bluuebunny/crossref_metadata_2025_split huggingface dataset:
c7E3 Fab Inhibits Low Shear Flow Modulated Platelet Adhesion to Endothelium and Surface-adsorbed Fibrinogen by Blocking Platelet GP IIb/IIIa as well as Endothelial Vitronectin Receptor
Authors: Agneskirchner Jens , Bode Patrick , Spannagl Michael , Wurzinger Laurenz , Reininger Armin 
Published: March 2018

SummaryThe c7E3 Fab reduces ischemic complications in patients undergoing high-risk coronary angioplasty or atherectomy. The present study investigated how c7E3 Fab inhibition of the platelet receptor glycoprotein IIb/IIIa and the endothelial vitronectin receptor affected platelet adhesion to endothelium and surface adsorbed fibrinogen under flow conditions.Platelet adhesion was examined using a stagnation point flow device with shear stress and shear rates up to 2.2 dynes/cm2 and 170 s−1, respectively. Ex vivo adhesion was compared between two groups of patients with acute myocardial infarction (AMI) treated with angioplasty and stent implantation and a group of healthy controls. Only one AMI group received c7E3 Fab therapy. Patients in both groups were administered acetyl salicylic acid (ASA) and heparin. In AMI patients c7E3 Fab reduced platelet adhesion to adsorbed fibrinogen by 79% compared to AMI patients without c7E3 Fab treatment and by 74% compared to healthy controls. Thirty hours after termination of c7E3 Fab infusion adhesion had slightly recovered with an inhibition of 61% and 52% still present, respectively. Additionally, in vitro platelet adhesion to intact endothelium and to adsorbed fibrinogen was measured during superfusion with ADP stimulated platelet rich plasma of healthy controls to which c7E3 Fab was added at a final concentration fc of 20 µg/ml. In spite of ADP stimulation c7E3 Fab completely blocked platelet adhesion to adsorbed fibrinogen and, moreover, to intact endothelium. Preincubation of endothelial cells with c7E3 (fc = 20 µg/ml) blocked adhesion of ADP-stimulated platelets by approximately 50%.Apart from the inhibition of platelet aggregation, c7E3 Fab added in vitro and given therapeutically in patients effectively blocks platelet adhesion to components of the injured as well as intact vessel wall under stagnation point flow conditions.
Oxidation of Human α-Thrombin by the Myeloperoxidase-H2O2-chloride System: Structural and Functional Effects
Authors: Landolfi Raffaele , De Cristofaro Raimondo 
Published: March 2018

SummaryThe myeloperoxidase-H2O2-chloride system (MPOS) is exploited by white blood cells to generate reactive oxygen species in many processes involved in the pathogenesis of inflammation and atherothrombosis. This study investigated the biochemical and functional effects of α-thrombin oxidation by MPOS. This system, in the presence of 100 µM L-tyrosine, caused in the thrombin molecule loss of tryptophan and lysine residues and formation of dityrosine, chloramine and carbonyl groups. The same changes could be directly induced by thrombin incubation with reagent HOCl, but not with H2O2 alone. Exposure to either MPOS or HOCl caused major functional abnormalities in human α-thrombin. The interaction of oxidized (ox-)thrombin with Protein C and antithrombin III-heparin complex were most sensitive to oxidation, being the kcat/Km value for Protein C hydrolysis roughly reduced 13-fold and the affinity for the antithrombin III-heparin complex decreased approximately 15-fold. Ox-thrombin interaction with small synthetic peptides showed several changes, arising from a perturbation of the S2-S3 specificity of the enzyme. Ox-thrombin was also characterized by a 5-fold decrease of the kcat/Km value for both fibrinopeptide A and B release from fibrinogen, a 5.8-fold increase of the EC50 value for platelet activation and a 2-fold decrease of binding affinity for thrombomodulin. The above results indicate a high sensitivity of thrombin to oxidative modifications by myeloperoxidase. Perturbed interactions with Protein C and the heparin-ATIII complex were the most relevant functional abnormalities of ox-thrombin.
Abbreviations: DNPH: 2,4-dinitrophenylhydrazine; FpA: fibrinopeptide A; FpB: fibrinopeptide B; FRS: Fibrinogen recognition Site; GpIb: Glycoprotein Ib; HBS: Heparin Binding Site; MPO: myeloperoxidase; MPOS: Myeloperoxidase System; Pip: Pipecolynic acid; PMN: polymorphonuclear cells; pNA: para-nitroaniline; PPACK: Phenylalanine-Proline-Arginine-Chloromethyl-Ketone; TM: Thrombomodulin; TNB: 5-thio-2-nitrobenzoic acid; TPCK: N-tosyl-L-Phenylalanine-Chloromethyl-Ketone. Thrombin residues are numbered according to the chymotrypsin numbering system (31).
Venous Thrombotic Risk in Family Members of Unselected Individuals with Factor V Leiden
Authors: Lensen R. P. M. , Bertina R. M. , de Ronde H. , Vandenbroucke J. P. , Rosendaal F. R. 
Published: March 2018

