IL-10 modulates cytokine gene transcription by protein synthesis-independent and dependent mechanisms in lipopolysaccharide-treated neutrophils.
We have recently reported that the ability of IL-10 to rapidly exert its anti-inflammatory effects on human neutrophils is dependent upon exposure of these cells to LPS for at least 3-4 h. Here, we demonstrate that, in neutrophils "preconditioned" by LPS, IL-10 primarily targets the transcription of TNF-alpha, CXCL8 and IL-1ra genes, as revealed by primary transcript real-time RT-PCR. We also show that IL-10-induced transcriptional repression of TNF-alpha and CXCL8 genes consists of two distinct phases: an early one, occurring rapidly and in a protein synthesis-independent manner, followed by a second phase, more delayed and dependent on protein synthesis. Interestingly, the protein synthesis dependence of the latter phase coincides with a reduced ability of IL-10 to induce STAT3 tyrosine phosphorylation. Importantly, inhibition of IL-10-induced STAT3 activation and IL-10-suppressive action by a prolonged exposure to cycloheximide (CHX) was observed to occur also in human monocytes and was caused by a defective IL-10-mediated activation of Jak1 and Tyk2 kinases. Taken together, our findings suggest that CHX interferes with the IL-10-mediated intracellular signaling pathway by interrupting events upstream of STAT3 activation. These data question the concept of the requirement of an IL-10-induced mediator as the unique mechanism to execute IL-10 anti-inflammatory program.
Rossato
Marzia
M
Department of Pathology, Division of General Pathology, University of Verona, Verona, Italy.
Cencig
Sabrina
S
Gasperini
Sara
S
Cassatella
Marco A
MA
Bazzoni
Flavia
F
D015153:Blotting, Western; D003513:Cycloheximide / Q000494:pharmacology; D016207:Cytokines / Q000096:biosynthesis*; D005434:Flow Cytometry; D015870:Gene Expression; D006801:Humans; D047468:Immunoprecipitation; D016753:Interleukin-10 / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000276:immunology*; D009504:Neutrophils / Q000276:immunology* / Q000378:metabolism; D014176:Protein Biosynthesis; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D050796:STAT3 Transcription Factor / Q000276:immunology / Q000378:metabolism; D015398:Signal Transduction / Q000276:immunology*; D014158:Transcription, Genetic / Q000276:immunology*
D016207:Cytokines; D008070:Lipopolysaccharides; D011500:Protein Synthesis Inhibitors; D050796:STAT3 Transcription Factor; D016753:Interleukin-10; D003513:Cycloheximide