Prostaglandin E2 downregulates TNF-alpha-induced production of matrix metalloproteinase-1 in HCS-2/8 chondrocytes by inhibiting Raf-1/MEK/ERK cascade through EP4 prostanoid receptor activation.
Matrix metalloproteinase-1 (MMP-1, collagenase-1) plays a pivotal role in the process of joint destruction in degenerative joint diseases. We have examined the regulation of MMP-1 production in human chondrocytic HCS-2/8 cells stimulated by tumor necrosis factor-alpha (TNF-alpha). In response to TNF-alpha, MMP-1 is induced and actively released from HCS-2/8 cells. The induction of MMP-1 expression correlates with activation of ERK1/2, MEK, and Raf-1, and is potently prevented by U0126, a selective inhibitor of MEK1/2 activation. In contrast, SB203580, a selective p38 mitogen-activated protein kinases (MAPK) inhibitor, had no effects on TNF-alpha-induced MMP-1 release. A serine/threonine kinase, Akt was not activated in TNF-alpha-stimulated HCS-2/8 cells. TNF-alpha stimulated the production of PGE(2) in addition to MMP-1 in HCS-2/8 cells. Exogenously added PGE(2) potently inhibited TNF-alpha-induced both MMP-1 production and activation of ERK1/2. The effects of PGE(2) were mimicked by ONO-AE1-329, a selective EP4 receptor agonist but not by butaprost, a selective EP2 agonist. In contrast, blockade of endogenously produced PGE(2) signaling by ONO-AE3-208, a selective EP4 receptor antagonist, enhanced TNF-alpha-induced MMP-1 production. Furthermore, the suppression of MMP-1 production by exogenously added PGE(2) was reversed by ONO-AE3-208. Activation of EP4 receptor resulted in cAMP-mediated phosphorylation of Raf-1 on Ser259, a negative regulatory site, and blocked activation of Raf-1/MEK/ERK cascade. Taken together, these findings indicate that Raf-1/MEK/ERK signaling pathway plays a crucial role in the production of MMP-1 in HCS-2/8 cells in response to TNF-alpha, and that the produced PGE(2) downregulates the expression of MMP-1 by blockage of TNF-alpha-induced Raf-1 activation through EP4-PGE(2) receptor activation.
Fushimi
Kazunari
K
Department of Orthopaedic Surgery, Gifu University Graduate School of Medicine, 1-1 Yanagido, Gifu, Japan.
Nakashima
Shigeru
S
You
Fukka
F
Takigawa
Masaharu
M
Shimizu
Katsuji
K
D045744:Cell Line, Tumor; D019902:Chondrocytes / Q000187:drug effects* / Q000378:metabolism; D015232:Dinoprostone / Q000502:physiology*; D015536:Down-Regulation / Q000502:physiology*; D006801:Humans; D007124:Immunoenzyme Techniques; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D020781:Matrix Metalloproteinase 1 / Q000096:biosynthesis*; D010766:Phosphorylation; D019908:Proto-Oncogene Proteins c-raf / Q000737:chemistry / Q000378:metabolism; D018078:Receptors, Prostaglandin E / Q000187:drug effects* / Q000378:metabolism; D058309:Receptors, Prostaglandin E, EP4 Subtype; D012694:Serine / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
C545650:PTGER4 protein, human; D018078:Receptors, Prostaglandin E; D058309:Receptors, Prostaglandin E, EP4 Subtype; D014409:Tumor Necrosis Factor-alpha; D012694:Serine; D019908:Proto-Oncogene Proteins c-raf; D020781:Matrix Metalloproteinase 1; D015232:Dinoprostone