bluuebunny/crossref_metadata_2025_split huggingface dataset:
125I-Fibrinogen Binding to Platelets in Myeloproliferative Disease
Authors: Mistry R , Cahil M , Chapman C , Wood J K , Barnett D B 
Published: July 2018

SummaryRecent reports have suggested a variation in the density and affinity of fibrinogen binding sites in platelets from patients with myeloproliferative disorders (MPD) which may reflect platelet functional abnormalities in these subjects. We have investigated the binding of 125I-fibrinogen (125I-Fb) to gel-filtered platelets from a large relatively homogeneous group of patients with MPD compared to normal age matched controls. Twenty-two of the patients investigated had polycythaemia vera and four essential thrombocythaemia.The maximal density and affinity (Kd) of 125I-Fb binding was assessed by saturation analysis in gel-filtered platelets (GFP) stimulated with either 10 εM ADP or 150 mU thrombin. In addition the functional significance of the binding sites was studied by evaluating the response of GFP from the two experimental groups by assessing the effects of increasing concentrations of added fibrinogen on the response to 10 εM ADP using standard light transmission aggregometry.In both groups the density of fibrinogen binding sites expressed in response to thrombin stimulation was significantly higher (approximately 2-3 fold) than that found in response to ADP. However, fewer binding sites were detected in the MPD group as compared with the control group in response to both ADP and thrombin. The Kd for 125I-Fb was similar for both agonists in normal controls and was significantly lower than that found in the MPD subjects.Although the 125I-Fb binding study results indicate a significant reduction in both the number and affinity of fibrinogen binding sites in patients with myeloproliferative disorders, the clinical and functional significance of these findings remain uncertain.
LY53857, a 5HT2 Receptor Antagonist, Delays Occlusion and Inhibits Platelet Aggregation in a Rabbit Model of Carotid Artery Occlusion
Authors: Wilson Harve C , Coffman William , Killam Anne L , Cohen Marlene L 
Published: July 2018

SummaryThe present study was designed to evaluate the effectiveness of the ergoline 5HT2 receptor antagonist, LY53857 in a rabbit model of vascular arterial occlusion. LY53857 (1 and 10 εM) inhibited serotonin amplified platelet aggregation responses to threshold concentrations of ADP in rabbit platelets in vitro. LY53857 (1 εM) not only inhibited the serotonin component of rabbit platelet aggregation, but also inhibited in vitro aggregation induced by ADP (48.7 ± 16.7% inhibition), collagen (76.1 ± 15.9% inhibition) and U46619 (65.2 ± 12.3% inhibition). The effectiveness of this ergoline 5HT2 receptor antagonist in blocking aggregation to ADP, collagen and U46619 may be related to its ability to inhibit a serotonin component of platelet aggregation since rabbit platelets possess high concentrations of serotonin that may be released during aggregation produced by other agents. Based on the effectiveness of LY53857 to inhibit rabbit platelet aggregation, we explored the ability of LY53857 to extend the time to carotid artery occlusion in rabbits following electrical stimulation of the artery. Reproducible carotid artery occlusion was induced in rabbits by moderate stenosis coupled to arterial cross clamping, followed by electrical stimulation. With this procedure, occlusion occurred at 47.0 ± 7 min (n = 30) after initiation of the electrical stimulation. Animals pretreated with LY53857 (50 to 500 εg/kg i.v.) showed a delay in the time to carotid artery occlusion (at 100 εg/kg i.v. occlusion time extended to 164 ± 16 min). Furthermore, ex vivo platelet aggregation from animals treated with LY53857 (300 εg/kg i.v.) resulted in 40.5% inhibition of platelet aggregation in response to the combination of ADP (1 εM) and serotonin (1 εM). These studies document the ability to obtain reproducible arterial occlusion in the rabbit and showed that intravenously administered LY53857 prolonged the time to carotid artery occlusion. Prolongation of carotid artery occlusion time was accompanied by inhibition of serotonin-amplified ADP-induced aggregation in rabbit platelets, an effect observed both in vitro and ex vivo. Thus, the rabbit is a useful model for studying the effectiveness of 5HT2 receptor antagonists in prolonging vascular occlusion induced by insult of the carotid artery.
The Role of GTP-Binding Proteins in Platelet Activation
Authors: Manning David R , Brass Lawrence F 
Published: July 2018

