title: Intracolonic infusion of fecal supernatants from ulcerative colitis patients triggers altered permeability and inflammation in mice: role of cathepsin G and protease-activated receptor-4.
abstract: BACKGROUND
  Cathepsin G (Cat-G) is a neutrophil serine-protease found in the colonic lumen of ulcerative colitis (UC) patients. Cat-G is able to activate protease-activated receptor-4 (PAR(4) ) located at the apical side of enterocytes, leading to epithelial barrier disruption. However, the mechanisms through which Cat-G triggers inflammation are not fully elucidated. The aims of our study were to evaluate in vivo the effects of UC fecal supernatants and Cat-G on epithelial barrier function and inflammation, and the connection between these two parameters.
  
  
  METHODS
  Male balb/c mice were used in this study. We evaluated the effect of a 2-hour intracolonic infusion of 1) fecal supernatants from UC patients pretreated or not with specific Cat-G inhibitor (SCGI); 2) PAR(4) -activating peptide (PAR(4) -AP); and 3) Cat-G on colonic myeloperoxidase (MPO) activity and paracellular permeability (CPP). The involvement of PAR(4) was assessed by pretreating animals with pepducin P4pal-10, which blocks PAR(4) signaling. We investigated the role of myosin light chain (MLC) kinase by using its inhibitor, ML-7, and we determined phosphorylated MLC (pMLC) levels in mice colonic mucosa.
  
  
  RESULTS
  UC fecal supernatants, Cat-G, and PAR(4) agonist increased both CPP and MPO activity in comparison with healthy subjects fecal supernatants. ML-7 inhibited the CPP increase triggered by Cat-G by 92.3%, and the enhanced MPO activity by 43.8%. Intracolonic infusion of UC fecal supernatant determined an increased phosphorylation level of MLC.
  
  
  CONCLUSIONS
  These observations support that luminal factors such as Cat-G play an important proinflammatory role in the pathogenesis of colitis, mainly depending on CPP increase by MLC phosphorylation.
authors.affiliation: INRA, UMR 1054, Neuro-Gastroenterology & Nutrition Unit, Toulouse, France.
authors.forename: Marta
authors.initials: M
authors.lastname: Dabek
authors.forename: Laurent
authors.initials: L
authors.lastname: Ferrier
authors.forename: Anita
authors.initials: A
authors.lastname: Annahazi
authors.forename: Valérie
authors.initials: V
authors.lastname: Bézirard
authors.forename: Arnaud
authors.initials: A
authors.lastname: Polizzi
authors.forename: Christel
authors.initials: C
authors.lastname: Cartier
authors.forename: Mathilde
authors.initials: M
authors.lastname: Leveque
authors.forename: R
authors.initials: R
authors.lastname: Roka
authors.forename: Tibor
authors.initials: T
authors.lastname: Wittmann
authors.forename: Vassilia
authors.initials: V
authors.lastname: Theodorou
authors.forename: Lionel
authors.initials: L
authors.lastname: Bueno
mesh_terms: D000285:Administration, Rectal; D000293:Adolescent; D000328:Adult; D000368:Aged; D000818:Animals; D015153:Blotting, Western; D056649:Cathepsin G / Q000502:physiology*; D002463:Cell Membrane Permeability / Q000502:physiology; D003092:Colitis / Q000209:etiology* / Q000503:physiopathology; D003093:Colitis, Ulcerative / Q000209:etiology* / Q000503:physiopathology; D003106:Colon / Q000503:physiopathology; D005243:Feces; D006801:Humans; D007413:Intestinal Mucosa / Q000503:physiopathology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008875:Middle Aged; D009195:Peroxidase / Q000378:metabolism; D018179:Receptors, Thrombin / Q000502:physiology*; D055815:Young Adult
chemical_list: D018179:Receptors, Thrombin; D009195:Peroxidase; D056649:Cathepsin G; C112908:protease-activated receptor 4