Mediation by NF-kappa B of cytokine induced expression of intercellular adhesion molecule 1 (ICAM-1) in an intestinal epithelial cell line, a process blocked by proteasome inhibitors.

Abstract

The gene promoter for the intercellular adhesion molecule ICAM-1 possesses binding sites for several transcriptional factors, including nuclear factor kappa B (NF-kappa B). The role of NF-kappa B in ICAM-1 gene regulation was therefore examined by using different proteasome inhibitors in tumour necrosis factor alpha (TNF-alpha) stimulated IEC-6 rat intestinal epithelial cells. ICAM-1 expression was analysed by enzyme linked immunosorbent assay (ELISA), reverse transcriptase polymerase chain reaction, and immunohistochemistry. Steady state levels of cytoplasmic I kappa B protein were evaluated by western blot, and nuclear translocation of NF-kappa B was determined by electrophoretic mobility shift assay and immunofluorescence staining. Cell adhesion was assayed by measuring the binding of fluorescence labelled MOLT-4 cells. TNF-alpha induced ICAM-1 mRNA and protein expression in IEC-6 cells, which was followed by increased adhesion of MOLT-4 lymphocytes. Blocking TNF-alpha induced I kappa B alpha degradation with proteasome inhibitors reduced TNF-alpha induced NF-kappa B activation and ICAM-1 gene induction and notably decreased MOLT-4 cell adhesion without affecting Jun N-terminal kinase (JNK/SAPK) activity or de novo protein synthesis. TNF-alpha induction of ICAM-1 expression is mediated by the transcription factor NF-kappa B and can be inhibited by blocking I kappa B alpha degradation. Thus the I kappa B/NF-kappa B system is a promising target for pharmacological modulation of the expression of adhesion molecules and other inflammatory genes in the intestine.