title: Establishment of reporter platforms capable of detecting NF-κB mediated immuno-modulatory activity.
abstract: Cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) are transcriptional targets of nuclear factor kappa B (NF-κB) that are involved in inflammatory responses. The aim of this study is to develop a method for efficiently detecting inflammation modulatory activities. Here we established RAW264.7 macrophage cells stably expressing a luciferase reporter gene directed by iNOS or COX-2 promoter. Lipopolysaccharide (LPS) treatment stimulated the luciferase activity which paralleled with increased iNOS and COX-2 mRNA levels determined by RT-q-PCR. The LPS-stimulated luciferase activity was blocked by NF-κB inhibitor CAPE and by nobiletin, an anti-inflammatory natural product from citrus peels. We have applied the platforms to screen various mushroom species; analysis by scatter plot revealed a strong correlation to the results obtained by ELISA-based detection of TNF-α. Together we have established luciferase reporter systems sensitive to NF-κB-dependent iNOS and COX-2 activation, which provides an alternative screening method for identifying food components with immune-modulatory activities.
authors.affiliation: Institute of Food Science and Technology, National Taiwan University , Taipei, Taiwan, R.O.C.
authors.forename: Chung-Huang
authors.initials: CH
authors.lastname: Wang
authors.forename: Ju-Hwa
authors.initials: JH
authors.lastname: Lin
authors.forename: Ting-Jang
authors.initials: TJ
authors.lastname: Lu
authors.forename: An-Na
authors.initials: AN
authors.lastname: Chiang
authors.forename: Shih-Ting
authors.initials: ST
authors.lastname: Chiou
authors.forename: Yi-An
authors.initials: YA
authors.lastname: Chen
authors.forename: Min-Hsiung
authors.initials: MH
authors.lastname: Pan
authors.forename: Shu-Chen
authors.initials: SC
authors.lastname: Hsieh
mesh_terms: D000818:Animals; D000893:Anti-Inflammatory Agents / Q000032:analysis* / Q000276:immunology; D001681:Biological Assay / Q000295:instrumentation / Q000379:methods*; D002957:Citrus / Q000737:chemistry*; D051546:Cyclooxygenase 2 / Q000235:genetics / Q000276:immunology; D017930:Genes, Reporter; D008264:Macrophages / Q000187:drug effects / Q000201:enzymology / Q000276:immunology; D051379:Mice; D016328:NF-kappa B / Q000032:analysis / Q000276:immunology*; D052247:Nitric Oxide Synthase Type II / Q000235:genetics / Q000276:immunology; D010936:Plant Extracts / Q000494:pharmacology*
chemical_list: D000893:Anti-Inflammatory Agents; D016328:NF-kappa B; D010936:Plant Extracts; D052247:Nitric Oxide Synthase Type II; D051546:Cyclooxygenase 2