, systemic effect than conventional therapy with streptokinase.