Karlheinz Ehrlich

Selective cyclo-oxygenase-2 inhibitors and their influence on the protective effect of a mild irritant in the rat stomach

1 The effects of the non‐selective cyclo‐oxygenase (COX) inhibitor indomethacin and the selective COX‐2 inhibitors, N‐[2‐(cyclohexyloxy)‐4‐nitrophenyl] methanesulphonamide (NS‐398), 5‐methanesulphonamido‐6‐(2,4‐difluorothio‐phenyl)‐1‐indanone (L‐745,337) and 5,5‐dimethyl‐3‐(3‐fluorophenyl)‐4‐(4‐methylsulphonyl) phenyl‐2(5H)‐furanone (DFU), on the protection induced by the mild irritant 20% ethanol were investigated in the rat stomach. 2 Instillation of 20% ethanol (1 ml, p.o.) effectively protected against gastric mucosal injury induced by subsequent instillation of 70% or 96% ethanol (1 ml, p.o.). 3 Oral administration of indomethacin (1.25–20 mg kg−1) dose‐dependently counteracted the protective effect of 20% ethanol (ID50: 3.5 mg kg−1). 4 Likewise, NS‐398 (0.1–1 mg kg−1), L‐745,337 (0.2–2 mg kg−1) and DFU (0.02–0.2 mg kg−1) inhibited the protective effect of 20% ethanol in a dose‐dependent manner with ID50 values of 0.3 mg kg−1, 0.4 mg kg−1 and 0.06 mg kg−1, respectively. 5 Inhibition of mild irritant‐induced protection was also found when NS‐398 (1 mg kg−1) was administered s.c. or when 96% ethanol was used to damage the mucosa. 6 Pretreatment with 16,16‐dimethyl‐prostaglandin (PG)E2 at 4 ng kg−1, a dose that did not protect against ethanol (70%)‐induced mucosal damage when given alone, completely reversed the effect of the selective COX‐2 inhibitors on the mild irritant‐induced protection. 7 Pretreatment with dexamethasone (3 mg kg−1, 24 and 2 h before instillation of 20% ethanol) did not affect the protective activity of the mild irritant, indicating that enzyme induction is not involved. 8 Indomethacin (20 mg kg−1, p.o.) did not prevent the protection conferred by sodium salicylate (100 mg kg−1), dimercaprol (30 μg kg−1), iodoacetamide (50 mg kg−1) and lithium (20 mg kg−1). Likewise, the protective effect of these agents was not counteracted by NS‐398 (1 mg kg−1, p.o.). 9 Whereas indomethacin (20 mg kg−1, p.o.) near‐maximally inhibited gastric mucosal formation of PGE2, 6‐keto‐PGF1α and thromboxane (TX) B2 as well as platelet TXB2 release, the selective COX‐2 inhibitors were ineffective. 10 The findings show that selective COX‐2 inhibitors, although lacking in ulcerogenic activity, prevent the protection conferred by a mild irritant. Prostaglandis generated by a constitutive COX‐2 could thus contribute to physiological functions involved in gastric homeostasis, although at present a non‐COX‐2‐related mechanism underlying the effect of the selective COX‐2 inhibitors tested on mild irritant‐induced protection cannot be completely excluded.

Selective cyclo‐oxygenase‐2 inhibitors aggravate ischaemia‐reperfusion injury in the rat stomach

