Citrobacter rodentium is a bacterial pathogen that causes a murine infectious colitis equivalent to enterohemorrhagic Escherichia coli infection in humans. Colonic luminal fluid from C. rodentiuminfected mice, but not from sham-infected mice, contains active serine proteinases that can activate proteinase-activated receptor-2 (PAR2). We have identified granzyme A and murine trypsins to be present in C. rodentium-infected luminal fluid, as determined by mass spectrometry and Western blot analysis. Inflammatory indices (colonic mucosa macroscopic damage score, increased intestinal wall thickness, granulocyte infiltration, and bacterial translocation from the colonic lumen to peritoneal organs) were all increased in C. rodentium-infected mice, compared with shaminfected mice. Soybean trypsin inhibitor-treated wild-type mice and untreated PAR2-deficient (PAR2 ؊/؊ ) mice (compared with their wild-type littermates) both had substantially reduced levels of C. rodentium-induced inflammation. These data point to an important role for both pathogen-induced host serine proteinases and PAR2 in the setting of infectious colitis.colitis ͉ inflammation ͉ trypsin ͉ granzyme

Objective. To investigate the role of proteinaseactivated receptor 4 (PAR-4) in mediating joint inflammation and pain in mice.Methods. Knee joint blood flow, edema, and pain sensitivity (as induced by thermal and mechanical stimuli) were assessed in C57BL/6 mice following intraarticular injection of either the selective PAR-4 agonist AYPGKF-NH 2 or the inactive control peptide YAPGKF-NH 2 . The mechanism of action of AYPGKF-NH 2 was examined by pretreatment of each mouse with either the PAR-4 antagonist pepducin P4pal-10 or the bradykinin antagonist HOE 140. Finally, the role of PAR-4 in mediating joint inflammation was tested by pretreating mice with acutely inflamed knees with pepducin P4pal-10.Results. PAR-4 activation caused a long-lasting increase in joint blood flow and edema formation, which was not seen following injection of the control peptide. The PAR-4-activating peptide was also found to be pronociceptive in the joint, where it enhanced sensitivity to a noxious thermal stimulus and caused mechanical allodynia and hyperalgesia. The proinflammatory and pronociceptive effects of AYPGKF-NH 2 could be inhibited by pepducin P4pal-10 and HOE 140. Finally, pepducin P4pal-10 ameliorated the clinical and physiologic signs of acute joint inflammation.Conclusion. This study demonstrates that local activation of PAR-4 leads to proinflammatory changes in the knee joint that are dependent on the kallikreinkinin system. We also show for the first time that PARs are involved in the modulation of joint pain, with PAR-4 being pronociceptive in this tissue. Thus, blockade of articular PAR-4 may be a useful means of controlling joint inflammation and pain.

Background and purpose: Changes in extracellular fluid osmolarity, which occur after tissue damage and disease, cause inflammation and maintain chronic inflammatory states by unknown mechanisms. Here, we investigated whether the osmosensitive channel, transient receptor potential vanilloid 4 (TRPV4), mediates inflammation to hypotonic stimuli by a neurogenic mechanism. Experimental approach: TRPV4 was localized in dorsal root ganglia (DRG) by immunofluorescence. The effects of TRPV4 agonists on release of pro-inflammatory neuropeptides from peripheral tissues and on inflammation were examined. Key results: Immunoreactive TRPV4 was detected in DRG neurones innervating the mouse hindpaw, where it was co-expressed in some neurones with CGRP and substance P, mediators of neurogenic inflammation. Hypotonic solutions and 4a-phorbol 12,13-didecanoate, which activate TRPV4, stimulated neuropeptide release in urinary bladder and airways, sites of neurogenic inflammation. Intraplantar injection of hypotonic solutions and 4a-phorbol 12,13-didecanoate caused oedema and granulocyte recruitment. These effects were inhibited by a desensitizing dose of the neurotoxin capsaicin, antagonists of CGRP and substance P receptors, and TRPV4 gene knockdown or deletion. In contrast, antagonism of neuropeptide receptors and disruption of TRPV4 did not prevent this oedema. TRPV4 gene knockdown or deletion also markedly reduced oedema and granulocyte infiltration induced by intraplantar injection of formalin. Conclusions and implications: Activation of TRPV4 stimulates neuropeptide release from afferent nerves and induces neurogenic inflammation. This mechanism may mediate the generation and maintenance of inflammation after injury and during diseases, in which there are changes in extracellular osmolarity. Antagonism of TRPV4 may offer a therapeutic approach for inflammatory hyperalgesia and chronic inflammation.

Thus, as opposed to a previously shown proinflammatory role for PAR1 in a TH1 cytokine-mediated colitis, our new data show anti-inflammatory role for PAR1 activation in the setting of TH2 cytokine colitis model.

No abstract