Functional mechanism underlying cyclooxygenase-2 expression in rat small intestine following administration of indomethacin: relation to intestinal hypermotility

BACKGROUND AND AIM: We recently reported that cyclooxygenase (COX)-2 is upregulated in the rat small intestine after administration of indomethacin, and this may be the key to non-steroidal anti-inflammatory drug (NSAID)-induced intestinal damage. The present study investigated the mechanism for COX-2 expression induced in the rat small intestine by indomethacin, in relation with ulcerogenic processes. METHODS: Animals were given indomethacin or SC-560 p.o., and the intestinal mucosa was examined 24 h later. RESULTS: Indomethacin caused hemorrhagic lesions in the small intestine, accompanied with an increase in intestinal motility, bacterial invasion and inducible nitric oxide synthase (iNOS) activity, as well as the expression of COX-2 mRNA in the mucosa. Although SC-560 did not cause any damage, this agent caused intestinal hypermotility, the bacterial invasion and the upregulation of COX-2 expression. The mucosal PGE2 content was decreased by SC-560 at 3 h but recovered 12 h later, and this recovery of PGE2 was attenuated by both atropine and ampicillin, in addition to rofecoxib. The intestinal hypermotility response to indomethacin was prevented by both 16,16-dimethyl PGE2 and atropine, but not ampicillin. Yet all these agents inhibited not only the bacterial invasion but also the expression of COX-2 and iNOS activity in the intestinal mucosa following indomethacin treatment, resulting in the prevention of intestinal lesions. CONCLUSION: These results suggest that COX-2 expression in the intestinal mucosa following the administration of indomethacin is associated with intestinal hypermotility and bacterial invasion. The intestinal hypermotility caused by COX-1 inhibition may be a key to COX-2 expression after administration of NSAIDs and their intestinal ulcerogenic properties.     

Functional mechanism underlying cyclooxygenase-2 expression in rat small intestine following administration of indomethacin: Relation to intestinal hypermotility

Background and Aim:  We recently reported that cyclooxygenase (COX)-2 is upregulated in the rat small intestine after administration of indomethacin, and this may be the key to non-steroidal anti-inflammatory drug (NSAID)-induced intestinal damage. The present study investigated the mechanism for COX-2 expression induced in the rat small intestine by indomethacin, in relation with ulcerogenic processes.
Methods:  Animals were given indomethacin or SC-560 p.o., and the intestinal mucosa was examined 24 h later.
Results:  Indomethacin caused hemorrhagic lesions in the small intestine, accompanied with an increase in intestinal motility, bacterial invasion and inducible nitric oxide synthase (iNOS) activity, as well as the expression of COX-2 mRNA in the mucosa. Although SC-560 did not cause any damage, this agent caused intestinal hypermotility, the bacterial invasion and the upregulation of COX-2 expression. The mucosal PGE₂ content was decreased by SC-560 at 3 h but recovered 12 h later, and this recovery of PGE₂ was attenuated by both atropine and ampicillin, in addition to rofecoxib. The intestinal hypermotility response to indomethacin was prevented by both 16,16-dimethyl PGE₂ and atropine, but not ampicillin. Yet all these agents inhibited not only the bacterial invasion but also the expression of COX-2 and iNOS activity in the intestinal mucosa following indomethacin treatment, resulting in the prevention of intestinal lesions.
Conclusion:  These results suggest that COX-2 expression in the intestinal mucosa following the administration of indomethacin is associated with intestinal hypermotility and bacterial invasion. The intestinal hypermotility caused by COX-1 inhibition may be a key to COX-2 expression after administration of NSAIDs and their intestinal ulcerogenic properties.     