SummaryThe factor V Leiden mutation (FVL) leads to a seven-fold increased risk of venous thromboembolism (VTE). In thrombophilic families, 25% of carriers have experienced thrombosis before the age of 40 years. Aim of our study was to assess the association of FVL with VTE in first-degree family members of unselected symptomatic and asymptomatic carriers of FVL.We tested 197 relatives of consecutive thrombosis patients with FVL and 36 relatives of asymptomatic carriers on the presence of FVL and the occurrence of VTE.The incidence of VTE in relatives with FVL of symptomatic carriers was 0.34%/year. This was similar to the incidence in relatives with FVL of asymptomatic carriers. Kaplan Meier analysis in relatives of symptomatic propositi showed that at the age of 58 years, thrombosisfree survival was reduced to 75% in carriers and 93% in non-carriers (P < 0.05). Carriers of FVL had a three times higher thrombotic risk than non-carriers. In combination with environmental risk factors, FVL clearly adds to the risk of VTE. The thrombotic incidence rate in these unselected relatives with FVL, however, is considerably lower than was seen in carriers of thrombophilic families (1.7%/year). Therefore, special care should be paid to individuals with a positive family history of venous thrombosis while exposed to these risk factors.
Tissue Factor and Homocysteine Levels in Ischemic Heart Disease Are Associated with Angiographically Documented Clinical Recurrences after Coronary Angioplasty
Authors: Prisco Domenico , Brunelli Tamara , Pepe Guglielmina , Gori Anna , Fedi Sandra , Capanni Monia , Simonetti Ignazio , Federici Giorgio , Pastore Anna , Abbate Rosanna , Gensini Gian , Marcucci Rossella 
Published: March 2018

Summary
Background In ischemic heart disease (IHD) patients high plasma levels of Tissue Factor (TF), the trigger of coagulation cascade, are present. Homocysteine (Hcy) is a risk factor for coronary artery disease, and several different pathophysiological mechanisms by which Hcy may play a role in thrombus formation have been postulated in “in vitro” studies. We investigated the “in vivo” role of Hcy in affecting plasma levels of TF, its inhibitor Tissue Factor Pathway Inhibitor (TFPI) and hypercoagulability. Methods and Results We investigated 119 IHD patients who underwent PTCA and compared them with 103 healthy subjects. TF, TFPI, Thrombin-Antithrombin complexes (TAT) and Hcy levels were significantly higher in the patients than in the controls. A positive correlation was found between Hcy and TF (r = 0.54; p < 0.0001), Hcy and TFPI (r = 0.26; p < 0.05) as well as Hcy and TAT (r = 0.33; p < 0.0001) levels. An inverse correlation existed between folate intake and Hcy levels (r = −0.28; p = 0.001). Hcy levels within the first quartile and in the highest quartile were associated with a lower (p < 0.001) and higher (p <0.0001) rate of clinical recurrences, respectively. Patients with TF values in the first quartile had a lower rate of angiographically documented clinical recurrences as compared to those in the fourth quartile (p < 0.01); those in the highest quartile of TF showed a higher rate of recurrences (p = 0.001). Multivariate analysis confirmed these results (first quartile of Hcy: OR 0.02, Cl 0.002–0.27; fourth quartile of Hcy: OR 36.5, Cl 3.6-365/first quartile of TF: OR 0.006, Cl 0.001–0.44; fourth quartile of TF: OR 16.4, Cl 3.0 – 90.0), also after adjustment for risk factors and Hcy and TF respectively. Conclusions In this study we show that TF, TFPI and TAT levels are correlated with Hcy plasma levels in IHD patients, providing evidence of an “in vivo” pathophysiological mechanism of hyperhomocysteinemia. The observed association between angiographically documented clinical recurrences and TF and Hcy values awaits confirmation in studies designated to evaluate this issue on a larger number of patients.
Clinical Evaluation of a Monoclonal Antibody-based Enzyme Immunoassay for Fibrin Degradation Products in Patients with Clinically Suspected Pulmonary Embolism
Authors: de Monyé Wouter , Lijmer Jeroen , Nieuwenhuizen Willem , Büller Harry , Huisman Menno , Brandjes Dees , Gillavry Melvin R. ,   the ANTELOPE-Study Group
Published: March 2018

