bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification of Novel Peptide Antagonists for von Willebrand Factor Binding to the Platelet Glycoprotein Ib Receptor from a Phage Epitope Library
Authors: Ricca George A , South Victoria , Searfoss George H , French Stephen , Cheadle Christopher , Murray Edward , Howk Richard , Jaye Michael 
Published: July 2018

SummaryWe have constructed a fusion phage epitope library in the filamentous bacteriophage fuse5. The library was made by inserting a degenerate oligonucleotide which encodes 15 variable amino acids into the NH2-terminal region of the phage gene III protein. This library, containing over 107different epitope bearing phage, has been used in an attempt to identify inhibitors of the von Willebrand factor (vWF)-platelet Glycoprotein lb interaction. The library was screened with a monoclonal antibody (RG46) that recognizes the GPIb binding domain of vWF (amino acids 445-733). A total of 30 clones falling into 8 classes have been identified that react with the RG46 antibody. Isolates from all 8 classes are positive by immunoblot analysis. The amino acid sequence of the gene III fusion protein from positive clones showed a strong homology to the known RG46 epitope. Peptides identified from the screen were synthesized and used to demonstrate that some of the synthetic peptides exhibited inhibitory activity towards ristocetin induced binding of vWF to the GPIb receptor. Thus, we have demonstrated that screening a fusion phage epitope library with a monoclonal antibody that inhibits vWF binding to the GPIb receptor can be a useful tool not only for mapping antibody recognizing determinants, but also can serve as a source for identifying novel peptides that are antagonists for vWF binding to the platelet GPIb receptor.
Definition of the Bleeding Tendency in Factor XI-Deficient Kindreds–A Clinical and Laboratory Study
Authors: Bolton-Maggs P H B , Patterson D A , Wensley R T , Tuddenham E G D 
Published: July 2018

SummaryIndividuals with severe factor XI deficiency are prone to excessive bleeding after injury or surgery, but the existence of a haemorrhagic tendency in partial factor XI deficiency is controversial. In this study, 172 members of 30 kindreds (20 non-Jewish) transmitting factor XI deficiency in North West England were interviewed and a bleeding history questionnaire completed. Blood was taken for coagulation assays. The questionnaires were categorised independently by two assessors to determine presence or absence of a bleeding tendency, in the absence of information about the factor XI level or family history. Analysis shows that 48% of heterozygotes have a bleeding tendency. Eighteen (60%) families came to attention because of bleeding problems in heterozygotes. Comparison of histories between partially deficient and non-deficient individuals demonstrated a higher incidence of menstrual problems, an increase in significant bruising, and an increased likelihood of excessive bleeding after tonsillectomy and dental extractions.The incidence of von Willebrand’s disease was not increased, but individuals with heterozygous factor XI deficiency who were bleeders tended to have lower levels of factor VIIIc and von Willebrand factor, and were more commonly of blood group 0. These features may contribute to the bleeding tendency. There was no evidence of alteration in factor VII activity (as defined by the ratio of activity to antigen) between the bleeders and non-bleeders.This is convincing evidence for abnormal bleeding in factor XI deficiency which is not confined to severely deficient patients.
Platelet Consumption in Thrombocythemia Complicated by Erythromelalgia: Reversal by Aspirin
Authors: Genderen Perry J J Van , Michiels Jan J , Strik Roel Van , Lindemans Jan , Van Vliet Huub H D M 
Published: July 2018

SummaryThe involvement of platelets in the pathogenesis of erythromelalgia, a frequent and characteristic microvascular thrombotic manifestation in patients with primary thrombocythemia and polycythemia rubra vera, was investigated by measuring the survival and turnover of 51Cr labeled autologous platelets in 10 patients with thrombocythemia complicated by erythromelalgia, in 10 asymptomatic thrombocythemia patients and in 6 subjects with reactive thrombocytosis. The mean platelet survival time of the erythromelalgia patients was 4.2 ± 0.2 days, which is significantly decreased as compared with asymptomatic thrombocythemia patients (6.6 ± 0.3 days, p <0.001) and patients with reactive thrombocytosis (8.0 ± 0.4 days, p <0.001). The mean platelet survival time of asymptomatic thrombocythemia patients was significantly decreased (p <0.01) as compared with reactive thrombocytosis patients. Treatment of erythromelalgia with aspirin increased the mean platelet survival time from 4.0 ± 0.3 days to 6.9 ± 0.4 days (p <0.001) and was associated with an elevation of the platelet count of 216 ± 30 × 109 platelets per liter (p <0.001). Coumadin failed to improve platelet survival or symptoms caused by erythromelalgia. The increased platelet consumption in erythromelalgia is attributed to the formation of platelet thrombi in the arterial microvasculature. This conclusion is supported by the ability of aspirin to interrupt platelet consumption and clinical features of erythromelalgia.
Complete Inhibition of Endotoxin-Induced Coagulation Activation in Chimpanzees with a Monoclonal Fab Fragment against Factor VII/VIIa
Authors: Biemond Bart J , Levi Marcel , ten Cate Hugo , Soule Howard R , Morris Lori D , Foster David L , Bogowitz Cheryl A , van der Poll Tam , Büller Harry R , ten cate Jan Wouter 
Published: July 2018

