Induction of NF-κB, IκB-α, and iNOS in rat gastric mucosa during endotoxemia

Kenneth S. Helmer, Lily Chang, Yan Cui, David W. Mercer

Research output: Contribution to journalArticle

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Abstract

Upregulation of inducible nitric oxide synthase (iNOS) contributes to the development of gastric injury during endotoxemia. The molecular mechanisms related to its induction are unknown. Because the transcription factor nuclear factor-kappa B (NF-κB) regulates inflammatory genes in response to endotoxemia, we hypothesized that its activity in gastric mucosa would increase while its inhibitor IκB-α would decrease commensurate with changes in iNOS expression. Rats were given intraperitoneal saline or lipopolysaccharide (LPS; 20 mg/kg) for 5, 15, or 30 min, or for 1, 3, or 5 h, and killed, and the gastric mucosa was prepared for determination of iNOS and IκB-α by Western immunoblotting, iNOS mRNA by quantitative real-time RT-PCR, and NF-κB by electrophoretic mobility-shift assay. LPS caused a significant increase in iNOS mRNA and protein immunoreactivity at 1, 3, and 5 h compared to controls. NF-κB-binding activity increased in the nuclear fraction of gastric mucosa at 1 h and steadily increased over time after LPS administration. The activated NF-κB consisted mainly of p50 with a lesser amount of p65 subunits as demonstrated by a supershift assay. IκB-α decreased in gastric cytosolic fractions over time, consistent with its degradation. These data suggest that during endotoxemia expression of the inflammatory mediator iNOS in the gastric mucosa may be upregulated by degradation of IκB-α and subsequent translocation of NF-κB into the nucleus and increased NF-κB activity.

Original languageEnglish (US)
Pages (from-to)46-52
Number of pages7
JournalJournal of Surgical Research
Volume104
Issue number1
DOIs
StatePublished - Jan 1 2002

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Endotoxemia
NF-kappa B
Nitric Oxide Synthase Type II
Gastric Mucosa
Stomach
Messenger RNA
Electrophoretic Mobility Shift Assay
Lipopolysaccharides
Real-Time Polymerase Chain Reaction
Transcription Factors
Up-Regulation
Western Blotting
Wounds and Injuries
Genes
Proteins

ASJC Scopus subject areas

  • Surgery

Cite this

Induction of NF-κB, IκB-α, and iNOS in rat gastric mucosa during endotoxemia. / Helmer, Kenneth S.; Chang, Lily; Cui, Yan; Mercer, David W.

In: Journal of Surgical Research, Vol. 104, No. 1, 01.01.2002, p. 46-52.

Research output: Contribution to journalArticle

Helmer, Kenneth S. ; Chang, Lily ; Cui, Yan ; Mercer, David W. / Induction of NF-κB, IκB-α, and iNOS in rat gastric mucosa during endotoxemia. In: Journal of Surgical Research. 2002 ; Vol. 104, No. 1. pp. 46-52.
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