NF-κB inhibition enhances peroxynitrite-induced enterocyte apoptosis

Douglas A. Potoka, Jeffrey S. Upperman, Evan P. Nadler, Catarina T. Wong, Xin Zhou, Xiao Ru Zhang, Henri R. Ford

Research output: Contribution to journalArticlepeer-review

31 Scopus citations

Abstract

Background. Sustained overproduction of nitric oxide and peroxynitrite (ONOO-) in conditions such as necrotizing enterocolitis and inflammatory bowel disease may promote gut barrier failure by inducing enterocyte apoptosis. NF-κB is upregulated in the gut during inflammation and, in addition to its proinflammatory effects, may upregulate protective or anti-apoptotic factors such as inhibitor of apoptosis proteins (IAPs). We have previously demonstrated that NF-κB inhibition increases cytokine-induced enterocyte apoptosis; however, the effect of NF-κB on ONOO--induced enterocyte apoptosis is unknown. Materials and methods. Rat intestinal epithelial cells (IEC-6) were transfected with the adenoviral vector AdIκB or AdlacZ. AdIκB contains a mutated form of IκB which functions as a superrepressor of NF-κB. Cells were then treated with 50 μM ONOO- or decomposed ONOO-. Apoptosis was then determined by flow cytometry with annexin V-FITC and propidium iodide staining. Caspase activation and IAP, Bcl-2, Bad, and Bax expression were examined using Western blot analysis, and NF-κB activation was determined via electrophoretic mobility shift assay (EMSA). Results. Inhibition of NF-κB with AdIκB significantly enhanced ONOO--induced apoptosis in IEC-6 cells. ONOO- treatment did not activate NF-κB in IEC-6 cells as determined by EMSA. There was no difference in IAP, Bcl-2, Bad, and Bax expression between nontransfected, AdlacZ-transfected, and AdIκB-transfected cells. Baseline procaspase 3 activation was increased in AdIκB-transfected cells. Conclusions. NF-κB inhibition enhances ONOO--induced enterocyte apoptosis, suggesting that NF-κB upregulates a protective factor. This protective factor does not appear to be an IAP or Bcl-2 family member and may be expressed constitutively, since ONOO- did not activate NF-κB over baseline levels of activation.

Original languageEnglish (US)
Pages (from-to)7-14
Number of pages8
JournalJournal of Surgical Research
Volume106
Issue number1
DOIs
StatePublished - Jan 1 2002
Externally publishedYes

Keywords

  • Apoptosis
  • Caspases
  • Enterocyte
  • Inhibitor of apoptosis proteins
  • NF-κB
  • Peroxynitrite

ASJC Scopus subject areas

  • Surgery

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