SummaryGram-negative sepsis is oftentimes complicated by activation of coagulation with disseminated intravascular coagulation and microthrombosis. This may contribute to the associated morbidity, multiple organ failure and death. Recent studies have established that the tissue factor-dependent pathway of blood coagulation has a significant participatory role in the initial endotoxin-induced activation of coagulation. Tissue factor (TF), expressed on the surface of activated monocytes and endothelial cells forms cell surface complexes with free circulating factors VII and VIIa. The latter complex proteolytically activates factors X and IX. Recent in vivo experiments have shown that a rapidly neutralizing TF monoclonal antibody prevents and arrests the endotoxin- induced activation of coagulation and similar studies have shown to reduce mortality in baboons. In this study we describe the preparation of a factor VII/VIIa neutralizing monoclonal Fab fragment and characterize its effect on in vivo activation of coagulation during experimental endotoxemia in chimpanzees.Four chimpanzees received a bolus intravenous injection of 4 ng/kg endotoxin in combination with Fab fragments of a factor VII/VIIa neutralizing murine monoclonal antibody (12D10) at a dose of either 50 μg/kg (n = 2) or 100 μfig/kg (n = 2). Four control animals received a bolus injection of endotoxin alone. Administration of the 12D10 Fab fragments, immediately preceding the endotoxin bolus injection, effectively blocked the endotoxin-induced activation of coagulation. Plasma levels of products of in vivo activation, namely F1+2, TAT complexes and FpA remained at baseline values. The administration of 12D10 resulted in a rapid decline in factor VII/VIIa antigen levels which remained below 5 ng/ml for 180-240 min, followed by a rapid return to baseline levels. Endotoxin administration resulted in activation of the fibrinolytic system as reflected by a rapid increase in plasma plasmin- α2-antiplasmin complexes. Administration of 12D10 was without effect on the endotoxin-induced fibrinolytic activation.In conclusion, this study confirmed the importance of the TF: VII complex in the initial, endotoxin-induced activation of coagulation which was completely blocked by neutralizing all free and tissue factor-complexed factor VII/VIIa by a specific monoclonal Fab fragment. Activation of fibrinolysis was not influenced. Therefore, neutralization of factor VII/VIIa might be a promising therapeutic option in preventing endotoxin-induced microthrombosis during Gram-negative sepsis.
Internalization and Degradation of Recombinant Human Coagulation Factor VIIa by the Human Hepatoma Cell Line HuH7
Authors: Chang Glenn T G , Kisiel Walter 
Published: July 2018

