bluuebunny/crossref_metadata_2025_split huggingface dataset:
Safety Profile of Porcine Factor VIII and Its Use as Hospital and Home-Therapy for Patients with Haemophilia-A and Inhibitors: the Results of An International Survey
Authors: Hay C R M , Lozier J N , Lee C A , Laffan M , Tradati F , Santagostino E , Ciavarella N , Schiavoni M , Fukui H , Yoshioka A , Teitel J , Mannucci P M , Kasper C K 
Published: July 2018

SummaryA multicentre retrospective survey was conducted to re-assess the use of porcine factor VIII (HYATE:C), its side effects and the selection of patients for regular or home-therapy. 15,152,000 units of HYATE:C were used by 154 patients. The median inhibitor cross-reactivity to porcine VIIIC of 137 patients was 15%, 27% of patients lacking cross-reactivity. An absent, intermediate or brisk specific antiporcine anamnestic response was observed in 29, 40 and 31% of patients respectively. Seven patients were treated on-demand as home-therapy for a median 6.2, range 1.5-13 years. 23 further patients were treated regularly in hospital for a median of 3, range 2-7 years. This group used 8,319,000 U of porcine VIIIC for 2,000 bleeding episodes.The incidence of transfusion reactions was 0.001%, 0.64% and 2.3%, for domiciliary infusions, infusions in multiply treated inpatients, and unselected in-patient infusions, respectively. The risk of reactions was dose-related. A post-infusion fall in platelet count was common, but usually transient and clinically insignificant. This was also dose-related (r = -0.64, p = 0.002). Marked reductions in platelet count were occasionally seen, usually with intensive replacement therapy. The relative lack of side effects observed amongst patients treated at home is attributable to the low, median 33 U/kg, dose used by this group.A subgroup of inhibitor patients, identifiable by their absent or modest anamnestic response to porcine factor VIII may be treated regularly and safely with this product in small doses, over a period of years.
β2-Glycoprotein I Modulates the Anticoagulant Activity of Activated Protein C on the Phospholipid Surface
Authors: Mori Tatsuyuki , Takeya Hiroyuki , Nishioka Junji , Gabazza Esteban C , Suzuki Koji 
Published: July 2018

SummaryThe objective of this study was to determine whether (β2-glycoprotein I (β2GPI) has procoagulant activity by inhibiting the anticoagulant activity of activated protein C (APC). β2GPI inhibited significantly the APC-catalyzed inactivation of factor Va in an assay using factor V-deficient plasma and physiological levels of protein S and factor Va. This inhibitory effect was diminished by the addition of increasing concentrations of phospholipids, suggesting that β2GPI competitively inhibits the binding of APC to the phospholipid surface. β2GPI inhibited weakly factor Va- and phospholipid-dependent prothrombinase activity at concentrations similar to those to inhibit APC activity. The depletion of β2GPI from plasma led to only a slight shortening of the diluted Russell’s viper venom-dependent clotting time, but to a strong and significant potentiation of the anticoagulant activity of APC. These results suggest that under certain physiological conditions β2GPI has procoagulant property by inhibiting the phospholipid-dependent APC anticoagulant activity.
Quantification of Circulating Activated Protein C in Human Plasma by Immunoassays - Enzyme Levels are Proportional to Total Protein C Levels
Authors: España Francisco , Zuazu Isabel , Vicente Vicente , Estellés Amparo , Marco Pascual , Aznar Justo 
Published: July 2018