SummaryPlatelet activation begins with the binding of an agonist to the cell surface and culminates in the events of platelet aggregation, secretion and clot formation. Recent studies have identified two large families of GTP-binding proteins in platelets that are thought to participate in the events of platelet activation. The first of these are the G proteins, heterotrimeric proteins which are best known for their ability to mediate the interaction between agonist receptors and intracellular enzymes such as adenylyl cyclase, phospholipase C and phospholipase A2. To date, at least six G proteins have been identified in platelets: Gs, Gz, three variants of Gi and either Gq or G11 (or both). An additional, pertussis toxinresistant G protein, Gq, may also be present. The second group of GTP-binding proteins present in platelets is substantially smaller than the heterotrimeric G proteins, ranging in size from 21 to 28 kDa. At least 15 such low molecular weight GTP-binding proteins have been identified in platelets, many of which are homologous to the products of the ras proto-oncogenes. In cells other than platelets, low molecular weight GTP-binding proteins have been implicated in protein transport, cell activation events and malignant transformation. Their role in platelets is unknown.
Deep Vein Thrombosis and Fibrinolysis
Authors: Levi Marcel , Lensing Anthonie W A , Büller Harry R , Prandoni Paolo , Dooijewaard Gerard , Cuppini Stefano , ten Cate Jan Wouter 
Published: July 2018

SummaryIn the present study 57 consecutive patients with a first episode of venographically proven deep vein thrombosis were investigated to evaluate the release of tissue-type plasminogen activator (t-PA) and of urokinase-type plasminogen activator (u-PA) in response to DDAVP stimulation as well as the resting plasminogen activator inhibitor (PAI) concentration, comparing this to the results obtained in 66 similar patients with a clinical suspicion of thrombosis but with a normal venogram. All assays were performed without knowledge of the patient's status.Four patients in the deep vein thrombosis-group (7%) had an absent u-PA antigen response upon DDAVP infusion, while a normal response was observed in all control subjects. Patients and controls showed similar increases in t-PA antigen level upon DDAVP. High resting PAI antigen levels were encountered in 5 patients in the deep vein thrombosis-group (9%) and in 6 subjects in the control group (9%).The results from this controlled study indicate that a defective release of u-PA may occur in patients with deep vein thrombosis and may have pathogenetic significance. Furthermore it is concluded that elevation of PAI levels cannot be considered as a specific risk factor for venous thrombosis.
Low Molecular Weight Heparin-Catalyzed Inactivation of Factor Xa and Thrombin by Antithrombin III – Effect of Platelet Factor 4
Authors: Schoen Pieter , Lindhout Theo , Franssen Jo , Hemker H Coenraad 
Published: July 2018

SummaryLow molecular weight (LMW) heparin preparations have unknown distributions of ATIII-binding material, so mean molecular weights as such might bear little information on their anti-factor Xa and anti-thrombin activities, and on the neutralization of these activities by platelet factor 4 (PF4). These properties were investigated in pure systems with proteins of human origin. Pseudo-first order rate constants of inactivation of factor Xa and thrombin by antithrombin III were determined as function of heparin concentration, in the presence of 4.0 mM CaCl2. Despite a large variation in the mean molecular weights, the ratios of the anti-factor Xa over the anti-thrombin activities were essentially the same for the 4th International Standard for heparin (0.46), the 1st International Standard for LMW heparin (0.32), CY216 (0.42) and enoxaparin (0.50). The ultra LMW heparin CY222 had only a 2-times higher ratio (0.98). Analysis of CY216 subfractions, obtained by gel filtration, showed that the heparin molecules of the upper region of the molecular weight distribution are responsible for the anti-thrombin, but also to a large extent for the anti-factor Xa activities. The results indicate that depolymerization of unfractionated heparin does not result in an increased anti-factor Xa/anti-thrombin ratio, because in the presence of Ca2+-ions the rate constants of inactivation of factor Xa are lowered as compared to those of native heparin. PF4-dependent neutralization of anti-factor Xa and anti-thrombin activities of fixed concentrations of the LMW heparins was studied by measuring rate constants as function of PF4 concentration. All anti-thrombin and 50% of the anti-factor Xa activities were readily neutralized. Excess PF4 was required to neutralize another 35-50% of the anti-factor Xa activities. At PF4 levels obtained at maximal release of the content of platelet α-granules, all anti-thrombin and most (≥85%) of the anti-factor Xa activities can be neutralized.
Assays for Phospholipid-Dependent Formation of Thrombin and Xa: A Potential Method for Quantifying Lupus Anticoagulant Activity
Authors: Brandt John T 
Published: July 2018

