Helicobacter pylori attaches to NeuAc alpha 2,3Gal beta 1,4 glycoconjugates produced in the stomach of transgenic mice lacking parietal cells.
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Helicobacter pylori infection of the human stomach is associated with altered acid secretion, loss of acid-producing parietal cells, and, in some hosts, adenocarcinoma. We have used a transgenic mouse model to study the effects of parietal cell ablation on H. pylori pathogenesis. Ablation results in amplification of the presumptive gastric epithelial stem cell and its immediate committed daughters. The amplified cells produce sialylated oncofetal carbohydrate antigens that function as receptors for H. pylori adhesins. Attachment results in enhanced cellular and humoral immune responses. NeuAc alpha 2,3Gal beta 1,4 glycoconjugates may not only facilitate persistent H. pylori infection in a changing gastric ecosystem, but by promoting interactions with lineage progenitors and/or initiated cells contribute to tumorigenesis in patients with chronic atrophic gastritis. (+info)
Morphological and histochemical variations of mucous and oxynticopeptic cells in the stomach of the seps, Chalcides chalcides.
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Mucous and oxynticopeptic cells in the gastric mucosa of the seps, Chalcides chalcides (Linnaeus, 1758) were examined by standard histochemical staining methods and by lectin histochemistry. The epithelial mucous cells lining the surface of the stomach and the mucous cells of the fundic glands elaborated mainly neutral glycoproteins with beta(1,4)GlcNAc oligomers, GalNAc glycosidic residues and Gal beta1,3GalNAc terminal sequences. The mucous cells of the fundic glands were stained specifically with the Paradoxical Con A method. The mucosecreting cells of the pyloric glands produced neutral glycoproteins, with beta(1,4)GlcNAc oligomers, GalNAc residues and Gal beta1,3GalNAc terminal sequences. Terminal L-fucose bound to the penultimate GlcNAc residues, and/or difucosylated oligosaccharides were also present. The pyloric glands did not stain with the Paradoxical Con A procedure. The morphology of the oxynticopeptic cells changes from the oral to the aboral region of the fundic mucosa. In the oral fundic tract the oxynticopeptic cells showed cytoplasm filled with zymogen granules, while in the aboral fundic region these cells contained few zymogen granules and showed cytoplasm full of empty vesicles, typical of the acid secreting cells. A secretion gradient of proteolytic enzymes and hydrochloric acid along the fundic mucosa of the seps can be hypothesised. (+info)
Effects of growth factors and trefoil peptides on migration and replication in primary oxyntic cultures.
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Restitution, the lateral migration of cells over an intact basement membrane, maintains mucosal integrity. We studied the regulation of migration and proliferation of enzyme-dispersed canine oxyntic mucosa cells in primary culture. Confluent monolayers were wounded and cultured in serum-free medium, and cells migrating into the wound were counted. [3H]thymidine incorporation into DNA was studied using subconfluent cultures. Considerable migration occurred in untreated monolayers; however, epidermal growth factor (EGF), transforming growth factor (TGF)-alpha, basic fibroblast growth factor (bFGF), insulin-like growth factor I (IGF-I), two trefoil peptides, and interleukin (IL)-1beta further enhanced migration. The specific EGF receptor (EGFR) monoclonal antibody, MAb-528, inhibited both basal and TGF-alpha- or IL-1beta-stimulated migration, but not the response to trefoil peptide, bFGF, or IGF-I. Exogenous TGF-beta inhibited cell proliferation but did not alter migration. Immunoneutralization with anti-TGF-beta blocked the response to exogenous TGF-beta and produced a small enhancement of basal thymidine incorporation but did not attenuate basal or TGF-alpha-stimulated migration. In conclusion, endogenous EGFR ligands regulate proliferation and migration. TGF-beta inhibits mitogenesis; it did not upregulate migration in these cultures. Although bFGF, IGF-I, and IL-1beta enhance gastric epithelial migration, only IL-1beta acted in a TGF-alpha-dependent fashion. (+info)
Parietal cells express high levels of Na-K-2Cl cotransporter on migrating into the gastric gland neck.
