Protease-activated receptor 2, dipeptidyl peptidase I, and proteases mediate Clostridium difficile toxin A enteritis.
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.
Cottrell
Graeme S
GS
Center for the Neurobiology of Digestive Disease, Department of Surgery, University of California, San Francisco, San Francisco, California 94143-0660, USA.
Amadesi
Silvia
S
Pikios
Stella
S
Camerer
Eric
E
Willardsen
J Adam
JA
Murphy
Brett R
BR
Caughey
George H
GH
Wolters
Paul J
PJ
Coughlin
Shaun R
SR
Peterson
Anders
A
Knecht
Wolfgang
W
Pothoulakis
Charalabos
C
Bunnett
Nigel W
NW
Grady
Eileen F
EF
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
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