PAR2 activation alters colonic paracellular permeability in mice via IFN-gamma-dependent and -independent pathways.
Activation of colonic proteinase-activated receptor-2 (PAR(2)) caused inflammation and increased mucosal permeability in mouse colon. The present study was aimed at characterizing the possible links between these two phenomena. We evaluated the effects of intracolonic infusion of PAR(2)-activating peptide, SLIGRL, on colonic paracellular permeability and inflammation at two different doses, 5 and 100 microg per mouse, in an attempt to discriminate between both PAR(2)-mediated effects. We further investigated the possible involvement of interferon gamma (IFN-gamma) and calmodulin-dependent activation of myosin light chain kinase (MLCK), and alterations of zonula occludens-1 (ZO-1) localization in PAR(2)-induced responses. Thus, at the lower dose, SLIGRL increased colonic permeability without causing inflammation. Western blotting showed phosphorylation of mucosal myosin light chain (MLC) expression after both doses of SLIGRL. Moreover, while the MLCK inhibitor, ML-7, abolished the permeability effects of the low dose of SLIGRL, it only partially inhibited that of the high dose. In IFN-gamma-deficient mice (B6 ifng(-/-)), the increases in permeability were similar for both doses of SLIGRL and prevented by ML-7. In addition, MLCK immunoprecipitation revealed an increase of calmodulin binding to MLCK in the mucosa of mice treated with either dose of SLIGRL. Finally, we have shown that direct activation of PAR(2) on enterocytes is responsible for increased permeability and ZO-1 disruption. Moreover, SLIGRL at a dose that does not produce inflammation increases permeability via calmodulin activation, which binds and activates MLCK. The resulting tight junction opening does not depend upon IFN-gamma secretion, while the increased permeability in response to the high dose of PAR(2) agonist involves IFN-gamma secretion.
Cenac
Nicolas
N
INRA, Neuro-Gastroenterology and Nutrition Unit, 180 Chemin de Tournefeuille, BP.3, 31931 Toulouse cedex 9, France.
Chin
Alex C
AC
Garcia-Villar
Rafael
R
Salvador-Cartier
Christel
C
Ferrier
Laurent
L
Vergnolle
Nathalie
N
Buret
André G
AG
Fioramonti
Jean
J
Bueno
Lionel
L
D000818:Animals; D002463:Cell Membrane Permeability / Q000187:drug effects; D002478:Cells, Cultured; D003106:Colon / Q000166:cytology / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D007371:Interferon-gamma / Q000096:biosynthesis / Q000172:deficiency*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009842:Oligopeptides / Q000494:pharmacology; D044464:Receptor, PAR-2 / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*
D009842:Oligopeptides; D044464:Receptor, PAR-2; C099570:seryl-leucyl-isoleucyl-glycyl-arginyl-leucine; D007371:Interferon-gamma