SummaryWe prospectively evaluated the diagnostic accuracy of the Fibrinostika® FbDP assay in 304 consecutive patients with suspected pulmonary embolism and examined potentially useful cut-off points at which the disease can be excluded. The prevalence of pulmonary embolism was 31%. The assay generated an area under the Receiver Operating Characteristic curve of 0.79 (95% CI 0.73–0.84). A cut-off point of 0.05 µ,g/ml yielded a sensitivity, specificity, negative predictive value and an exclusion efficiency of 100% (95% CI 96–100), 5% (95% CI 2–9), 100% (95% CI 69–100) and 3% (95% CI 2–6), respectively. A clinically useful cut-off point seems to be 0.11 µ,g/ml which corresponded with a sensitivity, specificity, negative predictive value and an exclusion efficiency of 96% (95% CI 90–99), 27% (95% CI 24–28), 93% (95% CI 84–98) and 20% (95% CI 16–25), respectively. We conclude that the assay has potential clinical utility for the exclusion of pulmonary embolism, but it cannot be used as a sole test.
Elastase Mediated Fibrinolysis in Acute Promyelocytic Leukemia
Authors: Oudijk Erik-Jan , Nieuwenhuis H. , Bos Rogier , Fijnheer Rob 
Published: March 2018

SummaryThe bleeding syndrome of acute promyelocytic leukemia (APL) is complex and consists of disseminated intravascular coagulation (DIC) and hyperfibrinolysis. Elastase, derived from malignant promyelocytes, is believed to mediate the fibrinogeno- and fibrinolysis by aspecific proteolysis. In this study we measured the role of elastase in fifteen patients with APL by using an assay for elastase degraded fibrin(ogen) and the results were compared with those obtained in patients with sepsis induced DIC.High levels of elastase were observed in sepsis and APL. The levels of fibrinogen and fibrin degradation products were significantly higher in APL patients compared to patients with sepsis induced DIC. Nevertheless, the level of elastase degraded fibrin(ogen) was higher in the sepsis group (635.3 ng/ml, compared to 144.3 ng/ml in APL; p <0.0001). So, the enormous increase in fibrin and fibrinogen degradation products in APL cannot be explained by elastase activity. This study suggests a minor role for elastase mediated proteolysis in the hemorrhagic diathesis in APL patients.
Fibrinogen, Fibrin and Crosslinking in Aging Arterial Thrombi
Authors: McBane Robert , Ford Monique , Karnicki Krzysztof , Stewart MaryLou , Owen Whyte 
Published: March 2018

