Indomethacin may injure the small intestine through a TLR4/MyD88-dependent pathway.

We examined the roles of cyclooxygenase (COX) isozymes, prostaglandins (PGs), and their receptors in the mucosal defense against ischemia/reperfusion (I/R)-induced gastric lesions in mice. Male C57BL/6 mice, including wild-type animals and those lacking prostaglandin E 2 (EP)1, EP3, or prostaglandin I 2 (IP) receptors, were used after 18 h of fasting. Under urethane anesthesia, the celiac artery was clamped (ischemia) for 30 min, and then reperfusion was achieved for 60 min through the removal of the clamp, and the stomach was examined for lesions. I/R produced hemorrhagic gastric lesions in wild-type mice. The severity of lesions was significantly increased by pretreatment with indomethacin (a nonselective COX inhibitor) and rofecoxib (a selective COX-2 inhibitor) but not 5-(4-chlorophenyl)-1-(4-methoxyphenyl)-3-(trifluoromethyl)-1H-pyrazole (SC-560; a selective COX-1 inhibitor). The expression of COX-2 mRNA was up-regulated in the stomach following I/R but not by sham operation or ischemia alone. The ulcerogenic response was markedly aggravated in IP receptor knockout mice but not those lacking EP1 or EP3 receptors. I/R increased the levels of 6-keto-PGF 1␣ and PGE 2 in the stomach of wildtype mice, and this response was attenuated by indomethacin and rofecoxib but not SC-560. Pretreatment of wild-type mice with iloprost, a prostacyclin (PGI 2 ) analog, significantly prevented the I/R-induced gastric lesions in the absence and presence of indomethacin or rofecoxib. PGE 2 also reduced the severity of I/R-induced gastric lesions, yet the effect was much less pronounced than that of iloprost. These results suggest that endogenous PGs derived from COX-2 play a crucial role in gastric mucosal defense during I/R, and this action is mainly mediated by PGI 2 through the activation of IP receptors.

The effects of capsaicin and mucosal acidification on gastric HCO 3 Ϫ secretion were compared in wild-type and prostacyclin (PGI 2 ) IP receptor or prostaglandin E receptor EP1 or EP3 knockout C57BL/6 mice as well as rats. Under urethane anesthesia, the stomach was mounted on an ex vivo chamber, perfused with saline, and the secretion of HCO 3 Ϫ was measured at pH 7.0 using the pH-stat method. Capsaicin or 200 mM HCl was applied to the chamber for 10 min. Capsaicin increased the secretion of HCO 3 Ϫ in rats and wild-type mice, the response at 0.3 mg/ml being equivalent to that induced by acidification. This effect of capsaicin in rats was abolished by ablation of capsaicin-sensitive afferent neurons and attenuated by indo- Ϫ , and the effect of bradykinin was blocked by indomethacin and L-NAME as well as FR172357. The stimulatory effect of capsaicin disappeared in IP (Ϫ/Ϫ) mice, whereas that of acidification disappeared in EP1 (Ϫ/Ϫ) mice. Intragastric application of capsaicin increased mucosal PGI 2 but not PGE 2 levels in the rat stomach. These results suggested that both capsaicin and acid increase gastric HCO 3 Ϫ secretion via a common pathway, involving PG and NO as well as capsaicin-sensitive afferent neurons, yet their responses differ concerning TRPV1 or prostanoid receptor dependence.

Aim: We investigated the roles of NO/NOS isoforms in the pathogenesis of ischemia/reperfusion (I/R)-induced gastric injury in mouse stomachs. Methods: Under urethane anesthesia, the celiac artery was clamped, and then reperfusion was established 30 min later by removal of the clamp. After a 60-min reperfusion, the stomach was examined for macroscopic lesions. Results: Following I/R, hemorrhagic lesions were generated in the mucosa, although ischemia alone caused no visible damage. Prior administration of L-NAME (a nonselective NOS inhibitor) significantly aggravated these lesions, in a L-arginine-inhibitable manner. By contrast, the selective iNOS inhibitor 1400W significantly prevented the occurrence of I/R-induced gastric lesions. The mucosal MPO activity was increased after I/R, and this response was enhanced and attenuated by prior administration of L-NAME and 1400W, respectively. Interestingly, the later treatment with L-NAME, given 10 min before reperfusion, significantly reduced the severity of the I/R-induced gastric damage, in a L-arginine-dependent manner. The expression of iNOS mRNA was up-regulated in the stomach following I/R, with an increase of mucosal NO content, and the NO production was significantly inhibited by both L-NAME and 1400W. Conclusion: Endogenous NO plays a dual role in the pathogenesis of IR-induced gastric damage; NO/cNOS is protective while NO/iNOS is proulcerogenic during I/R.

Toll-like receptors (TLRs) recognize microbial components and trigger the signaling cascade that activates innate and adaptive immunity. Recent studies have shown that the activation of TLR-dependent signaling pathways plays important roles in the pathogenesis of ischemia-reperfusion (I/R) injuries in many organs. All TLRs, except TLR3, use a common adaptor protein, MyD88, to transduce activation signals. We investigated the role of MyD88 in I/R injury of the small intestine. MyD88 and cyclooxygenase-2 (COX-2) knockout and wild-type mice were subjected to intestinal I/R injury. I/R-induced small intestinal injury was characterized by infiltration of inflammatory cells, disruption of the mucosal epithelium, destruction of villi, and increases in myeloperoxidase activity and mRNA levels of TNF-α and the IL-8 homolog KC. MyD88 deficiency worsened the severity of I/R injury, as assessed using the histological grading system, measuring luminal contents of hemoglobin (a marker of intestinal bleeding), and counting apoptotic epithelial cells, while it inhibited the increase in mRNA expression of TNF-α and KC. I/R significantly enhanced COX-2 expression and increased PGE(2) concentration in the small intestine of wild-type mice, which were markedly inhibited by MyD88 deficiency. COX-2 knockout mice were also highly susceptible to intestinal I/R injury. Exogenous PGE(2) reduced the severity of injury in both MyD88 and COX-2 knockout mice to the level of wild-type mice. These findings suggest that the MyD88 signaling pathway may inhibit I/R injury in the small intestine by inducing COX-2 expression.