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 Ford

Research output: Contribution to journalArticle

31 Citations (Scopus)

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

Fingerprint

Peroxynitrous Acid
Enterocytes
Inhibitor of Apoptosis Proteins
Apoptosis
Electrophoretic Mobility Shift Assay
Up-Regulation
Necrotizing Enterocolitis
Caspase Inhibitors
Propidium
Fluorescein-5-isothiocyanate
Annexin A5
Inflammatory Bowel Diseases
Caspase 3
Flow Cytometry
Nitric Oxide
B-Lymphocytes
Western Blotting
Epithelial Cells
Staining and Labeling
Cytokines

Keywords

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

ASJC Scopus subject areas

  • Surgery

Cite this

Potoka, D. A., Upperman, J. S., Nadler, E. P., Wong, C. T., Zhou, X., Zhang, X. R., & Ford, H. (2002). NF-κB inhibition enhances peroxynitrite-induced enterocyte apoptosis. Journal of Surgical Research, 106(1), 7-14. https://doi.org/10.1006/jsre.2002.6423

NF-κB inhibition enhances peroxynitrite-induced enterocyte apoptosis. / Potoka, Douglas A.; Upperman, Jeffrey S.; Nadler, Evan P.; Wong, Catarina T.; Zhou, Xin; Zhang, Xiao Ru; Ford, Henri.

In: Journal of Surgical Research, Vol. 106, No. 1, 01.01.2002, p. 7-14.

Research output: Contribution to journalArticle

Potoka, DA, Upperman, JS, Nadler, EP, Wong, CT, Zhou, X, Zhang, XR & Ford, H 2002, 'NF-κB inhibition enhances peroxynitrite-induced enterocyte apoptosis', Journal of Surgical Research, vol. 106, no. 1, pp. 7-14. https://doi.org/10.1006/jsre.2002.6423
Potoka DA, Upperman JS, Nadler EP, Wong CT, Zhou X, Zhang XR et al. NF-κB inhibition enhances peroxynitrite-induced enterocyte apoptosis. Journal of Surgical Research. 2002 Jan 1;106(1):7-14. https://doi.org/10.1006/jsre.2002.6423
Potoka, Douglas A. ; Upperman, Jeffrey S. ; Nadler, Evan P. ; Wong, Catarina T. ; Zhou, Xin ; Zhang, Xiao Ru ; Ford, Henri. / NF-κB inhibition enhances peroxynitrite-induced enterocyte apoptosis. In: Journal of Surgical Research. 2002 ; Vol. 106, No. 1. pp. 7-14.
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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.",
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T1 - NF-κB inhibition enhances peroxynitrite-induced enterocyte apoptosis

AU - Potoka, Douglas A.

AU - Upperman, Jeffrey S.

AU - Nadler, Evan P.

AU - Wong, Catarina T.

AU - Zhou, Xin

AU - Zhang, Xiao Ru

AU - Ford, Henri

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N2 - 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.

AB - 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.

KW - Apoptosis

KW - Caspases

KW - Enterocyte

KW - Inhibitor of apoptosis proteins

KW - NF-κB

KW - Peroxynitrite

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