title: Increased expression and function of integrins in enterocytes by endotoxin impairs epithelial restitution.
abstract: OBJECTIVE
  Experimental necrotizing enterocolitis (NEC) is characterized by circulating endotoxin (lipopolysaccharide [LPS]) and impaired enterocyte migration. We hypothesized that LPS increases integrin function and cell-matrix adhesion, leading to impaired enterocyte migration in the pathogenesis of NEC.
  
  
  METHODS
  NEC-like intestinal injury was induced in newborn rats by hypoxia/gavage feedings, and restitution was determined by assessing bromodeoxyuridine-labeled enterocytes along the crypt-villus axis. Newborn mice were injected with 5 mg/kg LPS. IEC-6 cells were treated with LPS +/- LY294002 or wortmannin, and beta 1- and alpha 3-integrins were assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunofluorescence. Beta 1-integrin function was determined by adherence of fibronectin beads to IEC-6 monolayers. Migration of IEC-6 cells into a scraped wound was measured by time-lapse microscopy.
  
  
  RESULTS
  Newborn intestinal injury was associated with decreased intestinal restitution and increased alpha 3- and beta 1-integrin expression in the ileal mucosa, which also was observed after LPS injection. In IEC-6 cells, LPS caused an increase in the expression of alpha 3- and beta 1-integrins, a shift of beta 1-integrins from the cytoplasm to the plasma membrane and an increase in fibronectin bead adhesion during which beta 1-integrins accumulated underneath attached beads. These effects could be reversed with LY294002 or wortmannin, suggesting phosphatidylinositol-3-phosphate kinase (PI3K) dependence. The increased integrin-matrix adhesion by LPS led to an inhibition of enterocyte migration, which could be reversed by anti-beta 1-antibodies.
  
  
  CONCLUSIONS
  Enterocyte migration is inhibited by LPS through increased expression and function of alpha 3- and beta 1-integrins. Modulation of enterocyte migration via integrins may provide novel insights into the pathogenesis of NEC, in which intestinal restitution is impaired.
authors.affiliation: Division of Pediatric Surgery, Children's Hospital Pittsburgh, University of Pittsburgh, PA 15213, USA.
authors.forename: Faisal G
authors.initials: FG
authors.lastname: Qureshi
authors.forename: Cynthia
authors.initials: C
authors.lastname: Leaphart
authors.forename: Selma
authors.initials: S
authors.lastname: Cetin
authors.forename: Jun
authors.initials: J
authors.lastname: Li
authors.forename: Anatoly
authors.initials: A
authors.lastname: Grishin
authors.forename: Simon
authors.initials: S
authors.lastname: Watkins
authors.forename: Henri R
authors.initials: HR
authors.lastname: Ford
authors.forename: David J
authors.initials: DJ
authors.lastname: Hackam
mesh_terms: D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002465:Cell Movement / Q000187:drug effects; D002867:Chromones / Q000494:pharmacology; D004731:Endotoxins / Q000494:pharmacology*; D020345:Enterocolitis, Necrotizing / Q000378:metabolism / Q000503:physiopathology*; D020895:Enterocytes / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D019012:Integrin beta1 / Q000378:metabolism; D016023:Integrins / Q000378:metabolism*; D007413:Intestinal Mucosa / Q000378:metabolism / Q000503:physiopathology*; D051379:Mice; D009025:Morpholines / Q000494:pharmacology; D051381:Rats; D000077191:Wortmannin; D014945:Wound Healing / Q000187:drug effects*
chemical_list: D000730:Androstadienes; D002867:Chromones; D004731:Endotoxins; D004791:Enzyme Inhibitors; D019012:Integrin beta1; D016023:Integrins; D009025:Morpholines; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D000077191:Wortmannin