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Na-K-2Cl cotransport and Cl/HCO3 exchange are prominent mechanisms for Cl- uptake in Cl--secreting epithelial cells. We used immunofluorescence microscopy to delineate the distributions of Na-K-2Cl cotransporter-1 (NKCC1) and anion exchanger-2 (AE2) proteins in rat gastric mucosa (zymogenic zone). Parietal cells (PCs) above the neck of the gastric gland contained abundant AE2 but little or no NKCC1, whereas those in the neck and base contained high NKCC1 but diminished AE2. Lower levels of NKCC1 were detected in surface mucous cells and in cells comprising the blind ends of all glands. Pulse labeling of proliferating cells with bromodeoxyuridine indicated that new PCs originate in the isthmus with scant NKCC1; the subset of PCs that migrate downward expresses NKCC1 abruptly on entering the neck, within 7 days of cell division. Our results suggest that downwardly migrating PCs replace one mechanism for Cl- entry (Cl/HCO3 exchange) with another (Na-K-2Cl cotransport). (+info)
Species-specific distribution of alpha-galactosyl epitopes on the gastric H/K ATPase beta-subunit: relevance to the binding of human anti-parietal cell autoantibodies.
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The gastric H/K ATPase beta-subunit, an abundant glycoprotein of the secretory membranes of gastric parietal cells, is the major autoantigen recognized by human parietal cell autoantibodies in gastric autoimmunity. Our previous studies demonstrated that the human autoantibodies recognize the H/K ATPase beta-subunit from a number of species and that glycosylation of the beta-subunit with complex N-glycans is required for autoantibody binding. The N-glycans of the beta-subunit contain polylactosamine chains. The lactosamine chains of the rabbit beta-subunit are terminated with alpha-linked galactosyl residues (alpha-galactosyl epitope) (Tyagarajan et al., Biochemistry, 1996, 35, 3238-3246). Here we have investigated the expression of alpha-galactosyl epitopes on the H/K ATPase beta-subunit from a number of species. Using the alpha-galactosyl binding lectin, BS1-IB4, and naturally occurring anti-alpha-galactosyl antibodies, we have demonstrated that the rat H/K ATPase beta-subunit also contains terminal alpha-galactosyl residues, but not the beta-subunit from pig, dog, and mouse, indicating species-specific differences in the terminal saccharide sequences of the beta-subunit. We also investigated the potential contribution of the alpha-galactosyl epitopes to the binding by human sera. The reactivity of human pernicious anemia serum with gastric parietal cells could not be inhibited with saccharide inhibitors and, in addition, no binding was observed with normal human sera. We conclude that the H/K ATPase beta-subunit oligosaccharides from rabbit and rat are terminated with alpha-galactosyl epitopes, and although the presence of this epitope does not contribute to binding by human parietal cell autoantibodies at the concentrations routinely used, it is recommended that neither rat or rabbit stomachs be used for screening human sera. (+info)
Carbachol activates ERK2 in isolated gastric parietal cells via multiple signaling pathways.
