Nuclear factor kappa B activation by NADPH oxidases

Robert A. Clark, Anthony J. Valente

Research output: Contribution to journalArticle

60 Scopus citations

Abstract

Reactive oxygen species (ROS) initiate activation of the transcription factor NF-κB in a variety of cell systems. Perhaps the most potent biological source of ROS is the NADPH oxidase of phagocytic cells, a multi-component system that catalyzes the formation of superoxide anion. Although phagocytes use this oxidase to kill ingested microorganisms, the products also mediate a broad range of biological oxidation reactions and some evidence exists for activation of NF-κB through this mechanism. Moreover, the components of the phagocyte NADPH oxidase are present in certain non-phagocytic cells and recently discovered homologues of the catalytic component gp91 phox are expressed in a number of tissues. We explored the hypothesis that the products of NADPH oxidases cause the activation of NF-κB. K562 human erythrokeukemia cells transfected with constructs for expression of gp91 phox, plus other essential NADPH oxidase components generated substantial amounts of superoxide when activated with phorbol ester, lesser amounts with arachidonic acid exposure, and none with TNFα. Gel shift assays demonstrated induction of NF-κB in K562 cells exposed to TNFα and specificity was shown by oligonucleotide competition. Supershift assays demonstrated the presence in nuclear complexes of the NF-κB components p65/RelA and p50. Nuclear complexes of identical electrophoretic mobility were induced in phorbol ester-stimulated K562 cells that expressed the complete NADPH oxidase system, but not in cells lacking one of the essential oxidase components. K562 cells were relatively resistant to NF-κB induction by exogenous peroxide, but certain other cell types (HEK293 and HeLaS3) demonstrated such induction upon exposure to reagent hydrogen peroxide or glucose oxidase plus glucose and this was blocked by catalase. Finally, we found a biphasic pattern of gp91 phox expression in rat liver during aging. High levels observed in young animals decreased in middle age, but increased again in old age. Collectively, these studies demonstrate the potential for NADPH-dependent induction of NF-κB and raise the possibility of a role for this pathway in the biology of aging.

Original languageEnglish (US)
Pages (from-to)799-810
Number of pages12
JournalMechanisms of Ageing and Development
Volume125
Issue number10-11 SPEC. ISS.
DOIs
Publication statusPublished - Oct 1 2004

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Keywords

  • Ageing
  • NADPH oxidase
  • NF-κB
  • NOX
  • Superoxide

ASJC Scopus subject areas

  • Aging
  • Developmental Biology

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