SummaryPrevious studies demonstrated that several normal and transformed cultured human cell lines specifically bind human coagulation factors VII and VIIa via tissue factor. In the present study, we show that 125I- radiolabeled recombinant human factor VIIa (125I-rFVIIa) binds to a human hepatoma cell line (HuH7). In the presence of rabbit polyclonal anti-human tissue factor apoprotein IgG, binding of 125I-rFVIIa to the HuH7 cells was decreased ∼60%, suggesting the presence of tissue factor-independent binding sites for 125I-rFVIIa on these cells. The binding isotherm of 125I-rFVIIa for the HuH7 cells in the presence of anti-tissue factor IgG exhibited a hyperbolic profile and was time-, temperature- and calcium-dependent. Furthermore, binding at 4° C was specific, dose-dependent and saturable. Scatchard analysis of the binding data demonstrated a single class of binding sites with a dissociation constant (Kd) of 3.2 nM and 27,000 binding sites per cell. At 4° C, 125I-rFVIIa bound to, and eluted from, the cell was indistinguishable from offered 125I-rFVIIa as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by autoradiography.The molecular properties of the tissue factor-independent binding protein were studied by using the cleavable cross-linking agent 3, 3’- dithiobis(sulfosuccinimidylpropionate). A cross-linking product of 125I-rFVIIa and a cell surface protein with an apparent Mr ∼100,000 was observed. The cross-linking reaction was strongly inhibited by a 100-fold molar excess of unlabeled rFVIIa, but not by rabbit polyclonal anti-human tissue factor apoprotein IgG, indicating that cross-linking does not involve the extracellular domain of tissue factor.After binding, internalization of 125I-rFVIIa by the HuH7 cells was observed at 37° C in the presence of rabbit polyclonal anti-human tissue factor apoprotein IgG. Internalization of 125I-rFVIIa proceeds at a rate of 0.4 fmol 125I-rFVIIa/min/106 cells, reaching a steady state after 2 h. Following binding and internalization, 125I-rFVIIa reappeared in the culture medium within 30 min, at approximately the same rate as internalized 125I-rFVIIa. This 125I-rFVIIa probably represents degradation of 125I-rFVIIa into small peptides, since it could not be precipitated by rabbit polyclonal affinity-purified anti-FVII IgG or by trichloroacetic acid. Chloroquine treatment of the HuH7 cells inhibited the degradation of 125I-rFVIIa, suggesting that degradation presumably occurs via a lysosomal-dependent pathway. These studies demonstrate that the liver may play an important role in the clearance mechanism(s) of FVIIa.
Antinociceptive Properties of Protein C in a Model of Inflammatory Hyperalgesia in Rats
Authors: Pichler L , Schramm W , Ulrich W , Varadi K , Schwarz H P 
Published: July 2018

SummaryWe investigated the role of human protein C in an animal model of inflammatory hyperalgesia. Pain was induced by intraplantar injection of carrageenan (3 mg) into the hind paw of rats. The pain threshold was measured by exerting increasing amounts of pressure (in mmHg) on the paw until a struggle reaction was observed. Protein C (8-800 IU/kg) was administered intravenously immediately after carrageenan. Controls received either intraplantar injections of saline (100 µ1) instead of carrageenan or carrageenan alone. Effects on pain threshold were expressed in percent of the pretreatment value. Carrageenan alone lowered the mean pain threshold after 3 h to 33.2 ± 2.2% of the pretreatment level. Addition of protein C resulted in a dose-dependent rise in pain threshold towards the level observed in control animals treated with saline instead of carrageenan (pain threshold after 800 IU/kg protein C = 62.9 ± 2.3% of pretreatment level), demonstrating an antinociceptive effect. Protein C had no effect in animals not preconditioned with intraplantar carrageenan. Thus protein C clearly antagonized the inflammatory pain induced by carrageenan. The antinociceptive action of protein C was antagonized by injection of a monoclonal antibody against protein C, providing additional evidence that the effect was protein C-mediated.
ARIC Hemostasis Study-IV. Intraindividual Variability and Reliability of Hemostatic Factors
Authors: Nguyen Nghia D , Ghaddar Habib , Stinson Valarie , Chambless Lloyd E , Wu Kenneth K ,   for the Atherosclerosis Risk in Communities (ARIC) Study Investigators
Published: July 2018

SummaryWe have recently reported the short-term intraindividual variability of several coagulation factors and inhibitors included in the ARIC study (Chambless et al. Ann Epidemiol 1992; 2:723). In this paper, we reported the intraindividual variability results of additional hemostatic factors. Blood samples were collected for hemostatic assays three times at 1-2-week intervals from 39 subjects recruited from 4 ARIC field centers. The contributions of within-person, processing and assay (designated “method”) and between-person variances to the total variance were estimated and from them the reliability coefficient, R, was computed as the proportion of total variance in the between-person component. The R value was high for (β-thromboglobulin and tissue- plasminogen activator: 0.83 and 0.81, respectively; and intermediate for D-dimer and plasminogen activator inhibitor-1: 0.73 and 0.72, respectively. Protein S (total and free) and platelet factor 4 had low repeatability (R<0.50) derived mostly from “method” variability while low R value (0.03) for fibrinopeptide A was attributed to high “method” and “within-person” variability. Gender, age and the level of hemostatic factors did not influence the intraindividual variability.
Up-Regulated Expression of Plasminogen Activator Inhibitor-1 in Hep G2 Cells: Interrelationship between Insulin and Insulin-Like Growth Factor 1
Authors: Anfosso F , Alessi M C , Nalbone G , Chomiki N , Henry M , Juhan-Vague I 
Published: July 2018