SummaryWe have developed a simple assay that measures the circulating activated protein C (APC) in plasma. The assay requires collection of duplicate blood samples, one in citrate plus heparin and the other in citrate plus inhibitors of the enzyme. In the heparin tube, APC reacts completely and irreversibly with its major plasma inhibitors, protein C inhibitor (PCI) and α1-antitrypsin (α1AT), and the complexes formed are measured by ELISAs. The amount of circulating APC is calculated from the difference between the total amount of complexed APC (sample in citrate plus heparin) and the amount of APC complexed in vivo (sample in citrate plus inhibitor). Over 95% of the APC added to blood collected with heparin was recovered in the assay. The assay can easily be performed in four hours, and had a detection limit of 0.1 ng/ml APC. The mean APC level in 18 protein C heterozygous members from seven kindreds was significantly lower (0.6 ± 0.3 ng/ml) than in 20 healthy controls (1.1 ± 0.3 ng/ml) (p <0.001), whereas the mean level in 10 non-affected members from the kindreds studied was 1.5 ± 0.3 ng/ml. In the group of 12 nonanticoagulated heterozygous protein C-deficient individuals, the three patients with a history of venous thrombosis had a mean APC level significantly lower than the nine asymptomatic members (p <0.01), both subgroups showing similar protein C levels. There was a significant correlation in all groups between the levels of APC and the levels of protein C antigen (r = 0.758, p <0.0001) and activity (r = 0.745, p <0.0001), which means that APC circulating levels are proportional to protein C levels and suggests that the protein C level is the limiting factor in the rate of protein C activation in vivo.
A Two-Allele Polymorphism in Protein C Inhibitor with Varying Frequencies in Different Ethnic Populations
Authors: Radtke Klaus-P , Greengard Judith S , Fernández José A , Villoutreix Bruno O , Griffin John H 
Published: July 2018

SummarycDNAs for protein C inhibitor (PCI), prepared from human liver RNA, contained two forms of PCI, designated PCI*A and PCPB
1
. While PCI*A is identical to the published PCI sequence, PCPB differs in 4 of 1221 bp and two amino acids, A36V and K86E. Frequencies for the PCI*B allele, determined from genomic DNA, differed among ethnic groups. Frequency distribution and historical migration of modem man suggest that PCI*A arose from the PCI*B allele. Antigen levels in plasma homozygous for PCI*A or PCI*B equalled that of pooled normal plasma. K86E in PCI*B causes a charge alteration in helix D which is likely involved in heparin binding in antithrombin III but not likely involved in glycosaminoglycan binding in PCI. Kinetic studies showed that plasmas homozygous for PCI*A and PCPB are similar in their APC inhibiting properties and in their heparin sensitivity, consistent with the idea that helix D in PCI is not involved in heparin binding
Evaluation of a Soluble Fibrin Assay in Patients with Suspected Pulmonary Embolism
Authors: Ginsberg Jeffrey S , Siragusa Sergio , Douketis James , Johnston Marilyn , Moffat Karen , Donovan Dianne , McGinnis Joanne , Brill-Edwards Patrick , Panju Akbar , Patel Ameen , Weitz Jeffrey I 
Published: July 2018

SummaryIn order to determine the clinical utility of an enzyme immunoassay (EIA) for soluble fibrin in patients with suspected pulmonary embolism (PE), 195 unselected patients with suspected PE underwent blood sampling for measurement of plasma levels of soluble fibrin, and objective testing for PE. A soluble fibrin result of ≤ 0.75 Μg/ml showed a sensitivity of 100% for PE and a specificity of 12.8%, whereas a soluble fibrin result of ≤ 1.35 Μg/ml showed a sensitivity of 90.3% and a specificity of 49.4% for PE. The soluble fibrin assay has potential clinical utility in excluding PE.
Accumulation of Unphosphorylated Calponin in the Submembranous Cytoskeletons of Arachidonic Acid-stimulated Human Platelets
Authors: Meyer Thomas , Unterberg Christina , Kreuzer Heinrich , Buchwald Arnd B 
Published: July 2018

