bluuebunny/crossref_metadata_2025_split huggingface dataset:
Longitudinal Analysis of Factor VIII Inhibitors in a Previously Untreated Mild Haemophilia A Patient with an Arg593→Cys Substitution
Authors: Timmermans Simone , Turenhout Ellen , Bank Christine , Fijnvandraat Karin , Voorberg Jan , Peters Marjolein , van den Brink Edward 
Published: March 2018

SummaryRecent studies suggest that certain missense mutations associated with mild to moderate haemophilia A predispose to inhibitor development. In this study, we present a longitudinal analysis of the epitope specificity of an inhibitor that developed in a mild haemophiliac with an Arg593→Cys mutation. Immunoprecipitation studies revealed the presence of antibodies directed towards the light chain and A2 domain of factor VIII. Limited reactivity was observed with metabolically labelled C2 domain. Almost complete inhibitor neutralization was achieved upon addition of A2 domain. Binding of the inhibitor was not affected by the presence of the Arg593→Cys substitution in the recombinant A2 fragment. Evaluation of the epitope specificity of anti-factor VIII antibodies in plasma samples obtained at different time-points of inhibitor development revealed initial development of a low titre inhibitor directed towards the A2 domain and factor VIII light chain. A second period of factor VIII replacement therapy resulted in a dramatic rise in factor VIII inhibitor titre, which maintained their original epitope specificity. Based on the results of this and previous studies (Fijnvandraat et al., 1997; Thompson et al., 1997) it is argued that inhibitor development in patients with the Arg593→Cys mutation may proceed via a similar mechanism.
Identification of the Binding Site for an Alloantibody to von Willebrand Factor which Inhibits Binding to Glycoprotein Ib within the Amino-terminal Region Flanking the A1 Domain
Authors: Shibata Masaru , Fujimura Yoshihiro , Takahashi Yukihiro , Nakai Hiroaki , Sakurai Yoshihiko , Asatani Masayo , Nomura Akihiko , Take Hiromichi , Giddings John , Yoshioka Akira , Shima Midori 
Published: March 2018

SummaryAn alloantibody to von Willebrand factor (vWF) which developed in a Japanese boy with type 3 von Willebrand disease has been characterized. The antibody was non-precipitating IgG and the main subclasses were IgG2 and IgG4. The antibody inhibited completely ristocetin-induced platelet aggregation (RIPA) and high shear stress-induced platelet aggregation (SIPA). Its predominant inhibitory role was focused, therefore, on the interaction between vWF and platelet gycoprotein Ib. The antibody reacted with a 52/48 kDa tryptic fragment of vWF (residues 449-728). No reaction was seen, however, with either a 39/34 kDa dispase fragment (480-718) or a recombinant vWF fragment (residues 465-728). These findings suggested that the essential epitope resided in the amino-terminal flanking region of the A1 domain. We synthesized overlapping peptides corresponding to the region containing D3/A1 boundary. A peptide, residues 458-472, bound to the antibody and dose-dependently blocked the antibody binding to the 52/48 kDa fragment. The same peptide neutralized the inhibitory effect of the alloanti-body on SIPA. The data are consistent with the presence of an epitope within residues 458-472 which reacted with the 52/48 kDa fragment.Furthermore, the specific component of the antibody, directed against residues 458-472, blocked vWF binding to GPIb in absence of exogenous agonist. Our results suggest that the region flanking the A1 domain plays an important role in regulating vWF binding to GPIb.
A Synergistic Effect of Serotonin Transporter Gene Polymorphism and Smoking in Association with CHD
Authors: Ohtsuki Tsuyuka , Yamakawa-Kobayashi Kimiko , Amemiya Hiroshi , Fujiwara Hideomi , Kawata Ken-ichi , Ishiguro Hiroki , Hamaguchi Hideo , Arinami Tadao 
Published: March 2018

