[Activation of nuclear factor-kappaB signaling pathway in rat gastric mucosa during stress ulceration].

Objective: To investigate the time course of nuclear factor-kappaB (NF-kappaB) activation in the process of stress ulcer formation.

Methods: Model of stress ulcer was reproduced by subjecting male Sprague-Dawley rats to water-immersion restraint (WIR) stress. At indicated time after the beginning of WIR stress, animals were sacrificed and cytoplasmic and nuclear protein and total RNA were prepared from gastric corpus mucosal tissues. DNA-binding activity of NF-KB was assessed as an index of NF-kappaB activation with electrophoretic mobility shift assay. Degradation of IkappaBalpha and IkappaBbeta, the inhibitory proteins of NF-kappaB, was analyzed by Western blot analysis. Expression of NF-kappaB dependent genes including tumor necrosis factor-alpha (TNF-alpha) , interleukin-1beta (IL-1beta), cytokine-inducible neutrophil chemoattractant-1 ( CINC-1), intercellular adhesion molecule-1 (ICAM-1), and inducible nitric oxide synthase (iNOS) was detected with Northern blot analysis.

Results: WIR stress induced a rapid biphasic activation of gastric mucosal NF-kappaB within 15 min of the beginning of stress, peaking at 45 min and 360 min. Compared with baseline, NF-kappaB activation by stress was increased (10.6 +/- 1.3) and (8.9 +/- 1.2) fold at 45 min and 360 min, respectively (P < 0.01). Antibody supershift assays revealed that p50/p65 heterodimer was the major active component of mucosal NF-kappaB. Western blot analysis showed that degradation of IkappaBalpha and IkappaBbeta occurred at first and second wave of NF-kappaB activation. Corresponding with the rapid and persistent activation of NF-kappaB, the levels of TNF-alpha, IL-1beta, CINC-1 and ICAM-1 mRNA in gastric mucosa were markedly increased 15 to 30 min after stress, respectively. Up-regulation of iNOS mRNAs was observed 30 to 90 min after stress, and the expression of all of these genes was increased consistently until the end of stress.

Conclusion: NF-kappaB activation is an early event and may play an important role in proinflammatory gene over-expression in rat gastric mucosa during WIR stress.