SummaryLupus anticoagulants (LAs) are antibodies which interfere with phospholipid-dependent procoagulant reactions. Their clinical importance is due to their apparent association with an increased risk of thrombo-embolic disease. To date there have been few assays for quantifying the specific activity of these antibodies in vitro and this has hampered attempts to purify and characterize these antibodies. Methods for determining phospholipid-dependent generation of thrombin and factor Xa are described. Isolated IgG fractions from 7 of 9 patients with LAs were found to reproducibly inhibit enzyme generation in these assay systems, permitting quantitative expression of inhibitor activity. Different patterns of inhibitory activity, based on the relative inhibition of thrombin and factor Xa generation, were found, further substantiating the known heterogeneity of these antibodies. These systems may prove helpful in further purification and characterization of LAs.
Detection of the Glanzmann's Thrombasthenia Mutations in Arab and Iraqi-Jewish Patients by Polymerase Chain Reaction and Restriction Analysis of Blood or Urine Samples
Authors: Peretz H , Seligsohn U , Zwang E , Coller B S , Newman P J 
Published: July 2018

SummarySevere Glanzmann's thrombasthenia is relatively frequent in Iraqi-Jews and Arabs residing in Israel. We have recently described the mutations responsible for the disease in Iraqi-Jews – an 11 base pair deletion in exon 12 of the glycoprotein IIIa gene, and in Arabs – a 13 base pair deletion at the AG acceptor splice site of exon 4 on the glycoprotein IIb gene. In this communication we show that the Iraqi-Jewish mutation can be identified directly by polymerase chain reaction and gel electrophoresis. With specially designed oligonucleotide primers encompassing the mutation site, an 80 base pair segment amplified in healthy controls was clearly distinguished from the 69 base pair segment produced in patients. Patients from 11 unrelated Iraqi-Jewish families had the same mutation. The Arab mutation was identified by first amplifying a DNA segment consisting of 312 base pairs in controls and of 299 base pairs in patients, and then digestion by a restriction enzyme Stu-1, which recognizes a site that is absent in the mutant gene. In controls the 312 bp segment was digested into 235 and 77 bp fragments, while in patients there was no change in the size of the amplified 299 bp segment. The mutation was found in patients from 3 out of 5 unrelated Arab families. Both Iraqi-Jewish and Arab mutations were detectable in DNA extracted from blood and urine samples. The described simple methods of identifying the mutations should be useful for detection of the numerous potential carriers among the affected kindreds and for prenatal diagnosis using DNA extracted from chorionic villi samples.
Thrombin Generation and Fibrinolysis in the Thrombotic Thrombocytopenic Purpura and the Hemolytic-Uremic Syndrome
Authors: Monteagudo J , Pereira A , Reverter J C , Pijoan J , Tusell J , Puig L , Ordinas A , Castillo R 
Published: July 2018

SummaryProthrombin fragment 1+2 (F1+2) and thrombin-antithrombin complexes (TAT), as well as other coagulation and fibrinolysis parameters, were studied in a series of 13 patients affected by thrombotic thrombocytopenic purpura (TTP) or hemolytic-uremic syndrome (HUS). Fragment F1+2 was found to be increased in all patients at diagnosis (patients' range, 1.21-19.03 nmol/1; normal limits, 0.28-1.08 nmol/1), and remained also higher than normal after treatment with plasma exchange (patients' range, 1.5-4.01 nmol/1). Even though the analysis of fibrinolysis markers did not show a definite state of hypo or hyperfibrinolysis in the systemic circulation, enhanced circulating D-dimer levels (0.53-12.6 ug/ml, normal levels of 0.03-0.29 εg/ml) indicated that a certain grade of fibrin lysis was present at previously formed thrombi. Plasma PAI-1 activities either on admission (9.2-38.2 U/ml) and after plasma exchange therapy (2.6-38.6 U/ml) showed a behavior irrespective of t-PA: Ag changes, and post-plasmapheresis values remained high only in patients with fatal neurological outcome. Nevertheless, no correlations between clinical and laboratory data could be established useful for the TTP/HUS prognosis. We conclude that increased thrombin generation occurring in damaged areas is appropriately inhibited by antithrombin III in the systemic circulation, avoiding consumption coagulopathy to develop in uncomplicated patients. In addition, fibrinolysis data suggest that elevated PAI-1 may decisively favor the development of microvascular thrombi.
Lipoprotein Associated Coagulation Inhibitor, Factor VII, Antithrombin III, and Monocyte Tissue Factor Following Surgery
Authors: Carson Steven D , Haire William D , Broze George J , Novotny William F , Pirrucello Samuel J , Duggan Michael J 
Published: July 2018