SummarySerotonin induces vasoconstriction in the presence of atherosclerotic lesions. Platelets acquire serotonin from the extracellular space by serotonin transporter and release it following aggregation. There is a functional polymorphism in the serotonin transporter (5-HTT) gene promoter associated with transcriptional efficacy and plasma serotonin levels. To examine whether the polymorphism is associated with coronary heart disease (CHD) in the Japanese, we analyzed 144 male CHD patients with an onset age before 65 and 222 apparently healthy men. The L allele was observed significantly more frequently in the CHD patients (26%) than in the control subjects (19%); the odds ratio was 1.48 (p <0.03). A significant interaction between the polymorphism and smoking was observed for CHD (p = 0.03), suggesting that the two have a synergistic effect on CHD. Odds ratio of the combination of the L allele and smoking was 1.95 (p <0.003). The 5-HTT gene promoter polymorphism may play a role in susceptibility to CHD, particularly when it is combined with smoking.
Vegetarians and Cardiovascular Risk Factors: Hemostasis, Inflammatory Markers and Plasma Homocysteine
Authors: Muñoz Ximena , Martínez Carlos , Cuevas Ada , Panes Olga , Aranda Eduardo , Guasch Viviana , Strobel Pablo , Muñoz Blanca , Rodríguez Soledad , Pereira Jaime , Leighton Federico , Mezzano Diego 
Published: March 2018

SummaryWe studied hemostatic and inflammatory cardiovascular risk factors (CVRF), and total plasma homocysteine (tHcy) in 26 vegetarians (23 lacto- or ovolactovegetarians and 3 vegans), matched by age, sex and socioeconomic status with omnivorous controls. Vegetarians had significantly lower proportion of eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids in plasma lipids, significantly shortened bleeding time, and increased blood platelet count and in vitro platelet function (aggregation and secretion). Plasma levels of all coagulation or fibrinolytic factors and natural inhibitors synthesized in the liver were lower in vegetarians than in controls. Whereas for some factors this decrease was statistically significant (fibrinogen, factor VIIc, anti-thrombin III, protein S, plasminogen) for the remaining (factors VIIIc, Vc, prothrombin, protein C) a trend in the same direction was found. For hemostatic proteins of predominantly extrahepatic origin (von Willebrand factor, tPA, PAI-1) this tendency was not present. No significant differences in inflammatory proteins (C-reactive protein and α1-protease inhibitor) were detected in both groups. tHcy was significantly increased in vegetarians, and correlated only with cobalamin levels. The increased platelet function and tHcy found in vegetarians may counteract the known cardiovascular health benefits of vegetarian diet (VD).
Increased Thromboxane Metabolites Excretion in Liver Cirrhosis
Authors: Ferro Domenico , Basili Stefania , Iuliano Luigi , Camastra Caterina , Giammarresi Carlo , Santarone Stella , Rocca Bianca , Landolfi Raffaele , Ciabattoni Giovanni , Cordova Corrado , Violi Francesco , Davì Giovanni 
Published: March 2018

