Prostaglandin E2 induces interleukin-6 expression in human chondrocytes via cAMP/protein kinase A- and phosphatidylinositol 3-kinase-dependent NF-kappaB activation.
Elevated levels of prostaglandin (PG)E(2) and interleukin (IL)-6 have been reported in the cartilage and synovial fluid from patients with arthritic disorders. PGE(2) regulates IL-6 production in numerous different cells including macrophages and synovial fibroblasts. Although PGE(2) stimulates IL-6 expression in human chondrocytes, the underlying signaling pathway of this process has yet to be delineated. Here, we investigate the mechanism of IL-6 induction in human T/C-28a2 chondrocytes treated with exogenously added PGE(2). PGE(2) induces IL-6 mRNA and protein expression via a cAMP-dependent pathway, reaching maximal levels after 60 min of stimulation before declining to baseline levels at 6 h. Forskolin, an adenylyl cyclase activator, also stimulates IL-6 expression in human chondrocytes in a dose- and time-dependent fashion. Inhibition of downstream effectors of cAMP activity such as protein kinase A (PKA) or phosphatidylinositol 3 kinase (PI3K) blocks PGE(2)- and forskolin-induced IL-6 upregulation. Simultaneous inhibition of PKA and PI3K reduces IL-6 expression in stimulated chondrocytes well below the basal levels of untreated cells. Gel shift, supershift, and chromatin immunoprecipitation assays reveal the activation and binding of the nuclear factor (NF)-kappaB p65 subunit to the IL-6 promoter, which is markedly suppressed by selective PI3K or PKA pharmacological inhibitors. p65 knockdown completely abrogates IL-6 mRNA synthesis in PGE(2)- and forskolin-primed chondrocytes. Cumulatively, our data show that PGE(2) and forskolin induce IL-6 expression in human chondrocytes via cAMP/PKA and PI3K-dependent pathways, which in turn regulate the activation and binding of p65 to the IL-6 promoter.
Wang
Pu
P
Department of Chemical and Biomolecular Engineering, The Johns Hopkins University, Baltimore, MD 21218, USA.
Zhu
Fei
F
Konstantopoulos
Konstantinos
K
D000262:Adenylyl Cyclases / Q000378:metabolism; D000730:Androstadienes / Q000494:pharmacology; D001665:Binding Sites; D002478:Cells, Cultured; D019902:Chondrocytes / Q000187:drug effects / Q000201:enzymology* / Q000276:immunology; D002867:Chromones / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism*; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015232:Dinoprostone / Q000378:metabolism*; D004789:Enzyme Activation; D020536:Enzyme Activators / Q000494:pharmacology; D006801:Humans; D015850:Interleukin-6 / Q000096:biosynthesis* / Q000235:genetics; D009025:Morpholines / Q000494:pharmacology; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011401:Promoter Regions, Genetic; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D034622:RNA Interference; D012333:RNA, Messenger / Q000096:biosynthesis; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors; D051996:Transcription Factor RelA / Q000378:metabolism; D015533:Transcriptional Activation; D015854:Up-Regulation; D000077191:Wortmannin
D000262:Adenylyl Cyclases; D000730:Androstadienes; D002867:Chromones; D005576:Colforsin; D000242:Cyclic AMP; D017362:Cyclic AMP Response Element-Binding Protein; D017868:Cyclic AMP-Dependent Protein Kinases; D015232:Dinoprostone; D020536:Enzyme Activators; D015850:Interleukin-6; D009025:Morpholines; D016328:NF-kappa B; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger; D051996:Transcription Factor RelA; D000077191:Wortmannin; C115163:CREB1 protein, human; C508600:IL6 protein, human; C498143:RELA protein, human; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
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