The role of PGE2 in human atherosclerotic plaque on platelet EP3 and EP4 receptor activation and platelet function in whole blood

Abstract

Atherosclerosis has an important inflammatory component. Macrophages accumulating in atherosclerotic arteries produce prostaglandin E(2) (PGE(2)), a main inflammatory mediator. Platelets express inhibitory receptors (EP(2), EP(4)) and a stimulatory receptor (EP(3)) for this prostanoid. Recently, it has been reported in ApoE(-/-) mice that PGE(2) accumulating in inflammatory atherosclerotic lesions might contribute to atherothrombosis after plaque rupture by activating platelet EP(3), and EP(3) blockade has been proposed to be a promising new approach in anti-thrombotic therapy. The aim of our investigation was to study the role of PGE(2) in human atherosclerotic plaques on human platelet function and thrombus formation. Plaque PGE(2) might either activate or inhibit platelets depending on stimulation of either EP(3) or EP(4), respectively. We found that the two EP(3)-antagonists AE5-599 (300 nM) and AE3-240 (300 nM) specifically and completely inhibited the synergistic effect of the EP(3)-agonist sulprostone on U46619-induced platelet aggregation in blood. However, these two EP(3)-antagonists neither inhibited atherosclerotic plaque-induced platelet aggregation, GPIIb/IIIa exposure, dense and alpha granule secretion in blood nor reduced plaque-induced platelet thrombus formation under arterial flow. The EP(4)-antagonist AE3-208 (1-3 μM) potentiated in combination with PGE(2) (1 μM) ADP-induced aggregation, demonstrating that PGE(2) enhances platelet aggregation when the inhibitory EP(4)-receptor is inactivated. However, plaque-induced platelet aggregation was not augmented after platelet pre-treatment with AE3-208, indicating that plaque PGE(2) does not stimulate the EP(4)-receptor. We found that PGE(2) was present in plaques only at very low levels (15 pg PGE(2)/mg plaque). We conclude that PGE(2) in human atherosclerotic lesions does not modulate (i.e. stimulate or inhibit) atherothrombosis in blood after plaque rupture.

The EP3 Agonist Sulprostone Enhances Platelet Adhesion But Not Thrombus Formation Under Flow Conditions

“…Therefore, the authors conclude that PGE 2 , released from damaged vessels and atherothrombotic plaques, promotes thrombus formation via EP3 activation. However, whether human plaques produce biologically relevant amounts of PGE 2 and, therefore, atherothrombotic thrombus formation is increased via EP3 receptor activation has remained unclear: while Tilly et al [22] found bioactive PGE 2 , Schober et al [26] could not show an effect of plaque homogenates on platelet Pselectin expression, GPIIb/IIIa activation or thrombus formation. While in earlier studies a moderate increase in bleeding time was seen in EP3 knockout mice [16,17] , more recent studies in mice as well as in humans demonstrated that the EP3 antagonist prevented EP3-augmented platelet aggregation ex vivo without affecting bleeding time in vivo [21][22][23] .…”

PGE2reverses Gs-mediated inhibition of platelet aggregation by interaction with EP3 receptors, but adds to non-Gs-mediated inhibition of platelet aggregation by interaction with EP4 receptors

“…Previous research has identified that the effects of PGE 2 on platelet function are mediated mainly through interaction with the EP3 receptor, which promotes platelet function via a G i -linked mechanism, and through interaction with the EP4 receptor, which inhibits platelet function via a G s -linked mechanism [8][9][10][11][12][13][14]. In this study, we have examined the possibility that the action of PGE 2 at EP3 and EP4 receptors can explain the previous observation that this prostaglandin is able to reverse the inhibition of aggregation by agents that stimulate cAMP production via G s , but adds to inhibition provided by agents that raise cAMP through other mechanisms, including the inhibition of cAMP PDE and direct stimulation of adenylate cyclase [15].…”

“…Interaction of each agent with its specific receptor leads to activation of adenylate cyclase, via G s , resulting in an increase in cAMP and subsequent inhibition of platelet function. Another naturally occurring prostaglandin, prostaglandin E 2 (PGE 2 ), is able to raise cAMP in a similar manner by interaction with the G s -linked EP4 receptor [9][10][11][12]. However, PGE 2 is unusual in that it also interacts with a second receptor, the G i -linked EP3, to inhibit adenylate cyclase [9,13,14].…”

Understanding the role of prostaglandin E2 in regulating human platelet activity in health and disease

“…PGE 2 has been found to be the predominant prostanoid from endothelial cells in human microvessels [33] and rabbit coronary microvessels [34], and it likely plays a significant role in angiogenesis [35] and vascular tone [36, 37]. PGE 2 is also present in atherosclerotic plaque [38, 39]. Resident macrophages in atherosclerotic plaque constitutively express COX-1 and make small amounts of PGE 2 at baseline.…”

“…Stimulation of EP3 via the EP3-specific agonist sulprostone does not directly lead to platelet aggregation, but potentiates aggregation in response to agonists such as ADP and U46,619 in human [38, 56, 62, 72] and murine [73] platelet rich plasma. Additionally, PGE 2 -mediated stimulation of EP3 decreased cAMP, consistent with its primary signaling in platelets being coupled to G i [73].…”