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Turkish Journal of Haematology :... Jun 2016The aim of this study was to investigate the effects of imatinib mesylate on platelet aggregation and adenosine triphosphate (ATP) release in chronic myeloid leukemia...
OBJECTIVE
The aim of this study was to investigate the effects of imatinib mesylate on platelet aggregation and adenosine triphosphate (ATP) release in chronic myeloid leukemia patients.
MATERIALS AND METHODS
Platelet aggregation and ATP release induced by 5.0 mM adenosine diphosphate, 0.5 mM arachidonic acid, 1.0 mg/mL ristocetin, and 2 µg/mL collagen were studied by whole blood platelet lumi-aggregometer in 20 newly diagnosed chronic myeloid leukemia patients before and after imatinib mesylate treatment.
RESULTS
At the time of diagnosis, 17/20 patients had abnormal platelet aggregation results; 8 (40%) had hypoactivity, 6 (30%) had hyperactivity, and 3 (15%) had mixed hypo- and hyperactivity. Repeat platelet aggregation studies were performed after a mean of 19 months (min: 5 months-max: 35 months) in all patients who received imatinib mesylate during this period. After therapy, 18/20 (90%) patients had abnormal laboratory results; 12 (60%) had hypoactive platelets, 4 (20%) had mixed hypo- and hyperactive platelets, and 2 (10%) had hyperactive platelets. Three of the 8 patients with initial hypoactivity remained hypoactive, while 2 developed a mixed picture, 2 became hyperactive, and 1 normalized. Of the 6 patients with initial hyperactivity, 4 became hypoactive and 2 developed a mixed pattern. All of the 3 patients with initial hypo- and hyperactivity became hypoactive. Finally, 2 of the 3 patients with initial normal platelets became hypoactive while 1 remained normal. There was a significant decrease in ristocetin-induced platelet aggregation after therapy (p<0.001), while platelet aggregation and secretion induced by other agonists showed no difference after treatment (p>0.05).
CONCLUSION
These findings indicate that a significant proportion of chronic myeloid leukemia patients have different patterns of platelet function abnormalities and imatinib mesylate has no effect on these abnormalities, with a significant impairment in ristocetin-induced platelet aggregation.
Topics: Adenosine Triphosphate; Adult; Aged; Antineoplastic Agents; Biomarkers; Blood Platelets; Female; Humans; Imatinib Mesylate; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Male; Middle Aged; Platelet Aggregation; Platelet Function Tests; Protein Kinase Inhibitors; Treatment Outcome
PubMed: 26377244
DOI: 10.4274/tjh.2014.0213 -
Circulation Aug 2017Platelet cross-linking during arterial thrombosis involves von Willebrand Factor (VWF) multimers. Therefore, proteolysis of VWF appears promising to disaggregate...
BACKGROUND
Platelet cross-linking during arterial thrombosis involves von Willebrand Factor (VWF) multimers. Therefore, proteolysis of VWF appears promising to disaggregate platelet-rich thrombi and restore vessel patency in acute thrombotic disorders such as ischemic stroke, acute coronary syndrome, or acute limb ischemia. -Acetylcysteine (NAC, a clinically approved mucolytic drug) can reduce intrachain disulfide bonds in large polymeric proteins. In the present study, we postulated that NAC might cleave the VWF multimers inside occlusive thrombi, thereby leading to their dissolution and arterial recanalization.
METHODS
Experimental models of thrombotic stroke induced by either intra-arterial thrombin injection or ferric chloride application followed by measurement of cerebral blood flow using a combination of laser Doppler flowmetry and MRI were performed to uncover the effects of NAC on arterial thrombi. To investigate the effect of NAC on larger vessels, we also performed ferric chloride-induced carotid artery thrombosis. In vitro experiments were performed to study the molecular bases of NAC thrombolytic effect, including platelet aggregometry, platelet-rich thrombi lysis assays, thromboelastography (ROTEM), and high-shear VWF string formation using microfluidic devices. We also investigated the putative prohemorrhagic effect of NAC in a mouse model of intracranial hemorrhage induced by in situ collagenase type VII injection.
