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.
OBJECTIVE
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.


METHODS
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.


RESULTS
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.


CONCLUSIONS
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.
Department of Medicine, University of North Carolina at Chapel Hill 27599, USA.
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Jobin
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Hellerbrand
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Licato
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Brenner
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Sartor
D000818:Animals; D002460:Cell Line; D003513:Cycloheximide / Q000494:pharmacology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D005786:Gene Expression Regulation; D020672:I-kappa B Proteins; D018799:Intercellular Adhesion Molecule-1 / Q000235:genetics* / Q000378:metabolism; D007413:Intestinal Mucosa / Q000378:metabolism; D007422:Intestines / Q000187:drug effects*; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000502:physiology*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D051381:Rats; D015533:Transcriptional Activation; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D015853:Cysteine Proteinase Inhibitors; D004268:DNA-Binding Proteins; D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606206:Nfkbia protein, rat; D011500:Protein Synthesis Inhibitors; D014409:Tumor Necrosis Factor-alpha; D018799:Intercellular Adhesion Molecule-1; D000072000:NF-KappaB Inhibitor alpha; D003513:Cycloheximide