In vitro and ex vivo activation of the TLR5 signaling pathway in intestinal epithelial cells by a commensal Escherichia coli strain.
The capacity of non-pathogenic enteric bacteria to induce a pro-inflammatory response is under debate in terms of its effect on the symbiosis between the mammalian host and its commensal gut microflora. Activation of NF-kappaB and induction of interleukin-8 (IL-8) and CCL-20 by the commensal Escherichia coli strain MG1655 were first studied in vitro in the human intestinal epithelial cell (IECs) lines HT29-19A and Caco-2, transfected or not with plasmids encoding dominant negative Toll-like receptor (TLR) 5 and myeloid differentiation factor-88 (MyD88) adaptor protein. The response of enterocytes in situ was then assessed using murine ileal biopsies mounted in Ussing chambers. Commensal E. coli induced NF-kappaB DNA binding, NF-kappaB transcriptional activity, CCL-20 expression, and IL-8 secretion in the human IEC lines. E. coli MG1655 flagellin was necessary and sufficient to trigger this pro-inflammatory pathway via its interaction with TLR5 and the subsequent recruitment of the adaptor protein MyD88. Following epithelial cell polarization, signaling could be induced by live E. coli and flagellin on the apical side of HT29-19A. The in vivo relevance of our findings was confirmed, because immunohistochemical staining of murine ileum demonstrated expression of TLR5 in the apical part of enterocytes in situ. Furthermore, flagellin added on the mucosal side of murine ileal biopsies mounted in Ussing chambers induced a basolateral production of KC, a functional murine homolog of human IL-8. These findings provide strong evidence that flagellin released by flagellated commensal bacteria in the intestinal lumen can induce a pro-inflammatory response in enterocytes in vivo.
INSERM, EMI-0212, 156 rue de Vaugirard, 75015 Paris, France.
Jean-Christophe
JC
Bambou
Antoine
A
Giraud
Sandrine
S
Menard
Bernadette
B
Begue
Sabine
S
Rakotobe
Martine
M
Heyman
François
F
Taddei
Nadine
N
Cerf-Bensussan
Valérie
V
Gaboriau-Routhiau
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D000943:Antigens, Differentiation / Q000378:metabolism; D001706:Biopsy; D018938:Caco-2 Cells; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D054418:Chemokine CCL20; D019742:Chemokines, CC / Q000378:metabolism; D017077:Culture Media, Conditioned / Q000494:pharmacology; D016207:Cytokines / Q000378:metabolism; D020895:Enterocytes / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D004847:Epithelial Cells / Q000166:cytology; D004926:Escherichia coli / Q000378:metabolism*; D005408:Flagellin / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D007082:Ileum / Q000473:pathology; D007249:Inflammation; D016209:Interleukin-8 / Q000378:metabolism; D007413:Intestinal Mucosa / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000378:metabolism; D019402:Macrophage Inflammatory Proteins / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000378:metabolism*; D051379:Mice; D008809:Mice, Inbred C3H; D009154:Mutation; D053594:Myeloid Differentiation Factor 88; D016328:NF-kappa B / Q000378:metabolism; D010641:Phenotype; D010957:Plasmids / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000378:metabolism*; D011971:Receptors, Immunologic / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D013997:Time Factors; D051198:Toll-Like Receptor 5; D051193:Toll-Like Receptors; D014158:Transcription, Genetic; D014162:Transfection
D048868:Adaptor Proteins, Signal Transducing; D000943:Antigens, Differentiation; C108355:CCL20 protein, human; D054418:Chemokine CCL20; D019742:Chemokines, CC; D017077:Culture Media, Conditioned; D016207:Cytokines; D016209:Interleukin-8; D008070:Lipopolysaccharides; C507779:MYD88 protein, human; D019402:Macrophage Inflammatory Proteins; D008562:Membrane Glycoproteins; C507780:Myd88 protein, mouse; D053594:Myeloid Differentiation Factor 88; D016328:NF-kappa B; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D011971:Receptors, Immunologic; C495346:TLR5 protein, human; D051198:Toll-Like Receptor 5; D051193:Toll-Like Receptors; D005408:Flagellin