SummaryInsulin resistance represents a situation with a high risk of athero-thrombosis and is accompanied by increased plasma plasminogen activator inhibitor-1 (PAI-1) levels. Fasting insulin level is highly correlated with PAI-1 levels in plasma. It has been shown that insulin increases PAI-1 synthesis by the human hepatoma cell line Hep G2. Moreover when Hep G2 cells expressing a down-regulation of insulin receptors by incubation with 10-7 M insulin, were stimulated by 10-9 M insulin, an overexpression of PAI-1 synthesis was observed despite a reduced number of insulin receptors. Insulin-like growth factor 1 (IGF-1) shares many properties with insulin. The aim of the present study was to evaluate the effect of IGF-1 on PAI-1 synthesis by Hep G2 cells down-regulated either by insulin or IGF-1.Incubation of Hep G2 cells with increasing doses from 10-9 to 10-7 M IGF-1 induced a dose-dependent stimulation of PAI-1 synthesis up to 4.5-fold the control level. When cells were first pre-incubated with 10-7M IGF-1 for 18 h, acid washed, and then stimulated with 10-9 M IGF-1, the expression of IGF-1 receptors was greatly reduced (up to 70%). In contrast PAI-1 secretion was increased 3.4-fold the level of control cells and by 1.9-fold the level of cells first stimulated with 10-9M IGF-1. Both transcripts of PAI-1 mRNA were also increased. The overexpression of PAI-1 synthesis was observed irrespective of the hormone used in the down-regulation step (i.e. 10-9 M insulin or IGF-1) or in the stimulation step (i. e. 10-9 M insulin or IGF-1). The results showed an interrelationship between insulin and IGF-1 on PAI-1 synthesis in down-regulated Hep G2 cells. They also suggest that in the insulin resistant state, IGF-1 would be able to participate in the increase in PAI-1 plasma levels by stimulating down-regulated insulin target cells.
Acute Dynamic Exercise Increases Fibrinolytic Activity
Authors: Rankinen Tuomo , Väisänen Sari , Penttilä llkka , Rauramaa Rainer 
Published: July 2018

SummaryThe influence of acute bouts of dynamic exercise on plasma tissue plasminogen activator (tPA), plasminogen activator inhibitor (PAI) and fibrinogen was investigated on nine healthy non-smoking men aged 23 to 37 years. Subjects performed maximal and two submaximal (duration 30 min, intensity 50% [aerobic threshold] and 78% [anaerobic threshold] V02max) bicycle ergospirometry tests separated by seven days. The order of the submaximal tests was randomized. Blood samples were drawn before, immediately after and 24 h after each test. Plasma tPA and PAI activities were measured amidolytically, and fibrinogen concentration by thrombin method. All postexercise values were corrected for the change in plasma volume. The pre-exercise tPA and PAI activities and fibrinogen concentration were similar in each tests. tPA activity increased during each test (from 2.0 [SEM 0.24] IU/ml to 20.3 [3.14] during maximal, 2.0 [0.22] to 19.0 [2.59] during anaerobic threshold test, 1.8 [0.22] to 5.5 [0.82] during aerobic threshold; p <0.001 during all tests, p = 0.01 aerobic threshold vs maximal and anaerobic threshold). PAI activity decreased during maximal (from 6.6 [2.51] AU/ml to 2.0 [2.00], p <0.05) and anaerobic threshold (5.0 [2.07] to 0.2 [0.22], p <0.01) tests but not during aerobic threshold test (7.0 [3.69] to 4.5 [2.93], p = 0.123). One subject had clearly higher pre-exercise PAI activity and smaller tPA response to exercise as compared to other subjects. All 24 h post-exercise activities were similar to pre-exercise values. Plasma fibrinogen concentration did not change during any tests. Single acute bouts of dynamic exercise transiently increase tPA and decrease PAI activity without affecting plasma fibrinogen concentration in healthy young men.
Association of Variables of Coagulation, Fibrinolysis and Acute-phase with Atherosclerosis in Coronary and Peripheral Arteries and those Arteries Supplying the Brain
Authors: Heinrich Jürgen , Schulte Helmut , Schönfeld Rainer , Köhler Ekkehart , Assmann Gerd 
Published: July 2018

