The role of prostanoid receptors in mediating the effects of PGE₂on human platelet function

Abstract

The effects of prostaglandin E(2) (PGE(2)) on platelet function are believed to be the result of opposing mechanisms that lead to both enhancement and inhibition of platelet function. Enhancement of platelet function is known to be via EP3 receptors linked to G(i) and inhibition of adenylyl cyclase. However, the receptors involved in inhibition of platelet function have not been fully defined. Here we have used measurements of platelet aggregation, calcium signaling and P-selectin expression to assess platelet function induced by platelet activating factor (PAF), thrombin receptor activating peptide (TRAP-6) and the thromboxane A(2) mimetic U46619 respectively, to determine the effects of PGE(2) and of selective prostanoid receptor agonists on platelet function. Their effects on vasodilator-stimulated phosphoprotein (VASP) phosphorylation were also determined. We also assessed the ability of selective prostanoid receptor antagonists to modify the effects of PGE(2). The agonists and antagonists used were iloprost (IP agonist), ONO-DI-004 (EP1 agonist), ONO-AE1-259 (EP2 agonist), sulprostone (EP3 agonist), ONO-AE1-329 (EP4 agonist), CAY10441 (IP antagonist), ONO-8713 (EP1 antagonist), DG-041 (EP3 antagonist) and ONO-AE3-208 (EP4 antagonist). Using the agonists available to us we demonstrated that EP3, EP4 and IP receptors elicit functional responses in platelets. The EP3 receptor agonist promoted platelet aggregation, calcium signaling and P-selectin expression and this was associated with a reduction in VASP phosphorylation. Conversely agonists acting at IP and EP4 receptors inhibited platelet function and this was associated with an increase in VASP phosphorylation. The effects on platelet function and VASP phosphorylation of the selective prostanoid receptor antagonists used in conjunction with PGE(2) were consistent with PGE(2) interacting with EP3 receptors to enhance platelet function and with EP4 receptors (but not IP receptors) to inhibit platelet function. This is the first demonstration of the involvement of EP4 receptors in platelet responses to PGE(2).

Prostaglandin E2Differentially Modulates Human Platelet Function through the Prostanoid EP2 and EP3 Receptors

“…A negative effect of butaprost on PAF-induced platelet aggregation has also been reported (Matthews and Jones, 1993;Iyú et al, 2010). Butaprost did not modify intraplatelet Ca 2ϩ , with or without ADP, whereas it promoted VASP-P, indicating a cAMP-mediated effect, which is consistent with previous data on different cells (Regan et al, 1994).…”

“…The data of 17-phenyl-trinor PGE 2 (EP3 Ͼ EP1 agonist based on K i values; Kiriyama et al, 1997), mimicking 11d-16dm PGE 2 on VASP-P, microaggregates, and P-selectin expression, further support this hypothesis. Our results are consistent with and extend previous data (Heptinstall et al, 2008;Iyú et al, 2010), but we used diverse PGE 2 analogs, calculated the EC 50 values, ranked potency profiles on specific effects (enhancement of the secondary aggregation, stabilization of aggregates), and explored different platelet agonists at various concentrations. Furthermore, this is the first description of 11d-16dm PGE 2 -responsive receptors on platelets.…”

“…Early studies of PGE 2 showed its capacity to potentiate platelet aggregation in response to subthreshold concentrations of agonists (Shio and Ramwell, 1972;Bruno et al, 1974;Vezza et al, 1993). Upon the pharmacological characterization of EPs, EP3 receptor agonists have been shown to potentiate human platelet aggregation in response to low concentrations of various agonists (Matthews and Jones, 1993;Heptinstall et al, 2008;Iyú et al, 2010), and clinical development has begun for an EP3 antagonist (Singh et al, 2010). However, the human EP3 receptor has many splice variants that have different signaling pathways (Kotani et al, 1995;Schmid et al, 1995), and mRNAs for at least four EP3 splicing variants have been isolated from human platelets (Paul et al, 1998).…”

Adenosine Derived From ADP Can Contribute to Inhibition of Platelet Aggregation in the Presence of a P2Y 12 Antagonist

“…The amounts of the phosphorylated form of VASP present in the lysates were measured by flow cytometry using a cytometric bead assay, which has been described previously. 5,31 The assay was developed using Becton Dickinson Cytometric Bead Array products as described below.…”

The Novel Prostaglandin I2Mimetic ONO-1301 Escapes Desensitization in an Antiplatelet Effect Due to Its Inhibitory Action on Thromboxane A2Synthesis in Mice

“…1). Recent studies have shown that platelet function is regulated via several prostanoid receptors expressed in platelets, including EP 2 (Kuriyama et al, 2010;Smith et al, 2010), EP 3 (Fabre et al, 2001;Ma et al, 2001), EP 4 (Iyú et al, 2010;Kuriyama et al, 2010;Philipose et al, 2010;Smith et al, 2010), IP (Murata et al, 1997), and TP (Thomas et al, 1998). Among these receptors, EP 2 and EP 4 , in addition to IP, mediate inhibitory signaling in platelets.…”