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We previously reported that both carbachol and epidermal growth factor (EGF) are potent inducers of the extracellular signal-regulated protein kinases (ERKs) in isolated gastric canine parietal cells and that induction of these kinases leads to acute inhibitory and chronic stimulatory effects on gastric acid secretion. In this study we investigated the molecular mechanisms responsible for these effects. Both carbachol (100 microM) and EGF (10 nM) induced Ras activation. The role of Ras in ERK2 induction was examined by transfecting parietal cells with a vector expressing hemoagglutinin (HA)-tagged ERK2 (HA-ERK2) together with a dominantly expressed mutant (inactive) ras gene. HA-ERK2 activity was quantitated by in-gel kinase assays. Dominant negative Ras reduced carbachol induction of HA-ERK2 activity by 60% and completely inhibited the stimulatory effect of EGF. Since Ras activation requires the assembly of a multiprotein complex, we examined the effect of carbachol and EGF on tyrosyl phosphorylation of Shc and its association with Grb2 and the guanine nucleotide exchange factor Sos. Western blot analysis of anti-Shc immunoprecipitates with an anti-phosphotyrosine antibody demonstrated that both carbachol and EGF induced tyrosyl phosphorylation of a major 52-kDa shc isoform. Grb2 association with Shc was demonstrated by blotting Grb2 immunoprecipitates with an anti-Shc antibody. Probing of anti-Sos immunoprecipitates with an anti-Grb2 antibody revealed that Sos was constitutively bound to Grb2. To examine the functional role of Sos in ERK2 activation, we transfected parietal cells with the HA-ERK2 vector together with a dominantly expressed mutant (inactive) sos gene. Dominant negative Sos did not affect carbachol stimulation of HA-ERK2 but inhibited the stimulatory effect of EGF by 60%. We then investigated the role of betagamma-subunits in carbachol induction of HA-ERK2. Parietal cells were transfected with the HA-ERK2 vector together with a vector expressing the carboxy terminus of the beta-adrenergic receptor kinase 1, known to block signaling mediated by betagamma-subunits. In the presence of this vector, carbachol induction of HA-ERK2 was inhibited by 40%. Together these data suggest that, in the gastric parietal cells, carbachol activates the ERKs through Ras- and betagamma-dependent mechanisms that require guanine nucleotide exchange factors other than Sos. (+info)
Kex2 family endoprotease furin is expressed specifically in pit-region parietal cells of the rat gastric mucosa.
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The proprotein-processing endoprotease furin is localized in the gastric epithelial cells of the pit region in the rat gastric gland. The gastric pit is composed of several cell types, including gastric surface mucosal (GSM) cells and parietal cells. Furin converts many growth- or differentiation-related proproteins to their active forms. We examined identification of furin-positive cells by immunostaining of rat gastric mucosa and regulators of the furin expression by measuring the furin promoter activity by luciferase assay. Furin-positive cells were stained for H(+)-K(+)-ATPase, indicating that they are parietal cells. Furin-positive parietal cells were not stained for transforming growth factor-alpha (TGF-alpha) but were surrounded by TGF-alpha-positive GSM cells. In contrast, parietal cells below the proliferative zone were positive for TGF-alpha but not for furin. Furin-positive parietal cells expressed a high level of epidermal growth factor receptor (EGFR). TGF-alpha stimulated the furin promoter activity highly in a mouse GSM cell line GSM06. Thus we suggest that the parietal cells of the pit region have furin-mediated functions that can be stimulated by EGFR signaling. (+info)
Development of the actin and the cytokeratin cytoskeletons of parietal cells during differentiation of the rat gastric mucosa.
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Available evidence strongly suggests that microfilaments and cytokeratin intermediate filaments (IF) play a role in the reorganization of the luminal pole required for the secretion of acid by parietal cells. To correlate the organization of both cytoskeletal systems with the differentiation of the secretory membranes of parietal cells, the distribution of F-actin and cytokeratin was studied during the ontogenic development of the rat. Primitive parietal cells were detected with parietal cells autoantibodies and ultrastructurally by transmission electron microscopy (TEM). The distribution of IF and of F-actin in differentiating parietal cells was determined using anticytokeratin antibodies and FITC-phalloidin, respectively. Development of both cytoskeletal systems was followed by TEM. Ultrastructurally, parietal cells are identified from day 19 on, by the presence of an incipient canaliculus, which later enlarges and fills with microvilli. No intracellular tubulovesicular system is observed. Using parietal cells autoantibodies these cells are detected from day 20 on. Immunocytochemistry and TEM demonstrate that parietal cells possess organized cytokeratin and actin cytoskeletons, which develop further as differentiation proceeds. At birth, parietal cells show an ultrastructure and a distribution of IF and microfilaments similar to that of differentiated cells. In newly born rats, the F-actin cytoskeleton redistributes after suckling. This reorganization results from an enlargement of the canalicular lumen, filled with microvilli rich in actin. Thus, functional maturation of parietal cells is paralleled by the development of organized IF and F-actin cytoskeletons associated to the secretory surface. (+info)