Sodium fluoride induced activation of phospholipase C in intact human platelets does not depend on ADP, PAF or arachidonate products.
Explore the source record for details and available documents.
Adenosine Diphosphate↗
Biomedical subjects
J Arnout
Publications and source records attributed to J Arnout.
Retreatment with alteplase for early signs of reocclusion after thrombolysis. The European Cooperative Study Group.
Recurrent chest pain with new ST-segment elevation was observed in 26 of 652 patients (4%) with myocardial infarction in a clinical trial of alteplase (recombinant tissue-type plasminogen activator; 100 mg) and aspirin with or without heparin. Clinical and electrocardiographic signs of reocclusion were treated with a second dose of alteplase: 50 mg in 20 patients with signs of reocclusion < or = 24 hours after initial therapy, and 100 mg in 5 patients with signs between 24 and 77 hours, and in 1 patient with early signs of reocclusion. Pain and ST changes disappeared within 100 minutes (median 50). D-dimer determinations in 15 patients were increased, indicating activation of the coagulation system. Signs of reocclusion occurred despite adequate anticoagulation with heparin in 5 of 11 patients in whom coagulation measurements were available. No excess bleeding was observed in patients who received a second dose of alteplase. Retreatment with alteplase is feasible and provides an alternative for angioplasty in patients with clinical and electrocardiographic signs of reocclusion early after thrombolytic therapy.
Adult↗
A higher than expected incidence of factor VIII inhibitors in multitransfused haemophilia A patients treated with an intermediate purity pasteurized factor VIII concentrate.
In May 1990, 218 patients with haemophilia A regularly attending the Leuven Haemophilia Center were randomly assigned to a group receiving either of two newly introduced factor VIII concentrates: factor VIII-P, an intermediate purity pasteurized concentrate, or factor VIII-SD, a high purity concentrate treated with solvent-detergent for viral inactivation. Patients were followed from May 1990 until October 1991. Between August 1991 and October 1991 a clinically important factor VIII inhibitor was detected in five out of the 109 patients receiving factor VIII-P while none of the 109 patients receiving factor VIII-SD developed such antibodies. All patients acquiring an inhibitor had previously been clinically tolerant to transfused factor VIII with 200 to more than 1,000 days of exposure to factor VIII prior to May 1990. Patients with inhibitors were transfused daily with 30 U factor VIII-SD per kg body weight, which was associated with a gradual decline of the inhibitor level. In all patients the antibodies were relatively slow-acting and predominantly directed towards the light chain of factor VIII. This study demonstrates a higher than expected incidence of factor VIII inhibitors associated with the use of a specific factor VIII concentrate in multitransfused haemophilia A patients. It indicates the usefulness of evaluating newly introduced concentrates in prospective, randomized trials.
Adolescent↗
Increased thromboxane formation in diabetic pregnancy as a possible contributor to preeclampsia.
OBJECTIVE: Because pregnant women with diabetes have an increased risk of preeclampsia, we tested the hypothesis that urinary excretion of thromboxane metabolites is increased in diabetic pregnancies without evidence of preeclampsia at the time of testing. STUDY DESIGN: Urinary excretion of thromboxane A2 metabolites (either 2,3-dinor-thromboxane B2 or 11-dehydro-thromboxane B2) was measured in 24 type I pregnant diabetic individuals and in 20 women with normal pregnancies between 28 and 32 weeks' gestation. RESULTS: The amount of 2,3-dinor-thromboxane B2 and 11-dehydro-thromboxane B2 in the urine of pregnant women with diabetes (1727 +/- 415 and 827 +/- 276 pg/mg creatinine) was significantly higher than in women with normal pregnancies (638 +/- 218 and 178 +/- 145 pg/mg creatinine) (p < 0.002 and p < 0.001). CONCLUSION: Our findings support a role for thromboxane in the pathogenesis of preeclampsia.
Diabetes Mellitus, Type 1↗
MK-383 (L-700,462), a selective nonpeptide platelet glycoprotein IIb/IIIa antagonist, is active in man.