SummaryThe assumption that fibrin and crosslinked fibrin impart irreversibility to arterial thrombi is explored with procedure developed for measuring changes in platelet function, morphology and fibrinogen metabolism in aging occlusive thrombi, in which the condition of stasis is imposed uniformly. Arterial thrombi containing autologous 111In labeled platelets were generated in vivo by bilateral mechanical injury of porcine carotid arteries. Vessels containing the platelet-rich thrombi were harvested and incubated intact (37° C) for intervals ranging from 30 min to 12 h. The isolated vessels were then bisected and agitated in culture medium containing tick anticoagulant and hirudin for 60 min. Disaggregated platelets were evaluated for yield (from 111In radioactivity) viability (dense body ATP secretion) and morphology (electron microscopy). Western analysis of fibrin(ogen) in thrombus extracts was performed using anti-fibrinogen Bβ- and γ-chain monoclonal antibodies for thrombi at each time point. A stable recovery of nearly 50% of platelets was observed during 12 h of thrombus aging. As thrombi aged, viability of disaggregated platelets gradually decreased with platelet necrosis the predominant feature beyond 6 h. By western analysis of thrombus extracts, nearly 50% of fibrinogen was cleaved to fibrin and extensively crosslinked within 30 min of injury with no evidence of fibrinolysis. With the exception of a declining proportion of γ-monomer, these features remain relatively constant during 12 h of thrombus maturation. It is concluded that neither fibrin nor crosslinked fibrin are dominant factors imparting cohesion within platelet thrombi. Furthermore, under conditions of complete arterial occlusion imposed by this experimental design, there is no evidence of endogenous fibrinolysis.
Comparison of the O’Brien Filter Test and the PFA-100 Platelet Analyzer in the Laboratory Diagnosis of von Willebrand’s Disease
Authors: Schlammadinger Agota , Kerenyi Adrienne , Muszbek Laszlo , Boda Zoltan 
Published: March 2018

SummaryVon Willebrand’s disease (vWD) is the most common congenital haemorrhagic diathesis, characterized by the quantitative or qualitative disorder of von Willebrand factor (vWF). A number of methods have been used for the diagnosis of the disease, and the bleeding time determination is widely accepted as a screening test in spite of its low sensitivity. Our aim was to evaluate and compare the performance of two high shear systems (the O’Brien filter test and the PFA-100 device) in the screening and diagnosis of vWD. Thirty patients (n=13 type 1 with mild symptoms, n = 9 type 1 with severe symptoms, n = 2 type 2A, n = 3 type 2B and n = 3 type 3 vWD) and twenty controls were investigated. In mild vWD the platelet retention in the second phase of the filter test with citrated blood showed the highest sensitivity (91.6%). The sensitivity of the PFA-100 method with collagen-epinephrine cartridges in this group was 76.9%, while the bleeding time was prolonged only in 15.4% of the cases. In severe type 1, in type 2A and type 3 all functional tests reflected the bleeding tendency of the patients. In type 2B disease the bleeding time was prolonged only when the patient was thrombocytopenic, but both high shear systems revealed the disease independently of the presence of thrombocytopenia. The overall sensitivity of the bleeding time determination was 50% compared to the 80-90% sensitivity of the O’Brien filter test and the PFA-100 system. The sensitivity values of the filter test and the PFA-100 device with collagen-epinephrine cartridges were in the same range, but the collagen-ADP cartridges showed a lower (65.5%) sensitivity, though the results were specific and had high positive predictive value. We conclude that both high shear systems are suitable for the screening of vWD, and that they are superior to the traditional bleeding time determination in case of mild disease or type 2B vWD.
Role of the Leucine-Rich Domain of Platelet GPIbα in Correct Post-translational Processing – The Nancy I Bernard-Soulier Mutation Expressed on CHO Cells
Authors: Ulsemer Philippe , Baas Marie-Jeanne , Schwartz Agnès , Ravanat Catherine , Briquel Marie-Elisabeth , Cranmer Sue , Jackson Shaun , Cazenave Jean-Pierre , de la Salle Corinne , Lanza François 
Published: March 2018

