Serum amyloid A activates NF-kappaB and proinflammatory gene expression in human and murine intestinal epithelial cells

Abstract

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.