title: Lipopolysaccharide modulation of normal enterocyte turnover by toll-like receptors is mediated by endogenously produced tumour necrosis factor alpha.
abstract: BACKGROUND
  Circulating levels of endotoxin (or lipopolysaccharide (LPS)) and anti-endotoxin antibodies are increased in patients with inflammatory bowel disease, supporting the hypothesis of a role for endogenous bacterial products in the pathogenesis of these disorders.
  
  
  OBJECTIVE
  The aim of this study was to analyse the direct effects of LPS on intestinal epithelial cell turnover.
  
  
  RESULTS
  LPS significantly inhibited growth of the human non-transformed immature crypt cell line (HIEC), whereas IEC-6 cell proliferation was stimulated by LPS. As LPS is a physiological inducer of tumour necrosis factor alpha (TNFalpha) in various cell systems and this cytokine exerted similar anti-proliferative (HIEC) or growth stimulatory (IEC-6 cells) effects, the study thus tested the hypothesis that endogenously produced TNFalpha in response to LPS mediates this growth modulatory effect in an autoparacrine/paracrine way. Therefore, during LPS stimulation, the biological activity of TNFalpha was blocked using neutralising anti-TNFalpha antibodies, as well as inhibitory, antagonistic antibodies directed against the p55 TNF receptor, signalling the antimitotic TNFalpha effect in HIEC. Both experimental approaches completely abolished the growth modulatory effects of LPS in HIEC/IEC-6 cells. Production and secretion of TNFalpha by HIEC/IEC-6 cells in response to LPS was confirmed on mRNA and protein level by reverse transcription polymerase chain reaction (RT-PCR) and enzyme linked immunosorbent assay. LPS signalling was independent of CD14 in HIEC, as these cells lack this receptor. However, HIEC expressed TLR4 and MD2 resulting in a fully functional signalling complex as demonstrated by RT-PCR, western blot, and immunofluorescence analyses.
  
  
  CONCLUSIONS
  These results support the hypothesis that LPS induced changes of intestinal epithelial cell turnover may directly contribute to the pathogenesis of inflammatory epithelial cell lesions by endogenous TNFalpha production by enterocytes.
authors.affiliation: Laboratory of Gastrointestinal Immunology, Division of Gastroenterology, Department of Paediatrics, Children's Hospital Medical Centre, University of Bonn, Bonn, Germany. ruemmele@necker.fr
authors.forename: F M
authors.initials: FM
authors.lastname: Ruemmele
authors.forename: J F
authors.initials: JF
authors.lastname: Beaulieu
authors.forename: S
authors.initials: S
authors.lastname: Dionne
authors.forename: E
authors.initials: E
authors.lastname: Levy
authors.forename: E G
authors.initials: EG
authors.lastname: Seidman
authors.forename: N
authors.initials: N
authors.lastname: Cerf-Bensussan
authors.forename: M J
authors.initials: MJ
authors.lastname: Lentze
mesh_terms: D000911:Antibodies, Monoclonal / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D029721:Drosophila Proteins; D020895:Enterocytes / Q000187:drug effects / Q000276:immunology* / Q000473:pathology; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D015212:Inflammatory Bowel Diseases / Q000276:immunology* / Q000473:pathology; D007421:Intestine, Small; D018950:Lipopolysaccharide Receptors / Q000032:analysis / Q000378:metabolism; D008070:Lipopolysaccharides / Q000378:metabolism* / Q000494:pharmacology; D008562:Membrane Glycoproteins / Q000032:analysis / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D011956:Receptors, Cell Surface / Q000032:analysis / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000276:immunology*
chemical_list: D000911:Antibodies, Monoclonal; D029721:Drosophila Proteins; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; C495345:TLR4 protein, human; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors; D014409:Tumor Necrosis Factor-alpha