BACKGROUND: Fibrinogen-dependent cross-linking of glycoprotein (GP) IIb/IIIa on activated platelets is the final mechanism leading to platelet aggregation. Inhibition of this mechanism may result in a novel antithrombotic agent. We studied the activity of MK-383 (L-700,462), a new, nonpeptide GPIIb/IIIa antagonist, in vitro and in vivo, in man. METHODS AND RESULTS: MK-383, a nonpeptide tyrosine derivative, dose-dependently inhibited fibrinogen-dependent platelet aggregation, in vitro. Binding of 125I-labeled fibrinogen to activated platelets was prevented in a competitive manner with an IC50 of 10 +/- 4.2 nmol/L. The activity and tolerability of MK-383 were evaluated in a two-part double-blind, placebo-controlled, dose-escalation study in healthy male subjects using 1- and 4-hour intravenous infusions. Effects on ADP- and collagen-induced ex vivo platelet aggregation (APA or CPA) and template bleeding time (TBT) were evaluated. Twenty-four subjects participated in the 1-hour part. Six received placebo and 18 MK-383 in doses ranging from 0.05 to 0.40 microgram.kg-1 x min-1. MK-383 inhibited platelet aggregation and prolonged bleeding time in a dose-dependent manner. APA and CPA were totally inhibited at the end of infusion of 0.4 microgram.kg-1 x min-1 and returned to 55% and 89% of baseline, respectively, at 3 hours after infusion. TBT was prolonged at this dose from 5.0 +/- 1.3 minutes predose to 22.7 +/- 6 minutes at the end of the infusion (P < .01) and was normalized by 3 hours after infusion. In the 4-hour infusion part, 15 subjects received MK-383 (0.1 to 0.2 microgram.kg-1 x min-1), and five received placebo. Complete inhibition of ex vivo platelet aggregation was seen at 0.15 and 0.2 microgram.kg-1 x min-1. At 0.2 microgram.kg-1 x min-1, TBT was prolonged from 4.4 +/- 1.2 to 23.9 +/- 4.3 minutes at the end of infusion (P < .01) and remained slightly prolonged 3 hours after infusion (7.2 +/- 1.8 minutes). No adverse effects were observed in any of the 33 subjects receiving MK-383. CONCLUSIONS: The results from this study indicate that MK-383 appears to be well tolerated and active in man. It is the first nonpeptide GPIIb/IIIa antagonist that can be used to investigate the antithrombotic potential of this new class of antiplatelet agents.
Bleeding Time↗
A monoclonal antibody recognizes a von Willebrand factor domain within the amino-terminal portion of the subunit that modulates the function of the glycoprotein IB- and IIB/IIIA-binding domains.
We developed a monoclonal antibody, 1C1E7, against vWf that increases ristocetin-induced platelet aggregation in a dose-dependent manner and lowers the threshold concentration of ristocetin needed to obtain a full aggregatory response. The platelet aggregatory effect of asialo vWf (ASvWf) also is enhanced by 1C1E7, in the presence or absence of glycoprotein (GP) IIb/IIIa receptor antagonism. In the presence of ristocetin, both intact 1C1E7 and its Fab fragments enhance specific binding of 125I-vWf to platelets. With 1C1E7, the intermediate and higher molecular weight multimers of vWf are preferentially bound to both GP Ib and GP IIb/IIIa. Thrombin-induced 125I-vWf binding to GP IIb/IIIa also is increased by 1C1E7. Maximal binding of 1C1E7 to vWf corresponds to 0.97 mol/mol vWf monomer with a Kd of 4.7 x 10(-10) M. 1C1E7 reacts with a 34/36-kD tryptic fragment (III-T4) and a 34-kD plasmic fragment (P34), which localizes the epitope between amino acid residues 1 and 272; this was confirmed by NH2-terminal amino acid sequencing. Finally, platelet aggregation by ASvWf was associated with a sharp rise in intracellular Ca2+ only in the presence of 1C1E7. An antibody-mediated conformational change of vWf may result in an improved presentation of the GP Ib- and GP IIb/IIIa-binding domains of mainly the larger multimers; the increased density of vWf on the platelet surface leads to platelet activation. The antibody may thus recognize a domain of relevance for vWf physiology.
Amino Acid Sequence↗
Factor VIII solutions in pre-filled syringes.
Explore the source record for details and available documents.
Drug Contamination↗
Correlation between level of heparinization and patency of the infarct-related coronary artery after treatment of acute myocardial infarction with alteplase (rt-PA).
