SC-19220, antagonist of prostaglandin E2 receptor EP1, inhibits osteoclastogenesis by RANKL.
We examined the direct effect of SC-19220, an EP1 prostaglandin (PG) E2 receptor antagonist, on osteoclastogenesis induced by RANK/RANKL signaling in mouse cell cultures. We found that SC-19220 inhibited RANKL-induced osteoclastogenesis by suppression of the RANK/RANKL signaling pathway in osteoclast precursors.


BACKGROUND
Bone growth is accomplished by a dynamic equilibrium between formation by osteoblasts and resorption by osteoclasts, which are regulated by many systemic and local osteotropic factors that induce osteoclast formation from hematopoietic precursors through RANK/RANKL signaling. There are four subtypes of prostaglandin E (PGE) receptors, EP1, EP2, EP3, and EP4, and PGE2 facilitates bone resorption by a mechanism mediated by EP2/EP4. It is well known that SC-19220 is an EP1-specific antagonist. We previously found that SC-19220 inhibited osteoclastogenesis induced by osteotropic factors, including PGE2; however, the inhibitory mechanism is not clear. In this study, we investigated the inhibitory effects of SC-19220 on osteoclastogenesis induced by RANK/RANKL signaling in mouse cell cultures and analyzed the mechanism involved.


METHODS
A bone marrow culture system and bone marrow macrophages were used to examine the effects of SC-19220 on PGE2-, 11-deoxy-PGE1-, and RANKL-induced osteoclastogenesis. We analyzed RANKL expression in osteoblasts induced by PGE2 using RT-PCR. We also examined the effects of SC-19220 on the macrophage-colony-stimulating factor (M-CSF) receptor (c-Fms) and RANK expression in osteoclast precursors as well as RANK/RANKL signaling using RT-PCR and Western blotting analyses.


CONCLUSIONS
SC-19220 dose-dependently inhibited osteoclast formation induced by PGE2, 11-deoxy-PGE1, and RANKL in the mouse culture system; however, it had no influence on RANKL expression in osteoblasts induced by PGE2. Furthermore, the expression of RANK and c-Fms in osteoclast precursors was decreased by SC-19220 at the mRNA and protein levels. In RANK signaling networks, SC-19220 inhibited c-Src and NFAT2 expression. Our findings indicated that SC-19220 inhibits RANKL-induced osteoclastogenesis through the suppression of RANK, c-Fms, c-Src, and NFAT2, suggesting that this EP1-specific antagonist inhibits osteoclast formation induced by RANKL from the early stage of osteoclastogenesis.
Tsujisawa
Toshiyuki
T
Department of Oral Microbiology, Kyushu Dental College, Kitakyushu, Japan.
Inoue
Hiromasa
H
Nishihara
Tatsuji
T
D000527:Alprostadil / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D001854:Bone Marrow Cells / Q000187:drug effects; D000081247:CSK Tyrosine-Protein Kinase; D002352:Carrier Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000502:physiology; D004268:DNA-Binding Proteins / Q000378:metabolism; D012531:Dibenz(b,f)(1,4)oxazepine-10(11H)-carboxylic acid, 8-chloro-, 2-acetylhydrazide / Q000494:pharmacology*; D015232:Dinoprostone / Q000819:agonists / Q000037:antagonists & inhibitors / Q000494:pharmacology; D015536:Down-Regulation; D005260:Female; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000502:physiology; D051379:Mice; D008815:Mice, Inbred Strains; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins / Q000378:metabolism; D010010:Osteoclasts / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D011448:Prostaglandin Antagonists / Q000494:pharmacology*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D053245:RANK Ligand; D053246:Receptor Activator of Nuclear Factor-kappa B; D018078:Receptors, Prostaglandin E / Q000819:agonists / Q000037:antagonists & inhibitors*; D058306:Receptors, Prostaglandin E, EP1 Subtype; D014157:Transcription Factors / Q000378:metabolism; D019061:src-Family Kinases
D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D008562:Membrane Glycoproteins; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D011448:Prostaglandin Antagonists; C545636:Ptger1 protein, mouse; D053245:RANK Ligand; D053246:Receptor Activator of Nuclear Factor-kappa B; D018078:Receptors, Prostaglandin E; D058306:Receptors, Prostaglandin E, EP1 Subtype; C505687:Tnfrsf11a protein, mouse; C505689:Tnfsf11 protein, mouse; D014157:Transcription Factors; D012531:Dibenz(b,f)(1,4)oxazepine-10(11H)-carboxylic acid, 8-chloro-, 2-acetylhydrazide; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; D019061:src-Family Kinases; D000527:Alprostadil; D015232:Dinoprostone; C034008:11-deoxyprostaglandin E1