Nitric oxide production by a murine macrophage cell line (RAW264.7) stimulated with lipopolysaccharide from Actinobacillus actinomycetemcomitans.
The aim of this study was to determine whether Actinobacillus actinomycetemcomitans lipopolysaccharide (LPS-A. actinomycetemcomitans) could stimulate a murine macrophage cell line (RAW264.7 cells) to produce nitric oxide (NO). The cells were treated with LPS-A. actinomycetemcomitans or Escherichia coli LPS (LPS-Ec) for 24 h. The effects of N(G)-monomethyl-L-arginine (NMMA), polymyxin B and cytokines (IFN-gamma, TNF-alpha, IL-4 and IL-12) on the production of NO were also determined. The role of protein tyrosine kinase, protein kinase C and microtubulin organization on NO production were assessed by incubating RAW264.7 cells with genistein, bisindolylmaleide and colchicine prior to LPS-A. actinomycetemcomitans stimulation, respectively. NO levels from the culture supernatants were determined by the Griess reaction. The results showed that LPS-A. actinomycetemcomitans stimulated NO production by RAW264.7 cells in a dose-dependent manner, but was slightly less potent than LPS-Ec. NMMA and polymyxin B blocked the production of NO. IFN-gamma and IL-12 potentiated but IL-4 depressed NO production by LPS-A. actinomycetemcomitans-stimulated RAW264.7 cells. TNF-alpha had no effects on NO production. Genistein and bisindolylmalemaide, but not colchicine, reduced the production of NO in a dose-dependent mechanism. The results of the present study suggest that A. actinomycetemcomitans LPS, via the activation of protein tyrosine kinase and protein kinase C and the regulatory control of cytokines, stimulates NO production by murine macrophages.
Sosroseno
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W
Department of Oral Biology, School of Dental Science, University Sains Malaysia, Kelantan, Malaysia.
Barid
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I
Herminajeng
E
E
Susilowati
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H
D016976:Aggregatibacter actinomycetemcomitans / Q000378:metabolism*; D000704:Analysis of Variance; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D002460:Cell Line; D003078:Colchicine / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000378:metabolism; D016166:Free Radical Scavengers / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019833:Genistein / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D007371:Interferon-gamma / Q000494:pharmacology; D018664:Interleukin-12 / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism*; D008301:Maleimides / Q000494:pharmacology; D051379:Mice; D022101:Microtubule-Organizing Center / Q000187:drug effects; D008870:Microtubules / Q000187:drug effects; D009569:Nitric Oxide / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D011112:Polymyxin B / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D019323:omega-N-Methylarginine / Q000494:pharmacology
D000900:Anti-Bacterial Agents; D004791:Enzyme Inhibitors; D016166:Free Radical Scavengers; D007211:Indoles; D008070:Lipopolysaccharides; D008301:Maleimides; D014409:Tumor Necrosis Factor-alpha; D018664:Interleukin-12; D019323:omega-N-Methylarginine; D009569:Nitric Oxide; D007371:Interferon-gamma; D019833:Genistein; D011505:Protein-Tyrosine Kinases; D011493:Protein Kinase C; D011112:Polymyxin B; C088060:bisindolylmaleimide; D003078:Colchicine