Serum amyloid A activates NF-kappaB and proinflammatory gene expression in human and murine intestinal epithelial cells.
Serum amyloid A (SAA) is an acute-phase protein whose levels positively correlate with disease activity in inflammatory bowel diseases. In this study we investigated the impact of SAA on NF-kappaB signaling and proinflammatory gene expression in intestinal epithelial cells (IEC). Human HT-29 and Caco-2 monolayers were stimulated with recombinant SAA and NF-kappaB activation/NF-kappaB-dependent gene expression measured. Adenoviral dominant negative mutants IkappaB-alpha (Ad5IkappaBAA) were utilized to determine the contribution of NF-kappaB signaling pathway to SAA-dependent gene expression. Intestinal explant and primary IEC derived from kappaB-EGFP transgenic mice were exposed to SAA and NF-kappaB-dependent enhanced green fluorescent protein (EGFP) fluorescence measured. SAA induced IkappaB-alpha degradation, RelA serine 536 (S536) phosphorylation, NF-kappaB transcriptional activity, RelA recruitment to the IL-8 gene promoter and endogenous gene expression (IL-8, COX-2) in HT-29 cells. Further, Ad5IkappaBAA abrogated SAA-induced RelA nuclear translocation, NF-kappaB transcriptional activity and IL-8 gene expression. SAA-dependent IL-8 gene expression required activation of the MAPK ERK, p38 and JNK in HT-29 cells. Finally, SAA induced EGFP expression in intestinal explants isolated from kappaB-EGFP transgenic mice and enhanced RelA and IkappaBalpha phosphorylation in primary IEC. This indicates that SAA potentially participate in the inflammatory process by virtue of its ability to activate proinflammatory signaling in IEC.
Jijon
Humberto B
HB
Gastrointestinal Research Unit, University of Alberta, Edmonton, Canada.
Madsen
Karen L
KL
Walker
John W
JW
Allard
Brigitte
B
Jobin
Christian
C
D000818:Animals; D015153:Blotting, Western; D018938:Caco-2 Cells; D051546:Cyclooxygenase 2; D024202:Electrophoretic Mobility Shift Assay; D004789:Enzyme Activation / Q000187:drug effects* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression / Q000187:drug effects* / Q000276:immunology; D049452:Green Fluorescent Proteins / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D019073:HT29 Cells; D006801:Humans; D020672:I-kappa B Proteins / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D047468:Immunoprecipitation; D016209:Interleukin-8 / Q000276:immunology / Q000378:metabolism; D007413:Intestinal Mucosa / Q000187:drug effects* / Q000276:immunology; D048031:JNK Mitogen-Activated Protein Kinases / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D048670:MAP Kinase Kinase 4; D008565:Membrane Proteins; D051379:Mice; D008822:Mice, Transgenic; D020929:Mitogen-Activated Protein Kinase Kinases / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011451:Prostaglandin-Endoperoxide Synthases / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D000685:Serum Amyloid A Protein / Q000276:immunology / Q000494:pharmacology*; D051996:Transcription Factor RelA; D048051:p38 Mitogen-Activated Protein Kinases / Q000187:drug effects / Q000276:immunology / Q000378:metabolism
D020672:I-kappa B Proteins; D016209:Interleukin-8; D008565:Membrane Proteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; C000606205:Nfkbia protein, mouse; D000685:Serum Amyloid A Protein; D051996:Transcription Factor RelA; C485184:enhanced green fluorescent protein; D000072000:NF-KappaB Inhibitor alpha; D049452:Green Fluorescent Proteins; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases; D048031:JNK Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D048670:MAP Kinase Kinase 4; D020929:Mitogen-Activated Protein Kinase Kinases