RESUMO
BACKGROUND & AIMS: The homeostasis of the gastrointestinal epithelium relies on cell regeneration and differentiation into distinct lineages organized inside glands and crypts. Regeneration depends on Wnt/ß-catenin pathway activation, but to understand homeostasis and its dysregulation in disease, we need to identify the signaling microenvironment governing cell differentiation. By using gastric glands as a model, we have identified the signals inducing differentiation of surface mucus-, zymogen-, and gastric acid-producing cells. METHODS: We generated mucosoid cultures from the human stomach and exposed them to different growth factors to obtain cells with features of differentiated foveolar, chief, and parietal cells. We localized the source of the growth factors in the tissue of origin. RESULTS: We show that epidermal growth factor is the major fate determinant distinguishing the surface and inner part of human gastric glands. In combination with bone morphogenetic factor/Noggin signals, epidermal growth factor controls the differentiation of foveolar cells vs parietal or chief cells. We also show that epidermal growth factor is likely to underlie alteration of the gastric mucosa in the precancerous condition atrophic gastritis. CONCLUSIONS: Use of our recently established mucosoid cultures in combination with analysis of the tissue of origin provided a robust strategy to understand differentiation and patterning of human tissue and allowed us to draw a new, detailed map of the signaling microenvironment in the human gastric glands.
Assuntos
Padronização Corporal/efeitos dos fármacos , Proteína Morfogenética Óssea 4/farmacologia , Diferenciação Celular/efeitos dos fármacos , Fator de Crescimento Epidérmico/farmacologia , Células Epiteliais/efeitos dos fármacos , Mucosa Gástrica/efeitos dos fármacos , Proteínas de Transporte/farmacologia , Linhagem da Célula , Células Cultivadas , Microambiente Celular , Celulas Principais Gástricas/efeitos dos fármacos , Celulas Principais Gástricas/metabolismo , Celulas Principais Gástricas/ultraestrutura , Células Epiteliais/metabolismo , Células Epiteliais/ultraestrutura , Mucosa Gástrica/metabolismo , Mucosa Gástrica/ultraestrutura , Gastrite Atrófica/metabolismo , Gastrite Atrófica/patologia , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Organoides , Células Parietais Gástricas/efeitos dos fármacos , Células Parietais Gástricas/metabolismo , Células Parietais Gástricas/ultraestrutura , Via de Sinalização WntRESUMO
BACKGROUND/OBJECTIVE: The present study investigated the potent therapeutic effects of Ruscogenin, main steroid sapogenin of traditional Chinese plant called 'Ophiopogon japonicas', on chronic ulcer model established with acetic acid in rats. METHODS: 24 rats were attenuated to the sham (2 ml/kg/day isotonic solution), control (untreated ulcer) and treatment (3 ml/kg/day ruscogenin) groups. After treatment for 2 weeks, gastric tissues were collected and prepared for light microscopic (H&E), immunohistochemical (Collagen I, III and IV) and biochemical analysis [Epidermal growth factor (EGF), Prostaglandin E2 (PGE2), Tumor Necrosis Factor alpha (TNF-α), Interleukin 6 and 8 (IL-6 and IL-8), Lipid Peroxidase (LPO), Myeloperoxidase (MPO), Glutathione (GSH) and Glutathione Peroxidase (GSH-Px)] and transmission electron microscopy (TEM). RESULTS: Macroscopic scoring showed that the ulceration area of ruscogenin-treated group decreased compared with control group. Immunohistochemical analysis revealed ruscogenin ameliorated and restored the levels of Collagen I and IV to the levels of sham group. Tissue levels of EGF and PGE2 enhanced significantly in untreated ulcer group while were higher in treated ulcer group than the control group. TNF-α, IL-6, IL-8, LPO, MPO levels increased significantly in control group whereas decreased in treated rats after ruscogenin treatment. However, levels of GSH and GSH-Px increased significantly in treatment group. TEM showed chief cells and parietal cells of ulcer group having degenerated organelles while ruscogenin group had normal ultrastructure of cells. CONCLUSION: There are potent anti-inflammatory and anti-oxidant effects of ruscogenin on gastric ulcer and may be successfully used as a safe and therapeutic agent in treatment of peptic ulcer.
