Characteristics of insulin binding to isolated rat adipocytes were studied by use of fluorescein isothiocyanate (FITC)-labeled insulin and flow cytometry, and were compared with those of insulin-induced uptake of 3H-2-deoxy-D-glucose (3H-2-DG). FITC-labeled insulin bound to the adipocytes during the initial incubation, but washing once with Hanks' buffer completely removed the bound insulin from the cells. Uptake of 3H-2-DG by the adipocytes was induced by insulin but ceased when the cells were washed. 5-(N-ethyl-N-isopropyl)amiloride (EIPA), a Na+/H+ exchanger inhibitor, suppressed both the insulin-induced 3H-2-DG uptake and insulin-induced translocation of glucose transporter type 4 (Glut-4) from the microsomal membranes to the plasma membrane of the cells. In the absence of insulin, when the Nay in the Hanks' buffer was replaced with Li+[Na+(-)Li+(+) Hanks' buffer], the level of 3H-2-DG uptake by the cells was significantly higher than in the presence of Na+. The increase in the uptake of 3H-2-DG induced by Li+ was similar to the increase induced by insulin, but phosphorylation of the insulin receptors was not detected by Western blotting. The increased 3H-2-DG uptake induced by the Na+(-)Li+(F) Hanks' buffer solution was abolished by adding EIPA. This result suggests that the Na+/H+ exchanger influences Li+-induced 3H-2-DG uptake. Based on these results, we conclude that the observed insulin-induced glucose uptake was tightly regulated by the Na+/H+ exchanger.
The objective of the present study was to investigate the role of oxygen radicals in the acute gastric mucosal injury induced by taurocholate combined with serotonin (TCA-5HT) in rats. Just after taurocholate had been given intragastrically at a dose of 30mM/rat, serotonin was injected subcutaneously at a dose of 20mg/kg. Superficial hemorrhagic erosions were observed on the glandular stomach 2h after administration of TCA-5HT. The level of lipid peroxides and that of α-tocopherol in the gastric mucosa significantly increased and decreased, respectively, with time after TCA-5HT treatment. Myeloperoxidase (MPO) activity, an index of neutrophil infiltration, in the gastric mucosa also increased significantly. Pretreatment with the combination of recombinant human Cu, Zn-superoxide dismutase (hSOD) and catalase significantly inhibited the gastric mucosal injury, accumulation of lipid peroxides, and the decrease in α-tocopherol content in the gastric mucosa. Polaprezinc, a new synthetic antioxidant, significantly inhibited gastric mucosal injury and the accumulation of lipid peroxides in the gastric mucosa. All these results suggest that oxygen radicals and lipid peroxidation are involved in the pathogenesis of the acute gastric mucosal injury induced by TCA-5HT.
Erect of administering ginger (Zingiber officinale R.), one of the commonly consumed spices, on high-fat-fed rats was studied for a period of 10 weeks. Ginger was given at two different doses, 35mg and 70mg/kg body weight, made into a coarse solution with distilled water and administered orally by intragastric intubation daily. There was a significant decrease in the levels of cholesterol, phospholipids, and free fatty acids in the tissues (liver, intestine, kidney and aorta) and serum of the ginger-treated rats. Levels of serum triglyceride were also significantly reduced in the ginger-treated groups. Supplementation of the control and high-fat-fed rats with ginger increased the concentration of HDL and decreased the concentration of LDL and VLDL in the serum as compared with the levels in the rats not receiving the supplement. Thus, dietary intake of ginger was found to reduce the risk of atherosclerosis markedly by virtue of its hypolipidemic and antiatherogenic effects.
The effect of green tea extract on acute gastric mucosal damage induced by ischemia-reperfusion injury was investigated in rats. Ischemia-reperfusion injury was produced by applying a small vascular clamp to the celiac artery for 30min followed by removal of the clamp with reperfusion for 60min. An anti-ulcer effect of the green tea extract was demonstrated in this model. The increase seen in the lipid peroxide level in the gastric mucosa after ischemia-reperfusion was significantly inhibited by the extract. Tissue-associated myeloperoxidase activity, an index of neutrophil accumulation, was increased significantly in the gastric mucosa after reperfusion; this increase of activity was significantly inhibited by the green tea extract and paralleled the increase in the total area of gastric erosions. An electron spin resonance spin trapping study showed that the extract scavenged superoxide radicals generated by the hypoxanthine-xanthine oxidase system and that diphenyl-p-picryl-hydrazyl radicals were also eliminated in a concentration-dependent manner. In an in vitro study, the green tea extract significantly inhibited the increase in lipid peroxide in brain homogenates. Incubation of whole blood cells with interleukin-8 increased the expression of CD11b/CD18 by neutrophils, whereas co-incubation with the extract did not cause this upregulation. Human umbilical vein endothelial cells stimulated with interleukin-1β showed increased expression of E-selectin and intercellular adhesion molecule-1, but co-incubation with the extract significantly inhibited this upregulation. These results suggest that the protective effect of green tea extract against ischemia/reperfusion-induced gastric mucosal injury may be related to its antioxidant activity and inhibition of neutrophil accumulation.
To determine the peroxyl radical-scavenging activity of several biological substances, we developed a spin trapping assay using electron paramagnetic resonance (EPR). Peroxyl radicals, generated in a mixture of tert-butyl hydroperoxide and methemoglobin in sodium phosphate buffer, were trapped by 5, 5-dimethyl-1-pyrroline-N-oxide (DMPO). Scavenging activities were calculated from DMPO-OOR (tert-butyl peroxyl radical spin adduct) signal by a kinetic competition study. The scavenging activity, R, of a given substance was expressed as the relative second-order rate constant of the reaction with peroxyl radical as compared with that obtained for DMPO. Of the various biological substances evaluated, ascorbic acid (R=227), uric acid (R=28), glutathione (R=24), and α-tocopherol (R=14) were major peroxyl radical scavengers in phosphate buffer. This method will be useful for determining peroxyl radical-scavenging activities of biological materials.