SUMMARYAim: To investigate the inhibitory effects on gastric acid secretion of three proton pump inhibitors, omeprazole, lansoprazole and rabeprazole, using a three-way crossover design in healthy Helicobacter pylori-negative, S-mephenytoin 4¢-hydroxylase (CYP2C19) 1 homo-and hetero-extensive metabolizers. Methods: Eight healthy Japanese male volunteers were enrolled. After the administration of rabeprazole (10 mg ⁄ day), lansoprazole (30 mg ⁄ day) or omeprazole (20 mg ⁄ day), intragastric pH monitoring was commenced from 24 h before the first proton pump inhibitor dose, and continued for days 1-3 after proton pump inhibitor administration. The pH electrode was used for 48 h and changed just before pH monitoring on day 2. Results: For the administration of 10 mg ⁄ day rabeprazole, the mean ratios of the 24-h pH ‡ 3 holding time were 5.7 ± 1.1%,13.6 ± 2.2%, 35.3 ± 2.7% and 62.8 ± 3.1% for the pre-treatment day and days 1, 2 and 3, respectively. The same ratios for lansoprazole (30 mg ⁄ day) were 5.7 ± 0.7%, 7.4 ± 1.5%, 13.6 ± 3.4% and 26.6 ± 4.9%; the same ratios for 20 mg ⁄ day omeprazole were 5.9 ± 0.9%, 6.1 ± 1.2%, 11.4 ± 2.8% and 16.4 ± 4.6%. The mean ratio of the 24-h pH ‡ 3 holding time of days 1-3 increased significantly compared to the pre-treatment day (P < 0.01) with the administration of rabeprazole and lansoprazole. The magnitude of inhibition of gastric acid secretion after rabeprazole administration was stronger than that after lansoprazole. A significant elevation of the mean ratio of the 24-h pH ‡ 3 holding time was demonstrated on days 2 and 3 with omeprazole (P < 0.01). Conclusions: In H. pylori-negative CYP2C19 extensive metabolizers, rabeprazole (10 mg ⁄ day) shows a faster onset of rising intragastric pH and a stronger inhibition of gastric acid secretion than do lansoprazole (30 mg ⁄ day) or omeprazole (20 mg ⁄ day).

H¢lic~baeter pylorl has I~en identified as a ¢~ttaati~ a=enl in ,ctiw chronic $astritis, The rec=ptor for this baeteri=t, however, is not known, It is likely that the r~wptor molecules may b~ 81yt:osphinsolipid~.* as shown in th~ ea~cs orother bacteria. We explored this po~sihility by a thin-layer chromatography (~LC).immun~staininu method. Amontt lllyeo~phinllolipids extracted from hum;tn gastric mucosa, intact t1¢11¢.oba¢l¢t pylorl Sl~.cifleall~, bound to PSO~.O~tleer trod IPNeuAc.LacCer. whereas no specific bindin8 to neutral tll~sphint~olipids, ,,vhiel~ sh~re the same ¢era mide muiety with P$Os-OalCer or ! PNeuAe. LacCer, was demonstrated, $onieated bacteria could still bind to I PNeoAoLacCer with comparable affinity. In contrast, the btndinll of bacteria to PSO~.GaiCer was 8reatly diminished upon sonieation, Th~se results suS$est that each of the olisosaeehar.ide moieties mr I PNeuAc-LacCcr and PSO,-GalCer may be spccitlcally recol]nized by difl'eren~ liljand molecule= of llelieaba¢ter i)~,'larLH¢lirc~harr¢r pylorl: Gastric mucosa: Glycosphinsolipid; Bacterial adhesion

The FSSG score reflects the severity of the endoscopic findings of GERD.

The composition of the glycosphingolipids of the human gastrointestinal tract was studied. The major neutral glycosphingolipids were ceramide monohexosides (e.g., GalCer, GlcCer), LacCer, Gb3Cer, Gb4Cer and more polar ones with more than four sugars, whereas neither Gg3Cer nor Gg4Cer were present. The acidic glycosphingolipids consisted of sulfatides and gangliosides such as GM3, GM1, GD3 and GD1a. Also a large amount of sulfatides was found in the gastric mucosa and duodenum. The concentrations of sulfatides in the fundic mucosa, antral mucosa and duodenum amounted to 416.0, 933.8 and 682.9 nmol/g of dry weight, respectively, exceeding those in the gastric mucosa and kidney of other mammals. The major molecular species of the sulfatides were identified as I3SO3-GalCer with hydroxylated longer-chain fatty acids based on the analyses by gas-liquid chromatography and negative ion fast-atom bombardment mass spectrometry. In contrast, gangliosides in these regions showed a tendency to be lower than sulfatides, and the molar ratios of sulfatides to gangliosides were about 2.0, whereas those in other parts were less than 0.5. A high content of sulfatides in the gastric and duodenal mucosa, where mucosa is easily insulted by acid, pepsin and bile salts, may be closely related to their roles in mucosal protection.