RESULTS
We demonstrated that intravenous NAC administration promotes lysis of arterial thrombi that are resistant to conventional approaches such as recombinant tissue-type plasminogen activator, direct thrombin inhibitors, and antiplatelet treatments. Through in vitro and in vivo experiments, we provide evidence that the molecular target underlying the thrombolytic effects of NAC is principally the VWF that cross-link platelets in arterial thrombi. Coadministration of NAC and a nonpeptidic GpIIb/IIIa inhibitor further improved its thrombolytic efficacy, essentially by accelerating thrombus dissolution and preventing rethrombosis. Thus, in a new large-vessel thromboembolic stroke model in mice, this cotreatment significantly improved ischemic lesion size and neurological outcome. It is important to note that NAC did not worsen hemorrhagic stroke outcome, suggesting that it exerts thrombolytic effects without significantly impairing normal hemostasis.
CONCLUSIONS
We provide evidence that NAC is an effective and safe alternative to currently available antithrombotic agents to restore vessel patency after arterial occlusion.
Topics: Acetylcysteine; Animals; Blood Platelets; Chlorides; Disease Models, Animal; Ferric Compounds; Fibrinolytic Agents; Infarction, Middle Cerebral Artery; Male; Mice; Platelet Aggregation; Ristocetin; Thromboembolism; Thrombosis; Tissue Plasminogen Activator; von Willebrand Factor
PubMed: 28487393
DOI: 10.1161/CIRCULATIONAHA.117.027290 -
Journal of Thrombosis and Haemostasis :... Jul 2018Essentials Von Willebrand ristocetin cofactor activity (VWF:RCo) is not a completely reliable assay. Three automated VWF activity assays were compared within a von... (Comparative Study)
Comparative Study
UNLABELLED
Essentials Von Willebrand ristocetin cofactor activity (VWF:RCo) is not a completely reliable assay. Three automated VWF activity assays were compared within a von Willebrand disease (VWD) cohort. Raw values for all three assays were virtually the same. An overall problem within type 2A/IIE VWD using VWF:GPIb-binding activity/VWF:Ag was observed.
SUMMARY
Background von Willebrand disease (VWD) is an inherited bleeding disorder caused by quantitative (type 1 and 3) or qualitative (type 2) von Willebrand factor (VWF) defect. VWD diagnosis and classification require numerous laboratory tests. VWF: glycoprotein Ib (GPIb)-binding activity assays are used to distinguish type 1 from type 2 VWD. Objectives Three different automated VWF:GPIb-binding activity assays were compared. Patients and methods BC-VWF:RCo (Siemens Healthcare Diagnostics), HemosIL VWF:RCo (Instrumentation Laboratory) and INNOVANCE VWF:Ac (Siemens Healthcare Diagnostics) were performed in a well typed VWD cohort (n = 142). Results Based on the three most used VWD parameters (FVIII:C, VWF:Ag and VWF:GPIb-binding activity) and using a cut-off of <0.70 for type 2 VWD revealed sensitivity and specificity of, respectively, 92% and 72.4% for VWF:RCo/VWF:Ag, 84% and 89.7% for VWF:GPIbR/VWF:Ag, and 92% and 85.1% for VWF:GPIbM/VWF:Ag, whereas a lowered cut-off of < 0.60 resulted in reduced sensitivity with increased specificity for all assays. Conclusion VWD classification based on FVIII:C, VWF:Ag and VWF:GPIb-binding activity revealed an overall problem with normal VWF:GPIb-binding activity/VWF:Ag within type 2, especially type 2A/IIE. Although all assays were practically identical, BC-VWF:RCo had higher %CV compared with both new assays but comparable lower limit of quantification (LLOQ) ~4 IU dL . No clear improved distinction between type 1 and 2 VWD with new assays was seen.
BC-VWF
RCo and HemosIL are ristocetin dependent, whereas INNOVANCE does not rely upon ristocetin and is not influenced by VWF polymorphisms increasing VWF:GPIb-binding activity levels. INNOVANCE seems to be the best choice as a first-line VWF:GPIb-binding activity assay, providing the best balance between sensitivity and specificity for type 2 VWD.
Topics: Automation, Laboratory; Belgium; Biomarkers; Cross-Sectional Studies; Czech Republic; Equipment Design; Hematologic Tests; Humans; Platelet Glycoprotein GPIb-IX Complex; Predictive Value of Tests; Protein Binding; Reproducibility of Results; von Willebrand Diseases; von Willebrand Factor
PubMed: 29742318
DOI: 10.1111/jth.14145 -
Journal of Clinical Pathology Aug 1979A quantitative study of various aspects of platelet function was carried out in eight patients with typical hairy-cell leukaemia (HCL). In at least two patients platelet...
