Proteinase-activated receptors (PARs), G protein-coupled receptors, play critical roles in the alimentary system. Increasing evidence suggests that endogenous prostaglandins (PGs) mediate some of PARs' gastrointestinal functions. Systemic administration of the PAR1 agonist protects against gastric mucosal injury through PG formation in rats. PGs also appear to contribute, at least in part, to enhancement of gastric mucosal blood flow and suppression of gastric acid secretion by PAR1 activation. There is also evidence for involvement of PGs in modulation of gastrointestinal motility by PAR1 or PAR2. Importantly, modulation of ion transport by PAR1 or PAR2 in the intestinal mucosal epithelium is largely mediated by PGs. Studies using gastric and intestinal mucosal epithelial cell lines imply that the PAR1-triggered formation of PGs involves multiple signaling pathways including Src, EGF receptor trans-activation and activation of MAP kinases. Collectively, a functional linkage of PAR1 and/or PAR2 to PGs is considered important in the gastrointestinal system.
Proteinase-activated receptor-1 (PAR1), upon activation, exerts prostanoid-dependent gastroprotection, and increases prostaglandin E(2) (PGE(2)) release through cyclooxygenase-2 (COX-2) upregulation in rat gastric mucosal epithelial RGM1 cells. However, there is a big time lag between the PAR1-triggered PGE(2) release and COX-2 upregulation in RGM1 cells; that is, the former event takes 18 h to occur, while the latter rapidly develops and reaches a plateau in 6 h. The present study thus aimed at clarifying mechanisms for the delay of PGE(2) release after PAR1 activation in RGM1 cells. Although a PAR1-activating peptide, TFLLR-NH(2), alone caused PGE(2) release at 18 h, but not 6 h, TFLLR-NH(2) in combination with arachidonic acid dramatically enhanced PGE(2) release even for 1-6 h. TFLLR-NH(2) plus linoleic acid caused a similar rapid response. CP-24879, a Δ(5)/Δ(6)-desaturase inhibitor, abolished the PGE(2) release induced by TFLLR-NH(2) plus linoleic acid, but not by TFLLR-NH(2) alone. The TFLLR-NH(2)-induced PGE(2) release was not affected by inhibitors of cytosolic phospholipase A(2) (cPLA(2)), Ca(2+)-independent PLA(2) (cPLA(2)) or secretory PLA(2) (sPLA(2)), but was abolished by their mixture or a pan-PLA(2) inhibitor. Among PLA(2) isozymes, mRNA of group IIA sPLA(2) (sPLA(2)-IIA) was upregulated following PAR1 stimulation for 6-18 h, whereas protein levels of PGE synthases were unchanged. These data suggest that the delay of PGE(2) release after COX-2 upregulation triggered by PAR1 is due to the poor supply of free arachidonic acid at the early stage in RGM1 cells, and that plural isozymes of PLA(2) including sPLA(2)-IIA may complementarily contribute to the liberation of free arachidonic acid.
We previously showed that activation of proteinase‐activated receptor‐1 (PAR1) caused delayed (18 h later) PGE2 release in rat gastric mucosal epithelial RGM1 cells, although the maximal up‐regulation of COX‐2 protein was observed much earlier (6 h later). In this study, we analyzed mechanisms for the delay of PGE2 release caused by PAR1 stimulation in RGM1 cells. Stimulation with a PAR1‐activating peptide, TFLLR‐NH2 (TF), in combination with arachidonic acid (AA) or linoleic acid (LA) induced rapid and synergistic increase in PGE2 release within 3 h. CP‐24879, a Δ5/Δ6‐desaturase inhibitor, suppressed the PGE2 release induced by TF plus LA, but not by TF plus AA or TF alone, at 18 h. The TF‐induced delayed PGE2 release was suppressed by combined application of inhibitors of cPLA2, iPLA2 and sPLA2, but not by each of them. Protein levels of microsomal PGE synthase (mPGES)‐1, mPGES‐2 and cytosolic PGES were unchanged by stimulation with TF. These data suggest that the delay of PAR1‐triggered PGE2 release is due to slowly developing AA production in RGM1 cells, and that three isozymes of PLA2 may complementarily contribute to the AA production.
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