Characterization of the pro-inflammatory signaling induced by protein acetylation in microglia.
Protein acetylation regulates the extent of inflammatory responses and disturbances in protein acetylation have been proposed to play an important role in inflammatory and neurodegenerative diseases. We have recently observed that histone deacetylase inhibitors, such as trichostatin A (TSA) and SAHA, strongly potentiate the LPS induced inflammatory response in several rat and mouse inflammatory models. Our aim here was to characterise pro-inflammatory signaling mediated via increased protein acetylation and protein phosphorylation in microglial N9 cells. First we observed that TSA induced pro-inflammatory response was independent of the different Toll-like receptors activated, since LPS, flagellin and unmethylated CpG oligonucleotides, equally potentiated IL-6 secretion from N9 microglia. Next we compared the protein acetylation induced potentiation to that induced by okadaic acid, a well-known inducer of pro-inflammatory responses. The time scale of the IL-6 responses showed that the effects of okadaic acid were clearly early-response effects appearing as soon as 6h after exposure, whereas TSA evoked a significant inhibition in IL-6 secretion up to 12h but after that it induced an exponential increase in cytokine and nitric oxide production up to 24h. It seems that okadaic acid induces an early moderate response and TSA a late but exponential potentiation of microglial inflammatory responses. The pro-inflammatory responses of TSA and okadaic acid were both dependent on NF-kappaB signaling but independent on the DNA-binding capacity of nuclear NF-kappaB complexes. Interestingly, we observed that the transactivation of the NF-kappaB-Luc reporter gene was clearly activated during TSA induced pro-inflammatory potentiation. Our studies imply that the potentiation of the inflammatory response by increased acetylation is due to the enhancement of transactivation of NF-kappaB driven inflammatory genes. Our studies on signaling pathways revealed that PI3K inhibitors LY294002 and Wortmannin blocked the TSA induced pro-inflammatory response but surprisingly did not affect the okadaic acid induced response. Furthermore, LY294002 did not inhibit DNA-binding activity of NF-kappaB but still inhibited NF-kappaB-Luc reporter gene transactivation. These results indicate that PI 3-kinase regulates the transactivation efficiency of NF-kappaB-dependent transcription rather than transduction of NF-kappaB signaling.
Suuronen
Tiina
T
Department of Neuroscience and Neurology, University of Kuopio, P.O. Box 1627, FIN-70211 Kuopio, Finland.
Huuskonen
Jari
J
Nuutinen
Tapio
T
Salminen
Antero
A
D000107:Acetylation; D000818:Animals; D002867:Chromones / Q000494:pharmacology; D004247:DNA / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D006877:Hydroxamic Acids / Q000494:pharmacology; D066298:In Vitro Techniques; D007249:Inflammation / Q000503:physiopathology*; D018836:Inflammation Mediators / Q000494:pharmacology; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D017628:Microglia / Q000187:drug effects / Q000502:physiology*; D009025:Morpholines / Q000494:pharmacology; D016328:NF-kappa B / Q000235:genetics / Q000502:physiology; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism; D019319:Okadaic Acid / Q000494:pharmacology; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D051381:Rats; D051193:Toll-Like Receptors / Q000187:drug effects
D002867:Chromones; D004791:Enzyme Inhibitors; D006877:Hydroxamic Acids; D018836:Inflammation Mediators; D008070:Lipopolysaccharides; D009025:Morpholines; D016328:NF-kappa B; D009419:Nerve Tissue Proteins; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051193:Toll-Like Receptors; D019319:Okadaic Acid; D009569:Nitric Oxide; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C012589:trichostatin A; D004247:DNA; D007770:L-Lactate Dehydrogenase