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Adenosine suppresses lipopolysaccharide-induced tumor necrosis factor-alpha production by murine macrophages through a protein kinase A- and exchange protein activated by cAMP-independent signaling pathway. J Pharmacol Exp Ther 2009 Dec;331(3):1051-61

Date

09/15/2009

Pubmed ID

19749080

Pubmed Central ID

PMC2784717

DOI

10.1124/jpet.109.157651

Scopus ID

2-s2.0-73349106518   32 Citations

Abstract

Adenosine is generated during tissue hypoxia and stress, which reduces inflammation by suppressing the activity of most immune cells. Among its various actions, adenosine suppresses the production of proinflammatory cytokines including tumor necrosis factor (TNF)-alpha, through the cAMP-elevating A(2A) adenosine receptor (AR) subtype. In this study, we examined the signaling mechanisms by which A(2A)AR activation inhibits TNF-alpha production in thioglycollate-elicited mouse peritoneal macrophages. Pretreating murine macrophages with the nonselective AR agonist adenosine-5'-N-ethylcarboxamide (NECA), the A(2A)AR agonist 2-[p-(2-carboxyethyl)phenethylamino]-5'-N-ethylcarboxamidoadenosine (CGS 21680), or the cAMP-elevating agent forskolin reduced TNF-alpha production in response to lipopolysaccharide (LPS) by greater than 60%. All of these agents increased cAMP production in macrophages and activated protein kinase A (PKA). However, we were surprised to find that treating macrophages with three different PKA inhibitors or small interfering RNA-mediated knockdown of the exchange protein activated by cAMP (Epac-1) failed to block the suppressive actions of NECA or forskolin on LPS-induced TNF-alpha release. Instead, okadaic acid was effective at low concentrations that selectively inhibit protein serine/threonine phosphatases. Subsequent studies showed that NECA and forskolin decreased LPS-induced steady-state TNF-alpha mRNA levels; this effect was due to a decreased rate of transcription based on assays examining the rate of generation of primary TNF-alpha transcripts. Treatment with NECA or forskolin did not interfere with LPS-induced translocation or DNA binding of the RelA/p65 subunit of nuclear factor-kappaB or phosphorylation of inhibitor of nuclear factor-kappaB-alpha, extracellular signal-regulated kinase 1/2, c-Jun NH(2)-terminal kinase, or p38 kinase. Our results suggest that AR activation inhibits LPS-induced TNF-alpha production by murine macrophages at the level of gene transcription through a unique cAMP-dependent, but PKA- and Epac-independent, signaling pathway involving protein phosphatase activity.

Author List

Kreckler LM, Gizewski E, Wan TC, Auchampach JA

Author

John A. Auchampach PhD Professor in the Pharmacology and Toxicology department at Medical College of Wisconsin




MESH terms used to index this publication - Major topics in bold

Adenosine
Adenosine A2 Receptor Agonists
Animals
Blotting, Western
Cells, Cultured
Cyclic AMP
Cyclic AMP-Dependent Protein Kinases
Enzyme Activation
Guanine Nucleotide Exchange Factors
Lipopolysaccharides
Macrophages, Peritoneal
Mice
Mice, Inbred C57BL
Receptor, Adenosine A2A
Reverse Transcriptase Polymerase Chain Reaction
Signal Transduction
Tumor Necrosis Factor-alpha
jenkins-FCD Prod-486 e3098984f26de787f5ecab75090d0a28e7f4f7c0