A quantitative study of various aspects of platelet function was carried out in eight patients with typical hairy-cell leukaemia (HCL). In at least two patients platelet aggregation was convincingly reduced to more than one aggregating agent (ADP, adrenaline, collagen, thrombin, and ristocetin). Granular storage capacity for {(14)C} 5-HT was reduced in five of the six patients tested. The two patients with definitely abnormal aggregation had the greatest reduction in granular storage pool and the longest bleeding times of those tested but, like the other patients, they did not have a clinical haemostatic defect. It was concluded that a granular storage pool defect (SPD) was at least partly responsible for aggregation abnormalities in HCL since the platelet release reaction in response to thrombin appeared to be normal. All our patients ran a chronic course uncomplicated by any of the factors known to predispose to a platelet SPD acquired in the circulation. Although in the one patient tested before and after splenectomy there was some improvement in platelet aggregation after operation, there was no clear general relationship between defective platelet function and either previous splenectomy or platelet count. Since a direct involvement of the megakaryocytic series in the underlying cell proliferation of HCL seems unlikely, it is concluded that the platelet defect can most reasonably be attributed to the production of abnormal platelets as a result of marrow fibrosis and/or infiltration by hairy cells.
Topics: Bleeding Time; Blood Platelets; Cell Survival; Humans; Leukemia, Hairy Cell; Platelet Aggregation; Serotonin
PubMed: 512041
DOI: 10.1136/jcp.32.8.814 -
The Journal of International Medical... Apr 2019To investigate the role of alteplase, a widely-used thrombolytic drug, in platelet function.
OBJECTIVE
To investigate the role of alteplase, a widely-used thrombolytic drug, in platelet function.
METHODS
Human platelets were incubated with different concentrations of alteplase followed by analysis of platelet aggregation in response to adenosine diphosphate (ADP), collagen, ristocetin, arachidonic acid or epinephrine using light transmittance aggregometry. Platelet activation and surface levels of platelet receptors GPIbα, GPVI and αIIbβ3 were analysed using flow cytometry. The effect of alteplase on clot retraction was also examined.
RESULTS
This study demonstrated that alteplase significantly inhibited platelet aggregation in response to ADP, collagen and epinephrine in a dose-dependent manner, but it did not affect ristocetin- or arachidonic acid-induced platelet aggregation. Alteplase did not affect platelet activation as demonstrated by no differences in P-selectin levels and PAC-1 binding being observed in collagen-stimulated platelets after alteplase treatment compared with vehicle. There were no changes in the surface levels of the platelet receptors GPIbα, GPVI and αIIbβ3 in alteplase-treated platelets. Alteplase treatment reduced thrombin-mediated clot retraction.
CONCLUSIONS
Alteplase inhibits platelet aggregation and clot retraction without affecting platelet activation and surface receptor levels.
Topics: Blood Coagulation; Blood Platelets; Humans; Platelet Activation; Platelet Aggregation; Platelet Function Tests; Platelet Glycoprotein GPIIb-IIIa Complex; Platelet Glycoprotein GPIb-IX Complex; Platelet Membrane Glycoproteins; Tissue Plasminogen Activator
PubMed: 30799665
DOI: 10.1177/0300060519829991 -
Blood Jan 1987We have investigated whether megakaryocytes can be aggregated by ristocetin and bovine plasma and whether such aggregation can be used as a step in the purification of...
We have investigated whether megakaryocytes can be aggregated by ristocetin and bovine plasma and whether such aggregation can be used as a step in the purification of megakaryocytes from marrow cell suspensions. Guinea pig marrow cell suspensions were first enriched for megakaryocytes by density equilibrium centrifugation in continuous Percoll density gradients. The megakaryocyte-enriched marrow was stirred in a platelet aggregometer to which ristocetin or bovine plasma was added. Megakaryocytes were aggregated by both ristocetin and bovine plasma with the proportion aggregated being related to the concentration of ristocetin or bovine plasma. Maximal aggregation (greater than 90% of megakaryocytes) was achieved with 2.0 mg/mL ristocetin or 5% bovine plasma and required five minutes. All maturation stages of morphologically recognizable megakaryocytes were aggregated. The megakaryocyte aggregates were separated from the marrow suspension by sedimentation at 1 g and the megakaryocytes disaggregated by dilution with media (ristocetin aggregated) or addition of dextran sulfate (bovine plasma aggregated). Megakaryocyte purity and recovery were higher with bovine plasma than with ristocetin. A mean of 92% of the megakaryocytes in the bovine plasma aggregated cell suspensions were recovered with megakaryocytes constituting an average of 76% of the final cell suspensions. The viability as well as the diameters and DNA content distribution of these megakaryocytes were similar to those of the starting population. We conclude that guinea pig megakaryocytes behave like platelets in that they can be aggregated with ristocetin or bovine plasma and that megakaryocyte aggregation induced by ristocetin or bovine plasma provides a means to enrich these cells based on membrane rather than physical characteristics. This approach yields purified megakaryocyte populations that are representative of those in unfractionated marrow.
