5-Aza-2'-deoxycytidine stimulates inducible nitric oxide synthase induction in C6 astrocytoma cells.
The influence of a nucleoside analog 5-aza-2'-deoxycytidine (5-AzadC) on inducible nitric oxide synthase (iNOS)-dependent nitric oxide (NO) production in various rat cell types was investigated. In C6 astrocytoma cell line and primary astrocytes, 5-AzadC enhanced proinflammatory cytokine (IFN-gamma, TNF-alpha, IL-1)-triggered NO synthesis in a time- and dose-dependent manner. In contrast, 5-AzadC did not potentiate NO production in IFN-gamma-stimulated macrophages, fibroblasts, or endothelial cells. Blockade of transcription or translation in C6 cells abolished the observed effect, suggesting the iNOS gene expression, rather than its catalytic activity, as a target for the drug action. Accordingly, 5-AzadC upregulated IFN-gamma-induced expression of iNOS mRNA in C6 astrocytes. The effect of 5-AzadC on astrocyte NO release was blocked by the inhibitor of p44/42 mitogen activated protein kinase-dependent signaling. Finally, the observed stimulatory effect of 5-AzadC on iNOS expression was apparently independent of DNA demethylation, as DNA digestion with methylation-sensitive restriction enzyme HpaII showed that 5-AzadC failed to demethylate cellular DNA in conditions used for iNOS induction.
Cvetkovic
I
I
Laboratory for Immunology, Institute for Biological Research, 29. novembra 142, 11000 Belgrade, Serbia, Yugoslavia. branicevo@yahoo.com
Popadic
D
D
Vuckovic
O
O
Harhaji
Lj
Lj
Miljkovic
Dj
Dj
Trajkovic
V
V
D000818:Animals; D000831:Animals, Newborn; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D001253:Astrocytes / Q000187:drug effects*; D001254:Astrocytoma / Q000473:pathology; D001374:Azacitidine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D015152:Blotting, Northern; D002478:Cells, Cultured; D019175:DNA Methylation / Q000187:drug effects; D015254:DNA Modification Methylases / Q000037:antagonists & inhibitors*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D000077209:Decitabine; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D004790:Enzyme Induction / Q000187:drug effects; D015870:Gene Expression / Q000187:drug effects; D050856:Interferon Regulatory Factor-1; D007372:Interferons / Q000494:pharmacology; D007375:Interleukin-1 / Q000494:pharmacology; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000096:biosynthesis*; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D000964:Antimetabolites, Antineoplastic; D004268:DNA-Binding Proteins; D050856:Interferon Regulatory Factor-1; D007375:Interleukin-1; C494259:Irf1 protein, rat; D010750:Phosphoproteins; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D000077209:Decitabine; D007372:Interferons; D019001:Nitric Oxide Synthase; D015254:DNA Modification Methylases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D001374:Azacitidine