title: The spin trap 5,5-dimethyl-1-pyrroline N-oxide inhibits lipopolysaccharide-induced inflammatory response in RAW 264.7 cells.
abstract: OBJECTIVE
  Exposure of macrophages to lipopolysaccharide (LPS) induces oxidative and inflammatory stresses, which cause cell damage. Antioxidant and anti-inflammatory properties have been attributed to the nitrone spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO), commonly used in free radical analysis, but these aspects of DMPO have been little explored. In this study, we sought to establish the anti-inflammatory activity of DMPO, presumably by removing free radicals which otherwise help activate inflammatory response and damage cells.
  
  
  METHODS
  RAW 264.7 macrophages were treated with LPS and/or DMPO for different time points, cell damage, production of inflammatory mediators, inducible nitric oxide synthase (iNOS) expression, NF-κB p65 activation, phosphorylation of MAPKs and Akt, and intracellular reactive oxygen species (ROS) were determined.
  
  
  RESULTS
  After cells were treated with LPS and/or DMPO for 24 h, DMPO reduced the LPS-induced inflammatory response as indicated by downregulated iNOS expression and production of inflammatory mediators. Accordingly, DMPO protected cells from LPS-induced cytotoxicity. In order to understand the mechanistic basis of these DMPO effects, the NF-κB p65 activation and the phosphorylation of MAPKs and Akt were examined. We found, by assaying cells treated with LPS and/or DMPO for 15-60 min, that DMPO inhibited the phosphorylation of MAPKs, Akt, and IκBα, and reduced the NF-κB p65 translocation. Furthermore, we demonstrated that DMPO inhibited LPS-induced ROS production.
  
  
  CONCLUSIONS
  DMPO showed the anti-inflammatory activity and attenuated LPS-induced cell damage, most likely by reducing ROS production and thus preventing the subsequent inflammatory activation and damage.
authors.affiliation: Experimental Therapeutics Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK 73104, USA. zlzhai@yahoo.com
authors.forename: Zili
authors.initials: Z
authors.lastname: Zhai
authors.forename: Sandra E
authors.initials: SE
authors.lastname: Gomez-Mejiba
authors.forename: Hua
authors.initials: H
authors.lastname: Zhu
authors.forename: Florea
authors.initials: F
authors.lastname: Lupu
authors.forename: Dario C
authors.initials: DC
authors.lastname: Ramirez
mesh_terms: D000818:Animals; D000893:Anti-Inflammatory Agents / Q000494:pharmacology*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D003497:Cyclic N-Oxides / Q000494:pharmacology*; D007249:Inflammation / Q000139:chemically induced / Q000517:prevention & control*; D008070:Lipopolysaccharides / Q000633:toxicity; D008264:Macrophages / Q000187:drug effects; D051379:Mice; D052247:Nitric Oxide Synthase Type II / Q000378:metabolism; D009573:Nitrites / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013113:Spin Labels
chemical_list: D000893:Anti-Inflammatory Agents; D003497:Cyclic N-Oxides; D008070:Lipopolysaccharides; D009573:Nitrites; D017382:Reactive Oxygen Species; D013113:Spin Labels; C017245:5,5-dimethyl-1-pyrroline-1-oxide; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse