Effects of PGH2 and PGD2 on CRTH2 and DP receptors in primary cells and co-expressed in HEK293 cells

Abstract

Prostaglandin (PG) D2 is a PGH2 metabolite deriving from the cyclooxygenase pathway and the major prostanoid released from activated human mast cells. The biological effects of PGD2 are mediated by the G protein-coupled receptors (GPCRs) D-type prostanoid receptor (DP) and the chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2). Eosinophils, important effector cells in allergy, express both receptors. Activation of CRTH2 has been shown to result in pro-inflammatory responses, while the role of DP in allergic inflammation is still unclear. In this study we show that PGH2 selectively stimulates human peripheral blood eosinophils and basophils, but not neutrophils, and this effect is prevented by the CRTH2 receptor antagonist, Cay10471. In chemotaxis assays, eosinophils showed a pronounced migratory response towards PGH2, while eosinophil degranulation was inhibited by PGH2. Moreover, collagen-induced platelet aggregation was inhibited by PGH2 in platelet-rich plasma, which was abrogated in the presence of the DP antagonist, BWA868c. HEK293 cells transfected with either human CRTH2 or DP responded with Ca2+ flux, while untransfected HEK293 cells showed no response. These data indicate that PGH2 causes activation of the PGD2 receptors, CRTH2 and DP, even in the absence of functional PGD synthase. In further experiments, CRTH2 and DP receptors were stably co-expressed in HEK293 cells as a tool to explore receptor signalling and to investigate possible receptor heterodimerization. Data will be shown that demonstrate possible combinatorial effects of CRTH2 and DP to selective and non-selective agonists and antagonists in Ca2+ signalling assays.