Topics: Animals; Cattle; Cell Aggregation; Cell Separation; Cell Survival; DNA; Guinea Pigs; Megakaryocytes; Plasma; Ristocetin; von Willebrand Diseases; von Willebrand Factor
PubMed: 3491638
DOI: No ID Found -
Scientific Reports Jun 2021The interaction of platelet GPIbα with von Willebrand factor (VWF) is essential to initiate platelet adhesion and thrombosis, particularly under high shear stress... (Randomized Controlled Trial)
Randomized Controlled Trial
The interaction of platelet GPIbα with von Willebrand factor (VWF) is essential to initiate platelet adhesion and thrombosis, particularly under high shear stress conditions. However, no drug targeting GPIbα has been developed for clinical practice. Here we characterized anfibatide, a GPIbα antagonist purified from snake (Deinagkistrodon acutus) venom, and evaluated its interaction with GPIbα by surface plasmon resonance and in silico modeling. We demonstrated that anfibatide interferds with both VWF and thrombin binding, inhibited ristocetin/botrocetin- and low-dose thrombin-induced human platelet aggregation, and decreased thrombus volume and stability in blood flowing over collagen. In a single-center, randomized, and open-label phase I clinical trial, anfibatide was administered intravenously to 94 healthy volunteers either as a single dose bolus, or a bolus followed by a constant rate infusion of anfibatide for 24 h. Anfibatide inhibited VWF-mediated platelet aggregation without significantly altering bleeding time or coagulation. The inhibitory effects disappeared within 8 h after drug withdrawal. No thrombocytopenia or anti-anfibatide antibodies were detected, and no serious adverse events or allergic reactions were observed during the studies. Therefore, anfibatide was well-tolerated among healthy subjects. Interestingly, anfibatide exhibited pharmacologic effects in vivo at concentrations thousand-fold lower than in vitro, a phenomenon which deserves further investigation.Trial registration: Clinicaltrials.gov NCT01588132.
Topics: Animals; Blood Coagulation; Blood Platelets; Crotalid Venoms; Crotalinae; Fibrinolytic Agents; Healthy Volunteers; Humans; Lectins, C-Type; Models, Molecular; Platelet Adhesiveness; Platelet Aggregation; Platelet Count; Platelet Glycoprotein GPIb-IX Complex; Protein Binding; Protein Conformation; Ristocetin; Snake Venoms; Structure-Activity Relationship; Thrombin; Thrombosis; von Willebrand Factor
PubMed: 34083615
DOI: 10.1038/s41598-021-91165-8 -
EClinicalMedicine Sep 2021SARS-CoV-2 infection is associated with thrombotic and microvascular complications. The cause of coagulopathy in the disease is incompletely understood.
BACKGROUND
SARS-CoV-2 infection is associated with thrombotic and microvascular complications. The cause of coagulopathy in the disease is incompletely understood.
METHODS
A single-center cross-sectional study including 66 adult COVID-19 patients (40 moderate, 26 severe disease), and 9 controls, performed between 04/2020 and 10/2020. Markers of coagulation, endothelial cell function [angiopoietin-1,-2, P-selectin, von Willebrand Factor Antigen (WF:Ag), von Willebrand Factor Ristocetin Cofactor, ADAMTS13, thrombomodulin, soluble Endothelial cell Protein C Receptor (sEPCR), Tissue Factor Pathway Inhibitor], neutrophil activation (elastase, citrullinated histones) and fibrinolysis (tissue-type plasminogen activator, plasminogen activator inhibitor-1) were evaluated using ELISA. Tissue Factor (TF) was estimated by antithrombin-FVIIa complex (AT/FVIIa) and microparticles-TF (MP-TF). We correlated each marker and determined its association with severity. Expression of pulmonary TF, thrombomodulin and EPCR was determined by immunohistochemistry in 9 autopsies.