SummaryCalponin, a basic smooth-muscle protein capable of binding to F-actin, tropomyosin and calmodulin in vitro, was tested for its expression and subcellular localization in resting and stimulated human platelets. Using immunoblotting techniques calponin was revealed as a single protein band with a molecular weight of 34 kDa. Although calponin has been shown to be proteolytically degraded by calpain, in the presence of the calpain inhibitor E-64 and EGTA a significant hydrolysis of calponin could not be detected. Upon stimulation with 10 μM arachidonic acid calponin became increasingly incorporated into Triton X-100 insoluble cytoskeletal fractions reaching a plateau after 15 s. The accumulation of calponin in the cytoskeletons of stimulated platelets paralleled the polymerization of actin into newly formed microfilaments. Immunofluorescence microscopy revealed a sub-membranous co-localization of calponin and actin in aggregated platelets. Since isolated calponin is phosphorylated by protein kinase C and Ca2+/calmodulin-dependent protein kinase II thereby losing its inhibitory effect on the actomyosin MgATPase activity, we examined whether changes in cell shape due to platelet stimulation are accompanied by a phosphorylation of calponin. By performing immunoblotting analysis on either resting or stimulated platelets phosphorylation of calponin on tyrosine, serine or threonine residues could not be demonstrated. In line, [32P]radiolabeling experiments were unable to detect phosphate incorporation into calponin. These observations support the hypothesis that calponin plays a physiological role in regulating contraction and secretion of human platelets even in the absence of its phosphorylation.
High Density Lipoproteins Enhance the Na+/H+ Antiport in Human Platelets
Authors: Nofer Jerzy-Roch , Tepel Martin , Kehrel Beate , Walter Michael , Seedorf Udo , Assmann Gerd , Zidek Walter 
Published: July 2018

SummaryIn the present study, we investigated the effect of high density lipoproteins 3 (HDL3) on Na+/H+ exchanger activity and cytosolic pH (pHi) in human platelets. HDL3 alone failed to affect pHi? but preincubation with HDL3 significantly enhanced the Na+/H+ antiport activation brought about by acidification with 100 mM sodium propionate or stimulation with 0.05 U/ml thrombin. The stimulatory effect of HDL3 was unaffected by indomethacin excluding a role for cyclooxygenase products. The HDL3 effect was not mediated by Ca2+/calmodulin-dependent protein kinase as HDL3 failed to increase cytosolic free calcium concentration. However, the potentiating effect of HDL3 was completely blocked in the presence of the protein kinase C inhibitor, bisindoylmaleimide and the phosphatidylcholine-specific phospholi-pase C inhibitor, D609. Furthermore, the effect of HDL3 was abolished after covalent modification of HDL3 with dimethylsuberimidate and was not observed in platelets from Glanzmann thrombasthenia type 1 which do not express GP IIb/IIIa, as well as in platelets preincubated with anti-GP Ilb/IIIa polyclonal antibodies. We conclude that HDL3 enhances the sodium propionate- and thrombin-induced Na+/H+ antiport activity in human platelets via binding to GP Ilb/IIIa and activation of protein kinase C and phosphatidylcholine-specific phospholipase C.
Naturally Occurring IgG Autoantibodies to Platelet Cytoskeleton Tropomyosin
Authors: Hou Ming , Stockelberg Dick , Kutti Jack , Wadenvik Hans 
Published: July 2018

SummaryWe have observed that naturally occurring serum antibodies generated a 30 Kd band in a platelet immunoblot assay. The target protein had the same molecular weight (30 Kd) under nonreduced and reduced electrophoretic conditions, and could be immunoblotted from either autologous or homologous platelet lysates. Also, the 30 Kd reactive autoantibodies could be totally adsorbed by platelet cytoskeletons. From these data one likely candidate for the autoantibody target was the intracellular platelet protein tropomyosin. Indeed, a commercially available monoclonal antitropomyosin antibody reacted with proteins comigrating with this 30 Kd band; affinity purified human platelet tropomyosin was bound by the antibodies that recognized the 30 Kd protein. This body of evidence conclusively demonstrated that naturally occurring serum autoantibodies reacted with the platelet cytoskeleton protein - tropomyosin. These tropomyosin specific antibodies were found in roughly the same percentage of sera from patients with chronic idiopathic thrombocytopenic purpura (ITP) as from normal individuals.
Protein-Tyrosine Phosphorylation and p72
syk
Activation in Human Platelets Stimulated with Collagen Is Dependent upon Glycoprotein Ia/IIa and Actin Polymerization
Authors: Asazuma Naoki , Yatomi Yutaka , Ozaki Yukio , Qi Ruomei , Kuroda Kenji , Satoh Kaneo , Kume Shoji 
Published: July 2018