Pathogenesis of NSAID-induced gastric damage: importance of cyclooxygenase inhibition and gastric hypermotility

This article reviews the pathogenic mechanism of non-steroidal anti-inflammatory drug (NSAID)-induced gastric damage, focusing on the relation between cyclooxygenase (COX) inhibition and various functional events. NSAIDs, such as indomethacin, at a dose that inhibits prostaglandin (PG) production, enhance gastric motility, resulting in an increase in mucosal permeability, neutrophil infiltration and oxyradical production, and eventually producing gastric lesions. These lesions are prevented by pretreatment with PGE₂ and antisecretory drugs, and also via an atropine-sensitive mechanism, not related to antisecretory action. Although neither rofecoxib (a selective COX-2 inhibitor) nor SC-560 (a selective COX-1 inhibitor) alone damages the stomach, the combined administration of these drugs provokes gastric lesions. SC-560, but not rofecoxib, decreases prostaglandin E₂ (PGE₂) production and causes gastric hypermotility and an increase in mucosal permeability. COX-2 mRNA is expressed in the stomach after administration of indomethacin and SC-560 but not rofecoxib. The up-regulation of indomethacin-induced COX-2 expression is prevented by atropine at a dose that inhibits gastric hypermotility. In addition, selective COX-2 inhibitors have deleterious influences on the stomach when COX-2 is overexpressed under various conditions, including adrenalectomy, arthritis, and Helicobacter pylori-infection. In summary, gastric hypermotility plays a primary role in the pathogenesis of NSAID-induced gastric damage, and the response, causally related with PG deficiency due to COX-1 inhibition, occurs prior to other pathogenic events such as increased mucosal permeability; and the ulcerogenic properties of NSAIDs require the inhibition of both COX-1 and COX-2, the inhibition of COX-1 upregulates COX-2 expression in association with gastric hypermotility, and PGs produced by COX-2 counteract the deleterious effect of COX-1 inhibition.     

Dexamethasone Damages the Rat Stomach but Not Small Intestine During Inhibition of COX-1

We previously reported that inhibition of both COX-1 and COX-2 is required for the gastrointestinal ulcerogenic properties of nonsteroidal anti-inflammatory drugs (NSAIDs). Inhibition of COX-1 up-regulates COX-2 expression, and the prostaglandins (PGs) produced by COX-2 help to maintain the mucosal integrity during inhibition of COX-1. In the present study we investigated whether dexamethasone damages rat gastrointestinal mucosa during inhibition of COX-1 and further developed the idea that COX-2 expression is a key event in the ulcerogenic actions of NSAIDs. Dexamethasone was given p.o. in the absence or presence of SC-560 (a selective COX-1 inhibitor), and the stomach or intestine was examined 8 or 24 hr later, respectively. Neither dexamethasone nor SC-560 alone damaged the gastrointestinal mucosa. In the presence of SC-560, however, dexamethasone damaged the stomach but not small intestine. SC-560 decreased PGE₂ levels in both tissues, with a gradual recovery accompanying the up-regulation of COX-2 expression, and both the recovery of PGE₂ levels and the expression of COX-2 were inhibited by dexamethasone. In the animals treated with SC-560, iNOS expression was up-regulated in the intestinal but not the gastric mucosa, and this response was also inhibited by dexamethasone. These results suggest a risk from steroid therapy in the stomach when COX-2 expression is up-regulated. Dexamethasone does not provoke damage in the intestine, despite inhibiting the up-regulation of COX-2 expression under conditions of PG deficiency; at least one of the reasons is that this agent prevents the expression of iNOS, a major factor in the pathogenesis of intestinal lesions.     

Pathogenic importance of intestinal hypermotility in NSAID-induced small intestinal damage in rats

