Time-dependent modulation of thioredoxin reductase activity might contribute to sulforaphane-mediated inhibition of NF-kappaB binding to DNA.
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.
Heiss
Elke
E
Division C010, Toxicology and Cancer Risk Factors, Deutsches Krebsforschungszentrum, 69120 Heidelberg, Germany.
Gerhäuser
Clarissa
C
D000818:Animals; D002460:Cell Line; D054746:Cyclic AMP-Dependent Protein Kinase Type II; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D004137:Dinitrochlorobenzene / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005978:Glutathione / Q000378:metabolism; D017879:Isothiocyanates; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D016328:NF-kappa B / Q000037:antagonists & inhibitors* / Q000378:metabolism; D009569:Nitric Oxide / Q000096:biosynthesis; D052247:Nitric Oxide Synthase Type II / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D013454:Sulfoxides; D013861:Thiocyanates / Q000494:pharmacology*; D013880:Thioredoxin-Disulfide Reductase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013997:Time Factors
D004137:Dinitrochlorobenzene; D004791:Enzyme Inhibitors; D017879:Isothiocyanates; D008070:Lipopolysaccharides; D016328:NF-kappa B; D013454:Sulfoxides; D013861:Thiocyanates; D009569:Nitric Oxide; D004247:DNA; D052247:Nitric Oxide Synthase Type II; D013880:Thioredoxin-Disulfide Reductase; D054746:Cyclic AMP-Dependent Protein Kinase Type II; D017868:Cyclic AMP-Dependent Protein Kinases; C016766:sulforaphane; D005978:Glutathione