Abstract 5786: Prostaglandin E 2 and EP3 Activation Potentiate Human Platelet Responsiveness

Abstract

Activated human platelets synthesize prostaglandin (PG) E 2 , although at much lower rates than thromboxane A 2 . PGE 2 binds different receptors (EP1–4). EP3-deleted mice displayed lower platelet response to sub-threshold concentrations of agonists and less susceptibility to experimental thrombosis than wild type mice. However, the role of PGE 2 in human platelet function remains poorly characterized. We investigated the effect of PGE 2 and EP-agonists on human platelet responsiveness to other agonists. Platelets were incubated with increasing concentrations of PGE 2 or different EP agonists, and stimulated by aggregating doses of adenosine diphosphate (ADP), collagen or arachidonic acid (AA). Aggregation was measured by the optical method. EP expression was studied by immunohistochemistry in platelets and bone marrow megakaryocytes. PGE 2 in the microM range (1–200 microM) dose-dependently inhibited 10microM ADP-induced aggregation. However nanoM PGE 2 concentrations (2–200 nanoM), dose-dependently increased the slope (velocity) of the secondary wave of 8 microM ADP-induced platelet aggregation, with an EC 50 of 25±6 nanoM (n=15), without affecting maximal aggregation. At lower (4 microM) ADP concentration, PGE 2 reverted the reversible aggregation, with an EC 50 of 29±10 nanoM (n=6). In in vitro aspirinated platelets, 200 nanoM PGE 2 in part counteracted the effect of aspirin in abolishing the secondary wave of 8 microM ADP-induced aggregation. In addition, PGE 2 dose-dependently abolished shape change caused by 10 microg/ml collagen and 50 microg/ml AA, with an IC 50 of 157±4 nanoM. The EP3 agonist, 11-deoxy-16,16-dimetyl PGE 2 , mimicked the PGE 2 effect on the secondary wave of ADP-induced aggregation, with an EC 50 58±1 nanoM (n=10), while the EP2 agonist butaprost was ineffective. The EP3 agonist at microM concentrations, dose-dependently induced spontaneous platelet aggregation with an EC 50 of 5.5±1 microM. Immunostains revealed EP3 expression in human platelets and megakaryocytes. We conclude that PGE 2 -EP3 interaction potentiates human platelet response to physiological concentrations of common agonists. EP3 might represent a novel pharmacological target for inhibiting platelet responsiveness in atherothrombotic diseases.