BACKGROUND AND OBJECTIVES: The conjunctive use of intravenous heparin may influence the efficacy of alteplase for coronary thrombolysis in patients with acute myocardial infarction. In this study we examined the relation between the level of intravenous anticoagulation with heparin and sustained coronary artery patency in a subgroup of patients of the European Cooperative Study Group (ECSG) trial. METHODS: In the ECSG trial, patients treated with alteplase and aspirin were randomized to concomitant fixed doses of intravenous heparin (a bolus dose of 5,000 U followed by a continuous infusion of 1,000 U/h or placebo). The current study group comprised 149 of 324 ECSG patients allocated to heparin therapy and 132 of 320 ECSG patients allocated to placebo administration who had both an interpretable coronary angiogram obtained within 6 days of treatment and sufficient plasma samples to assess the level of anticoagulation. Activated partial thromboplastin times, fibrinogen and D-dimer levels were determined on plasma samples at baseline and at 45 min and 3, 12, 24 and 36 h after the start of alteplase administration. RESULTS: The coronary artery patency rate was higher in patients allocated to heparin therapy than in those allocated to placebo (80% and 71%, respectively, p = 0.05). Patients allocated to heparin were classified into three subgroups: 48 patients (32%) with all activated partial thromboplastin times at least twice their own baseline value (optimal anticoagulation), 40 patients (27%) with the lowest activated partial thromboplastin time at 3, 12, 24 or 36 h between 130% and 200% of the baseline value (suboptimal anticoagulation) and 61 patients with at least one activated partial thromboplastin time less than 130% of baseline (inadequate anticoagulation). In the heparin group, coronary artery patency correlated with the level of anticoagulation: 90%, 80% and 72%, respectively, in patients with optimal, suboptimal and inadequate anticoagulation (p = 0.02, optimal vs. inadequate anticoagulation). Heparin administration was associated with a smaller reduction in fibrinogen and a smaller increase in D-dimer level during and after alteplase administration. No correlation was found between fibrinogen or D-dimer levels and coronary artery patency. No intracerebral hemorrhage occurred in these patients; however, bleeding was more frequent in the subgroup with optimal anticoagulation (p = 0.05). CONCLUSIONS: Intense anticoagulation with intravenous heparin enhances coronary artery patency after alteplase treatment of acute myocardial infarction.
Adult↗
Quality control in haemostasis.
Laboratory investigation of the haemostatic system deserves particular procedures in the quality control of analytical variables as well as preanalytical variables. This paper reviews the precautions that have to be taken in the blood prelevement, the transport of the tubes and the performance of the laboratory tests aimed to investigate the haemostatic system in order to obtain reliable results.
Blood Coagulation Tests↗
A new lupus anticoagulant neutralization test based on platelet-derived vesicles.
We here present an easily standardizable and reproducible procedure which clearly separates lupus anticoagulants (LA) from coagulation factor inhibitors. This new LA neutralization test makes use of platelet-derived microvesicles which were prepared as follows: gel-filtered platelets (4 x 10(5)/microliters) were incubated with 60 microM of the calcium ionophore A23187 for 20 min at 37 degrees C. The vesicles were separated from the platelet aggregates by centrifugation at 1000 g for 10 min. The vesicle containing supernatant was then spun down at 15,000 g for 15 min, lyophilized and stored at -20 degrees C until used. The vesicles were resuspended in plasma from normal individuals, from patients with LA activity, from patients with factor VIII inhibitors, from patients with congenital factor deficiencies and from patients receiving oral anticoagulants or intravenous heparin. A kaolin clotting time was performed in the absence (KCT) or presence of these vesicles (KCTves) and the ratios of these times to their respective mean normal times were calculated. Segregation of LA patients from all remaining patients except heparinized ones could be made with a high degree of accuracy. A thrombin time was needed to separate LA from heparinized patients. The method was highly reproducible and only minor (negligible) differences in potencies were observed between different vesicle preparations. Both the intra-batch and the inter-batch coefficients of variations on the KCTves were lower than 6%.
Blood Coagulation Factors↗
A patient with von Willebrand's disease characterized by a compound heterozygosity for a substitution of Arg854 by Gln in the putative factor-VIII-binding domain of von Willebrand factor (vWF) on one allele and very low levels of mRNA from the second vWF allele.
We describe a patient with a lifelong bleeding disorder previously classified as von Willebrand's disease (vWD) type I. The factor VIII (FVIII) level in this patient was disproportionately low and we showed that this was due to a decreased factor VIII binding capacity of her vWF. To characterize the molecular defect in this type of vWD, a cDNA-dependent polymerase chain reaction (PCR) amplification was performed using platelet RNA as a template. Direct sequencing of the amplified fragment, which encodes for the FVIII-binding domain, showed a single nucleotide change in exon 20 at codon 854, resulting in the substitution of CAG glutamine (Gln) for CGG arginine (Arg). At the level of the cDNA only the mutated sequence was found, whereas at genomic DNA level the patient was heterozygous for this mutation. This patient is therefore a compound heterozygote for a point mutation resulting in a FVIII-binding defect and a vWF allele with low transcript levels.
Adult↗
Plasma fibrinopeptide A levels in patients with acute myocardial infarction treated with alteplase. Correlation with concomitant heparin, coronary artery patency, and recurrent ischemia. The European Cooperative Study Group.
