Kiyoshi Oketani

Inhibitions of acid secretion by E3810 and omeprazole, and their reversal by glutathione

A substituted benzimidazole ([4-(3-methoxypropoxy)-3-methylpyridine-2-yl]methylsulfinyl)- 1H-benzimidazole sodium salt (E3810), is a gastric proton pump (H+, K(+)-ATPase) inhibitor. E3810 and omeprazole inhibited acid accumulation dose dependently as measured with aminopyrine uptake in isolated rabbit gastric glands, their IC50 values being 0.16 and 0.36 microM, respectively. The addition of exogenous reduced glutathione (GSH) to the gland suspension reactivated dose dependently the acid secretion which had been inhibited by 2 microM E3810 or omeprazole as a function of the incubation time. Furthermore, GSH at 1 and 3 mM reversed the antisecretory effect of E3810 more quickly than it did that of omeprazole. The antisecretory effect of E3810 was slightly greater than that of omeprazole in histamine-stimulated fistula dogs in vivo. The duration of the antisecretory activity of E3810 at concentrations of 2 and 4 mg/kg was shorter than that of omeprazole at the same concentrations in pentagastrin-stimulated fistula dogs. The reversal of the antisecretory activity of the inhibitors in dogs is suggested to be due to the action of endogenous extracellular GSH, in addition to de novo synthesis of the proton pump, because bullfrog gastric mucosae were found in the present study to secrete GSH into the mucosal solution at the rate of about 0.25 nmol/min/g tissue.

Effect of E3040, an inhibitor of 5-lipoxygenase and thromboxane synthase, on rat bowel damage induced by lipopolysaccharide

Intravenous administration of lipopolysaccharide to rats that had been immunized with lipopolysaccharide induced hemorrhagic damage in the large intestine. We investigated the role of 5-lipoxygenase and thromboxane synthase products in the damage of the large intestine induced by lipopolysaccharide. In the large intestine of lipopolysaccharide-immunized rats, intravenous injection of lipopolysaccharide increased the vascular permeability, production of leukotriene B(4), leukotriene C(4)/D(4), thromboxane B(2) and prostaglandin E(2), and also increased the activity of myeloperoxidase, a marker enzyme of neutrophils. Oral administration of E3040 (6-hydroxy-5,7-dimethyl-2-(methylamino)-4-(3-pyridylmethyl)benzothiazole), a novel dual inhibitor of 5-lipoxygenase and thromboxane synthase, at 30 and 100 mg/kg inhibited the increase in vascular permeability induced by lipopolysaccharide in the large intestine. E3040 inhibited the production of leukotriene B(4) and thromboxane B(2) and tended to increase the production of prostaglandin E(2) in the large intestine. Sulfasalazine (500 mg/kg) and prednisolone (10 mg/kg), drugs used for the treatment of inflammatory bowel disease, had no significant effect on eicosanoid production and vascular permeability. These results indicate that E3040 inhibits the production of both leukotriene B(4) and thromboxane B(2) and prevents lipopolysaccharide-induced damage in the large intestine of lipopolysaccharide-immunized rats.

In vitro effects of E3040, a dual inhibitor of 5-lipoxygenase and thromboxane A2 synthetase, on eicosanoid production

In vitro pharmacological profiles of E3040, 6-hydroxy-5, 7-dimethyl-2-(methylamino)-4-(3-pyridylmethyl) benzothiazole were investigated. Against the 5-lipoxygenase activity of rat basophilic leukemia cells, E3040 and zileuton (a 5-lipoxygenase inhibitor) had an IC(50) of 0.23 and 0.93 microM, respectively. Against the thromboxane A(2) synthetase activity of human platelets, E3040 had an IC(50) of 0.01 microM, which was comparable to that of OKY-1581 (sodium (E)-3-[4-(3-pyridylmethyl) phenyl]-2-methylacrylate, a thromboxane A(2) synthetase inhibitor). Against cyclooxygenase activity of sheep seminal vesicles, E3040 showed no inhibition (IC(50), >300 microM). Sulfasalazine and 5-aminosalicylic acid, therapeutic drugs for inflammatory bowel disease, inhibited 5-lipoxygenase activity with an IC(50) of 293 and 970 microM, respectively. Sulfasalazine inhibited thromboxane A(2) synthetase activity with an IC(50) of 20 microM. In rat peritoneal leukocytes, E3040 inhibited leukotriene B(4) and thromboxane B(2) production with an IC(50) of 0.17 and 0.24 microM, respectively. E3040 inhibited leukotriene B(4) production in human neutrophils and thromboxane B(2) production in human platelets (IC(50) of 0.21 and 0.09 microM, respectively). These results indicated that E3040 potently inhibited 5-lipoxygenase and thromboxane A(2) synthetase and blocked leukotriene B(4) and thromboxane B(2) production in rat peritoneal and human blood cells.

Effects of the Proton Pump Inhibitor, E3810, on Gastric Secretion and Gastric and Duodenal Ulcers or Erosions in Rats

SummaryThe effects of E3810, a potent proton pump inhibitor, on gastric secretion and gastric or duodenal ulcers or erosions were studied in rats. Intraduodenal administration of E3810 dose dependently inhibited gastric secretion (volume and acid outputs) in pylorus-ligated rats. The effective dose required to inhibit volume output by 50% (ED50) was 12 mg/kg and that for acid output was 3.4 mg/kg. Acutely induced gastric ulcers or erosions, such as cold-restraint stress erosions, Shay ulcers and mercaptamine (cysteamine)-induced duodenal ulcers, were significantly inhibited by oral, intraduodenal or subcutaneous administration of E3810 at 1 to 100 mg/kg. The protective effects of E3810 in these acute models were compared with those of omeprazole at the same doses. At low doses, 10 mg/kg (intraduodenally) in Shay ulcers and 3 mg/kg (subcutaneously) in mercaptamine-induced ulcers, E3810 was significantly more effective than omeprazole. But at high doses, there was no significant difference between the 2 drugs in efficacy. In the cold stress erosion model, the drugs were equally effective.