Effects of indomethacin, the selective cyclo‐oxygenase (COX)‐2 inhibitors NS‐398 and DFU, and dexamethasone on gastric damage induced by 30 min ischaemia followed by 60 min reperfusion (I‐R) were investigated in rats. Modulation of gastric levels of COX‐1 and COX‐2 mRNA by I‐R was evaluated using Northern blot and reverse transcription‐polymerase chain reaction. I‐R‐induced gastric damage was dose‐dependently aggravated by administration of indomethacin (1–10 mg kg−1), NS‐398 (0.4–4 mg kg−1) or DFU (0.02–2 mg kg−1) as assessed macroscopically and histologically. Likewise, administration of dexamethasone (1 mg kg−1) significantly increased I‐R damage. Low doses of 16,16‐dimethyl‐prostaglandin(PG)E2, that did not protect against ethanol‐induced mucosal damage, reversed the effects of the selective COX‐2 inhibitors, indomethacin and dexamethasone. I‐R had no effect on gastric COX‐1 mRNA levels but increased COX‐2 mRNA levels in a time‐dependent manner. Dexamethasone inhibited the I‐R‐induced expression of COX‐2 mRNA. I‐R was not associated with a measurable increase in gastric mucosal formation of 6‐keto‐PGF1α and PGE2. PG formation was substantially inhibited by indomethacin (10 mg kg−1) but was not significantly reduced by NS‐398 (4 mg kg−1), DFU (2 mg kg−1) or dexamethasone (1 mg kg−1). The findings indicate that selective COX‐2 inhibitors and dexamethasone markedly enhance gastric damage induced by I‐R. Thus, whereas COX‐2 has no essential role in the maintenance of gastric mucosal integrity under basal conditions, COX‐2 is rapidly induced in a pro‐ulcerogenic setting and contributes to mucosal defence by minimizing injury. This suggests that in certain situations selective COX‐2 inhibitors may have gastrotoxic effects.

Peptidergic and cholinergic neurons and mediators in peptone-induced gastroprotection: role of cyclooxygenase-2

This study investigates the neural pathways, mediators, and cyclooxygenase isoenzymes involved in the gastroprotection conferred by peptone in rats. Intragastric perfusion with 8% peptone protected against gross and histological damage induced by subsequent perfusion with 50% ethanol. The gastroprotective effect of peptone was near maximally inhibited by gastrin immunoneutralization, inactivation of capsaicin-sensitive afferent neurons, calcitonin gene-related peptide (CGRP) immunoneutralization, blockade of gastrin receptors, CGRP, bombesin/gastrin-releasing peptide (GRP), or somatostatin receptors, and by the nitric oxide (NO) synthase inhibitor N G-nitro-l-arginine methyl ester and was partially (46%) counteracted by atropine. Indomethacin and the selective cyclooxygenase-2 inhibitors NS-398 and L-745,337 dose dependently (50% inhibitory dose, 4.2, 0.8, and 1.5 mg/kg, respectively) attenuated the peptone-induced protection. Dexamethasone was ineffective. These results indicate that protective effects of peptone involve endogenous gastrin and possibly somatostatin and are mediated by capsaicin-sensitive afferent, cholinergic, and bombesin/GRP neurons. CGRP, NO, and prostaglandins participate as essential mediators. The study provides evidence that prostaglandins derived from a constitutive cyclooxygenase-2 contribute to mucosal defense in the presence of ulcerogens and thus participate in homeostatic functions of the stomach.

Interaction of cyclooxygenase-2 inhibitors and salicylate in gastric mucosal damage

The interactions of sodium salicylate and the selective cyclooxygenase-2 inhibitors N-[2-(cyclohexyloxy)-4-nitrophenyl]-methanesulfonamide (NS-398) and 5.5-dimethyl-3-(3-fluorophenyl)-4-(4-methylsulfonyl)phenyl-2(5II)-furanone (DFU), dexamethasone and the nitric oxide (NO) synthase inhibitor N(G)-nitro-L-arginine methylester (L-NAME) were examined in ischaemia-reperfusion damage and adaptive protection in the rat stomach. Ischaemia-reperfusion damage was substantially aggravated by pretreatment with NS-398 (4 mg/kg), DFU (2 mg/kg), dexamethasone (1 mg/kg) or L-NAME (3 and 10 mg/kg). Salicylate (0.01-0.05 mg/kg) reversed the aggravating effect of NS-398, DFU and dexamethasone, while the effect of L-NAME was counteracted by L-arginine (twice 400 mg/kg) but not salicylate (0.05 or 10 mg/kg). Instillation of 20% ethanol prevented mucosal damage induced by 70% ethanol. This adaptive gastroprotection was abolished by pretreatment with NS-398 (1 mg/kg), DFU (0.2 mg/kg) or L-NAME (10 mg/kg). Salicylate (0.01-0.05 mg/kg) reversed the inhibition of protection by NS-398 and DFU, while the effect of L-NAME (10 mg/kg) was antagonized by L-arginine (100 mg/kg) but not salicylate (0.05 mg/kg). The precise mechanism of the functional antagonism between extremely low doses of salicylate and selective cyclooxygenase-2 inhibitors remains to be investigated.