SummaryIn human platelets treated with acetylsalicylic acid, collagen induced protein-tyrosine-phosphorylation of several proteins. The major 75 kDa band included cortactin and autophosphorylated p72
syk
. p72
syk
activity rapidly increased upon collagen stimulation, whereas p60c-src
activation was below detectable levels. A combination of inhibitors to remove the effects of extracellular and intracellular Ca2+, released ADP, and fibrinogen binding to GPIIb/IIIa delayed and attenuated the major 75 kDa band. By contrast, p72
syk
activation was not inhibited by these treatments. Cytochalasin D completely inhibited protein tyrosine phosphorylation and p72
syk
activation. It also potently inhibited aggregation and [Ca2+]i elevation. Anti-GPMIa/IIa MoAb in a concentration-dependent manner partially attenuated protein tyrosine phosphorylation and p72
syk
activation. Its inhibitory effects on intracellular Ca2+ mobilization, release of intracellular granule contents, and aggregation also were partial. No tyrosine kinase activity was coprecipitated with GPIa/IIa. These results suggest that p72
syk
activation lies upstream of protein tyrosine phosphorylation, Ca2+ mobilization, ADP release, thromboxane A2 production and aggregation. GPIa/IIa plays a key role in p72
syk
activation induced by collagen, but other collagen receptors may work in synergy to fully activate p72
syk
. Actin polymerization is a prerequisite for both p72
syk
activation and other intracellular signal transduction pathways.
Porcine von Willebrand Factor Binding to Human Platelet GPIb Induces Transmembrane Calcium Influx
Authors: Mazzucato Mario , De Marco Luigi , Pradella Paola , Masotti Adriana , Pareti Francesco I 
Published: July 2018

SummaryPorcine von Willebrand factor (P-vWF) binds to human platelet glycoprotein (GP) lb and, upon stirring (1500 rpm/min) at 37° C, induces, in a dose-dependent manner, a transmembrane flux of Ca2+ ions and platelet aggregation with an increase in their intracellular concentration. The inhibition of P-vWF binding to GP lb, obtained with anti GP lb monoclonal antibody (LJ-Ib1), inhibits the increase of intracellular Ca2+ concentration ([Ca2+]i) and platelet aggregation. This effect is not observed with LJ-Ib10, an anti GP lb monoclonal antibody which does not inhibit the vWF binding to GP lb. An anti GP Ilb-IIIa monoclonal antibody (LJ-CP8) shown to inhibit the binding of both vWF and fibrinogen to the GP IIb-IIIa complex, had only a slight effect on the [Ca2+]i rise elicited by the addition of P-vWF. No inhibition was also observed with a different anti GP IIb-IIIa monoclonal antibody (LJ-P5), shown to block the binding of vWF and not that of fibrinogen to the GP IIb-IIIa complex. PGE1, apyrase and indomethacin show a minimal effect on [Ca2+]i rise, while EGTA completely blocks it. The GP lb occupancy by recombinant vWF fragment rvWF445-733 completely inhibits the increase of [Ca2+]i and large aggregates formation. Our results suggest that, in analogy to what is seen with human vWF under high shear stress, the binding of P-vWF to platelet GP lb, at low shear stress and through the formation of aggregates of an appropriate size, induces a transmembrane flux of Ca2+, independently from platelet cyclooxy-genase metabolism, perhaps through a receptor dependent calcium channel. The increase in [Ca2+]i may act as an intracellular message and cause the activation of the GP IIb-IIIa complex.
Long Term Persistence of Biological Activity following Administration of Enoxaparin Sodium (Clexane) Is due to Sequestration of Antithrombin-binding Low Molecular Weight Fragments - Comparison with Unfractionated Heparin
Authors: Brieger David , Dawes Joan 
Published: July 2018