SummaryFifteen patients undergoing major surgical procedures were evaluated for lipoprotein associated coagulation inhibitor (LACI) antigen, factor VII (F VII), antithrombin III (AT III), and peripheral blood monocyte tissue factor (TF) activity immediately before surgery and on following days. A peak in monocyte TF activity occurred between the first and fifth days after surgery in 10 of the patients, while LACI, F VII, and AT III levels dropped in a qualitatively parallel manner in 8 of these patients. LACI, F VII, and AT III levels decreased after surgery in two additional patients even though TF activity also decreased after surgery in these patients. In the remaining 3 patients who developed infections during the study, TF activity rose within 2 days of the diagnosis of infection in addition to the postoperative peak. In two of these patients, LACI levels increased dramatically near the end of the study period without concomitant changes in F VII and AT III. Overall, the presurgical TF levels in disrupted monocytes varied 52-fold and the maximal TF activity varied 24-fold among patients. The TF response following surgery is therefore heterogenous in both temporal occurrence and magnitude of the postsurgical peak. The patients also varied considerably in the presurgical levels of monocyte TF activity. A possible association between the level of presurgical TF activity and the magnitude of the postsurgical peak was noted. Thirteen of the patients had malignant disease, and, as a group, had presurgical levels of LACI (83 ± 26; mean ± SD), F VII (124 ± 22) and AT III (111 ± 14) within the normal ranges established with pooled reference plasma and similar values for three normal donors studied over the same period (AT III 121 ± 6, LACI 95 ± 4, and F VII 112 ± 29). No cause-effect relationship could be established between changes in TF activity and changes in the other factors measured. Patient- or therapy-specific variables may determine postoperative variations in LACI, F VII, AT III, and monocyte TF.
The Factor VIII Bypassing Activity of Prothrombin Complex Concentrates: The Roles of Factor VIla and of Endothelial Cell Tissue Factor
Authors: Teitel Jerome M 
Published: July 2018

SummaryAn experimental model incorporating cultured endothelial cells (EC) was used to study the "factor VIII bypassing" activity of prothrombin complex concentrates (PCC), a property exploited in the treatment of hemophiliacs with alloantibodies to factor VIII. Two PCC preparations were ineffective as stimuli of tissue factor expression by EC. However, incubation with a combination of PCC plus endotoxin (lipopolysaccharide, LPS) or tumor necrosis factor (TNF) induced much greater tissue factor expression than was seen in response to either substance alone. PCC expressed an additional direct procoagulant activity at the EC surface, which could not be attributed to either thrombin or factor Xa, and which was diminished by an anti-tissue factor antibody. Therefore factor VIIa, which was detectable in both PCC preparations, likely provided this additional direct procoagulant activity at the EC surface. We also excluded the possibility that coagulation proteases contained in or generated in the presence of PCC are protected from inactivation by AT III. Therefore, PCC can indirectly bypass factor VIII by enhancing induced endothelial tissue factor expression, and also possess direct procoagulant activity, probably mediated by factor VIIa.
Pharmacokinetic and Hemostatic Properties of the Recombinant Plasminogen Activator BM 06.022 in Healthy Volunteers
Authors: Martin U , von Möllendorff E , Akpan W , Kientsch-Engel R , Kaufmann B , Neugebauer G 
Published: July 2018

SummaryIn a randomized, single-blind, placebo-controlled, cross-over Phase-I study pharmacokinetic and hemostatic properties of BM 06.022 were investigated in seven healthy, male human volunteers. The novel recombinant plasminogen activator BM 06.022 consists of the kringle 2 domain and the protease domain of human t-PA and is unglycosylated due to its expression in Escherichia coli cells. Vehicle or 6 MU (= 10.4 mg) BM 06.022 was administered as a single i.v. bolus injection of 10 ml over 2 min. BM 06.022 was well tolerated. Fibrinogen levels and clotting times remained unchanged at baseline levels after 6 MU BM 06.022; plasminogen and α2-antiplasmin (collected on chloromethylketone) decreased maximally to 83 ± 1% and 64 ± 3%, respectively, of baseline. D-dimers and fibrinogen degradation products increased to 1,006 ± 234 ng/ml and 555 ± 155 ng/ml, respectively, after BM 06.022. Half-life of BM 06.022-activity was 11.2 ± 0.4 min and of antigen was 13.9 ± 0.7 min, followed by a terminal half-life only for antigen of 173 ± 33 min. Plasma clearance of BM 06.022 was 371 ± 13 ml/min for activity and 183 ± 15 ml/min for antigen. Thus, BM 06.022 is not fibrinogenolytic at 6 MU and is a fibrinolytic agent with a longer half-life than t-PA.
Abnormal Plasminogen Maywood I
Authors: Robbins K C , Boreisha I G , Godwin J E 
Published: July 2018