BACKGROUND/AIM: Nonsteroidal anti-inflammatory drugs (NSAIDs) such as indomethacin produce damage in the small intestine as a major adverse reaction. We examined the effect of various NSAIDs on intestinal motility and investigated the pathogenic importance of motility changes in the intestinal ulcerogenic response to indomethacin in rats. METHODS: Animals without fasting were given various NSAIDs (indomethacin 10 mg/kg, diclofenac 40 mg/kg, flurbiprofen 20 mg/kg, naproxen 40 mg/kg) s.c., and in the case of indomethacin, the following parameters were examined in the small intestine 24 h later; the lesion score, the number of enterobacteria and myeloperoxidase (MPO) as well as inducible nitric oxide (iNOS) activity. Intestinal motility was monitored as intraluminal pressure recordings using a balloon under anesthesia. RESULTS: All NSAIDs tested decreased mucosal PGE(2) levels and produced hemorrhagic lesions in the small intestine, accompanied by intestinal hypermotility. As representative of NSAIDs, indomethacin also increased the extent of enterobacterial invasion and MPO as well as iNOS activity before the occurrence of intestinal damage, and the hypermotility response was observed earlier than the onset of any other event caused by this agent. The intestinal lesions induced by indomethacin were prevented by either supplementation with dmPGE(2), inhibition of bacterial invasion with ampicillin or inhibition of iNOS activity with aminoguanidine, while the hypermotility response was prevented by dmPGE(2) only. In addition, the observed effects of dmPGE(2) were all mimicked by atropine when the intestinal hypermotility was suppressed by this agent. CONCLUSION: These results suggest the pathogenic importance of intestinal hypermotility in the intestinal ulcerogenic response to NSAIDs in rats and show that this event is critical for the occurrence of enterobacterial invasion under PG deficiency, followed by various inflammatory changes and damage in the mucosa. This study also suggests that the antispasmodic drug is protective against NSAID-induced intestinal lesions.     

Functional Mechanism Underlying COX-2 Expression Following Administration of Indomethacin in Rat Stomachs: Importance of Gastric Hypermotility

The expression of COX-2 is up-regulated in the rat stomach after administration of indomethacin, and the inhibition of this enzyme may be a key to NSAID-induced gastric damage. The present study investigated the mechanism for COX-2 expression induced in the rat stomach by indomethacin, in relation with the ulcerogenic processes. The animals were given indomethacin or SC-560 p.o., and the gastric mucosa was examined 8 hr later. Indomethacin decreased the mucosal PGE2 content and produced gross damage with gastric hypermotility and the expression of COX-2 mRNA in the mucosa. Although SC-560 did not produce damage, this agent caused a decrease in the PGE2 content and an increase in gastric motility as well as the up-regulation of COX-2 expression, and provoked damage in the presence of rofecoxib. Gastric lesions induced by indomethacin were prevented by both atropine (even in the presence of exogenous HCl) and omeprazole, although the hypermotility response was inhibited only by atropine. The COX-2 expression induced by indomethacin or SC-560 was inhibited by atropine, even in the presence of exogenous HCl, while omeprazole had no effect. The mucosal PGE2 content was decreased by SC-560 at 2 hr but recovered 8 hr later, and this recovery of PGE2 was attenuated by both atropine and rofecoxib but not omeprazole. These results suggested that the COX-2 expression in the stomach following treatment with indomethacin is functionally associated with gastric hypermotility response induced by COX-1 inhibition. Luminal acid does not play a role in the up-regulation of COX-2 expression in the stomach following administration of indomethacin.     

Prevention by parenteral aspirin of indomethacin-induced gastric lesions in rats: mediation by salicylic acid

Nonsteroidal antiinflammatory drugs (NSAIDs) produce gastric damage in experimental animals, irrespective of the route of administration. However, aspirin (ASA) causes damage only when it is given orally. In the present study, we examined the gastric ulcerogenic effect of subcutaneously administered ASA in rats, in comparison with various NSAIDs, and investigated the reason why ASA does not cause damage in the stomach, in relation to its metabolite salicylic acid (SA). Since the antiinflammatory action of SA is known to be mediated, partly, by endogenous adenosine (AD), we also examined the possible involvement of AD in the protective action of SA. Various NSAIDs (indomethacin, flurbiprofen, naproxen, diclrofenac, ASA, SA) were administered subcutaneously, and the gastric mucosa was examined macroscopically 4 hr later. All NSAIDs tested, except ASA and SA, caused hemorrhagic lesions in the stomach, with a marked gastric hypermotility and a decrease of mucosal PGE₂ contents. These ulcerogenic and motility responses caused by NSAIDs were blocked by pretreatment with atropine or PGE₂. ASA, although inhibiting PGE₂ generation, caused neither hypermotility nor damage in the stomach. On the other hand, SA alone inhibited basal gastric motility without any effect on mucosal PGE₂ contents, and this agent, when given together with indomethacin, prevented gastric hypermotility and lesion formation in response to indomethacin, without affecting the reduced PGE₂ contents. Likewise, ASA inhibited these responses to indomethacin, yet the effects appeared later than those of SA. Following administration of ASA, the blood SA levels reached a peak within 30 min and remained elevated for 4 hr. In addition, the protective effect of SA was not significantly influenced by either the AD deaminase or the AD-receptor antagonists. These results suggest that the failure of parenteral ASA to induce gastric damage may be explained by a protective action of SA metabolized from ASA. SA has a cytoprotective action against NSAID-induced gastric lesions, and this action is not mediated by endogenous AD but may be functionally associated with inhibition of the gastric motility response.     

