Flow Cytometry Analysis of Intracellular VASP Phosphorylation for the Assessment of Activating and Inhibitory Signal Transduction Pathways in Human Platelets
- 1 January 1999
- journal article
- letter
- Published by Georg Thieme Verlag KG in Thrombosis and Haemostasis
- Vol. 82 (09) , 1145-1152
- https://doi.org/10.1055/s-0037-1614344
Abstract
Increased platelet adhesion or aggregation are key events in the pathogenesis of cardiovascular diseases. Exact determination of the platelet activation state is essential to recognize, prevent, and treat cardiovascular complications due to platelet dysfunction. Initial phases of platelet activation and inhibition are characterized by phosphorylation of specific intracellular proteins. However, methodological problems often prevent analysis of platelet protein phosphorylation under clinical conditions. A novel flow cytometry-based method using a phosphorylation-specific antibody was developed for fast and easy quantification of the phosphorylation state of a specific intracellular platelet protein. This method was used to analyze various platelet receptors and their intracellular signaling which may be impaired in genetic or acquired disorders or altered due to therapeutic interventions. In a first clinical application, the inhibitory effects of ticlopidine and clopidogrel on the platelet P2YAC ADP receptor were monitored. Abbreviations: ADP: adenosine 5’-diphosphate; cAMP: cyclic adenosine-3’,5’-monophosphate; cGMP: cyclic guanosine-3’,5’-monophosphate; HUVECs: human umbilical vein endothelial cells; MAPK: mitogen-activated protein kinase; PG-E1: prostaglandin E1; PRP: platelet-rich plasma; SNP: sodium nitroprusside; VASP: vasodilator-stimulated phosphoproteinKeywords
Funding Information
- BMBF
This publication has 20 references indexed in Scilit:
- Analysis and Regulation of Vasodilator-stimulated Phosphoprotein Serine 239 Phosphorylation in Vitro and in Intact Cells Using a Phosphospecific Monoclonal AntibodyJournal of Biological Chemistry, 1998
- Chapter 13 Integrin Signaling and the Platelet CytoskeletonPublished by Elsevier ,1996
- Clopidogrel: An Antithrombotic Drug Acting on the ADP-dependent Activation Pathway of Human PlateletsClinical and Applied Thrombosis/hemostasis, 1996
- Protein Kinase C Regulates Pleckstrin by Phosphorylation of Sites Adjacent to the N-terminal Pleckstrin Homology DomainPublished by Elsevier ,1995
- Regulation of protein tyrosine kinases in plateletsTrends in Biochemical Sciences, 1994
- Phosphorylation of Focal Adhesion Vasodilator‐Stimulated Phosphoprotein at Ser157 in Intact Human Platelets Correlates with Fibrinogen Receptor InhibitionEuropean Journal of Biochemistry, 1994
- p42 mitogen-activated protein kinase and p90 ribosomal S6 kinase are selectively phosphorylated and activated during thrombin-induced platelet activation and aggregation.Molecular and Cellular Biology, 1994
- Human red blood cells inhibit endothelium-derived relaxing factor (EDRF) activityEuropean Journal of Pharmacology, 1989
- Thrombin treatment induces rapid changes in tyrosine phosphorylation in platelets.Proceedings of the National Academy of Sciences, 1989
- Adenosine Diphosphate in Red Cells as a Factor in the Adhesiveness of Human Blood PlateletsNature, 1961