SummaryAn augmented systemic production of thromboxane (TX) A2, as assessed by urinary excretion of the thromboxane metabolites, has been described in severe liver cirrhosis. However, the significance of this finding remains unclear since in liver cirrhosis a number of phenomena i.e. altered hepatic TXA2 metabolism, increased intrasplenic platelet destruction, may affect TXA2 entry into systemic circulation as well as its metabolism. In order to further clarify this, we measured both major enzymatic metabolites of TXB2 in the urine of 44 patients affected by liver cirrhosis, subdivided in three classes on the basis of Child-Pugh criteria. Urinary 11-dehydro-TXB2 and 2,3-dinor-TXB2 were assayed with previously validated RIA techniques.The urinary excretion rate of 11-dehydro-TXB2 was significantly (p = 0.0001) increased in the cirrhotic patients (673.5 pg/mg cr, median) in comparison with the controls (275 pg/mg cr, median) but no significant difference could be demonstrated among the excretion rates of the three patient subgroups. The excretion rate of 2,3 dinor-TXB2 was also significantly (p = 0.0001) increased in the patients (824 pg/mg cr, median) in comparison with controls (175 pg/mg cr, median), with a significant (p <0.05) increase from class A (381 pg/mg cr) to class C (1337 pg/mg cr). The sum of the two enzymatic metabolites was significantly (p = 0.0001) increased in the cirrhotic patients in comparison to controls, with a progressive increase from class A (1003 pg/mg cr, median) to class C (2240 pg/mg cr, median). The urinary excretion of 2,3 dinor-TXB2 was significantly (p = 0.008) related to plasma prothrombin fragment 1+2 (F1+2).This study provides further evidence of increased thromboxane biosynthesis in liver cirrhosis. Moreover, we demonstrate intraliver shift of thromboxane metabolic disposition, due to progressive liver decompensation, because only the fraction undergoing β-oxidation to 2,3-dinor-TXB2 was progressively increased with the degree of liver failure. We, also, find a significant correlation between urinary excretion of 2,3-dinor-TXB2 and plasma F1+2, suggesting that clotting activation could partly account for in vivo platelet activation.
Evaluation of the Factors Contributing to Fibrin–dependent Plasminogen Activation
Authors: Siebenlist Kevin , Voskuilen Marijke , Nieuwenhuizen Willem , Mosesson Michael 
Published: March 2018

SummaryPolymerized fibrin strongly enhances tissue plasminogen activator (tPA)-mediated plasminogen activation, concomitant with exposure of ‘fibrin-specific’ epitopes at ‘Aα148-160’ and ‘γ312-324’. To investigate which aspects of polymerization are involved in these activities, we explored the fibrin polymerization process by evaluating the ability of factor XIIIa-crosslinked fibrinogen polymers to expose ‘fibrin-specific’ epitopes and enhance plasminogen activation. Crosslinked normal fibrinogen, fibrinogen with deficient [des Bβ1-42] or defective [Birmingham (AαR16H)] fibrin ‘D:E’ assembly sites (‘EA’), or with defective end-to-end self-association sites (‘D:D’) [Cedar Rapids (γR275C)], exposed both ‘fibrin-specific’ epitopes and enhanced tPA-dependent plasminogen activation, whereas non-crosslinked fibrinogens showed minimal or no such activities. Epitope expression in cross-linked fibrinogen was retained in the presence of the fibrin EA site peptide homolog, gly-pro-arg-pro (GPRP), which inhibits fibrin D:E association, except for the Aα148-160 epitope in des Bβ1-42 fibrinogen, which was not expressed. Fibrin prepared from crosslinked normal or abnormal fibrinogen, except for the des Bβ1-42 fibrin epitopes, which were reduced or absent, expressed ‘fibrin-specific’ epitopes even in the presence of GPRP, which otherwise impairs such expression in noncrosslinked fibrin. Epitope exposure in fibrin prepared from non-cross-linked fibrinogen was nearly normal in Cedar Rapids fibrin (heterozygous D:D defect), but reduced in Birmingham fibrin (heterozygous EA defect), nil in des Bβ1-42 fibrin (EA deficient), and absent in all cases in the presence of GPRP. In contrast, plasminogen activation stimula-tory activity that had been exposed in crosslinked normal fibrinogen or in crosslinked des Bβ1-42 or Cedar Rapids fibrin, was preserved to a large extent in the presence of GPRP, suggesting that once enhanced stimulatory activity and epitopes are exposed, they are not completely reversible. The findings indicate that end-to-end intermolecular associations (D:D) are not critical for ‘fibrin-specific’ epitope exposure, but that polymerization brought about in fibrinogen through factor XIIIa crosslinking, or in fibrin through ‘D:E’ interactions, is necessary for ‘fibrin-specific’ (more correctly, ‘polymerization-specific’) epitope exposure and enhancement of plasminogen activation.
The Distribution of the Secreted and Intracellular Forms of Plasminogen Activator Inhibitor 2 (PAI-2) in Human Peripheral Blood Monocytes Is Modulated by Serum
Authors: Ritchie Helen , Booth Nuala 
Published: March 2018