SummaryWe investigated the vessel status of coronary and peripheral arteries and those arteries supplying the brain in 929 consecutive male patients admitted to a coronary rehabilitation unit. The severity of coronary atherosclerosis was scored using coronary angiography. Changes in extracranial brain vessels and manifest cerebrovascular disease (CVD) were determined by B-mode ultrasound and Doppler examination. Peripheral arterial disease (PAD) was diagnosed using base-line and stress oscillography. We assessed variables of coagulation, fibrinolysis, and the acute phase response.There was a significant increase in plasma fibrinogen, plasminogen, d-dimer and C-reactive protein (CRP) with increasing severity of coronary heart disease. Compared to men with unaffected arteries, men with 3 diseased coronary arteries had 58% greater d-dimer concentrations. Patients with CVD and PAD, respectively, also had significantly higher fibrinogen, d-dimer and CRP concentrations. We did not find an association between plasminogen activator inhibitor activity and the severity of coronary atherosclerosis.In conclusion, plasma fibrinogen, d-dimer and CRP concentrations were significantly related to atherosclerosis in the coronary, peripheral and extracranial brain arteries.
Inhibition of Thrombin by Antithrombin III and Heparin Cofactor II In Vivo
Authors: Liu Longbin , Dewar Lori , Song Yingqi , Kulczycky Myron , Morris Blajchman A , Fenton John W , Andrew Maureen , Delorme M , Ginsberg Jeffrey , Preissner Klaus T , Ofosu Frederick A 
Published: July 2018

SummaryThe critical role of thrombin in the pathogenesis of venous and arterial thrombosis, and the effectiveness of glycosaminoglycans as antithrombotic drugs are well known. Antithrombin III is a major inhibitor of thrombin and augmentation of its inhibitory actions by heparin is the basis for the clinical uses of heparin. Recent clinical and experimental studies have demonstrated that another glycosaminoglycan, dermatan sulfate, is an effective antithrombotic drug. Dermatan sulfate catalyses the inhibition of thrombin by heparin cofactor II. The concentrations of heparin cofactor II are higher in the plasmas of individuals with congenital antithrombin III deficiency and pregnant women than controls. The role of heparin cofactor II as a physiologic thrombin inhibitor is unknown. Enzyme-linked immunosorbent assays were used to quantify thrombin-heparin cofactor II and thrombin-antithrombin III endogenous to the plasmas of adult antithrombin III-Hamilton deficient subjects, their siblings with normal antithrombin III levels, pregnant women at term and 3 to 5 days after delivery. Both thrombin-antithrombin III and thrombin-heparin cofactor II complexed with vitronectin were detected in all the plasmas. Significantly, the concentrations of thrombin-heparin cofactor II-vitronectin were higher in the plasmas of congenital antithrombin III deficient subjects and in pre- and post-delivery plasmas than those of normal subjects. In addition, the concentrations of thrombin-heparin cofactor II decreased 3 to 5 days after delivery, reflecting the disappearance of the catalytically active dermatan sulfate elaborated by the placenta. Thus, heparin cofactor II normally inactivates thrombin in vivo, with its role increasing in conditions associated with high levels of heparin cofactor II and/or dermatan sulfate.
Reproducibility of Fibrinolytic Response to Venous Occlusion in Healthy Subjects
Authors: Stegnar Mojca , Mavri Alenka 
Published: July 2018

SummaryFibrinolytic response to venous occlusion is used to assess the efficiency of the fibrinolytic system. Reproducibility of fibrinolytic variables after 20 min upper arm venous occlusion was investigated in 40 apparently healthy subjects tested twice in a period of 11-23 (mean 15) days. Resting and post-venous occlusion euglobulin clot lysis time, tissue-type plasminogen activator (t-PA) activity, t-PA antigen, plasminogen activator inhibitor (PAI) activity and plasminogen activator inhibitor type 1 (PAI-1) antigen determined on the two occasions did not differ significantly. Positive correlation coefficients for variables before (r=0.43-0.74, all p<0.01) and after venous occlusion (r=0.07-0.66) indicated low to moderate associations between repeated measurements. Differences between repeated measurements relative to the initial measurement were greater after venous occlusion than before venous occlusion and were for euglobulin activity 41 (0-826)%, t-PA activity 27 (2-398)%, and for t-PA antigen 27 (0-179)% (medians and ranges). Poor responses were defined by the lowest (euglobulin activity, t-PA activity and t-PA antigen) or the highest (PAI activity) fifth percentile of the distribution. Almost no agreement between poor responses was observed: poor responses at the first occasion were not determined in the same subjects when re-examined after two weeks. It was concluded that due to relatively low reproducibility of the variables measured after venous occlusion the test needs improvement in order to be potentially clinically useful.