BACKGROUND: Fibrin generation during and after therapy with alteplase may depend on the level of concomitant anticoagulation. The hypothesis that fibrinopeptide A (FPA) levels, as markers of ongoing in vivo fibrin formation, correlate with the angiographic and clinical outcome of thrombolysis is tested. METHODS AND RESULTS: Serial plasma FPA levels were determined in 334 patients of the randomized European Cooperative Study Group trial comparing heparin versus placebo plus alteplase and aspirin in patients with acute myocardial infarction. Median FPA levels (with the 10th to 90th percentiles) were 21 ng/ml (2-390 ng/ml) before treatment in placebo-allocated patients (n = 166) and increased to 49 (15-580), 34 (4-320), 27 (2-240), 29 (2-430), and 30 (3-390) ng/ml after 0.75, 3, 12, 24, and 36 hours, respectively. In heparin-allocated patients (n = 168), median baseline FPA values were 18 ng/ml (2-210 ng/ml) and decreased to 6 (1-110), 5 (1-75), 5 (1-60), 7 (1-100), and 10 (1-170) ng/ml at corresponding time points (p less than 0.0001 for the difference at each time point). Adequate anticoagulation, defined as no activated partial thromboplastin time value below twice the pretreatment value at 3, 12, 24, and 36 hours after initiation of treatment, was obtained in 48 patients assigned to heparin. It was associated with normal median FPA levels (less than or equal to 4 ng/ml) at all time points compared with 12 (2-80), 16 (2-240), and 15 (2-240) ng/ml at 12, 24, and 36 hours, respectively, in heparin-assigned but inadequately anticoagulated patients (n = 102, p less than 0.001 for each time point). In the heparin-treated group, median FPA values tended to be lower at all time points in patients with patent vessels than in patients with occluded arteries, but the difference was significant only at 24 hours (p = 0.04). FPA levels did not correlate with clinically apparent recurrent ischemia or with left ventricular thrombosis on two-dimensional echocardiography. CONCLUSIONS: During and after thrombolytic therapy with alteplase, the enhanced fibrin generation is suppressed by sustained concomitant anticoagulation with intravenous heparin. Adequate anticoagulation warrants individual titration of the heparin dose. High plasma FPA levels 24 hours after alteplase therapy are specific but insensitive markers of vessel occlusion in anticoagulated patients. They do not correlate with clinical outcome.
Coronary Vessels↗
Influence of hydroxyethyl starch on coagulation in patients during the perioperative period.
The perioperative use of hydroxyethyl starch (HES) has been implicated as a possible cause of intracranial bleeding. The purpose of this study was to compare the influence on blood coagulation of the isovolemic replacement of 1-L blood loss with either 6% HES (molecular weight [MW] average: 450,000) or 5% human albumin during neurosurgery or lower abdominal surgery. Twenty patients scheduled for brain tumor surgery and 20 patients undergoing transabdominal hysterectomy were studied. The activated partial thromboplastin time, prothrombin time, fibrinogen concentration, factor VIII coagulant, von Willebrand factor antigen, platelet count, and the activated clotting time were compared after induction of anesthesia, after administration of 500 and 1000 mL of colloid solution, and 24 and 48 h postoperatively. All measured coagulation variables remained within physiologic range. Changes in coagulation indices were identical in neurosurgical and hysterectomy patients, except for a larger increase in fibrinogen concentration 24 and 48 h after hysterectomy. The acute phase reaction of factor VIII coagulant and von Willebrand factor, which plays a role in postoperative hypercoagulability, was attenuated by the use of HES. We conclude that isovolemic replacement of 1-L blood loss with either 6% HES (MW average: 450,000) or 5% human albumin does not interfere with normal hemostasis during and after neurosurgery or lower abdominal surgery.
Adult↗
Is the antiphospholipid syndrome caused by antibodies directed against physiologically relevant phospholipid-protein complexes?
Explore the source record for details and available documents.
Antibodies↗
In vitro and in vivo evaluation of two factor VIII concentrates virally inactivated by solvent-detergent or by pasteurization.