SummaryPlasminogen activator inhibitor 2 (PAI-2) is produced by activated monocytes in two forms, intracellular and secreted. We have studied the distribution of these two forms in unstimulated human peripheral blood monocytes and after stimulation by thrombin. Fetal calf serum (FCS) in the culture medium was absolutely necessary for accumulation of intracellular PAI-2; but not for synthesis and secretion. Even at a concentration as low as 0.1%, FCS restored accumulation of intra-cellular PAI-2. Increasing concentrations of FCS resulted in an increase in the ratio of intracellular to secreted PAI-2. The factor that promoted accumulation of intracellular PAI-2 was not a platelet product. Failure of monocytes to accumulate PAI-2 did not reflect leakage due to cell death, as assessed by LDH in culture supernatants. We propose that accumulation of intracellular PAI-2 is not simply due to poor secretion, but is an active process that is modulated by factor(s) found in serum.
Rap1b Association with the Platelet Cytoskeleton Occurs in the Absence of Glycoproteins IIb/IIIa
Authors: Fischer Thomas , Duffy Christina , White Gilbert 
Published: March 2018

SummaryPlatelet membrane glycoproteins (GP) IIb/IIIa and rap1b, a 21 kDa GTP binding protein, associate with the triton-insoluble, activation-dependent platelet cytoskeleton with similar rates and divalent cation requirement. To examine the possibility that GPIIb/IIIa was required for rap1b association with the cytoskeleton, experiments were performed to determine if the two proteins were linked under various conditions. Chromatography of lysates from resting platelets on Sephacryl S-300 showed that GPIIb/IIIa and rap1b were well separated and distinct proteins. Immunoprecipitation of GPIIb/IIIa from lysates of resting platelets did not produce rap1b or other low molecular weight GTP binding proteins and immunoprecipitation of rap1b from lysates of resting platelets did not produce GPIIb/IIIa. Finally, rap1b was associated with the activation-dependent cytoskeleton of platelets from a patient with Glanzmann’s thrombasthenia who lacks surface expressed glycoproteins IIb and IIIa. Based on these findings, we conclude that no association between GPIIb/IIIa and rap1b is found in resting platelets and that rap1b association with the activation-dependent cytoskeleton is at least partly independent of GPIIb/IIIa.
Oligosaccharide Configuration of Fibrinogen Kaiserslautern: Electrospray Ionisation Analysis of Intact γ Chains
Authors: Loreth Ralph , George Peter , Brennan Stephen 
Published: March 2018

SummaryElectrospray ionisation mass spectrometry was used to probe the structure of the new N-linked oligosaccharide in fibrinogen Kaiserslautern (γ 380 Lys→Asn). The mass increase of 2177 Da in the new γ chain indicated the attachment of a fully sialylated biantennary oligosaccharide on the new Asn residue; the expected increase for this change being 2192 Da. Some 95% of the new oligosaccharide was in the disialylated state while only 5% of the endogenous γ chain carbohydrate was disialylated in the control. Mass measurements of intact Kaiserslautern γ chains after neuraminidase treatment of the native fibrinogen confirmed a total of three residues of sialic acid in the dominant isoform. Incubation with endoglycosidase F showed that the new oligosaccharide was more resistant to hydrolysis than the endogenous one. Recent X-ray analyses of covalently linked D domains show that position γ 380 is distant from both the GPR binding pocket and the D-D interface. It appears that the polymerisation defect of this fibrinogen results from electrostatic repulsion between condensing protofibrils and that this is induced by the two new residues of sialic acid that are present on the new γ chain.
Differences in Platelet α-granule Release between Normals and Immune Thrombocytopenic Patients and between Young and Old Platelets
Authors: Tracey Jayne , Recht Michael , DeCastro Laura , Rinder Christine , McHugh Colleen , Smith Brian , Rinder Henry 
Published: March 2018

