Kazuhiko Kubota

Inhibitory effect of neopterin on NADPH-dependent superoxide-generating oxidase of rat peritoneal macrophages

The effect of the oxidized form of neopterin (NP) on the NADPH‐dependent superoxide‐generating oxidase (NADPH‐oxidase) was investigated in both whole‐cell and cell‐free activation systems by using peritoneal macrophages of rats which had received an intraperitoneal injection of mineral oil. In the whole‐cell system, NP remarkably inhibited the generation of Superoxides in macrophages stimulated with phorbol myristate acetate (PMA). NP also showed an significant suppression of the activation of superoxide‐generating NADPH‐oxidase in the cell‐free system using solubilized membranes and sodium dodecyl sulfate (SDS) as a stimulant. The 50%‐inhibitory concentration (IC50) of NP was about 1 μM in both assay systems. In a kinetic study, competitive inhibition of the NADPH‐oxidase by NP was observed in the cell‐free system with a calculated inhibition constant (K i) of 6.50 μM. These findings suggest that NP may play an important role in the suppression of Superoxide generation via the inhibition of the NADPH‐oxidase in phagocytes.

Neopterin as an endogenous antioxidant

The in vitro potency of neopterin (NP) as an antioxidant and its in vivo activity to suppress alloxan‐induced diabetes were investigated. The reduced form of neopterin, 5,6,7,8‐tetrahydroneopterin (NPH‐4), showed an extremely high superoxide anion radical scavenging activity in two assay systems, i.e. xanthine/xanthine oxidase‐ and macrophage/phorbol myrislate acetate (PMA)‐reaction systems. NPH‐4 also inhibited the oxidation of linoleic acid about as effectively as uric acid. Furthermore, NPH‐4 and NP effectively suppressed alloxan‐induced mouse diabetes. These results suggest that pteridines play an important role as endogenous antioxidant.

Cholecystokinin antagonism by benzodiazepines in the contractile response of the isolated guinea-pig gallbladder

Three benzodiazepines, chlordiazepoxide (CDP), diazepam (DZP) and medazepam (MZP), inhibited the contractile response of circular muscle strips from the isolated guinea-pig gallbladder to sulfated cholecystokinin octapeptide (CCK8) in the presence of atropine. The dose-response curves for CCK8 were shifted in parallel to the right by 10(-6) to 10(-5) M of the three benzodiazepines, although the maximum response to CCK8 was depressed by higher concentrations. Schild plot analysis of the antagonism gave pA2 values of 6.70 for CDP and 7.33 for DZP and slopes of the regression lines close to 1.0, suggesting that the antagonism was competitive in nature. The responses to acetylcholine, histamine and bradykinin were also depressed by 10(-5) M of DZP, but no parallel shift of their dose-response curves was observed. The non-specific inhibitory action of the benzodiazepines on these agonists was presumed to be due to the calcium antagonist-like action of the benzodiazepines. The antagonism between CCK8 and the benzodiazepines in the gallbladder was unaffected by GABA.

General pharmacological studies of fusarenon-X, a trichothecene mycotoxin from Fusarium species.

Abstract Fusarenon-X (F-X) is one of the trichothecene mycotoxins. In the present work, pharmacological properties of F-X were examined. (1) F-X induced hypothermia but did not produce appreciable behavioral changes of mice. (2) In anesthetized rats, F-X caused a rise in the blood pressure and a decrease in the respiratory rate but induced no significant change in the heart rate. (3) In isolated tissues such as fundus muscle, vas deferens, tracheal muscle, ileum, and atrium, F-X had no detectable effects on spontaneous activity and the responsiveness to agonists. (4) Subplantar administration of F-X into rat hindpaw induced edema. F-X had little influence on the histamine release from mast cells and the membrane stability of erythrocytes. (5) F-X decreased the spontaneous peristalsis of intestine but increased the intestinal propulsion of charcoal meal. (6) F-X induced vomiting in dogs which was suppressed by preliminary administration of chlorpromazine and metoclopramide. F-X may induce vomiting by stimulating the chemoreceptor trigger zone in dogs.

Cholecystokinin antagonism by benzodiazepines in the food intake in mice

Three benzodiazepines, chlordiazepoxide, diazepam and flurazepam, were demonstrated to reverse the suppressed food intake in mice in response to cholecystokinin octapeptide (CCK8). CCK8 (200 ng) was administered intracisternally, and the benzodiazepines intraperitoneally at doses of 0.1 to 1 mg/kg. The three benzodiazepines slightly depressed the feeding by themselves, but significantly reversed the satiety effects of CCK8. Naloxone (2 mg/kg) decreased the food intake but failed to reverse the CCK8 satiety action. The benzodiazepines were considered to antagonize the satiety action of CCK8 in the central nervous system through unknown mechanisms.