SummaryThe mechanisms governing the biosynthesis and surface expression of platelet adhesive receptors on parent megakaryocytes are as yet poorly understood. In particular, the assembly and processing of the multisubunit glycoprotein (GP) Ib-IX-V complex, a receptor for von Willebrand factor (vWf) is not fully understood. In the present work, these questions were addressed by reproducing a natural mutation of GPIbα found in a variant case of Bernard-Soulier syndrome (Nancy I), due to the deletion of leucine 179 in the seventh leucine-rich repeat of the polypeptide. Wild type and mutated GPIbα were transfected into CHO cells expressing GPIbβ and GPIX. Flow cytometry showed surface expression of the three subunits of both GPIb-IX complexes, but GPIbαΔLeu was present at lower levels (20-40%) and was recognized only by a sub class of monoclonal antibodies which epitopes were not modified by the mutation. These properties reproduce the defect found in the patient’s platelets, demonstrating the causative nature of the mutation and validate the use of the CHO cells model. Biochemical studies were performed in an attempt to elucidate the mechanism of the conformational change of GPIbαΔLeu. They unexpectedly revealed a major glycosylation deficiency of the mutated GPIbα leading to a 40% decrease in molecular weight. The other two subunits of the complex were however normal and present at the plasma membrane. The deletion led to complete functional deficiency with lack of vWf binding of CHOαΔLeu transfected cells in the presence of botrocetin and defective adhesion to a vWf coated surface under static conditions. Finally, in contrast to normal CHOαβIX cells, which displayed rolling and deceleration when perfused over a vWf surface, CHOαΔLeuβIX cells were unable to roll over or attach to a vWf substratum. These results show that the integrity of the leucine-rich region of GPIbα is essential for normal processing and function of the GPIb-IX complex. In addition, these results obtained in a cellular system supported the suspected role of the macroglycopeptide region of GPIbα in maintaining a suitable conformation of this multisubunit receptor to perform its adhesive function.
Homozygous Pro74 → Arg Mutation in the Platelet Glycoprotein Ibβ Gene Associated with Bernard-Soulier Syndrome
Authors: Tomiyama Yoshiaki , Honda Shigenori , Fukunishi Mariko , Hara Junichi , Inoue Chikako , Kamiya Tadashi , Saito Hidehiko , Kunishima Shinji 
Published: March 2018

SummaryBernard-Soulier syndrome (BSS) is an autosomal recessive bleeding disorder due to quantitative or qualitative abnormalities in the glycoprotein (GP) Ib/IX/V complex, the platelet receptor for von Willebrand factor. This complex is composed of four subunits, GPIbα, GPIbβ, GPIX and GPV. We describe here the genetic basis of the disorder in a patient with BSS. Flow cytometric analysis of the patient’s platelets showed greatly reduced GPIbα and GPIX surface expression. Immunoblot analysis disclosed absence of GPIbα, GPIbβ and GPIX in the platelets. DNA sequencing analysis revealed a novel missense mutation in the GPIbβ gene that converts Pro (CCG) to Arg (CGG) at residue 74. Homozygosity of the mutation was confirmed by allele-specific restriction analysis, chromosome 22 microsatellite analysis and quantitative Southern blotting. The mutant GPIbβ was normally transcribed. Transient transfection studies confirmed that mutant GPIbβ impairs surface expression of GPIb/IX, showing that the mutation is responsible for a BSS phenotype observed in the patient.
Persistently Elevated Levels of von Willebrand Factor Antigen in HIV Infection
Authors: Bjørnsen S. , Lunden B. , Otterdal K. , Ng E. C. , Ameln W. , Ueland T. , Müller F. , Solum N. O. , Brosstad F. , Frøland S. S. , Aukrust P. 
Published: March 2018

