Abstract
Many trypsin-like serine proteases such as β-tryptase are involved in the pathogenesis of colitis and inflammatory bowel diseases. Inhibitors of individual proteases show limited efficacy in treating such conditions, but also probably disrupt digestive and defensive functions of proteases. Here, we investigate whether masking their common target, protease-activated receptor 2 (PAR2), is an effective therapeutic strategy for treating acute and chronic experimental colitis in rats. A novel PAR2 antagonist (5-isoxazoyl-Cha-Ile-spiro[indene-1,4′-piperidine]; GB88) was evaluated for the blockade of intracellular calcium release in colonocytes and anti-inflammatory activity in acute (PAR2 agonist-induced) versus chronic [2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced] models of colitis in Wistar rats. Disease progression (disease activity index, weight loss, and mortality) and postmortem colonic histopathology (inflammation, bowel wall thickness, and myeloperoxidase) were measured. PAR2 and tryptase colocalization were investigated by using immunohistochemistry. GB88 was a more potent antagonist of PAR2 activation in colonocytes than another reported compound, N1-3-methylbutyryl-N4-6-aminohexanoyl-piperazine (ENMD-1068) (IC50 8 μM versus 5 mM). Acute colonic inflammation induced in rats by the PAR2 agonist SLIGRL-NH2 was inhibited by oral administration of GB88 (10 mg/kg) with markedly reduced edema, mucin depletion, PAR2 receptor internalization, and mastocytosis. Chronic TNBS-induced colitis in rats was ameliorated by GB88 (10 mg/kg/day p.o.), which reduced mortality and pathology (including colon obstruction, ulceration, wall thickness, and myeloperoxidase release) more effectively than the clinically used drug sulfasalazine (100 mg/kg/day p.o.). These disease-modifying properties for the PAR2 antagonist in both acute and chronic experimental colitis strongly support a pathogenic role for PAR2 and PAR2-activating proteases and therapeutic potential for PAR2 antagonism in inflammatory diseases of the colon.
Footnotes
Funding was provided by the Australian National Health and Medical Research Council [Grants 569595, 1000745]; the Australian Research Council [Grants FF668733, DP1093245]; and an Australian Research Council Federation Fellowship (to D.P.F.).
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
↵ The online version of this article (available at http://jpet.aspetjournals.org) contains supplemental material.
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ABBREVIATIONS:
- CD
- Crohn's disease
- UC
- ulcerative colitis
- IBD
- inflammatory bowel disease
- PAR2
- protease-activated receptor 2
- TNBS
- 2,4,6-trinitrobenzenesulfonic acid
- ANOVA
- analysis of variance
- ENMD-1068
- N1-3-methylbutyryl-N4-6-aminohexanoyl-piperazine
- MPO
- myeloperoxidase
- DAI
- disease activity index
- H&E
- hematoxylin and eosin
- ELISA
- enzyme-linked immunosorbent assay
- GB88
- 5-isoxazoyl-Cha-Ile-spiro[indene-1,4′-piperidine].
- Received August 24, 2011.
- Accepted October 24, 2011.
- Copyright © 2012 by The American Society for Pharmacology and Experimental Therapeutics
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