E3040 sulphate, a novel thromboxane synthase inhibitor, blocks the Cl− secretion induced by platelet‐activating factor in isolated rat colon

E3040 (6‐hydroxy‐5,7‐dimethyl‐2‐methylamino‐4‐(3‐pyridylmethyl)benzothiazole), is a novel dual inhibitor of 5‐lipoxygenase (5‐LOX) and thromboxane synthase (Tx synthase). Here, we examined the effects of E3040 sulphate, a sulphate conjugate of E3040, on these enzyme activities in cell‐free systems and on the thromboxane A2 (TxA2)‐mediated Cl− secretion induced by platelet‐activating factor (PAF) in isolated rat colons. E3040 sulphate inhibited Tx synthase activity in a concentration‐dependent manner (IC50=0.013 μM), whereas it induced little effects on 5‐LOX and cyclo‐oxygenase activities (IC50>100 μM) with the cell‐free enzyme assay. With isolated rat colonic mucosa, E3040 sulphate in a concentration‐dependent manner (IC50=1.8 μM) inhibited the Cl− secretion induced by 10 μM PAF. On the other hand, E3040 sulphate (30 μM) induced no effect on the prostaglandin E2 (0.5 μM)‐ and leukotriene D4 (1 μM)‐induced Cl− secretion in the colon. PAF (10 μM) increased a release of TxB2, a stable metabolite of TxA2, from the colonic mucosa. This increase was significantly inhibited by subsequent addition of E3040 sulphate (30 μM). Probenecid (100 μM), an inhibitor of organic anion transporter, abolished the inhibitory effect of E3040 sulphate on the PAF‐induced Cl− secretion. Another inhibitor, sulphobromophthalein (30 μM) partially but significantly attenuated the effect of E3040 sulphate. p‐Aminohippuric acid (1 mM) had no effect. These findings suggest that E3040 sulphate is a novel Tx synthase inhibitor, and that E3040 sulphate taken up into the colonic cells by organic anion transporters inhibits the PAF‐induced Cl− secretion by blocking a release of TxA2.

Effects of taurocholic acid/HCl alone or after pretreatment with geranylgeranylacetone on phospholipid metabolism in rat gastric mucosa

Changes in phospholipid metabolism in gastric mucosa caused by instillation of taurocholic acid (TCA)/HCl (80 mM/300 mM) into the stomach of rats and the effects of pretreatment with an antiulcer agent, geranylgeranylacetone (GGA), were studied after intravenous injection of radioisotope-labeled precursors. The instillation of TCA/HCl rapidly reduced the incorporation of labeled fatty acids and glycerol into phosphatidylcholine and phosphatidylethanolamine, indicating the inhibition of de novo synthesis of phospholipids. These changes were restored by 120-150 min after the TCA/HCl treatment. Pretreatment with GGA enhanced the incorporation of precursors into phosphatidylcholine immediately after the instillation of TCA/HCl. Experiments in which the mucosal lipids were labeled with fatty acids prior to the instillation of TCA/HCl showed that the degradation of cellular lipids and release of the products into the gastric lumen were induced by TCA/HCl and that these changes were not prevented by GGA. Since GGA almost completely inhibited the gastric lesions induced by TCA/HCl, the enhancement of synthesis of mucosal phosphatidylcholine induced by GGA may be involved in the prevention of gastric damage. The incorporation of labeled fatty acids into free fatty acid fraction and diacylglycerol was increased quickly by the TCA/HCl treatment, suggesting early damage to the blood vessels of the gastric mucosa; these changes were inhibited significantly by GGA.

Effect of secretin on stress-induced gastric bleeding in rats

The effect of secretin on gastric bleeding induced by water (20°C) immersion stress was studied by means of either perfusion of hydrochloric acid (0.13 N) or instillation of hydrochloric acid solution (0.4 N) into the rat stomach. When the stomach was perfused with acid solution, gastric bleeding occurred 20 min after the stress of water immersion and the amount of bleeding increased with time. It was found that integrated amounts of gastric bleeding after stress were more significantly decreased in the secretin-treated group (7.5 clinical units/kg/hr) than the saline control. On the other hand, intraduodenal administration of cimetidine (100 mg/kg) or propantheline (30 mg/kg) did not affect gastric bleeding. It was also revealed that the total amount of glycoprotein in the 4-hr perfusate was decreased 30% by the water immersion, but that secretin treatment prevented the 30% decrease in glycoprotein in the perfusate. In experiments on intragastric instillation of hydrochloric acid solution, it was found that gastric bleeding was seen only when the concentration of hydrochloric acid solution was higher than 0.4 N. However, the bleeding was also significantly prevented by treatment with secretin (2.5 and 7.5 clinical units/kg/hr). In conclusion, secretin prevents stress-induced gastric bleeding at least in part by increasing glycoprotein secretion in the gastric mucosa in the rat.