COX and NOS isoforms involved in acid-induced duodenal bicarbonate secretion in rats

Duodenal HCO₃ ^– secretion increases in response to luminal acid, mediated by endogenous nitiric oxide (NO) as well as prostaglandins (PGs). In this study, we examined the effects of various inhibitors of cyclooxygenase (COX) or NO synthase (NOS) on the acid-induced HCO₃ ^– secretion in rats and determined the enzyme isoforms responsible for this response. A proximal duodenal loop was perfused with saline under urethane anesthesia, and the HCO₃ ^– secretion was measured at pH 7.0 using a pH-stat method and by adding 10 mM HCl. Mucosal acidification was performed by exposing the loop to 10 mM HCl for 10 min. Indomethacin, SC-560 (a selective COX-1 inhibitor) and rofecoxib (a selective COX-2 inhibitor) were given intraduodenally 1 hr before exposure to 10 mM HCl, while N ^G-nitro-l-arginine methyl ester (l-NAME: a nonselective NOS inhibitor) and aminoguanidine (a relatively selective inhibitor of iNOS) were given subcutaneously 3 hr before the acidification. The mucosal acidification increased the HCO₃ ^– secretion, with a rise in mucosal PGE₂ content and luminal release of NO. The HCO₃ ^– secretory and PGE₂ biosynthetic responses were significantly inhibited by indomethacin and SC-560, while rofecoxib had no effect on these responses. On the other hand, l-NAME, but not aminoguanidine, attenuated NO release following the acidification, resulting in inhibition of the acid-induced HCO₃ ^– secretion in a l-arginine-sensitive manner. Neither COX-2 nor iNOS mRNAs were observed in the mucosa before and 1 hr after acidification, while the gene expression of COX-1 and nNOS was constitutively detected in the mucosa and appeared to be slightly up-regulated after the acid stimulation. These results suggest that COX-1 and cNOS play as the respective key enzyme responsible for producing PG and NO following the duodenal acidification, both of which are involved in the mechanism for the acid-induced HCO₃ ^– secretion in the duodenum.     

Prophylactic Effect of Irsogladine Maleate Against Indomethacin-Induced Small Intestinal Lesions in Rats

The effect of irsogladine maleate, a widely used antiulcer drug in Japan, on indomethacin-induced small intestinal lesions was examined in rats. Animals without fasting were given indomethacin (10 mg/kg, s.c.) and sacrificed 24 h later. Irsogladine (1-10 mg/kg) or 16,16-dimethyl prostaglandin E(2) (dmPGE(2) 0.03 mg/kg) was given p.o. twice, 0.5 before and 6 h after indomethacin, while ampicillin (800 mg/kg) was given twice, 18 and 0.5 h before. Indomethacin caused severe lesions in the small intestine, mainly the jejunum and ileum, accompanied by intestinal hypermotility, the up-regulation of inducible nitric oxide synthase (iNOS) expression, and an increase of myeloperoxidase (MPO) activity as well as enterobacterial invasion in the mucosa. These events were all prevented by both dmPGE(2) and ampicillin, except the intestinal hypermotility which was only prevented by dmPGE(2). Likewise, irsogladine also significantly and dose-dependently prevented these lesions at >1 mg/kg. This agent alone increased mucus secretion and significantly suppressed the decreased mucus response to indomethacin, resulting in a suppression of the bacterial invasion as well as the increase in MPO activity and iNOS expression. The protective effect of irsogladine was mimicked by isobutylmethylxanthine, a nonselective inhibitor of phosphodiesterase (PDE), as well as rolipram, a selective PDE4 inhibitor. These results suggest that irsogladine protects the small intestine against indomethacin-induced lesions, and this effect may be associated with the increased mucus secretion, probably due to the inhibitory actions of PDE, resulting in suppression of enterobacterial invasion and iNOS expression.     