SummaryThe risk of serious bleeding in patients with immune thrombocytopenic purpura (ITP) appears to be less than in comparably thrombocytopenic patients with megakaryocytic hypoplasia. It has been proposed that this difference is due to enhanced hemostatic activity of young platelets, which are increased in the circulation during ITP. We examined α-granule release in reticulated platelets (RP), which are thought to be the youngest circulating platelets, and in older non-reticulated platelets (non-RP) in normal human controls and ITP patients. Normal controls had a mean RP of 7%, compared with 42% in ITP patients. The mean concentration of thrombin receptor agonist peptide (TRAP) causing 50% of control RP to express CD62P (EC50) was 0.82 ± 0.08 μM (SEM), significantly higher than the TRAP CD62P EC50 for RP in ITP, 0.57 ± 0.06 uM (p = 0.04). Similarly, the TRAP EC 50 for non-RP in controls, 0.84 ± 0.09 μM, was significantly higher than in ITP, 0.56 ± 0.07 μM (p = 0.03), suggesting that all platelets in ITP have an enhanced α-granule threshold response to TRAP compared with controls, while RP and older platelets within each patient group have similar threshold sensitivities to TRAP. By contrast, high-dose TRAP caused RP to express twice as much mean and total CD62P as non-RP in both ITP patients and controls (p <0.05 for both comparisons). We conclude that compared with controls, all platelets in ITP are primed to undergo α-granule release to TRAP, while in both ITP and controls, the newly circulating, reticulated platelets have the potential to contribute greater amounts of CD62P surface ligand compared with older platelets (non-RP) after stimulation. Both the increased RP% and enhanced platelet response to agonist in ITP may contribute to maintenance of hemostasis despite thrombocytopenia.
SR 121787, a New Orally Active Fibrinogen Receptor Antagonist
Authors: Savi Pierre , Badorc Alain , Lalé Alain , Bordes Marie-Françoise , Bornia Josette , Labouret Catherine , Bernat André , Cointet Paul de , Hoffmann Peter , Maffrand Jean-Pierre , Herbert Jean-Marc 
Published: March 2018

SummaryThe aim of this study was to describe the pharmacological properties of SR 121787, a new antiaggregating drug which is metabolized in vivo into SR 121566, a potent non-peptide antagonist of Gp IIb/IIIa. In vitro, SR 121566 antagonized the binding of [125I]-fibrinogen (IC50 = 19.8 ± 6.3 nM) and of [125I]-L-692,884, an RGD-containing peptide (IC50 = 291 ± 96 nM) to activated human platelets. SR 121566 inhibited the aggregation of human platelets induced by ADP, collagen, thrombin, arachidonic acid and PAF at concentrations lower than 0.1 μM. Adhesion of human platelets to adhesive proteins was inhibited by SR 121566 (IC50 = 40.3 ± 2.5 nM) only when Gp IIb/IIIa and fibrinogen were involved. No effect was found with regard to other adhesive proteins and/or other integrins. SR 121787 demonstrated a potent and sustained antiaggregating effect when administered intravenously to baboons at a dose 50 μg/kg, and eight hours after the administration of 100 μg/kg, ADP-induced aggregation was still strongly inhibited (more than 80%). A single oral administration of 2 mg/kg of SR 121787 produced a nearly complete inhibition of platelet aggregation for up to 8 h (ED50 at 8 h = 193 ± 20 μg/kg), a significant residual antiaggregating activity being still observed 24h after the administration. When administered orally to rabbits, SR 121787 exhibited a potent antiaggregating (ED50 = 2.3 ± 0.3 mg/kg) and antithrombotic activity in an arterio-venous shunt thrombosis model (ED50 = 10.4 ± 0.8 mg/kg). After oral and IV administration, SR 121787 was well tolerated suggesting that SR 121787, the most potent and long lasting orally active Gp IIb/IIIa antagonist described to date, is a promising antithrombotic compound.
Heparin Regulates ICAM-1 Expression in Human Endothelial Cells: An Example of Non-Cytokine-Mediated Endothelial Activation
Authors: Hoggatt A. M. , Faulk W. Page , Miller Steven 
Published: March 2018

