Annexin V binding to platelets is agonist, time and temperature dependent.
Platelets bind annexin V when stimulated with combinations of platelet agonists such as collagen and thrombin. Previous studies have demonstrated significant heterogeneity of platelets binding annexin V. The relative role of the thrombin protease-activated receptors (PARs), PAR1 and PAR4, together with different methods of platelet preparation on annexin V binding to platelets is unclear. We therefore investigated the role of PAR1- and PAR4-activating peptides in combination with collagen-related peptide on annexin V binding. In diluted whole blood, PAR1- and PAR4-activating peptides were as effective as thrombin in inducing annexin V binding. However, in washed platelets, PAR-activating peptides were less potent than thrombin at inducing annexin V binding. This difference was more pronounced when experiments were performed at 37 degrees C compared to room temperature. In studies of diluted whole blood, platelet rich plasma and washed platelets, platelets incubated at room temperature bound more annexin V than platelets incubated at 37 degrees C. We also saw a significant effect of time on annexin V binding, in that more annexin V bound to platelets with longer incubation times. In conclusion, PAR1 and PAR4-activating peptides were as effective as thrombin in inducing annexin V binding in combination with collagen-related peptide in diluted whole blood and platelet rich plasma, but not in washed platelets. In addition, incubation temperature and time has a strong influence on annexin V binding to platelets. Thus variations in these conditions may explain the differences observed between previous studies.
Biomedical Diagnostics Institute Programme, Molecular & Cellular Therapeutics, Royal College of Surgeons in Ireland, 123 St Stephens Green, Dublin 2, Ireland.
Sofia
S
Ramstrom
Sarah
S
O'Neill
Eimear
E
Dunne
Dermot
D
Kenny
D000328:Adult; D000818:Animals; D017304:Annexin A5 / Q000378:metabolism*; D001792:Blood Platelets / Q000378:metabolism*; D006801:Humans; D010455:Peptides / Q000378:metabolism; D011485:Protein Binding; D044463:Receptor, PAR-1 / Q000819:agonists* / Q000378:metabolism; D018179:Receptors, Thrombin / Q000819:agonists* / Q000378:metabolism; D013696:Temperature; D013917:Thrombin / Q000378:metabolism; D013997:Time Factors
D017304:Annexin A5; D010455:Peptides; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin; D013917:Thrombin; C112908:protease-activated receptor 4