SummaryWe have previously reported (Brieger D, Dawes J. Thromb Haemost 1994; 72: 275-80) that the prolonged anti-Xa amidolytic activity following intravenous administration of the low molecular weight heparin Enoxaparin sodium is mediated by small molecules derived from the injected drug, and an antithrombin binding penta/hexasaccharide can be detected in the circulation as late as 1 week after administration. To investigate the mechanism underlying this persistence we administered 125I-labelled fractions of Enoxaparin sodium and unfractionated 125I-heparin to rabbits. Both 125I-heparin and the radiolabeled high molecular weight (>6000 Da) Enoxaparin sodium were more effectively cleared from the circulation than the smaller components of LMW heparin. However, our data suggest that the circulating biologically active penta/hexasaccharide was not an unmodified component of the injected drug but was derived from a subpopulation of molecules of intermediate molecular weight (1800-6000 Da) which was retained in the tissues. Significant quantities of both Enoxaparin sodium and unfractionated heparin were retained in the internal organs. We propose that the sequestered subpopulations of Enoxaparin sodium and unfractionated heparin follow different catabolic routes. After administration of both unfractionated and LMW heparin additional antithrombin binding material was released into the circulation by a bolus dose of heparin. This material was not contained on circulating blood cells and was probably sequestered on the endothelium.
The Effects of Dermatan Sulfate at Submicrogram/ml Concentrations on In Vitro Thrombin Generation
Authors: Delorme Michael  A , Tam Andrew S  K , Xu Ling 
Published: July 2018

SummaryDermatan sulfate is an antithrombotic glycosaminoglycan which has been shown to be effective in preventing deep venous thrombosis in general surgery patients when present at concentrations less than 1 |ig/ml. It has also been found to circulate physiologically in similar concentrations in pregnant women at term and in cord blood. We investigated the ability of dermatan sulfate added to plasma at 0.2, 0.5 and 1.0 μg/ml to inhibit thrombin generation initiated by low concentrations of recombinant human tissue factor in defibrinated plasma. A dose dependent decrease in thrombin potential was demonstrated at therapeutically relevant concentrations of dermatan sulfate (0.5 and 1.0 μg/ml) but there was no induction of a lag phase in thrombin generation. We were unable to demonstrate a significant effect on thrombin potential of dermatan sulfate at a concentration similar to that found in pregnancy plasma (0.2 μg/ml). This indicates that either the dermatan sulfate concentration found in pregnancy plasma is not physiologically relevant or that our experimental system (which lacks platelets and fibrin) does not accurately reflect physiologic conditions. The effect on the thrombin potential was somewhat greater at the lowest concentration of tissue factor and amounted to a maximum inhibition of approximately 50% at 1 μg/ml dermatan sulfate. A dose dependent increase in formation of thrombin-heparin cofactor II complexes and a decrease in thrombin-antithrombin complex formation with increasing dermatan sulfate concentration were observed at all dermatan sulfate concentrations. Prothrombin consumption was not changed by any dose of dermatan sulfate. We conclude that dermatan sulfate, at the concentrations tested, catalyses inhibition of free thrombin by heparin cofactor II but not efficiently enough to inhibit prothrombinase formation.
Production of Tissue-Type Plasminogen Activator (t-PA) and Type-1 Plasminogen Activator Inhibitor (PAI-1) in Mildly Cirrhotic Rat Liver
Authors: Seki Taiichiro , Imai Hideo , Uno Shigeyuki , Ariga Toyohiko , Gelehrter Thomas D 
Published: July 2018