Studies on mechanisms of diarrhea induced by fusarenon-X, a trichothecene mycotoxin from Fusarium species

Abstract Cardinal signs of red mold toxicosis in man and farm animals are vomiting, nausea, diarrhea, and food refusal. The red mold toxicosis has been suggested to be induced by trichothecenes which are produced by Fusarium fungi. Fusarenon-X (F-X) is one of the trichothecene mycotoxins. The ip injection of F-X to rats caused watery diarrhea 36 to 60 hr after injection. Autopsy done 24 hr after an injection of F-X revealed an expansion of the small intestine but no macroscopic bleeding into the lumen of the intestine. F-X increased the absorption rate of d -xylose from the intestine in vitro , caused leakage of intravenously injected Evans blue dye into the intestinal lumen, and decreased the serum sodium level. However, F-X did not increase leakage of erythrocytes into the intestinal lumen. In histological observations, F-X shortened intestinal villi and caused an infiltration of erythrocytes into the intestinal lamina propria. These findings suggest that F-X may increase the permeability of the blood vessel wall and intestinal mucosal epithelium and thereby cause leakage of plasma contents into the intestinal lumen which eventually leads to diarrhea.

Differences in contractile responses to acetylcholine and serotonin in rat stomach fundus strips

The differences in contractile responses to acetylcholine (ACh) and serotonin (5-HT) were studied in the rat fundus strips. At low temperatures, the contractile response to 5-HT was reduced more noticeably than to ACh. The contraction induced by ACh at 4 degrees C was not inhibited by La3+. Removal of extracellular Ca2+ reduced the response to 5-HT more significantly than that to ACh.. La3+, verapamil and D-600 inhibited the 5-HT response more than the ACh response. In Ca2+-free solution the contractile response to ACh of a fundus strip preloaded with Ca2+ was lar ger than that to 5-HT. These results suggest that the Ca2+ movements induced by ACh and by 5-HT are different, and that the difference in Ca2+ movements may cause the difference in contractile responses to the two agonists at low temperatures.

Studies on mechanisms of diarrhea induced by fusarenon-X, a trichothecene mycotoxin from Fusarium species: Fusarenon-X-induced diarrhea is not mediated by cyclic nucleotides

Cardinal signs of red mold toxicosis in man and farm animals are vomiting, nausea, diarrhea, and food refusal. The red mold toxicosis has been suggested to be induced by trichothecenes, which are produced by Fusarium fungi. Fusarenon-X (F-X) is one of the trichothecene mycotoxins. The ip injection of F-X to rats causes an expansion of the small intestine and watery diarrhea. In this study, we measured the concentrations of protein, sodium, potassium, and calcium in the serum of rat treated with F-X for the sake of demonstrating the loss of serum protein and the decreases of serum sodium and calcium by F-X. Since it is well known that some diarrheal diseases are due to the increase of cyclic nucleotide level in the intestinal mucosa, we also measured cyclic AMP and cyclic GMP levels in the intestinal mucosa. It was demonstrated that F-X did not increase the cyclic AMP and cyclic GMP levels in the jejunal and the ileal mucosa at 8 and 24 hr after F-X treatment. The results obtained in this work suggest that F-X-induced diarrhea is not mediated by the cyclic nucleotide system.

Protective effects of tetrahydroneopterin against free radical-induced injury

The therapeutic potency of 5,6,7,8-tetrahydroneopterin (NPH4) was investigated in ischemic paw edema, Adriamycin (ADR)-induced cardiotoxicity, and endotoxin [lipopolysaccharide (LPS)]- and carbon tetrachloride (CCl4)-induced hepatotoxicity in mice. Ischemic paw edema was completely suppressed by pre-administration of NPH4. ADR-induced cardiotoxicity and LPS-induced hepatotoxicity were significantly decreased by post-administration of NPH4. Furthermore, NPH4 ameliorated CCl4-induced hepatotoxicity. These results suggest that NPH4 may be useful for the treatment of free radical, especially superoxide radical, -related tissue injury.

Actions of bile salts and of papaverine and intracellular cyclic AMP in isolated rat uterus.

Both antioxytocin and antiphosphodiesterase activities of desoxycholate were potentiated by lowering the pH of the medium. The results suggest the possibility that it is the bile salt in the non-ionized form which exerts the antioxytocin and antiphosphodiesterase action. The concentration of tissue cyclic AMP was measured at different times during relaxation of the uterus. The results show a significant increase in tissue cyclic AMP concentration at a time when the muscle was just beginning to relax in response to papaverine, but not in response to chenodesoxycholate. The intracellular level of cyclic AMP during relaxation of the uterus produced by papaverine or chenodesoxycholate was, however, significantly increased when relaxation was about 90% complete. The difference between the mode of action of papaverine and that of the bile salts is discussed.