bluuebunny/crossref_metadata_2025_split huggingface dataset:
Molecular Characterisation and Three-Dimensional Structural Analysis of Mutations in 21 Unrelated Families with Inherited Factor VII Deficiency
Authors: Peyvandi Flora , Jenkins Vincent , Mannucci Pier , Billio Atto , Zeinali Sirous , Perkins Stephen , Perry David 
Published: March 2018

SummaryFactor VII (FVII) is a four-domain glycoprotein that plays a critical role in the initiation of blood coagulation. Hereditary deficiencies of this plasma protein results in a bleeding diathesis that varies in severity amongst affected patients. We have analysed the FVII gene in 27 patients with FVII deficiency from 21 unrelated families predominantly of Middle-Eastern extraction. A total of 19 different mutations were identified, of which 12 were novel and 7 had been previously reported. Nine of the 12 novel mutations were missense mutations located in the Gla domain (Ser23Pro), the second epidermal growth factor domain (Cys135Arg) and the catalytic serine protease domain (Arg247Cys, Arg277Cys, Ser282Arg, Pro303Thr, Ser363Ile, Trp364Cys, Trp364Phe), of which five are homozygous. Three novel splice mutations were identified in intron 1a (IVS1a+5), intron 2 (IVS2+1) and intron 6 (IVS6+1). Of the seven previously reported mutations, five were missense mutations of which three are homozygous (Gln100Arg, Arg152Gln, Arg304Gln, Cys310Phe and Thr359Met), one was a 17 bp deletion (10585del17bp) and one was a splice site mutation within intron 7 (IVS7+7). This study has significantly extended the current database of FVII mutations, including the number of known homozygous mutations. Conformational analyses of crystal structures for FVIIa and the FVIIa-tissue factor complex provided likely explanations for the effect of the missense mutations on FVIIa secretion or function. In particular, since 23 missense mutations were located to the serine protease domain, mostly to the region between the catalytic triad and the contact surface with tissue factor, this showed that the orientation of the serine protease domain relative to bound tissue factor in the complex is crucial for functional activity.
Discrimination of von Willebrands Disease (VWD) Subtypes: Direct Comparison of von Willebrand Factor:Collagen Binding Assay (VWF:CBA) with Monoclonal Antibody (MAB) Based VWF-capture Systems
Authors: Henniker Anthony , Facey David , Hertzberg Mark , Favaloro Emmanuel 
Published: March 2018

SummaryDiscrimination of von Willebrand’s Disease (VWD) subtypes is important since it influences management. Qualitative [ie Type 2A, 2B, 2M] defects exhibit von Willebrand factor (VWF) discordance and give high VWF:Ag to VWF:’activity’ ratios. Classically, VWF:’activity’ is assessed using the VWF:RCof assay. The VWF:CBA is an ELISAbased VWF-functional adhesive assay which has consistently proved to be superior to VWF:RCof. A commercially available monoclonal antibody (MAB) based ELISA assay system claimed to mimic a VWF:RCof-like activity has also been recently described (‘SE’), as has the production and characterisation of a large number [n = 10] of locally generated anti-VWF MAB. In the current study, we have adapted these MAB to in-house ELISA assays to assess their utility for VWD diagnosis and subtype discrimination, and to compare them with other assay systems. Thus, the VWF:CBA, VWF:RCof by agglutination, the SE assay, and in-house MAB based assays have been directly compared for their ability to discriminate Type 1 [n = 9] from Type 2 VWD samples [phenotypes 2A and 2B; n = 11]. In summary, MAB-based systems can be used to measure VWF and confirm a diagnosis of VWD, as well as exhibiting some VWD-subtype-discriminatory capabilities. However, better evidence of VWF-discordance was usually achieved using the VWF:RCof (agglutination) assay, while the greatest degree of VWFdiscordance was consistently observed using the VWF:CBA assay. In conclusion, the VWF:CBA assay proved to offer the best diagnostic predictive tool for a Type 2 VWD defect, while MAB-based systems appear to be less effective in this regard.
Abbreviations: HMW: High Molecular Weight [VWF], MAB: Monoclonal antibody (/antibodies), PNP: Pooled Normal Plasma, RIPA: Ristocetin induced platelet aggregation, VWD: von Willebrands disease (/disorder), VWF: von Willebrand Factor, VWF:Ag: von Willebrand Factor Antigen (assay), VWF:CBA: Collagen Binding [Activity] Assay for VWF, VWF:RCof: Ristocetin Cofactor Assay for VWF
Sequence Alignment between vWF and Peptides Inhibiting the vWF-collagen Interaction Does not Result in the Identification of a Collagen-binding Site in vWF
Authors: van der Plas R. M. , Vandecasteele G. , Vauterin S. , Huizinga E. G. , Sixma J. J. , Deckmyn H. , Vanhoorelbeke K. 
Published: March 2018