Aggravation by Selective COX-1 and COX-2 Inhibitors of Dextran Sulfate Sodium (DSS)-Induced Colon Lesions in Rats

We examined the effect of cyclooxygenase (COX) inhibitors on dextran sulfate sodium (DSS)-induced ulcerative colitis in rats and investigated the role of COX isozymes in the pathogenesis of this model. Experimental colitis was induced by treatment with 2.5% DSS in drinking water for 6 days. Indomethacin (a nonselective COX inhibitor), SC-560 (a selective COX-1 inhibitor), or celecoxib (a selective COX-2 inhibitor) was given PO twice daily for 6 days, during the first 3 or last 3 days of the experimental period. Daily treatment with 2.5% DSS for 6 days caused damage to the colon, with a decrease in body weight gain and colon length as well as an increase of myeloperoxidase (MPO) activity. All COX inhibitors given for 6 days significantly worsened the severity of DSS-induced colonic damage with increased MPO activity. The aggravation was also observed by SC-560 given for the first 3 days or by celecoxib given for the last 3 days. The expression of COX-2 mRNA in the colon was upregulated on day 3 during DSS treatment, with significant increase of prostaglandin E₂ PGE₂ production. The PGE₂ content on day 3 during DSS treatment was inhibited by both indomethacin and SC-560, but not by celecoxib; on day 6 it was suppressed by both indomethacin and celecoxib, but not SC-560. These results suggest that endogenous prostaglandins (PGs) afford protection against colonic ulceration, yet the COX isozyme responsible for the production of PGs differs depending on the stage of ulceration; COX-1 in the early stage and COX-2 in the late stage.     

Role of COX-2 in nonsteroidal anti-inflammatory drug enteropathy in rodents

Abstract

Objective. It is widely thought that cyclooxygenase 1 (COX-1) inhibition with consequential decreases in mucosal prostaglandins, along with concomitant inhibition of COX-2, is pivotal in nonsteroidal anti-inflammatory drug-induced (NSAID) enteropathy. We examined the role of COX-1, COX-2 and topical effects of drugs in NSAID enteropathy. Material and methods. We quantified small intestinal damage and prostaglandin E₂ levels in wild-type, COX-1 and COX-2 deficient mice after administration of R-2-phenylpropionic acid (which has the same topical characteristics as conventional NSAIDs but does not affect the COX enzymes), the conventional NSAIDs flurbiprofen and the selective COX-2 inhibitor celecoxib. We also measured intestinal permeability and inflammation in rats given the selective COX-1 inhibitor SC-560 and NSAIDs. The parameters were assessed at baseline and after administration of the drugs. Results. R-2-phenylpropionic acid caused small intestinal damage in COX-2^-/- and wild-type mice given celecoxib, but not in wild type or COX-1^-/- mice. PGE₂ levels in mice dosed with R-2-phenylpropionic acid were elevated. Indomethacin raised permeability and caused inflammation in rats. Conclusions. The combination of COX-2 absence (or inhibition) and the topical effect of NSAIDs lead to changes characteristic of NSAID enteropathy without concomitant COX-1 inhibition and/or associated decreases in mucosal prostaglandins. COX-2 appears to be more important for maintaining small bowel integrity than COX-1.     

