Effect Of Protease Activated Receptor-2Agonist On Gastrointestinal Motility In Mice

Objective:Proteinase-activated receptor-2(PAR-2) is a G protein-coupledreceptor with seven trans-membrane domain, PAR-2could be activated bytrypsin and tryptase,PAR-2could be activated by SLIGRL-NH2andSLIGKV-NH2too.it was highly expressed throughout the gut,correlated withionic transport of endothelial cell,viscera sensitivity and regulation ofgastrointestinal motility,but the specific mechanism was not identify.in thisexperiment,the expression of PAR-2mRNA and PAR-2protein in the gut（stomach,duodenum,jejunum and ileum）were be detected,the effect ofPAR-2agonist on gastrointestinal motility in mice was be investigated invivo.the effect and mechanism of PAR-2on contraction of isolated ileumlongitudinal strips in mice was also be emphasize study in vitro.Methods:1the level of PAR-2mRNA was tested by RT-PCR,theimmunore-activity of PAR-2was examined by IHC.2In vivo:120BABL/c female mices were randomly divided into sixgroups: control group,amastain group,SLIGRL-NH2(1μmol/kg)+amastaingroup, SLIGRL-NH2(2.5μmol/kg)+amastain group,SLIGRL-NH2(5μmol/kg)+amastain group,LRGILS-NH2(5μmol/kg)+amastain group.Anddetermine the gastric emptying rate and intestinal propulsion rate of eachgroup.3In vitro:to tested the influence in the contraction amplitude of ileumlongitudinal strips of trypsin（10-8-10-4M）,SLIGRL-NH（-8210-10-4M）and otherrelevant reagents(atropine,TTX,capsaicin,NK1antagon,NK2antagon).Results:1PAR-2mRNA was expressed throughout the gut（stomach,duodenum,jejunum and ileum）,PAR-2positive immunore-activity was present in mucous layer,submucous layer and muscular layer of gut（stomach,duodenum,jejunumand ileum）,cell membranes were positive expression；2in vivo:PAR-2agonist effect on the gastric emptying rate and intestinalpropulsion rate.2.1amastain group did not affect the gastric emptying rate and intestinalpropulsion rate compaired with control group (p>0.05).2.2the gastric emptying rate and intestinal propulsion rate of SLIGRL-NH2(2.5μmol/kg,5μmol/kg)+amastain group were markedly increased compairedwith control group(p<0.001).2.3LRGILS-NH2(5μmol/kg)+amastain group did not affect the gastricemptying rate and intestinal propulsion rate compaired with controlgroup(p>0.05).3in vitro:the influence of PAR-2agonist and relevant reagents in thecontraction amplitude of ileum longitudinal strips3.1the PAR-2agnoist （trysin,SLRGIL-NH2）could evoked dependentcontractions of ileum longitudinal strips.LRGILS-NH-52（10-9-10M）was nosignificant effect on the contraction amplitude of isolated ileum longitudinalstrips.3.2atropine was no significant enhancement effect on PAR-2agonist-inducedcontraction（p>0.05）.3.3TTX effect on the contraction of PAR-2agonist-induced:TTX（2uM）+trypsin group could apparently depress the contraction amplitude of isolatedileum longitudinal strips compaired with trypsin group（p<0.05）,TTX（2uM）+SLRGIL-NH2group could apparently depress the contraction amplitude ofisolated ileum longitudinal strips compaired with SLRGIL-NH2group（p<0.05）.34capsaicin effect on the contraction of PAR-2agonist-induced:capsaicin（10-4M）+trypsin group could apparently depress the contraction amplitudeof isolated ileum longitudinal strips compaired with trypsin group（p<0.05）,capsaicin（10-4M）+SLRGIL-NH2group could apparently depressthe contraction amplitude of isolated ileum longitudinal strips compaired with SLRGIL-NH2group（p<0.05）.3.5NK1antagon and NK2antagons effect on the contraction of PAR-2agonist-induced:sc-203095（1uM）+trypsin group could apparently depress thecontraction amplitude of isolated ileum longitudinal strips compaired withtrypsin group （p<0.05）,MEN10376（1uM）+trypsin group could alsoapparently depress the contraction amplitude of isolated ileum longitudinalstrips compaired with trypsin group（p<0.05）,sc-203095（1uM）+MEN10376（1uM）+trypsin group could also apparently depress the contractionamplitude of isolated ileum longitudinal strips compaired with trypsin group（p<0.05）,sc-203095（1uM）+SLIGRL-NH2group could apparently depressthe contraction amplitude of isolated ileum longitudinal strips compaired withSLIGRL-NH2group（p<0.05）,MEN10376（1uM）+SLIGRL-NH2group couldalso apparently depress the contraction amplitude of isolated ileumlongitudinal strips compaired with SLIGRL-NH2group（p<0.05）,sc-203095（1uM）+MEN10376（1uM）+SLIGRL-NH2group could also apparentlydepress the contraction amplitude of isolated ileum longitudinal stripscompaired with SLIGRL-NH2group（p<0.001）.Conclusion:1.PAR-2was highly expressed throughout the gut（stomach,duodenum,jejunum and ileum）.2.activation of PAR-2could markedly promote the gastrointestinal motility inmice in vivo.3.PAR-2agnoist has apparent excitatory effect on contraction of isolated ileumlongitudinal strips in vitro,We concluded this effect was mediated by nerves,the excitatory effect is also dependent on TRPV1sensitive sensory neural,andrequires both NK1and NK2receptors.