SummaryWe previously found that two peptides (N- and Q-peptide) selected by phage display for binding to an anti-vWF antibody, were able to inhibit vWF-binding to collagen (1). The sequence of those peptides could be aligned with the sequence in vWF at position 1129-1136 just outside the A3-domain. As the peptides represent an epitope or mimotope of vWF for binding to collagen we next wanted to study whether the alignment resulted in the identification of a new collagen binding site in vWF. We mutated the 1129-1136 VWTLPDQC sequence in vWF to VATAPAAC. Expressing this mutant vWF (7.8-vWF) in a fur-BHK cell line resulted in well processed 7.8-vWF containing a normal distribution of molecular weight multimers. However, binding studies of this mutant vWF to rat tail, human and calf skin collagens type I, to human collagen types III and VI, revealed no decrease in vWF-binding to any of these collagens. Thus, although the N-and Q-peptides did inhibit the vWF-collagen interaction, the resulting alignment with the vWF sequence did not identify a collagen binding site, pointing out that alignments (although with a high percentage of identity) do not always result in identification of binding epitopes. However, suprisingly removal of the A3-domain or changing the vWF sequence at position 1129-1136 resulted in an increase of vWF-binding to human collagen type VI and to rat tail collagen type I, implying that these changes result in a different conformation of vWF with an increased binding to these collagens as a consequence.
Levels of Activated FXII in Survivors of Myocardial Infarction – Association with Circulating Risk Factors and Extent of Coronary Artery Disease
Authors: Kohler Hans , Carter Angela , Stickland Max , Grant Peter 
Published: March 2018

SummaryLittle data is available regarding the activated form of factor XIIa (FXIIa) in survivors of myocardial infarction. 292 Caucasian patients characterised for extent of coronary atheroma by angiography and for a past history of myocardial infarction and 77 healthy controls were included in the study. To investigate the relationship between coronary artery disease, activated factor XII and other circulating factors, we studied levels of FXIIa, cholesterol, triglycerides, fasting insulin, fibrinogen, FVII:C, t-PA antigen and PAI-1 antigen. Factor XIIa levels were higher in all patients [2.5 (2.3-2.6) ng/ml] and in patients with a history of MI [2.6 (2.4-2.9) ng/ml] than in controls [1.9 (1.7-2.1) ng/ml], p <0.0001. In patients, FXIIa levels positively correlated with FVII:C, BMI, cholesterol, insulin, PAI-1 antigen, t-PA antigen and triglycerides. In controls FXIIa levels only correlated with PAI-1 antigen and triglycerides. FXIIa levels were strongly associated with extent of coronary stenosis: 2.8 (2.6-3.1) ng/ml and 2.6 (2.3-2.9 ng/ml) in those with 2 and 3 vessels stenosed compared to 2.1 (1.9-2.3) ng/ml in those with 0 vessel stenosed (p = 0.0004). Activated FXII relates to both extent of coronary atheroma and to a past history of myocardial infarction and clusters with features of insulin resistance.
Attenuation of Thrombolysis-induced Increase of Plasminogen Activator Inhibitor-1 by Intravenous Enalaprilat
Authors: Wagner Andreas , Gwechenberger Marianne , Herkner Harald , Müllner Marcus , Woisetschläger Christian , Laggner Anton , Hirschl Michael 
Published: March 2018

SummaryWe examined the effect of intravenous enalaprilat on the course of PAI-1 plasma levels in 23 patients with acute myocardial infarction undergoing thrombolytic therapy. All patients received 100 mg aspirin, 1000 IU/h heparin, thrombolysis with 100 mg rt-PA within 90 min, and betablockers. Eleven out of 23 patients received 5 mg enalaprilat intravenously prior to thrombolysis. Blood samples for determination of PAI-1 plasma levels were collected on admission, 2, 4, 6, 12, and 24 h after thrombolysis. PAI-1 plasma levels in patients receiving enalaprilat were similar to those of the control patients before thrombolysis (5 ng/ml, 95% confidence interval: 2-10 vs. 7 ng/ml, 95% confidence interval: 2-10; p = 0.5). The PAI-1AUC was 9 ng/ml/h (95% confidence interval: 5-10) in the enalaprilat group and 19 ng/ml/h (95% confidence interval: 13-26) in the control group (p = 0.0006). The maximum difference was observed 6 h after thrombolysis (enalaprilat: 13 ng/ml, 95% confidence interval: 5-25, control: 42 ng/ml, 95% confidence interval: 18-55; p = 0.003).Our study clearly demonstrates that application of intravenous enalaprilat prior to thrombolysis attenuates the thrombolysis-related increase of PAI-1. This finding may suggest a possible therapeutic approach to influence the fibrinolytic system in patients with acute myocardial infarction after thrombolysis.
Ecarin Clotting Time: a Predictive Coagulation Assay for the Antithrombotic Activity of Argatroban in the Rat
Authors: Lunven Catherine , Girardot Christine , Lechaire Irène , Girard Denise , Charles Marie-Christine , Ferrari Patrice , O’Brien Donogh , Berry Christopher 
Published: March 2018