Assuntos
Fitoterapia , Espirostanos/administração & dosagem , Úlcera Gástrica/tratamento farmacológico , Animais , Doença Crônica , Colágeno/metabolismo , Citocinas/metabolismo , Dinoprostona/metabolismo , Modelos Animais de Doenças , Fator de Crescimento Epidérmico/metabolismo , Feminino , Microscopia Eletrônica de Transmissão , Ophiopogon/química , Células Parietais Gástricas/patologia , Células Parietais Gástricas/ultraestrutura , Peroxidases/metabolismo , Ratos Sprague-Dawley , Espirostanos/isolamento & purificação , Úlcera Gástrica/metabolismo , Úlcera Gástrica/patologia , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Digestion in the stomach depends on acidification of the lumen. Histamine-elicited acid secretion is triggered by activation of the PKA cascade, which ultimately results in the insertion of gastric H,K-ATPases into the apical plasma membranes of parietal cells. Our recent study revealed the functional role of PKA-MST4-ezrin signaling axis in histamine-elicited acid secretion. However, it remains uncharacterized how the PKA-MST4-ezrin signaling axis operates the insertion of H,K-ATPases into the apical plasma membranes of gastric parietal cells. Here we show that MST4 phosphorylates ACAP4, an ARF6 GTPase-activating protein, at Thr545 Histamine stimulation activates MST4 and promotes MST4 interaction with ACAP4. ACAP4 physically interacts with MST4 and is a cognate substrate of MST4 during parietal cell activation. The phosphorylation site of ACAP4 by MST4 was mapped to Thr545 by mass spectrometric analyses. Importantly, phosphorylation of Thr545 is essential for acid secretion in parietal cells because either suppression of ACAP4 or overexpression of non-phosphorylatable ACAP4 prevents the apical membrane reorganization and proton pump translocation elicited by histamine stimulation. In addition, persistent overexpression of MST4 phosphorylation-deficient ACAP4 results in inhibition of gastric acid secretion and blockage of tubulovesicle fusion to the apical membranes. Significantly, phosphorylation of Thr545 enables ACAP4 to interact with ezrin. Given the location of Thr545 between the GTPase-activating protein domain and the first ankyrin repeat, we reason that MST4 phosphorylation elicits a conformational change that enables ezrin-ACAP4 interaction. Taken together, these results define a novel molecular mechanism linking the PKA-MST4-ACAP4 signaling cascade to polarized acid secretion in gastric parietal cells.
Assuntos
Proteínas do Citoesqueleto/metabolismo , Proteínas Ativadoras de GTPase/metabolismo , ATPase Trocadora de Hidrogênio-Potássio/metabolismo , Modelos Biológicos , Células Parietais Gástricas/metabolismo , Processamento de Proteína Pós-Traducional , Proteínas Serina-Treonina Quinases/metabolismo , Substituição de Aminoácidos , Animais , Membrana Celular/enzimologia , Membrana Celular/metabolismo , Membrana Celular/ultraestrutura , Polaridade Celular , Células Cultivadas , Biologia Computacional , Proteínas do Citoesqueleto/química , Proteínas do Citoesqueleto/genética , Bases de Dados de Proteínas , Proteínas Ativadoras de GTPase/química , Proteínas Ativadoras de GTPase/genética , Microscopia Eletrônica de Transmissão , Mutação , Células Parietais Gástricas/citologia , Células Parietais Gástricas/ultraestrutura , Fosforilação , Conformação Proteica , Multimerização Proteica , Proteínas Serina-Treonina Quinases/química , Proteínas Serina-Treonina Quinases/genética , Transporte Proteico , Coelhos , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/metabolismoRESUMO
We hypothesized that basic helix-loop-helix (bHLH) MIST1 (BHLHA15) is a "scaling factor" that universally establishes secretory morphology in cells that perform regulated secretion. Here, we show that targeted deletion of MIST1 caused dismantling of the secretory apparatus of diverse exocrine cells. Parietal cells (PCs), whose function is to pump acid into the stomach, normally lack MIST1 and do not perform regulated secretion. Forced expression of MIST1 in PCs caused them to expand their apical cytoplasm, rearrange mitochondrial/lysosome trafficking, and generate large secretory granules. Mist1 induced a cohort of genes regulated by MIST1 in multiple organs but did not affect PC function. MIST1 bound CATATG/CAGCTG E boxes in the first intron of genes that regulate autophagosome/lysosomal degradation, mitochondrial trafficking, and amino acid metabolism. Similar alterations in cell architecture and gene expression were also caused by ectopically inducing MIST1 in vivo in hepatocytes. Thus, MIST1 is a scaling factor necessary and sufficient by itself to induce and maintain secretory cell architecture. Our results indicate that, whereas mature cell types in each organ may have unique developmental origins, cells performing similar physiological functions throughout the body share similar transcription factor-mediated architectural "blueprints."