SummaryActivated endothelial cells up-regulate the expression of several molecules on their plasma membranes, including intercellular adhesion molecule-1 (ICAM-1). The role of heparin in regulating endothelial cell gene expression is unclear. We thus have investigated the ability of heparin to regulate ICAM-1 gene expression by using flow cytometry and the ribonuclease protection assay with human umbilical vein and aortic endothelial cells cultured in growth medium supplemented with 90 μg/ml heparin (heparin-sufficient, HS) or in growth medium without added heparin (heparin-deficient, HD). We found that HD medium increased plasma membrane protein and mRNA for ICAM-1 but not for HLA-DR, even though both ICAM-1 and HLA-DR protein and mRNA were inducible by gamma interferon (IFN-γ). In addition, phorbol ester and IFN-γ increased the expression of plasma membrane ICAM-1 or ICAM-1 and HLA-DR, respectively, more in HD medium than in HS medium. We found that the HD-mediated increase of ICAM-1 mRNA was reversible by the addition of heparin, and that the half-life of ICAM-1 mRNA was the same in both HS- and HD-treated cells. Also, heparin was found to suppress increases in ICAM-1 mRNA at a concentration as low as 5 μg/ml. These findings indicate that heparin deficiency induces endothelial activation characterized by increased ICAM-1, and that such induction is not dependent on cytokines or endotoxin. The modulation of ICAM-1 expression by heparin appears to occur at the transcriptional level. Thus, heparin may have a role in regulating endothelial function by affecting the expression of ICAM-1, thereby impacting upon the trans-endothelial trafficking of leukocytes.
Fibrin-Rich and Platelet-Rich Thrombus Formation on Neointima: Recombinant Tissue Factor Pathway Inhibitor Prevents Fibrin Formation and Neointimal Development following Repeated Balloon Injury of Rabbit Aorta
Authors: Asada Yujiro , Hara Seiichiro , Tsuneyoshi Atsushi , Hatakeyama Kinta , Kisanuki Atsushi , Marutsuka Kousuke , Sato Yuichiro , Kamikubo Yu-ichi , Sumiyoshi Akinobu 
Published: March 2018

SummaryThrombus formation and neointimal growth are the critical events in restenosis after balloon angioplasty. However, the responses of diseased vessels to injuries caused by balloon angioplasty have not been well examined. We investigated the thrombus formation and neointimal development following the balloon injury to the previously induced neointima in the rabbit aorta and the effects of recombinant tissue factor pathway inhibitor (rTFPI) on these responses. Rabbit thoracic aortas were subjected to injury with a Fogarty 4F balloon catheter at 1.75 atm (first injury), and 4 weeks later the same vessels were subjected to the second injury with a Swan-Ganz 5F balloon catheter at 1.4 atm (mild-injury group) or 1.8 atm (severe-injury group), and immediately after that a retrograde bolus injection of rTFPI (100 μg/kg body weight) or saline was performed into the injured segments via the central tube of the Swan-Ganz catheter. Twenty minutes after the second injury, the injured surfaces were covered with platelet-rich thrombi in the mild-injury group and with fibrin-rich thrombi in the severe-injury group. Damaged intimal smooth muscle cells, which were immunohistochemically positive for tissue factor (TF), were observed beneath the fibrin-rich thrombi. The neointima 4 weeks after the second injury was significantly thicker in the severe-injury group than in the mild-injury group. The bolus infusion of rTFPI markedly inhibited fibrin formation on the injured surfaces, and significantly reduced the neointimal development in the severe-injury group at 4 weeks after the second injury. These results indicate that TF-dependent coagulation pathway is primarily responsible for fibrin-rich thrombus formation and may play an important role in neointimal development following the balloon injury to the rabbit aortic neointima. Additionally the bolus administration of rTFPI to the injured vessels could prevent mural thrombus formation and neointimal growth after balloon angioplasty.