The present study compares two new factor VIII preparations currently used in the treatment of haemophilia. A Factor VIII was partially purified from plasma obtained from unpaid voluntary Belgian donors in the blood transfusion centers of Lille and Amsterdam and virus-inactivated by exposure to solvent-detergent (FVIII-SD) or by pasteurization (FVIII-P) respectively. The factor VIII content and the purity of both preparations were assessed in vitro, whereas in vivo we studied the recovery and the plasma half-life of both concentrates. The higher purity of FVIII-SD was confirmed. Factor VIII-P preparations contained more factor VIII than mentioned on the label. Both preparations gave good in vivo recoveries and half-lives. Patients who did not have antibodies to hepatitis B, hepatitis C or HIV at the initial screening, remained negative after six months treatment with the new concentrates. No patients developed neutralizing factor VIII antibodies. Furthermore patients appreciated the ease of administration of both preparations. In conclusion both FVIII concentrates are suited for the treatment of haemophilia A patients.
Antibodies, Viral↗
Measurement of von Willebrand factor antigen in plasma and platelets with an enzyme-linked immunosorbent assay based on two murine monoclonal antibodies.
Two murine monoclonal antibodies, raised against von Willebrand factor (vWF), were used to construct an enzyme-linked immunosorbent assay (ELISA), for quantitation of vWF antigen (vWFAg) in human plasma and platelets. This assay had a lower limit of sensitivity of 0.0001 IU/ml in buffer, and thus is one to two orders of magnitude more sensitive than other ELISA assays which have been reported. The intraassay, interassay and interdilution coefficients of variation were 4.1, 10.4 and 9.9%, respectively. In normal plasma (n = 20), the vWFAg level was 0.83 (range: 0.42-1.25) IU/ml. In normal washed platelets (n = 10), 0.35 (0.25-0.49) IU/10(9) platelets was found. In plasma obtained from various patient groups the following vWFAg levels (geometric mean and range) were observed: von Willebrand's disease (n = 19): 0.18 (0.02-0.77) IU/ml; patients with liver cirrhosis (n = 20): 3.73 (1.68-9.20) IU/ml; patients with pregnancy-induced hypertension (n = 20): 4.14 (2.28-7.44) IU/ml and patients with malignant disease (n = 10), 2.54 (1.51-5.60) IU/ml. A linear correlation was found between vWFAg levels measured with a polyclonal antibody based Laurell electroimmunoassay (r = 0.92, n = 58) or with a polyclonal antibody based ELISA (r = 0.94, n = 64). The present assay is based on stable and reproducible reagents and allows the specific measurement of vWFAg in plasma and in platelets. This assay may constitute a useful tool for the further investigation of clinical conditions associated with changes in vWFAg levels. In addition, its high sensitivity may facilitate a more detailed study of platelet vWFAg in normal and in pathological conditions.
Animals↗
Detection of lupus-like anticoagulants by an enzyme-linked immunosorbent assay using a partial thromboplastin as antigen; a comparative study.
Clotting assays allow qualitative rather than quantitative detection of the lupus anticoagulant. We have therefore studied the usefulness of an ELISA using a commercial partial thromboplastin, Throombofax, as antigen; the results obtained on 146 selected patient plasmas were compared to the results of coagulation tests (kaolin clotting time, tissue thromboplastin inhibition test, activated partial thromboplastin time) and of ELISAs using cardiolipin or phosphatidylserine as antigen. While satisfactory agreement was found within the group of coagulation tests or that of ELISAs, only a moderate agreement was obtained between clotting tests and ELISAs, the best being with the partial thromboplastin ELISA using low plasma dilutions. The study further indicates that ELISA techniques cannot entirely replace coagulation tests for the detection of a lupus anticoagulant, even when a partial thromboplastin is used as antigen. On the other hand, coagulation tests are less sensitive than ELISAs for the detection of antiphospholipid antibodies.
Antigens↗
Ridogrel, a combined thromboxane synthase inhibitor and receptor blocker, decreases elevated plasma beta-thromboglobulin levels in patients with documented peripheral arterial disease.
The combination of thromboxane synthase inhibition with thromboxane receptor antagonism has been shown to result in a strong inhibition of platelet aggregation and a prolongation of the bleeding time (Gresele et al., J. Clin Invest 1987; 80: 1435-45). Ridogrel is a single molecule that efficiently achieves both inhibitions in human volunteers. The present study was performed in patients with obstructive peripheral arterial disease and elevated plasma beta-thromboglobulin levels. Patients were treated with either 2 x 300 mg ridogrel or 2 x 300 mg placebo per day for 2 1/2 days, according to a double blind randomised parallel design. Plasma beta-thromboglobulin decreased significantly throughout active treatment starting within 2 h after administration; serum and urinary immunoreactive TxB2 levels and urinary 11-dehydro-TxB2 excretion were significantly lower and serum PGE2 and 6-keto-PGF1 alpha levels significantly higher with ridogrel; no changes were observed in the placebo-treated group. In conclusion this study demonstrates a reduction of platelet activation in vivo by ridogrel.
Aged↗