title: Protease-activated receptor 2, dipeptidyl peptidase I, and proteases mediate Clostridium difficile toxin A enteritis.
abstract: OBJECTIVE
  We studied the role of protease-activated receptor 2 (PAR(2)) and its activating enzymes, trypsins and tryptase, in Clostridium difficile toxin A (TxA)-induced enteritis.
  
  
  METHODS
  We injected TxA into ileal loops in PAR(2) or dipeptidyl peptidase I (DPPI) knockout mice or in wild-type mice pretreated with tryptase inhibitors (FUT-175 or MPI-0442352) or soybean trypsin inhibitor. We examined the effect of TxA on expression and activity of PAR(2) and trypsin IV messenger RNA in the ileum and cultured colonocytes. We injected activating peptide (AP), trypsins, tryptase, and p23 in wild-type mice, some pretreated with the neurokinin 1 receptor antagonist SR140333.
  
  
  RESULTS
  TxA increased fluid secretion, myeloperoxidase activity in fluid and tissue, and histologic damage. PAR(2) deletion decreased TxA-induced ileitis, reduced luminal fluid secretion by 20%, decreased tissue and fluid myeloperoxidase by 50%, and diminished epithelial damage, edema, and neutrophil infiltration. DPPI deletion reduced secretion by 20% and fluid myeloperoxidase by 55%. In wild-type mice, FUT-175 or MPI-0442352 inhibited secretion by 24%-28% and tissue and fluid myeloperoxidase by 31%-71%. Soybean trypsin inhibitor reduced secretion to background levels and tissue myeloperoxidase by up to 50%. TxA increased expression of PAR(2) and trypsin IV in enterocytes and colonocytes and caused a 2-fold increase in Ca(2+) responses to PAR(2) AP. AP, tryptase, and trypsin isozymes (trypsin I/II, trypsin IV, p23) caused ileitis. SR140333 prevented AP-induced ileitis.
  
  
  CONCLUSIONS
  PAR(2) and its activators are proinflammatory in TxA-induced enteritis. TxA stimulates existing PAR(2) and up-regulates PAR(2) and activating proteases, and PAR(2) causes inflammation by neurogenic mechanisms.
authors.affiliation: Center for the Neurobiology of Digestive Disease, Department of Surgery, University of California, San Francisco, San Francisco, California 94143-0660, USA.
authors.forename: Graeme S
authors.initials: GS
authors.lastname: Cottrell
authors.forename: Silvia
authors.initials: S
authors.lastname: Amadesi
authors.forename: Stella
authors.initials: S
authors.lastname: Pikios
authors.forename: Eric
authors.initials: E
authors.lastname: Camerer
authors.forename: J Adam
authors.initials: JA
authors.lastname: Willardsen
authors.forename: Brett R
authors.initials: BR
authors.lastname: Murphy
authors.forename: George H
authors.initials: GH
authors.lastname: Caughey
authors.forename: Paul J
authors.initials: PJ
authors.lastname: Wolters
authors.forename: Shaun R
authors.initials: SR
authors.lastname: Coughlin
authors.forename: Anders
authors.initials: A
authors.lastname: Peterson
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Knecht
authors.forename: Charalabos
authors.initials: C
authors.lastname: Pothoulakis
authors.forename: Nigel W
authors.initials: NW
authors.lastname: Bunnett
authors.forename: Eileen F
authors.initials: EF
authors.lastname: Grady
mesh_terms: D000818:Animals; D001427:Bacterial Toxins / Q000494:pharmacology; D020789:Cathepsin C / Q000172:deficiency / Q000378:metabolism*; D002478:Cells, Cultured; D003106:Colon / Q000166:cytology / Q000378:metabolism; D004751:Enteritis / Q000139:chemically induced* / Q000209:etiology / Q000378:metabolism / Q000473:pathology; D004768:Enterotoxins / Q000494:pharmacology; D006098:Granulocytes / Q000473:pathology; D007079:Ileitis / Q000209:etiology; D007082:Ileum / Q000378:metabolism; D007413:Intestinal Mucosa / Q000378:metabolism; D007422:Intestines / Q000473:pathology; D051379:Mice; D018345:Mice, Knockout; D009420:Nervous System / Q000378:metabolism; D010447:Peptide Hydrolases / Q000378:metabolism*; D009195:Peroxidase / Q000378:metabolism; D044464:Receptor, PAR-2 / Q000172:deficiency / Q000378:metabolism*; D018040:Receptors, Neurokinin-1 / Q000378:metabolism; D014357:Trypsin / Q000378:metabolism; D014361:Trypsin Inhibitors / Q000494:pharmacology; D053802:Tryptases / Q000494:pharmacology; D015854:Up-Regulation
chemical_list: D001427:Bacterial Toxins; D004768:Enterotoxins; D044464:Receptor, PAR-2; D018040:Receptors, Neurokinin-1; D014361:Trypsin Inhibitors; C041348:tcdA protein, Clostridium difficile; D009195:Peroxidase; D010447:Peptide Hydrolases; D020789:Cathepsin C; C040749:PRSS3 protein, human; D014357:Trypsin; D053802:Tryptases