Sildenafil, an Inhibitor of Phosphodiesterase Subtype 5, Prevents Indomethacin-Induced Small-Intestinal Ulceration in Rats via a NO/cGMP-Dependent Mechanism

We examined the effect of sildenafil, an inhibitor of phosphodiesterase subtype 5, that catalyzes hydrolysis of 3′,5′-cyclic guanosine monophosphate (cGMP), on indomethacin-induced small-intestinal ulceration in rats and investigated the mechanism of this action, especially in relation to endogenous nitric oxide (NO). Animals without fasting were given indomethacin (10 mg/kg) s.c. and then killed 24 h later. Indomethacin produced hemorrhagic lesions in the small intestine, accompanied by a promotion of enterobacterial invasion and the expression of inducible NO synthase (iNOS) as well as myeloperoxidase (MPO) activity in the mucosa. Sildenafil (3–20 mg/kg), given p.o. 30 min before indomethacin, dose-dependently reduced the severity of these lesions, with concomitant suppression of the increase in MPO activity, iNOS expression and bacterial invasion. These effects were attenuated by the prior administration of the nonselective NOS inhibitor, N ^G-nitro-l-arginine methyl ester, in an l-arginine-reversible manner. Indomethacin also decreased the secretion of mucus and fluid (enteropooling) and enhanced intestinal motility, but these responses were all prevented by the prior administration of sildenafil. Likewise, pretreatment of the animals with NOR-3, a NO donor, also reversed the functional changes caused by indomethacin, followed by suppression of bacterial invasion and iNOS expression, and prevented the development of intestinal lesions. These results suggest that sildenafil prevents indomethacin-induced small-intestinal ulceration in rats, via a NO/cGMP-dependent mechanism, and this effect is functionally associated with an increase in the secretion of mucus/fluid and a decrease of hypermotility, resulting in the suppression of bacterial invasion and iNOS expression following indomethacin treatment.     

16,16-Dimethyl Prostaglandin E2 Inhibits Indomethacin-Induced Small Intestinal Lesions Through EP3 and EP4 Receptors

We evaluated the effect of various PGE analogs specific to EP receptor subtypes on indomethacin-induced small intestinal lesions in rats and investigated the relationship of EP receptor subtype with the PGE action using EP receptor knockout mice. Animals were administered indomethacin subcutaneously, and they were killed 24 hr later. 16,16-dimethyl prostaglandin E₂ (dmPGE₂) or various EP agonists were administered intravenously 10 min before indomethacin. Indomethacin caused hemorrhagic lesions in the rat small intestine, accompanied with an increase in intestinal motility and the number of enteric bacteria as well as iNOS and MPO activities. Prior administration of dmPGE₂ dose-dependently prevented intestinal lesions, together with inhibition of those functional changes. These effects of dmPGE₂ were mimicked by prostanoids (ONO-NT-012 and ONO-AE1-329), only specific to EP3 or EP4 receptors, although the intestinal motility was inhibited only by ONO-AE1-329. Intestinal mucus secretion and fluid accumulation were decreased by indomethacin but enhanced by dmPGE₂, ONO-NT-012, and ONO-AE1-329 at the doses that prevented intestinal lesions. Indomethacin also caused intestinal lesions in both wild-type and knockout mice lacking EP1 or EP3 receptors, yet the protective action of dmPGE₂ was observed in wild-type and EP1 receptor knockout mice but not the mice lacking EP3 receptors. These results suggest that the intestinal cytoprotective action of PGE₂ against indomethacin is mediated by EP3/EP4 receptors and that this effect is functionally associated with an increase of mucus secretion and enteropooling as well as inhibition of intestinal hypermotility, the former two processes mediated by both EP3 and EP4 receptors, and the latter by EP4 receptors.     

Healing impairment effect of cyclooxygenase inhibitors on dextran sulfate sodium-induced colitis in rats

We examined the effects of various cyclooxygenase (COX) inhibitors on the healing of colonic lesions induced by dextran sulfate sodium (DSS) in the rat. Colonic lesions were induced by 2.5% DSS in the drinking water for 7 days, and then the animals were fed with tap water for subsequent 7 days. Indomethacin (a nonselective COX inhibitor), SC-560 (a selective COX-1 inhibitor), or rofecoxib (a selective COX-2 inhibitor) was given orally twice daily after termination of the DSS treatment. DSS treatment caused severe colonic lesions with a decrease in body weight gain and colon length as well as an increase in myeloperoxidase activity and thiobarbituric acid reactant levels. The severity of colitis gradually reduced, with an improvement of morphological and histological alterations. Daily administration of indomethacin and rofecoxib significantly delayed the healing of colitis with deleterious influences on histological restitution as well as mucosal inflammation, while SC-560 had no effect. Although COX-1 mRNA was expressed in the colon without much alteration during the test period, the expression of COX-2 was upregulated with a peak on day 3 and decreased thereafter. The mucosal prostaglandin E2 content in the colon showed a biphasic change, in parallel with that of the COX-2 expression. The increased prostaglandin E2 production in the injured mucosa was attenuated by indomethacin and rofecoxib, but not by SC-560. These results suggest that endogenous prostaglandins produced by COX-2 play an important role in the healing of DSS-induced colonic lesions. Caution should be paid to the use of selective COX-2 inhibitors as well as nonsteroidal anti-inflammatory drugs in patients with colitis.     