SummaryWe studied the use of the Ecarin Clotting Time (ECT) as a predictive assay of the antithrombotic effects of argatroban in a new tissue factor-dependent model of venous thrombosis and a model of arterial thrombosis in the rat. Heparin was used as a reference anticoagulant.Infusions of argatroban dose-dependently increased the ECT across the range of doses required for antithrombotic activity in models of venous and arterial thrombosis (1.25-40 μg/kg/min). The TT was only useful as a marker in the case of venous thrombosis, since, in the arterial thrombosis model, the clotting times were >200 s in the majority of animals receiving antithrombotic doses. The aPTT is not sufficiently sensitive to be predictive of an antithrombotic effect in the venous model, and shows only modest increases in the arterial thrombosis model. Heparin did not significantly increase the ECT at antithrombotic doses in the venous thrombosis model, and only increased the ECT by 53% at 40 μg/kg/min in the arterial model, despite a marked antithrombotic effect. Both the TT and aPTT were dose-dependently increased by heparin at doses active in the venous model, whereas both parameters were >200 s at doses active in the arterial thrombosis model.Thus, the ECT provides a predictive marker for the antithrombotic activity of argatroban in both venous and arterial thrombosis, at least in the rat.
Expression of Thrombospondin 1 on the Surface of Activated Platelets Mediates their Interaction with the Heavy Chains of Human Kininogens through Lys 244-Pro 254
Authors: Kunapuli Satya , Walz Daniel , Colman Robert , DeLa Cadena Raul 
Published: March 2018

SummaryPlatelet thrombospondin (TSP1) forms a complex with high (HK) and low (LK) molecular weight kininogens. We isolated a proteolytic fragment from HK and LK heavy chains (12 kDa) recognized by TSP1 with a N-terminal sequence, K244ICVGCPRDIP254. Lys244-Pro254 oxidized to cyclic form prevented binding of 125I-LK to TSP1. This effect was abolished by reduction and alkylation. Oxidized peptide KICVGCPRDIP (100 μM) reversed the known inhibitory effects of LK or HK (1 μM), on thrombin-induced platelet activation, suggesting this peptide forms part of the cell binding site on HK and LK for activated platelets. KICVGCPRDIP completely inhibited the binding of 125I-LK to activated platelets. However, the peptide only partially inhibited binding of 125I-HK to platelets, suggesting an additional binding site on the HK light chain. Fluorescein-labeled KICVGCPRDIP bound directly and specifically to activated platelets. A monoclonal antibody directed to TSP1 partially inhibited the binding of 125I-HK to activated but not inactivated platelets. We conclude residues Lys244-Pro254 on kininogen heavy chain is responsible for binding to thrombospondin on the surface of activated platelets.
The Effects of Low-intensity Warfarin on Coagulation Activation in Patients with Antiphospholipid Antibodies and Systemic Lupus Erythematosus
Authors: Crowther Mark , Julian James , Stewart Katheryne , Donovan Dianne , Kaminska Elzbieta , Laskin Carl , Ginsberg Jeffrey , Douketis James 
Published: March 2018

SummaryThe optimal intensity of oral anticoagulant therapy for the prevention of thromboembolism in patients with antiphospholipid antibodies (APLA) and systemic lupus erythematosus is controversial. Retrospective studies have suggested that patients with APLA are resistant to oral anticoagulant therapy, with a targeted International Normalization Ratio (INR) of 2.0 to 3.0, and that a higher intensity of anticoagulation (INR: 2.6 to 4.5) is required to prevent recurrent thromboembolism. To investigate if patients with APLA are resistant to the anticoagulant effect of low intensities of warfarin therapy, we performed a randomized trial in which 21 patients with APLA and systemic lupus erythematosus were allocated to receive one of three intensities of warfarin (INR: 1.1 to 1.4, 1.5 to 1.9 or 2.0 to 2.5) or placebo for four months. The main outcome was the effect of each intensity of warfarin therapy on prothrombin fragment 1+2 level (F1+2), that was used as a marker of coagulation activation. When F1+2 levels in patients allocated to the three warfarin intensities were compared to F1+2 levels in the placebo group, there was a statistically significant decrease (p <0.05) in the patient group receiving warfarin with a targeted INR of 2.0 to 2.5 at two, three and four months, and in the patient group with a targeted of INR 1.5 to 1.9 at three months. We conclude that in patients with APLA and systemic lupus erythematosus, warfarin therapy, with a targeted INR of 2.0 to 2.5, is effective in suppressing coagulation activation, and therefore, might be effective in preventing thromboembolism.
Surface-dependent Conformations of Human Fibrinogen Observed by Atomic Force Microscopy under Aqueous Conditions
Authors: Sit P. Sidney , Marchant Roger 
Published: March 2018

