title: Microbial cell components induced tolerance to flagellin-stimulated inflammation through Toll-like receptor pathways in intestinal epithelial cells.
abstract: In the intestine, bacterial components activate innate responses that protect the host. We hypothesize that bacterial components reduce Interleukin-8 (IL-8) production in intestinal epithelial cells stimulated by flagellin via the Toll-like receptor (TLR) signaling pathway. Caco-2 cells were pretreated with various doses of lipopolysaccharide (LPS), lipoteichoic acid (LTA), or low-dose flagellin (LDFL) for 24h. Cells were then treated with flagellin (FL) 500 ng/ml (HDFL) for another 48 h. IL-8 production was measured in the cell culture medium by ELISA. Eighty-four genes in the TLR pathway were evaluated by RT Profiler PCR Array. Pathway Studio 8.0 software was used for altered pathway analysis. HDFL induced IL-8 production by 19-fold (p<0.01). Pretreatment with LDFL at 20, 10 or 1 ng/ml reduced HDFL-induced IL-8 production by 61%, 52% and 40%, respectively (p<0.05). LPS at 50 μg/ml decreased HDFL-induced IL-8 production by 38% (p<0.05). HDFL up-regulated CXCL10, IL1B, IL-8, IRAK2, NF-κB1 and I-κB (all p<0.05). Pathway Studio analysis showed that HDFL induced cell processes including inflammation, cell death and apoptosis. Pretreatment with LDFL at 10 ng/ml down-regulated FADD, FOS, MAP4K4, MyD88, TLR2, TLR3 and TNFERSF1A compared to HDFL (all p<0.05). These down-regulated genes are integral for numerous cell functions including inflammatory response, cell death, apoptosis and infection. These results demonstrate that LPS and LDFL provoke tolerance to HDFL-induced IL-8 production. This tolerance effect was accompanied by a complex interaction of multiple genes related to inflammatory as well as other responses in the TLR pathway rather than a single gene alteration.
authors.affiliation: Department of Pediatrics, University of Florida, 1600 SW Archer Road, Gainesville, FL 32610, USA.
authors.forename: Nan
authors.initials: N
authors.lastname: Li
authors.forename: Maria C
authors.initials: MC
authors.lastname: Quidgley
authors.forename: Firas H
authors.initials: FH
authors.lastname: Kobeissy
authors.forename: Jessica
authors.initials: J
authors.lastname: Joseph
authors.forename: Josef
authors.initials: J
authors.lastname: Neu
mesh_terms: D017209:Apoptosis; D018938:Caco-2 Cells; D054357:Chemokine CXCL10 / Q000378:metabolism; D015536:Down-Regulation; D004847:Epithelial Cells / Q000276:immunology; D004926:Escherichia coli; D053200:Fas-Associated Death Domain Protein / Q000378:metabolism; D005408:Flagellin / Q000276:immunology*; D020869:Gene Expression Profiling; D006801:Humans; D020672:I-kappa B Proteins / Q000378:metabolism; D007249:Inflammation; D053592:Interleukin-1 Receptor-Associated Kinases / Q000378:metabolism; D053583:Interleukin-1beta / Q000378:metabolism; D016209:Interleukin-8 / Q000096:biosynthesis*; D007413:Intestinal Mucosa / Q000276:immunology* / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D008070:Lipopolysaccharides / Q000276:immunology*; D053594:Myeloid Differentiation Factor 88 / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D016760:Proto-Oncogene Proteins c-fos / Q000378:metabolism; D015398:Signal Transduction; D013211:Staphylococcus aureus; D013682:Teichoic Acids / Q000276:immunology*; D051195:Toll-Like Receptor 2 / Q000378:metabolism; D051196:Toll-Like Receptor 3 / Q000378:metabolism; D051193:Toll-Like Receptors / Q000378:metabolism*; D015854:Up-Regulation
chemical_list: C114753:CXCL10 protein, human; D054357:Chemokine CXCL10; C505533:FADD protein, human; D053200:Fas-Associated Death Domain Protein; D020672:I-kappa B Proteins; D053583:Interleukin-1beta; D016209:Interleukin-8; D047908:Intracellular Signaling Peptides and Proteins; D008070:Lipopolysaccharides; C507779:MYD88 protein, human; D053594:Myeloid Differentiation Factor 88; D016328:NF-kappa B; D016760:Proto-Oncogene Proteins c-fos; C495342:TLR2 protein, human; C495344:TLR3 protein, human; D013682:Teichoic Acids; D051195:Toll-Like Receptor 2; D051196:Toll-Like Receptor 3; D051193:Toll-Like Receptors; D005408:Flagellin; C009900:lipoteichoic acid; C117090:MAP4K4 protein, human; C507771:IRAK3 protein, human; D053592:Interleukin-1 Receptor-Associated Kinases; D017346:Protein Serine-Threonine Kinases