title: Prostaglandin D(2) induces apoptosis of human osteoclasts through ERK1/2 and Akt signaling pathways.
abstract: In a recent study we have shown that prostaglandin D2 (PGD2) induces human osteoclast (OC) apoptosis through the activation of the chemoattractant receptor homologous molecule expressed on T-helper type 2 cell (CRTH2) receptor and the intrinsic apoptotic pathway. However, the molecular mechanisms underlying this response remain elusive. The objective of this study is to investigate the intracellular signaling pathways mediating PGD2-induced OC apoptosis. OCs were generated by in vitro differentiation of human peripheral blood mononuclear cells (PBMCs), and then treated with or without the selective inhibitors of mitogen-activated protein kinase-extracellular signal-regulated kinase (ERK) kinase, (MEK)-1/2, phosphatidylinositol3-kinase (PI3K) and NF-κB/IκB kinase-2 (IKK2) prior to the treatments of PGD2 as well as its agonists and antagonists. Fluorogenic substrate assay and immunoblotting were performed to determine the caspase-3 activity and key proteins involved in Akt, ERK1/2 and NF-κB signaling pathways. Treatments with both PGD2 and a CRTH2 agonist decreased ERK1/2 (Thr202/Tyr204) and Akt (Ser473) phosphorylation, whereas both treatments increased β-arrestin-1 phosphorylation (Ser412) in the presence of naproxen, which was used to eliminate endogenous prostaglandin production. In the absence of naproxen, treatment with a CRTH2 antagonist increased both ERK1/2 and Akt phosphorylations, and reduced the phosphorylation of β-arrestin-1. Treatment of OCs with a selective MEK-1/2 inhibitor increased caspase-3 activity and OC apoptosis induced by both PGD2 and a CRTH2 agonist. Moreover, a CRTH2 antagonist diminished the selective MEK-1/2 inhibitor-induced increase in caspase-3 activity in the presence of endogenous prostaglandins. In addition, treatment of OCs with a selective PI3K inhibitor decreased ERK1/2 (Thr202/Tyr204) phosphorylation caused by PGD2, whereas increased ERK1/2 (Thr202/Tyr204) phosphorylation by a CRTH2 antagonist was attenuated with a PI3K inhibitor treatment. The DP receptor was not implicated in any of the parameters evaluated. Treatment of OCs with PGD2 as well as its receptor agonists and antagonists did not alter the phosphorylation of RelA/p65 (Ser536). Moreover, the caspase-3 activity was not altered in OCs treated with a selective IKK2/NF-κB inhibitor. In conclusion, endogenous or exogenous PGD2 induces CRTH2-dependent apoptosis in human differentiated OCs; β-arrestin-1, ERK1/2, and Akt, but not IKK2/NF-κB are probably implicated in the signaling pathways of this receptor in the model studied.
authors.affiliation: Department of Pharmacology, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, 3001 12e Avenue Nord, Sherbrooke, Quebec J1H 5N4, Canada; Division of Rheumatology, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, 3001 12e Avenue Nord, Sherbrooke, Quebec J1H 5N4, Canada. Electronic address: Li.Yue@USherbrooke.ca.
authors.forename: Li
authors.initials: L
authors.lastname: Yue
authors.affiliation: Division of Rheumatology, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, 3001 12e Avenue Nord, Sherbrooke, Quebec J1H 5N4, Canada. Electronic address: Sonia.Haroun@USherbrooke.ca.
authors.forename: Sonia
authors.initials: S
authors.lastname: Haroun
authors.affiliation: Department of Pharmacology, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, 3001 12e Avenue Nord, Sherbrooke, Quebec J1H 5N4, Canada; Division of Rheumatology, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, 3001 12e Avenue Nord, Sherbrooke, Quebec J1H 5N4, Canada. Electronic address: Jean-Luc.Parent@USherbrooke.ca.
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Parent
authors.affiliation: Department of Pharmacology, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, 3001 12e Avenue Nord, Sherbrooke, Quebec J1H 5N4, Canada; Division of Rheumatology, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, 3001 12e Avenue Nord, Sherbrooke, Quebec J1H 5N4, Canada. Electronic address: Artur.Fernandes@USherbrooke.ca.
authors.forename: Artur J
authors.initials: AJ
authors.lastname: de Brum-Fernandes
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D019390:Arrestins / Q000378:metabolism; D002417:Cattle; D002454:Cell Differentiation / Q000187:drug effects; D006801:Humans; D051550:I-kappa B Kinase / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism; D010010:Osteoclasts / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology*; D010766:Phosphorylation / Q000187:drug effects; D010768:Phosphoserine / Q000378:metabolism; D015230:Prostaglandin D2 / Q000494:pharmacology*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D011971:Receptors, Immunologic / Q000378:metabolism; D011982:Receptors, Prostaglandin / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D000071559:beta-Arrestin 1; D000071557:beta-Arrestins
chemical_list: C000605253:ARRB1 protein, human; D019390:Arrestins; D016328:NF-kappa B; D011971:Receptors, Immunologic; D011982:Receptors, Prostaglandin; D000071559:beta-Arrestin 1; D000071557:beta-Arrestins; D010768:Phosphoserine; D051057:Proto-Oncogene Proteins c-akt; D051550:I-kappa B Kinase; C496560:IKBKB protein, human; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D015230:Prostaglandin D2; C036293:prostaglandin D2 receptor
keywords: Akt; Apoptosis; ERK1/2; Osteoclasts; Prostaglandin D(2); β-Arrestin