Pharmacological analysis of cyclooxygenase-1 in inflammation

The enzymes cyclooxygenase-1 and cyclooxygenase-2 (COX-1 and COX-2) catalyze the conversion of arachidonic acid to prostaglandin (PG) H₂, the precursor of PGs and thromboxane. These lipid mediators play important roles in inflammation and pain and in normal physiological functions. While there are abundant data indicating that the inducible isoform, COX-2, is important in inflammation and pain, the constitutively expressed isoform, COX-1, has also been suggested to play a role in inflammatory processes. To address the latter question pharmacologically, we used a highly selective COX-1 inhibitor, SC-560 (COX-1 IC₅₀ = 0.009 μM; COX-2 IC₅₀ = 6.3 μM). SC-560 inhibited COX-1-derived platelet thromboxane B₂, gastric PGE₂, and dermal PGE₂ production, indicating that it was orally active, but did not inhibit COX-2-derived PGs in the lipopolysaccharide-induced rat air pouch. Therapeutic or prophylactic administration of SC-560 in the rat carrageenan footpad model did not affect acute inflammation or hyperalgesia at doses that markedly inhibited in vivo COX-1 activity. By contrast, celecoxib, a selective COX-2 inhibitor, was anti-inflammatory and analgesic in this model. Paradoxically, both SC-560 and celecoxib reduced paw PGs to equivalent levels. Increased levels of PGs were found in the cerebrospinal fluid after carrageenan injection and were markedly reduced by celecoxib, but were not affected by SC-560. These results suggest that, in addition to the role of peripherally produced PGs, there is a critical, centrally mediated neurological component to inflammatory pain that is mediated at least in part by COX-2.     

Preconditioning Stress Prevents Cold Restraint Stress-Induced Gastric Lesions in Rats: Roles of COX-1, COX-2, and PLA<Subscript>2</Subscript>

We investigated the protective effect of mild stress on gastric lesions induced by cold-restraint stress, especially concerning prostaglandins (PGs)/cyclo-oxygenase (COX) isozymes. Rats were exposed to severe stress (cold-restraint stress at 10°C for 6 hr) or mild stress (cold-restraint stress at 10°C for 30 min and kept at room temperature for 60 min) followed by severe stress. Severe stress induced gastric lesions, with a concomitant decrease in body temperature (BT). The ulcerogenic response was inhibited by atropine but worsened by indomethacin and SC-560 but not rofecoxib, although none of these agents had any effect on the change in BT. Mild stress suppressed the gastric ulceration and the decrease in BT induced by severe stress, and these effects were reversed by both COX-1 and COX-2 inhibitors. The expression of COX-2 in the stomach was up-regulated from 4 hr after severe stress and this response was slightly expedited by mild stress. COX-2 was also expressed in the hypothalamus under normal and stressed conditions. Quinacrine (phospholipase A₂ inhibitor) attenuated the protective effect of mild stress on the ulceration and decrease in BT caused by severe stress. TA-0910 (TRH analogue) at a low dose also prevented the gastric ulceration and the decrease in BT induced by severe stress. These results suggest that mild stress protects against cold-restraint stress-induced gastric ulceration, and the effect is peripherally and centrally mediated by PGs derived from both COX-1 and COX-2 through the activation of phospholipase A₂. TRH may also be involved in the protective effect of mild stress, probably through regulation of the thermogenic system.     

