title: Clostridium difficile toxin A triggers human colonocyte IL-8 release via mitochondrial oxygen radical generation.
abstract: OBJECTIVE
  Clostridium difficile toxin A causes mitochondrial dysfunction resulting in generation of oxygen radicals and adenosine triphosphate (ATP) depletion. We investigated whether mitochondrial dysfunction is involved in nuclear factor kappaB (NF-kappaB) activation and interleukin (IL)-8 release from toxin A-exposed enterocytes.
  
  
  METHODS
  NF-kappaB activation and IL-8 release in response to toxin A were correlated with reactive oxygen intermediate (ROI) generation and ATP production in HT-29 monolayers or HT-29 cells exposed to ethidium bromide (EB) to inhibit mitochondrial function.
  
  
  RESULTS
  HT-29 cells exposed to EB showed damaged mitochondria and diminished resting levels of ATP. ROI production in EB-treated cells exposed to toxin A for 30 minutes was significantly reduced. Exposure of wild-type HT-29 cells to toxin A resulted in increased oxygen radical generation and IL-8 production (P < 0.01 vs. control) that was inhibited by antioxidant pretreatment. Degradation of IkappaB was observed within 30 minutes of toxin exposure, before ras homologue (Rho) glucosylation, and was followed by nuclear translocation of NF-kappaB. Toxin A did not increase IL-8 levels in EB-treated cells, whereas IL-8 release in response to IL-1beta was not affected.
  
  
  CONCLUSIONS
  Our data support an early role for mitochondria-derived ROIs in stimulation of IL-8 release from colonocytes by toxin A. ROI generation is independent of Rho inactivation and involves nuclear translocation of NF-kappaB before release of IL-8.
authors.affiliation: Division of Gastroenterology and Department of Surgery, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts 02215, USA.
authors.forename: Dan
authors.initials: D
authors.lastname: He
authors.forename: Stavros
authors.initials: S
authors.lastname: Sougioultzis
authors.forename: Susan
authors.initials: S
authors.lastname: Hagen
authors.forename: Jennifer
authors.initials: J
authors.lastname: Liu
authors.forename: Sarah
authors.initials: S
authors.lastname: Keates
authors.forename: Andrew C
authors.initials: AC
authors.lastname: Keates
authors.forename: Charalabos
authors.initials: C
authors.lastname: Pothoulakis
authors.forename: J Thomas
authors.initials: JT
authors.lastname: Lamont
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D001427:Bacterial Toxins / Q000494:pharmacology*; D004268:DNA-Binding Proteins / Q000378:metabolism; D004768:Enterotoxins / Q000494:pharmacology*; D006031:Glycosylation; D019073:HT29 Cells; D006801:Humans; D020672:I-kappa B Proteins; D016209:Interleukin-8 / Q000378:metabolism*; D008928:Mitochondria / Q000378:metabolism*; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D020741:rho GTP-Binding Proteins / Q000378:metabolism
chemical_list: D001427:Bacterial Toxins; D004268:DNA-Binding Proteins; D004768:Enterotoxins; D020672:I-kappa B Proteins; D016209:Interleukin-8; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D017382:Reactive Oxygen Species; C041348:tcdA protein, Clostridium difficile; D000072000:NF-KappaB Inhibitor alpha; D000255:Adenosine Triphosphate; D020741:rho GTP-Binding Proteins