Nuclear factor-kappaB is activated in intestinal mucosa during endotoxemia.

BACKGROUND The transcription factor nuclear factor-kappaB (NF-kappaB) regulates a large number of genes involved in the inflammatory response to critical illness. The intestinal mucosa plays an active role in the inflammatory and metabolic response to sepsis and endotoxemia, but it is not known if NF-kappaB is activated in the mucosa during these conditions. OBJECTIVE To test the hypothesis that endotoxemia in mice activates NF-kappaB in intestinal mucosa. METHODS Mice were injected subcutaneously with lipopolysaccharide, 12.5 mg/kg, or a corresponding volume of saline. At various intervals following injection, jejunal mucosa was harvested and nuclear and cytoplasmic fractions were prepared. The nuclear fractions were analyzed by electrophoretic mobility shift assay for NF-kappaB activation and by Western blot analysis for the NF-kappaB subunits p50 and p65. Cytoplasmic fractions were analyzed by Western blotting for the NF-kappaB inhibitory proteins IkappaB-alpha and IkappaB-beta. RESULTS Electrophoretic mobility shift assay showed that NF-kappaB was activated in jejunal mucosa 1 hour after injection of lipopolysaccharide and persisted for at least 4 hours. The NF-kappaB subunits p50 and p65 were present in nuclear fractions of mucosa from endotoxemic mice at the corresponding time points. Cytoplasmic levels of the inhibitory proteins IkappaB-alpha and IkappaB-beta decreased during endotoxemia, and the proteins were nearly absent 60 minutes after injection of lipopolysaccharide. CONCLUSIONS The results suggest that IkappaB is degraded and NF-kappaB is activated in intestinal mucosa during endotoxemia. The findings support the concept that the intestinal mucosa is an important component of the inflammatory response to sepsis and endotoxemia.