Interaction Between NSAIDs and Steroid in Rat Stomach

The relative risk of development of peptic ulcer with the use of nonsteroidal antiinflammatory drugs (NSAIDs) has been reported to increase when these drugs are administered in combination with steroids. We investigated the ulcerogenic potential of a combination of NSAIDs and steroids in rats and the underlying pathogenic mechanisms. Indomethacin alone produced gastric lesions, and the severity of the lesions markedly increased with concomitant administration of prednisolone. However, nimesulide, even in excessive doses, did not produce any gastric lesions, regardless of concomitant administration with prednisolone. Furthermore, we showed that the ulcerogenic potential of indomethacin administered in combination with prednisolone may be related to the induction of physiological changes, such as endogenous prostaglandin deficiency, an increase in neutrophil activation, and gastric hypermotility, by indomethacin and alteration of normal epithelial renewal by the steroid. These results suggest that the ulcerogenic potential of preferential a COX-1 inhibitor increases following concomitant administration with a steroid, whereas, nimesulide, a preferential COX-2 inhibitor, is nonulcerogenic, even when administered concomitantly with a steroid, and is therefore a clinically useful antiinflammatory agent.     

Effect of Different Cyclooxygenase Inhibitors on Gastric Adaptive Cytoprotection Induced by 20% Ethanol

In this study, we evaluated the effect of two different dosages of therapeutically prescribed nonsteroidal anti-inflammatory drugs (NSAIDs), ibuprofen, diclofenac, nimesulide, meloxicam, and celecoxib (ED80 for COX-1 and COX-2) on normal gastric mucosa and mucosa, previously exposed to 20% ethanol. At COX-2-inhibiting dosages, the NSAIDs tested were nonulcerogenic, and the same response profile was observed in “adapted” stomachs. Interestingly, low doses of nimesulide and celecoxib increase the levels of Prostaglandin E₂ and COX-2, and protect against subsequent 100% ethanol exposition, suggesting that these drugs may act as “mild irritants” to gastric mucosa. The ulcerogenic response to NSAIDs was prevented by the previous 20% ethanol exposition, probably the result of nitric oxide synthesis, because PGE₂ levels in gastric mucosa were reduced by these agents and a concomitant nitric oxide blockade reversed this protection. Supported by FAPESP, Brazil.     

Survivin: a novel target for indomethacin-induced gastric injury

BACKGROUND & AIMS: Nonsteroidal anti-inflammatory drugs (NSAIDs) cause gastrointestinal erosions and ulcers. Apoptosis is one of the mechanisms. The role of survivin, an antiapoptosis protein, in NSAID-induced gastric injury is unknown. We examined the role of survivin in NSAID-induced gastric mucosal and gastric cell injury. METHODS: We examined: (1) the effects of indomethacin (nonselective NSAID), celecoxib and NS-398 (cyclooxygenase [COX]-2-selective NSAIDs), SC-560 (a COX-1-selective NSAID), and SC-560 plus celecoxib on survivin expression and extent of injury in rat gastric mucosa; (2) the effects of indomethacin, NS-398, SC-560, and SC-560 plus NS-398 on survivin expression and injury in gastric epithelial (RGM-1) cells; and (3) the effects of survivin suppression with small interfering RNA (siRNA) on RGM-1 cell integrity at baseline and following indomethacin injury. RESULTS: Indomethacin treatment dose-dependently reduced survivin protein levels and caused severe injury of gastric mucosa and RGM-1 cells. Suppression of survivin expression with siRNA in RGM-1 cells caused cell damage and increased susceptibility to injury by indomethacin. Celecoxib treatment caused exfoliation of the mucosal surface epithelium, but neither caused deep erosions or altered survivin expression. Neither NS-398 nor SC-560 treatment altered survivin levels or produced injury in vivo or in vitro. COX-1 and COX-2 inhibitor combination caused injury in vivo and in vitro but did not decrease survivin expression. CONCLUSIONS: (1) Indomethacin, but not selective COX-1 or COX-2 inhibitors alone or in combination, reduces survivin expression in gastric mucosal cells and (2) significant reduction of survivin precedes greater severity of gastric injury.