SummaryLevels of circulating von Willebrand factor (vWf) antigen are thought to reflect endothelial involvement in various disorders. In the present study we found markedly elevated plasma levels of vWf in HIV-infected patients demonstrated on both cross-sectional and longitudinal testing. Notably, we found that a persistent rise in vWf antigen was associated with progression of HIV-related disease. This elevation of vWf antigen represented functionally normal vWf as evaluated by plasma FVIII, ristocetin cofactor assay and vWf multimer analyses. While HIV-infected patients showed enhanced platelet activation, platelets did not contribute substantially to the increased vWf levels. The high vWf levels were significantly correlated with high viral load, and during HAART, the pronounced decline in HIV RNA levels was accompanied by a corresponding decrease in vWf. The persistent elevation of functionally normal vWf during HIV infection, most probably reflecting a persistent endothelial cell activation, may have an important role in the pathogenesis of HIV infection.
Normalisation of Tissue Factor Pathway Inhibitor Activity after Glycaemic Control Optimisation in Type 1 Diabetic Patients
Authors: Rigla Mercedes , Mateo Jose , Fontcuberta Jordi , Souto Juan , de Leiva Alberto , Pérez Antonio 
Published: March 2018

SummaryIncreased plasma concentrations of various markers of endothelial damage have been observed in type 1 diabetic patients, particularly in those with microangiopathy. Objective. To evaluate the effect of nearnormalisation of glycaemic control on different markers of endothelial injury involved in haemostasis in poorly-controlled type 1 diabetic patients. Material and Methods. TFPI, thrombomodulin (TM), plasminogen activator inhibitor, tissue-type plasminogen activator and von Willebrand factor were measured in 14 poorly-controlled type 1 diabetic patients free of diabetes-related complications (8 men, 6 women; mean age 29.8 ± 9.9 years) before (baseline) and after 3 months of intensive therapy and in 14 sex-, age-and BMI-matched control subjects. Results. After a mean follow-up of 107 ± 49 days (56-210), Hb A1c decreased from 11.2 ± 2.3 to 6.7 ± 0.7% (p <0.0001). TFPI activity at baseline was higher than in the control group (126.9 ± 34 vs 92.0 ± 13%, p <0.005) and decreased after good glycaemic control was achieved (p <0.005), becoming similar to that in the control group (91.0 ± 16.5%). The TFPI descent correlated with the variations observed in HbA1c (p <0.05; r = 0.54). TM levels at baseline were significantly higher than in the control group (42.3 ± 9.1 vs 29.00 ± 10.9; p <0.005) and did not change. The remaining parameters studied were similar between patients and controls and did not change after glycaemic optimisation. Conclusions. Optimisation of glycaemic control normalises the increased activity of TFPI but not the higher TM levels observed in poorly-controlled type 1 diabetic patients without chronic complications.
Activity of a Sub-cutaneously Administered Novel Mixed Micellar Formulation of Argatroban in Rat and Rabbit Models of Venous Thrombosis
Authors: Visconte Catherine , Lecoffre Catherine , Lochot Sylvette , Girard Denise , Berry Christopher 
Published: March 2018

SummaryWe studied the antithrombotic activity of a mixed micellar formulation containing 14 mg/ml argatroban administered by the subcutaneous (s. c.) route in rat and rabbit models of venous thrombosis. The effects on bleeding time in the rat tail transection bleeding time test were also studied. In a tissue factor-dependent arterio-venous shunt model, argatroban treatment led to dose-dependent reduction in thrombus weight with an estimated ID50 of 1.8 mg/kg s. c. In the same model, heparin had an estimated ID50 of 179 IU/kg. The antithrombotic activity of argatroban was accompanied by increases in the thrombin and ecarin clotting times but not the aPTT, whereas heparin increased the thrombin time and aPTT but not the ecarin clotting times. Argatroban also inhibited thrombus formation in a rabbit model of thromboplastin + stasis induced thrombosis in the rabbit jugular vein with an estimated ID50 of 1 mg/kg s. c. When tested in the rat tail transection bleeding time test, the mixed micellar formulation of argatroban caused significant increases in the bleeding time as from 8 mg/kg s. c., while heparin significantly increased the bleeding time at 800 U/kg. Mixed micellar argatroban appears to have a superior safety margin to heparin in terms of antithrombotic efficacy and bleeding risk.Thus, a mixed micellar formulation of argatroban, which markedly enhances its solubility, could be useful as a potential antithrombotic agent for subcutaneous administration.