Time-dependent modulation of thioredoxin reductase activity might contribute to sulforaphane-mediated inhibition of NF-kappaB binding to DNA

Antioxid Redox Signal. Nov-Dec 2005;7(11-12):1601-11. doi: 10.1089/ars.2005.7.1601.

Abstract

The chemopreventive agent sulforaphane (SFN) exerts anti-inflammatory activity by thiol-dependent inhibition of nuclear factor kappaB (NF-kappaB) DNA binding. To further analyze the underlying mechanisms, we focused on the thioredoxin/thioredoxin reductase (TrxR) system as a key redox mechanism regulating NF-kappaB DNA binding. Using cultured Raw 264.7 mouse macrophages as a model, 1-chloro-2,4-dinitrobenzene (CDNB), a known inhibitor of TrxR, was identified as an inhibitor of lipopolysaccharide (LPS)-mediated nitric oxide (NO) production and of NF-kappaB DNA binding. CDNB and SFN acted synergistically with respect to inhibition of LPS-induced NO release, and we consequently identified SFN as a novel inhibitor of TrxR enzymatic activity in vitro. Short-term treatment of Raw macrophages with SFN or CDNB resulted in the inhibition of TrxR activity in vivo with half-maximal inhibitory concentration of 25.0 +/- 3.5 microM and 9.4 +/- 3.7 microM, respectively, whereas after a 24-h treatment with 25 microM SFN, TrxR activity was >1.5-fold elevated. In additional experiments, we could exclude that inhibition of trans-activating activity of NF-kappaB contributed to the reduced expression of pro-inflammatory proteins by SFN, based on transient transfection experiments with a (kappaB)(2)- chloramphenicol acetyltransferase construct and a lack of inhibition of protein kinase A activity. These findings further emphasize the importance of redox modulation or thiol reactivity for the regulation of NF-kappaB-dependent transcription by SFN. Antioxid. Redox Signal. 7, 1601-1611.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Line
  • Cyclic AMP-Dependent Protein Kinase Type II
  • Cyclic AMP-Dependent Protein Kinases / metabolism
  • DNA / metabolism*
  • Dinitrochlorobenzene / pharmacology
  • Enzyme Inhibitors / pharmacology
  • Glutathione / metabolism
  • Isothiocyanates
  • Lipopolysaccharides / pharmacology
  • Macrophages / drug effects
  • Macrophages / metabolism
  • Mice
  • NF-kappa B / antagonists & inhibitors*
  • NF-kappa B / metabolism
  • Nitric Oxide / biosynthesis
  • Nitric Oxide Synthase Type II / metabolism
  • Protein Binding / drug effects
  • Thiocyanates / pharmacology*
  • Thioredoxin-Disulfide Reductase / antagonists & inhibitors
  • Thioredoxin-Disulfide Reductase / metabolism*
  • Time Factors

Substances

  • Dinitrochlorobenzene
  • Enzyme Inhibitors
  • Isothiocyanates
  • Lipopolysaccharides
  • NF-kappa B
  • Thiocyanates
  • Nitric Oxide
  • DNA
  • Nitric Oxide Synthase Type II
  • Thioredoxin-Disulfide Reductase
  • Cyclic AMP-Dependent Protein Kinase Type II
  • Cyclic AMP-Dependent Protein Kinases
  • sulforafan
  • Glutathione