Synergistic IL-10 induction by LPS and the ceramide-1-phosphate analog PCERA-1 is mediated by the cAMP and p38 MAP kinase pathways.
Expression of the anti-inflammatory cytokine IL-10 can be induced either by TLR agonists such as lipopolysaccharide (LPS), or by various endogenous stimuli, in particular those acting via a cAMP-dependent signaling pathway. We have previously reported that the synthetic phospho-ceramide analogue-1 (PCERA-1) increases cAMP level and subsequently down-regulates production of TNFalpha and up-regulates production of IL-10 in LPS-stimulated macrophages. The objective of this study was to determine the mechanism of activity of PCERA-1 and the role of cAMP in LPS-induced IL-10 production. We show here that PCERA-1 induces IL-10 production in synergism with various TLR agonists in mouse RAW264.7 macrophages. Cooperativity is evident both at the mRNA and protein levels. IL-10 production by LPS and PCERA-1 is mediated by the cAMP pathway and by the p38 MAP kinase. Phosphorylation of p38 is cooperatively accomplished by LPS and PCERA-1 or other cAMP inducers. Furthermore, the activity of PCERA-1 can be partially mimicked by a cell-permeable analog of cAMP, and blocked by the protein kinase A (PKA) inhibitor H89. Finally, in the absence of PCERA-1, the residual IL-10 induction by LPS depends on the basal cAMP level as it can be largely elevated by the phosphodiesterase (PDE)-4 inhibitor rolipram. Our results thus indicate that IL-10 induction by LPS critically depends on basal cAMP level, and that a co-stimulus by a TLR agonist and a cAMP-elevating agent results in synergistic PKA-dependent and p38-dependent IL-10 production.
Department of Biochemistry, Life Sciences Institute, Tel-Aviv University, Tel-Aviv 69978, Israel.
Meir
M
Goldsmith
Dorit
D
Avni
Orna
O
Ernst
Yifat
Y
Glucksam
Galit
G
Levy-Rimler
Michael M
MM
Meijler
Tsaffrir
T
Zor
D000818:Animals; D002518:Ceramides / Q000494:pharmacology*; D000242:Cyclic AMP / Q000378:metabolism*; D054703:Cyclic Nucleotide Phosphodiesterases, Type 4 / Q000378:metabolism; D004357:Drug Synergism; D004789:Enzyme Activation / Q000187:drug effects; D016753:Interleukin-10 / Q000096:biosynthesis* / Q000235:genetics; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008264:Macrophages / Q000187:drug effects / Q000201:enzymology; D051379:Mice; D010755:Organophosphates / Q000494:pharmacology*; D014158:Transcription, Genetic / Q000187:drug effects; D015854:Up-Regulation / Q000187:drug effects; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*
C541119:1-methyl-2-(3-methoxyphenyl)-2-(octanoylamino)ethyl phosphate; D002518:Ceramides; D008070:Lipopolysaccharides; D010755:Organophosphates; C065576:ceramide 1-phosphate; D016753:Interleukin-10; D000242:Cyclic AMP; D048051:p38 Mitogen-Activated Protein Kinases; D054703:Cyclic Nucleotide Phosphodiesterases, Type 4; C095813:Pde4a protein, mouse