SummaryConformational differences in human fibrinogen under aqueous conditions on hydrophobic, positively charged and negatively charged surfaces, were examined by atomic force microscopy (AFM). Hydrophobic and positively charged surfaces were prepared by depositing octadecyltrichlorosilane (OTS) and 3-aminopropyltriethoxysilane (APTES) respectively on cleaned glass coverslips forming self-assembled monolayers. The negatively charged surface was prepared by freshly cleaving muscovite mica. AFM operated in fluid tapping mode with an ultrasharp carbon spike probe was used to obtain the molecular scale images. Fibrinogen displayed a characteristic trinodular structure on all three surfaces, although additional U-shaped conformations were observed on mica. In its native hydrated state, fibrinogen is well represented by three connected ellipsoids in close proximity. Quantitative dimensional analysis, which yielded structural information in three dimensions, indicates that surface-dependent structural deformation or spreading of fibrinogen increases according to the order: mica < APTES < OTS. Molecular length, and D and E domain widths of fibrinogen are increased, while the corresponding heights are decreased. The results provide direct evidence that material surface properties affect the conformational state of interacting fibrinogen.
Platelet-vessel Wall Interactions with Third-generation Oral Contraceptives: No Evidence of Detrimental Effects
Authors: Fruzzetti Franca , Giannessi Daniela , Ricci Cabiria , Bernini Walter , Puntoni Roberto , Genazzani Andrea Riccardo , De Caterina Raffaele 
Published: March 2018

SummaryBecause of the association of oral contraceptives (OC) and cigarette smoking with an increased thrombotic risk, we evaluated thromboxane (TX) and prostacyclin urinary (u) metabolites, as in vivo indices of platelet-vessel wall interactions, in women assigned to third generation OC. Twenty-eight women (15 smokers) underwent a 6-month trial of 30 μg ethinylestradiol plus 0.150 mg desogestrel. Cotinine plasma levels were elevated only in persons classified as smokers and serum TXB2 determination confirmed the absence of cyclooxygenase inhibition throughout the study. u-TXB2 and 11-dehydro-TXB2 were higher in smokers than in non-smokers. OC decreased u-11-dehydro-TXB2 both in smokers (from (pg/μmol creatinine) 35.1 ± 6.9 to 15.8 ± 2.8; P <0.025) and non-smokers (from 31.7 ± 9.8 to 20.6 ± 4.8, P = N.S.). u-6-keto-prostaglandin(PG)F1α excretion, also higher in smokers compared to non-smokers, was also reduced after OC in smokers (from (pg/μmol creatinine) 24.3 ± 5.2 to 14.8 ± 2.3; P <0.05). Smokers also had a trend to higher u-2,3-dinor-6-keto-PGF1α, marginally reduced by OC.Thus, the OC regimen used here improves – if anything – platelet vessel wall interactions as assessed by prostanoid production in vivo. The prothrombotic tendency associated with the use of OC in smokers does not appear to be mediated by changes in platelet-vessel wall interactions.
The Factor II G20210A and Factor V G1691A Gene Transitions and Coronary Heart Disease
Authors: Arsic Tanja , Katz Norbert , Tillmanns Harald , Hehrlein Friedrich , Haberbosch Werner , Gardemann Andreas 
Published: March 2018