A lectin from the fruiting body of Psathyrella velutina has been used as a specific probe for non-reducing terminal Nacetylglucosamine residues. We reveal in this report that P. velutina lectin recognizes a non-reducing terminal N-acetylneuraminic acid residue in glycoproteins and oligosaccharides. Binding of biotinyl P. velutina lectin to N-acetylneuraminic acid residues was prevented by desialylation of glycoconjugates and was distinguished from the binding to N-acetylglucosamine. Sialooligosaccharides were retarded or bound and eluted with Nacetylglucosamine on a P. velutina lectin column, being differentiated from each other and also from the oligosaccharides with non-reducing terminal N-acetylglucosamine which bound more strongly to the column.z 1999 Federation of European Biochemical Societies.

To evaluate the role of the histamine H2 receptor C terminus in signaling, desensitization, and agonist-induced internalization, canine H2 receptors with truncated C termini were generated. Wild-type (WT) and truncated receptors were tagged at their N termini with a hemagglutinin (HA) epitope and expressed in COS7 cells. Most of the C-terminal intracellular tail could be truncated (51 of 70 residues, termed T 308 mutant) without loss of functions: cAMP production, tiotidine binding, and plasma membrane targeting. In fact, the T 308 mutant produced more cAMP than the WT when cellsurface expression per cell was equivalent. As observed in a number of guanine nucleotide-binding protein-coupled receptors (GPCRs), 1 cAMP responses occurring via the histamine H2 receptor are rapidly desensitized after agonist stimulation (1-5). The H2 receptor also exhibits an agonist-induced internalization from the cell surface (2, 6). However, the mechanisms underlying these phenomena have yet to be identified. Recently, a number of studies focusing on GPCRs have examined the role of the C terminus in agonistinduced desensitization and internalization, with varying results. For example, C-terminal truncation of the angiotensin II receptor inhibited agonist-induced internalization of this receptor, but not desensitization of the calcium response mediated via the receptor (7). On the other hand, C-terminal truncation of ␤ 2 -adrenergic, ␣ 1B -adrenergic, lutropin/choriogonadotropin, platelet-activating factor, and neurokinin-2 receptors resulted in impairment of homologous desensitization (8 -12). Thus, the C termini of GPCRs have functional importances, which may differ among receptors. The present study was designed to analyze the role of this important portion of the histamine H2 receptor in signaling, desensitization, and agonist-induced internalization. To this end, we constructed H2 receptor cDNAs, devoid of either the 71 or the 51 amino acids at the C terminus, and expressed these cDNAs in COS7 cells. Herein, we present evidence that the 51 C-terminal amino acids of the histamine H2 receptor are important for agonist-induced internalization, but not for either signaling or homologous desensitization of cAMP response, which occurs via this receptor. In addition, these amino acids exert inhibitory effects on cAMP production via the H2 receptor.

A histamine H2 receptor, which had been mutated at its glycosylation site and tagged at its N-terminus with an HA tag (HA-H2 receptor), was expressed in Sf9 cells and COS7 cells. Immunoprecipitation and immunoblotting of HA-H2 receptors with otHA antibody revealed four bands of 31.5 ±2.5 kDa, 59.0 ± 6.0 kDa, 80.5 ± 4.5 kDa and 120 kDa. These bands were also detected by immunoblot using anti-H2 receptor serum (C-terminus). In addition, H2 receptors without the HA-tag coimmunoprecipitated with HA-tagged H2 receptors devoid of the 51 C-terminal amino acids, via immunoprecipitation with otHA antibody, when the two receptors were coexpressed. These results suggest that H2 receptors are present as receptor oligomers and that the C-terminal portion is not involved in the formation of these oligomers. © 1997 Federation of European Biochemical Societies.

G-protein-coupled receptors generally share a similar structure containing seven membrane-spanning domains and extracellular site(s) for N-glycosylation. The histamine H2 receptor is a member of the family of G-protein-coupled receptors, and has three extracellular potential sites for N-glycosylation (Asn-4, Asn-162 and Asn-168). To date, however, no information has been presented regarding N-glycosylation of the H2 receptor. To investigate the presence, location and functional roles of N-glycosylation of the H2 receptor, site-directed mutagenesis was performed to eliminate the potential site(s) for N-glycosylation singly and collectively. The wild-type and mutated H2 receptors were expressed stably in Chinese hamster ovary (CHO) cells or transiently in COS7 cells. Immunoblotting of the wild-type and mutated H2 receptors with an antiserum directed against the C-terminus of the H2 receptor showed that mutation at Asn-162, but not at Asn-168, resulted in a substantial decrease in the molecular mass. A mutation at Asn-4 led to a further decrease in the molecular mass. Tunicamycin treatment of the transfected cells yielded a sharp band with a molecular mass identical to that of the mutant devoid of all three potential sites for N-glycosylation. These findings indicate that the H2 receptor is N-glycosylated, and that N-glycosylation takes place mainly at two sites, Asn-4 and Asn-162. Neither the affinity for tiotidine nor that for histamine was affected by the mutagenesis. Immunocytochemistry and tiotidine binding showed that the mutated receptors were exclusively distributed on the cell surface in a fashion similar to that of the wild-type. In addition, the glycosylation-defective receptor was capable of activating adenylate cyclase and elevating the intracellular Ca2+ concentration in response to histamine in stable CHO cell lines. Thus N-glycosylation of the H2 receptor is not required for cell surface localization, ligand binding or functional coupling to G-protein(s).