FINDINGS
Comorbidities were frequent in both groups, with older age associated with severe disease. All patients were on prophylactic anticoagulants. Three patients (4.5%) developed pulmonary embolism. Mortality was 7.5%. Patients presented with mild alterations in the coagulogram (compensated state). Biomarkers of endothelial cell, neutrophil activation and fibrinolysis were elevated in severe moderate disease; AT/FVIIa and MP-TF levels were higher in severe patients. Logistic regression revealed an association of -dimers, angiopoietin-1, vWF:Ag, thrombomodulin, white blood cells, absolute neutrophil count (ANC) and hemoglobin levels with severity, with ANC and vWF:Ag identified as independent factors. Notably, postmortem specimens demonstrated epithelial expression of TF in the lung of fatal COVID-19 cases with loss of thrombomodulin staining, implying in a shift towards a procoagulant state.
INTERPRETATION
Coagulation dysregulation has multifactorial etiology in SARS-Cov-2 infection. Upregulation of pulmonary TF with loss of thrombomodulin emerge as a potential link to immunothrombosis, and therapeutic targets in the disease.
FUNDING
John Hopkins University School of Medicine.
PubMed: 34377969
DOI: 10.1016/j.eclinm.2021.101069 -
Journal of Clinical Pathology Nov 1978The addition of the antibiotic ristocetin to plasma accelerated the thrombin clotting time (TCT) in 20 out of 22 subjects. Prior incubation of ristocetin with thrombin...
The addition of the antibiotic ristocetin to plasma accelerated the thrombin clotting time (TCT) in 20 out of 22 subjects. Prior incubation of ristocetin with thrombin or plasma did not alter its effect on the TCT. Ristocetin accelerated clotting greatly at low but not at high levels of thrombin. A simple linear correlation between heparin concentrations and the TCT was demonstrated when ristocetin at 2.5 mg per ml was added to plasma containing between 0.05 and 0.5 unit of heparin per ml. There are implications for assay procedures involving heparin and the TCT.
Topics: Blood Coagulation Tests; Blood Platelets; Heparin; Humans; Ristocetin; Thrombin; Time Factors
PubMed: 739058
DOI: 10.1136/jcp.31.11.1106 -
Archives of Medical Science : AMS Aug 2013It is generally assumed that cholesterol reduction by statins is the predominant therapeutic result underlying their beneficial effects in cardiovascular disease....
INTRODUCTION
It is generally assumed that cholesterol reduction by statins is the predominant therapeutic result underlying their beneficial effects in cardiovascular disease. However, the action of statins may be partially independent of their effects on plasma cholesterol levels, as they combine lipid lowering with positive effects on hemorheological conditions and endothelial function. We evaluated the impact of statin treatment on platelet adhesion to fibrinogen (spontaneous and ADP-activated), along with ADP, collagen or ristocetin-induced aggregation in type II hyperlipidemic patients.
MATERIAL AND METHODS
The study group included 70 persons: 50 patients affected by type II hyperlipidemia without concomitant diseases and 20 healthy volunteers. The effects of 8-week statin treatment (atorvastatin 10 mg/day, simvastatin 20 mg/day, or pravastatin 20 mg/day) on platelet activation were evaluated.
RESULTS
Regardless of the type of statin, a significant decrease in ADP-induced platelet aggregation was observed: for atorvastatin 50.6 ±12.8% vs. 41.1 ±15.8% (p < 0.05), for simvastatin 57.2 ±18.0% vs. 44.7 ±22.1% (p = 0.05), and for pravastatin 55.8 ±19.5% vs. 38.8 ±23.3% (p < 0.05). There was no significant effect of statins on collagen or ristocetin-induced platelet aggregation and adhesion.
CONCLUSIONS
Therapy with statins beneficially modifies ADP-induced platelet aggregation in patients with hyperlipidemia and does not affect spontaneous or ADP-induced platelet adhesion to fibrinogen and platelet aggregation induced by collagen or ristocetin.
PubMed: 24049520
DOI: 10.5114/aoms.2013.36905