SummaryBackground: G to A transitions at nucleotide position 20210 of the factor II (FII) gene and at 1691 of the factor V (FV) gene have been shown to be associated with an increased risk of venous thrombosis. Since it is still unclear whether both gene variations are also related to an increased risk of coronary heart disease (CHD), we studied the relation of both gene variations to coronary artery disease (CAD) and myocardial infarction (MI) in a sample of 2210 male individuals whose coronary anatomy were defined by coronary angiography.Results: In the total sample, the FII G20210A gene variation was not associated with the presence or the extent of CAD, the latter defined either by the degree of vessel disease or by a CHD score according to Gensini. However, individuals with unfavourable lipid profiles showed pronounced differences in CHD scores between GA heterozygotes and GG homozygotes; this observation applied in particular to younger patients (<62 years; mean age of total sample) who simultaneously had low apoAI/apoB ratios (< 1.19, mean value) and high Lp(a) plasma levels (>28 mg/dl; mean value). In addition, in subjects without acetylsalicylic acid treatment GA heterozygotes had clearly higher CHD scores than AA genotypes. Further restriction to smokers, to subjects with high fibrinogen plasma levels (>3.47 g/l; mean value) or to patients with high glucose concentrations (>112 mg/dl; mean value) tended to increase the difference in CHD score between FII G20210A genotypes. An association of the FII G20210A gene variation with nonfatal MI was not observed. In the total sample and in high and low risk subpopulations, an association of the FV G1691A gene variation was not detected neither with presence and extent of CAD or with nonfatal MI.Conclusion: The importance of the factor II G20210A gene variation for CHD may be restricted to individuals with major cardiovascular risk factors. In addition, the present study did not strengthen the hypothesis of the factor V G1691A transition as a risk factor of coronary heart disease neither in the total sample nor in subgroups of individuals who were at high or low risk of CHD.
Performances of the Fibrin Monomer Test for the Exclusion of Pulmonary Embolism in Symptomatic Outpatients
Authors: Bounameaux Henri , Perrier Arnaud , Moerloose Philippe , Reber Guido 
Published: March 2018

SummaryMany studies have shown that D-dimer determinations can be used for the exclusion of venous thromboembolism in symptomatic outpatients, depending however on the method of D-dimer measurement. Another related assay, the Fibrin Monomer test which measures soluble fibrin levels in plasma by ELISA, is now available.We have evaluated the performances of this assay for the exclusion of pulmonary embolism (PE) in 426 consecutive outpatients presenting at the emergency ward of our institution. Diagnosis of PE was made by D-dimer measurement, compression ultrasonography, lung scintigraphy, venography and pulmonary angiography. With a cut-off of 3 μg/ml, the sensitivity and the negative predictive value were both 100% (95% CI: 97.1-100 and 96.3-100 respectively) and the specificity 33% (95% CI: 25.7-38.1). With 4 μg/ml, the corresponding figures were 98.4 (95% CI: 94.4-99.8), 98.3 (95% CI: 94.1-99.8) and 39% (95% CI: 33.6-44.7) respectively. The prevalence of PE was 30%, the exclusion rates were 23 and 27% for either cut-off. When compared with a reference D-dimer assay (Asserachrom D-Di), a good correlation was observed. In conclusion, this is the first study suggesting the interest of this Fibrin Monomer test to rule out PE; these results, however, need to be confirmed by other studies.
Antithrombotic and Anticoagulant Activities of a Low Molecular Weight Fucoidan by the Subcutaneous Route
Authors: Jouault S. Colliec , Mauray S. , Theveniaux J. , Sternberg C. , Vidal Boisson , Fischer A. M. , Millet J. 
Published: March 2018

SummaryFucoidans (high-molecular-weight sulfated polysaccharides extracted from brown seaweeds) have anticoagulant and antithrombotic effects. They inhibit thrombin by catalyzing both serpins (antithrombin and heparin cofactor II) according to their chemical structures and origins. In this study, a low-molecular-weight (LMW) fucoidan of 8 kDa was obtained by chemical degradation of a high-molecular-weight fraction. The antithrombotic and anticoagulant activities of this new compound were compared to those of a low-molecular-weight heparin (LMWH), dalteparin, following subcutaneous administration to rabbits. This LMW fucoidan exhibited dose-related venous antithrombotic activity, with an ED80 of about 20 mg/kg, 2 h after a single subcutaneous injection. Its activity was comparable to that of dalteparin (close to 200 anti-Xa IU/kg) and was maximal 30 min after a single subcutaneous injection. The activity remained stable (about 70%) from 1 to 4 h after injection, but disappeared by 8 h. The antithrombotic activity was not associated with either a prolongation of the thrombin clotting time (TCT) or an increase in anti-Xa activity, contrary to dalteparin. A slight prolongation of APTT occurred with both compounds. This venous antithrombotic activity was associated with a decrease in ex vivo thrombin generation and with a significant increase in the lag phase in a thrombin generation test. LMW fucoidan thus has potent antithrombotic activity and a potentially weaker haemorrhagic effect (i.e. a smaller effect on coagulation tests and a smaller prolongation of the bleeding time) than dalteparin.