SummaryWe have studied the production of tissue-type plasminogen activator (t-PA) and type-1 plasminogen activator inhibitor (PAI-1) in liver of normal rats and in rats with mild cirrhosis induced by carbon tetrachloride inhalation, to demonstrate the production of these fibrinolytic components and their pathophysiologic role in the liver in vivo. Immunohistochemical study of paraffin-embedded liver sections and fibrin autography of frozen sections showed that the normal rat liver produces very little t-PA or PAI-1. On the contrary, striking t-PA activity and both t-PA and PAI-1 antigens were observed in the cirrhotic liver. Both t-PA and PAI-1 in plasma were also markedly increased in the cirrhotic rats. Because the hepatocyte can internalize t-PA or PA/PAI-1 complexes from circulation, Northern blot analysis of the total liver RNA was performed to demonstrate the endogenous synthesis of t-PA and PAI-1 in the liver. Although the normal liver hardly expresses either t-PA or PAI-1 mRNA, striking t-PA and PAI-1 mRNA expression was observed in the liver of rats with mild cirrhosis.These data demonstrate that t-PA and PAI-1 production is strongly upregulated in the liver in rats with mild cirrhosis. These fibrinolytic components, whose production is closely associated with liver failure, may play important roles in the regulation of hepatocyte proliferation and liver regeneration in vivo.
Aprotinin Has no Effect on Platelet Activation and Adhesion during Cardiopulmonary Bypass
Authors: Wahba Alexander , Black Gregor , Koksch Mario , Rothe Gregor , Preuner Jürgen , Schmitz Gred , Bimbaum Dietrich E 
Published: July 2018

SummaryAprotinin reduces blood loss following cardiopulmonary bypass operations (CPB) by the prevention of hyperfibrinolysis. Its influence on circulating platelets is uncertain. In this prospective trial we investigated activation, adhesion, and aggregation receptors on the platelet surface in 20 patients who underwent elective coronary artery bypass grafting. These patients were randomly assigned to receive either a high dose of aprotinin or placebo. Flow cytometry was performed to determine platelet activation [P-selectin, glycoprotein (GP) 53], adhesive (GP Ib), and aggregatory (GP IIb-IIIa) receptors on circulating platelets, before, during, and after CPB. Aprotinin had neither a significant effect on platelet activation nor on adhesive and aggregatory receptors. Plasma levels of D-dimers were measured before and after CPB to assess fibrinolytic activity. D-dimers following CPB and chest tube drainage were significantly less in the aprotinin group. We conclude that aprotinin reduces blood loss by its effect on fibrinolysis but has no direct influence on platelet function.
High Purity Factor IX and Prothrombin Complex Concentrate (PCC): Pharmacokinetics and Evidence that Factor IXa Is the Thrombogenic Trigger in PCC
Authors: Philippou H , Adami A , Lane D A , MacGregor I R , Tuddenam E G D , Lowe G D O , Rumley A , Ludlam C A 
Published: July 2018

SummaryRecent studies using assays for surrogate markers of thrombogenic-ity in man have demonstrated that activation of the coagulation system occurs following infusion of clinical doses of prothrombin complex concentrates (PCC) but not after the same doses of high-purity factor IX concentrates (HP-FIX) in patients with haemophilia B. Here we have investigated the mechanism of such thrombogenesis by applying assays that detect early-through to late-events in coagulation system activation in a pharmacokinetic cross-over study of 50 IU/kg PCC and a new HP-FIX product in haemophilia B patients. Satisfactory recoveries and half-lives were observed for both concentrates.HP-FIX caused no increases in thrombin-antithrombin III complex (TAT), prothrombin activation peptide fragment F1+2 (F1+2), factor X activation peptide (FXAP) or factor Vila (FVIIa). In contrast the same dose of factor IX in the form of PCC was followed by significant increases over pre-infusion levels of TAT, F1+2 and FXAP, but not FVIIa. Elevations of FIXAP occurred after both HP-FIX and PCC but did not reach normal levels and were attributed to normalisation of the FIX concentration in those patients whose levels of FIXAP were initially low. We conclude that the thrombogenic trigger associated with PCC infusion occurs at the level of factor X activation. In the absence of any increase in FVIIa, we would attribute this to the likely presence of FIXa in the PCC.