PB0858 - Effect of Agonist Stimulation and Antiplatelet Drugs on Platelet Contractility

Background: Within the developing thrombus, the dense packing of platelets and fibrin is believed to hinder the diffusion of soluble agonists, ultimately directing each platelets’ activation response and giving rise to the core-shell thrombus architecture. Defects in platelet contractility result in decreased overall thrombus stability and increased bleeding. How soluble agonists and anti-platelet drugs might direct the development of platelet traction forces is incompletely understood.

Aims: To evaluate the potency of platelet agonists to induce contractility in adherent platelets and the modulation of platelet contractility by antiplatelet agents.

Methods: Blood was obtained from healthy consenting volunteers as approved by the RCSI Research Ethics Committee (REC1405). Washed platelets were seeded onto fibrinogen-coated micropost arrays or glass coverslips, in the presence of various agonists or antiplatelet agents. Traction force microscopy and fluorescence image analysis were used to assess the mechanical (total force per cell, force per area) and morphological (spreading area, f-actin organisation) characteristics of single platelets.

Results: Agonist treatment resulted in contrasting fractions of contractile versus non-contractile cells, as well as in a variable coupling between platelet spreading and contractility. PAR agonists thrombin and PAR-4 activating peptide yielded the highest total forces per cell, followed by TRAP-6, CRP-XL and then ADP. Thrombin-activated platelets showed the least spreading and the highest force per area. Traction forces and spreading area correlated closely for CRP-XL and ADP but were largely decoupled for all PAR agonists. Morphological analysis of agonist-treated platelets revealed modified f-actin arrangements which potentially could aid the identification of specific contractile signatures. Antiplatelet agents blocking GPVI, thromboxane A2 and P2Yx signalling pathways influenced platelet contractility to varying degrees

Conclusion(s): Platelet contractility is controlled by integrating different extracellular signals, including the identity of soluble agonists and antiplatelet agents, as present at different locations in the developing thrombus.