title: Endotoxin triggers nuclear factor-kappaB-dependent up-regulation of multiple proinflammatory genes in the diaphragm.
abstract: BACKGROUND
  Sepsis-induced diaphragmatic force loss and failure are associated with an increased exposure of the muscle to proinflammatory mediators.
  
  
  OBJECTIVE
  Our objectives were to test the hypothesis that force-inhibiting mediators may arise in large part from the diaphragm itself and to evaluate the roles of mechanical stress, free radicals, and the nuclear factor (NF)-kappaB transcription factor pathway in endotoxin (LPS)-induced proinflammatory responses of the diaphragm.
  
  
  METHODS
  Murine diaphragm and limb muscle cells were exposed to LPS in vitro and in vivo. Proinflammatory gene expression was measured using RNase protection assays (tumor necrosis factor [TNF]-alpha, TNF-alpha receptor p55, interleukin [IL]-1alpha, IL-1beta, IL-6, macrophage inflammatory peptide-2, intercellular adhesion molecule-1, Fas ligand, and inducible nitric oxide synthase) and ELISAs (TNF-alpha, IL-6, and macrophage inflammatory peptide-2). Cyclical muscle cell stretch and free-radical scavengers (N-acetylcysteine and catalase) were used to alter mechanical and oxidative stress levels, respectively. Pharmacologic (pyrrolidinedithiocarbamate) and dominant-negative transfection strategies were used to inhibit the NF-kappaB pathway.
  
  
  RESULTS
  In primary diaphragm muscle cell cultures, modulation of mechanical stress levels or free-radical exposure did not alter responses to LPS stimulation. However, pharmacologic blockade of the NF-kappaB pathway and dominant-negative molecular inhibition of IKB kinase-beta strongly suppressed LPS-induced proinflammatory gene expression. In vivo, acute endotoxemia induced significantly greater mRNA and protein levels for proinflammatory mediators in the diaphragm as compared with limb muscle. Basal expression levels of proinflammatory genes were significantly higher in the diaphragm.
  
  
  CONCLUSIONS
  Constitutive and LPS-induced proinflammatory gene expression are exaggerated in the diaphragm compared with limb muscles and are critically dependent on the NF-kappaB pathway. We suggest the diaphragm may be relatively predisposed to proinflammatory responses.
authors.affiliation: Meakins-Christie Laboratories, McGill University; and Respiratory Division, McGill University Health Center, Montreal, Quebec, Canada.
authors.forename: Alexandre
authors.initials: A
authors.lastname: Demoule
authors.forename: Maziar
authors.initials: M
authors.lastname: Divangahi
authors.forename: Linda
authors.initials: L
authors.lastname: Yahiaoui
authors.forename: Gawiyou
authors.initials: G
authors.lastname: Danialou
authors.forename: Dusanka
authors.initials: D
authors.lastname: Gvozdic
authors.forename: Katherine
authors.initials: K
authors.lastname: Labbe
authors.forename: Weisheng
authors.initials: W
authors.lastname: Bao
authors.forename: Basil J
authors.initials: BJ
authors.lastname: Petrof
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D016207:Cytokines / Q000235:genetics*; D003964:Diaphragm / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D019446:Endotoxemia / Q000139:chemically induced / Q000378:metabolism* / Q000473:pathology; D004797:Enzyme-Linked Immunosorbent Assay; D066298:In Vitro Techniques; D008070:Lipopolysaccharides / Q000633:toxicity; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D016328:NF-kappa B / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics*; D017382:Reactive Oxygen Species / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects*
chemical_list: D016207:Cytokines; D008070:Lipopolysaccharides; D016328:NF-kappa B; D012333:RNA, Messenger; D017382:Reactive Oxygen Species