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Keywords:

  • platelet apoptosis and activation;
  • mitochondrial membrane depolarization;
  • P-selectin exposure;
  • ABT-737 and thrombin;
  • Ca2+-ionophore A23187

Summary

Anucleate platelets perform two fundamental processes, activation and apoptosis. We elaborated an approach for selective and concurrent stimulation of platelet apoptosis and/or activation, processes important in haemostasis and platelet clearance. Human platelets were treated with BH3 mimetic ABT-737, thrombin, calcium ionophore A23187 and matched diluents. Apoptosis was determined as mitochondrial inner membrane potential (ΔΨm) depolarization and activation as P-selectin exposure. At optimal treatment conditions (90–180 min, 37°C), ABT-737 predominantly induced apoptosis, when 77–81% platelets undergo only ΔΨm depolarization. The ABT-737 impact on ΔΨm depolarization is strongly time- and temperature-dependent, and much higher at 37°C than at room temperature. In contrast, when platelets were treated with thrombin for 15–90 min at either temperature, activation-only was predominantly (79–85%) induced, whereas A23187 triggers both apoptosis and activation (73–81%) when platelets were treated for 15–60 min at 37°C or 15–90 min at room temperature. These data demonstrate that, depending on the triggering stimulus, platelets predominantly undergo ΔΨm depolarization-only, P-selectin exposure-only, or both responses, indicating that platelet apoptosis and activation are different phenomena driven by different mechanisms. The described model provides a basis for studying differential pharmacological manipulation of platelet apoptosis and activation and their role in haemostasis, thrombosis and platelet clearance.