Role of Lipoxygenases and the Lipoxin A4/Annexin 1 Receptor in Ischemia-Reperfusion-Induced Gastric Mucosal Damage in Rats

Rat gastric mucosal damage was induced by ischemia-reperfusion. The 5-lipoxygenase inhibitors MK886 and A63162, the 12-lipoxygenase inhibitor baicalein, the 15-lipoxygenase inhibitor PD146176 and the lipoxin (LX) A4/annexin 1 antagonist Boc1 increased mucosal damage in a dose-dependent manner. Low doses of these compounds, which have no effects on mucosal integrity, cause severe damage when combined with low doses of indomethacin, celecoxib or dexamethasone. 16,16-Dimethylprostaglandin (PG) E2 and LXA4 can replace each other in preventing mucosal injury induced by either cyclooxygenase or lipoxygenase inhibitors. The results suggest that not only cyclooxygenases, but also lipoxygenases have a role in limiting gastric mucosal damage during ischemia-reperfusion.

Role of Lipoxygenases and Lipoxin A4/Annexin-1 Receptor in Gastric Protection Induced by 20% Ethanol or Sodium Salicylate in Rats

The role of cyclooxygenases and prostaglandins in experimental models of gastroprotection is well established. We investigated the effects of the 5-lipoxygenase inhibitor A63162, the 12-lipoxygenase inhibitor baicalein and the 15-lipoxygenase inhibitor PD146176 as well as the nonspecific lipoxin A4/annexin-1 antagonist Boc1 on adaptive protection induced by 20% ethanol against 70% ethanol, and on protection induced by sodium salicylate against the mucosal-damage-aggravating effects of celecoxib and dexamethasone during local ischemia-reperfusion in rats. It was found that both types of gastroprotection were antagonized by the lipoxygenase inhibitors and the lipoxin A4/annexin-1 antagonist in doses that have no direct damaging effect on gastric mucosa. The results suggest that not only cyclooxygenases, but also active lipoxygenases and, possibly, annexin-1 are required for these types of gastroprotection to occur.

Dexamethasone Impairs the Gastric Mucosal Integrity in Rats Treated with a Cyclooxygenase-1 but Not with a Cyclooxygenase-2 Inhibitor

In rats, neither the cyclooxygenase-1 inhibitor SC-560 nor the cyclooxygenase-2 inhibitor rofecoxib damages the gastric mucosa. Coadministration of dexamethasone induced injury in SC-560- but not in rofecoxib-treated rats. High levels of cyclooxygenase-1 protein occurred in the gastric mucosa of control rats, with no change after administration of SC-560. In contrast, the gastric cyclooxygenase-2 protein levels were low in control rats, but increased in a time-dependent manner after administration of SC-560. Dexamethasone prevented the increase in cyclooxygenase-2 protein levels. Our findings show that inhibition of cyclooxygenase-1 upregulates cyclooxygenase-2. When the upregulation is prevented by dexamethasone, gastric damage develops, suggesting that induction of cyclooxygenase-2 represents a compensatory mechanism that counteracts the injurious effect of cyclooxygenase-1 inhibition.