SummaryMaywood I is a dysfunctional plasminogen. It is described in a patient (W. Y) with a reduced plasma functional activity and with a low normal antigen level. Plasminogen was isolated from the patients plasma by affinity chromatography with L-lysine-substituted Sepharose. The protein yield was 86 mg/1, which was 88% of the plasma Plg antigen level; the specific activity was 24.4 IU/mg protein compared to 28.5 IU/mg protein for the native molecule. The protein was the Glu-form determined by SDS-PAGE and by isoelectric focusing. Six major isoelectric forms were found with isoelectric points between pH's 6.40 and 5.45. Titration of the equimolar plasminogen.streptokinase complex with p-nitrophenyl-p-guanidinobenzoate gave 85% active-sites indicating a homogenous population of molecules; therefore, the propositus is a homozygote. Four different plasminogen activators: a) streptokinase, b) urokinase c) the plasmin-derived light (B) chain-streptokinase complex, and d) tissue plasminogen activator (with soluble fibrin/CNBr-fibrinogen fragments) generated little plasmin from the variant plasminogen (4.5 to 45 nM), 5% or less than that generated from normal plasminogen. At 45 nM plasminogen, the molar ratio of plasminogen : activator was 3.0 for streptokinase, 3.9 for urokinase, 7.1 for the light (B) chain-streptokinase complex, and 155 for tissue plasminogen activator. In the equimolar variant plasminogen.streptokinase complex, the active-site was slowly developed, to a maximum of 85% in 40 min; in the normal complex, 100% active-sites were developed in 15 min. The variant plasminogen forms two equimolar complexes with streptokinase (I and II), with different mobilities in PAGE, in about equal amounts. About 10% of complex I (abnormal), with a slower mobility than complex II (normal), converts to plasmin.streptokinase in 60 min, whereas about 40% of complex II converts to plasmin.streptokinase in 60 min. A kinetic defect was also found with this variant plasminogen. It showed substantially lowered kinetics of activation second-order rate constants with both streptokinase and urokinase, primarily due to lower catalytic rate constants, and with streptokinase, a higher Michaelis constant. Crossed-immunoelectrophoresis, and preparative isoelectric focusing isoelectric points did not show a charge mutation. Maywood I may be classified as a dysplasminogenemia with active center and kinetic defects, Type 1c.
Effects of Hirudin and Heparin on the Binding of New Fibrin to the Thrombus in t-PA Treated Rabbits
Authors: Agnelli Giancarlo , Pascucci Claudia , Cosmi Benilde , Nenci Giuseppe G 
Published: July 2018

SummaryThe aim of this study was to compare the ability of heparin and recombinant hirudin (r-hirudin) in preventing accretion of new fibrin on thrombi during and after treatment with tissue-type plasminogen activator (t-PA) and in enhancing t-PA induced fibrinolysis in a rabbit jugular vein thrombosis model. Heparin and r-hirudin were infused at doses capable of doubling aPTT. In the fibrin accretion inhibition experiments t-PA was infused over 3 h at a dose of 0.2 mg/kg along with saline or heparin, 0.75 mg/ kg or r-hirudin, 1.25 mg/kg. In rabbits treated with t-PA plus saline, heparin or r-hirudin, an accumulation of 125I-fibrinogen on the thrombi of 52.5 ±5.1 εg, 49.5 ± 5.6 εg and 23.5 ± 3.5 εg was observed, respectively, the difference between r-hirudin and both saline and heparin being statistically significant (p <0.01). The inhibition of fibrin accretion on the thrombi induced by r-hirudin persists for at least 9 h after the end of the infusion. By that time r-hirudin has been cleared from the circulation and aPTT has returned to the baseline level for at least 8 h. t-PA, 0.2, 0.4, and 1 mg/kg, infused with saline produced 34 ± 6%, 52 ± 5% and 79 ± 8% lysis of pre-formed thrombi, respectively. The same doses of t-PA infused with heparin, 0.75 mg/kg, produced 32 ± 3%, 54 ± 5% and 78 ± 6% fibrinolysis, respectively and infused with r-hirudin, 1.25 mg/kg, 38 ± 3%, 57 ± 5% and 82 ± 8%, respectively. Thus, no differences in fibrinolysis were observed among the groups of rabbits treated with heparin, r-hirudin and saline receiving the same dose of t-PA. When thrombolysis was assessed by thrombus weight treatment with t-PA and r-hirudin was more effective than treatment with t-PA and saline or heparin. The positive effects of r-hirudin on t-PA induced thrombolysis we observed deserve to be confirmed in clinical settings.