Assuntos
Regulação da Expressão Gênica/genética , Fator de Crescimento de Hepatócito/genética , Fator de Crescimento de Hepatócito/metabolismo , Células Parietais Gástricas/citologia , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Via Secretória/genética , Células Acinares/citologia , Células Acinares/efeitos dos fármacos , Células Acinares/metabolismo , Animais , Antineoplásicos Hormonais/farmacologia , Linhagem Celular , Expressão Ectópica do Gene/efeitos dos fármacos , Deleção de Genes , Regulação da Expressão Gênica/efeitos dos fármacos , Camundongos , Células Parietais Gástricas/efeitos dos fármacos , Células Parietais Gástricas/metabolismo , Células Parietais Gástricas/ultraestrutura , Tamoxifeno/farmacologiaRESUMO
BACKGROUND: Obesity is associated with risk of adenocarcinoma in the proximal stomach. We aimed to identify the links between dietary fat and gastric premalignant lesions. METHODS: C57BL/6 mice were fed high fat diet (HFD), and gastric mucosa was histologically analysed. Morphological changes were also analysed using an electron microscope. Transcriptome analysis of purified parietal cells was performed, and non-parietal gastric corpus epithelial cells were subjected to single-cell gene-expression profiling. Composition of gastric contents of HFD-fed mice was compared with that of the HFD itself. Lipotoxicity of free fatty acids (FFA) was examined in primary culture and organoid culture of mouse gastric epithelial cells in vitro, as well as in vivo, feeding FFA-rich diets. RESULTS: During ~8-20 weeks of HFD feeding, the parietal cells of the stomach displayed mitochondrial damage, and a total of 23% of the mice developed macroscopically distinct metaplastic lesions in the gastric corpus mucosa. Transcriptome analysis of parietal cells indicated that feeding HFD enhanced pathways related to cell death. Histological analysis and gene-expression profiling indicated that the lesions were similar to previously reported precancerous lesions identified as spasmolytic polypeptide-expressing metaplasia. FFAs, including linoleic acid with refluxed bile acids were detected in the stomachs of the HFD-fed mice. In vitro, FFAs impaired mitochondrial function and decreased the viability of parietal cells. In vivo, linoleic acid-rich diet, but not stearic acid-rich diet induced parietal-cell loss and metaplastic changes in mice. CONCLUSIONS: Dietary lipids induce parietal-cell damage and may lead to the development of precancerous metaplasia.
Assuntos
Dieta Hiperlipídica/efeitos adversos , Gorduras na Dieta/efeitos adversos , Ácidos Graxos/efeitos adversos , Mucosa Gástrica/patologia , Células Parietais Gástricas/patologia , Lesões Pré-Cancerosas/patologia , Animais , Ácidos e Sais Biliares/metabolismo , Morte Celular/genética , Células Cultivadas , Gorduras na Dieta/administração & dosagem , Células Epiteliais/patologia , Ácidos Graxos/administração & dosagem , Ácidos Graxos/metabolismo , Ácidos Graxos não Esterificados/metabolismo , Suco Gástrico/metabolismo , Perfilação da Expressão Gênica , Ácido Linoleico/administração & dosagem , Ácido Linoleico/efeitos adversos , Ácido Linoleico/metabolismo , Masculino , Metaplasia/genética , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Mitocôndrias/ultraestrutura , Células Parietais Gástricas/metabolismo , Células Parietais Gástricas/ultraestrutura , Lesões Pré-Cancerosas/genética , Cultura Primária de Células , Ácidos Esteáricos/administração & dosagem , Ácidos Esteáricos/efeitos adversosRESUMO
Ezrin, an adaptor protein that cross-links plasma membrane-associated proteins with the actin cytoskeleton, is concentrated on apical surfaces of epithelial cells, especially in microvilli of the small intestine and stomach. In the stomach, ezrin is predominantly expressed on the apical canalicular membrane of parietal cells. Transgenic ezrin knockdown mice in which the expression level of ezrin was reduced to <7% compared with the wild-type suffered from achlorhydria because of impairment of membrane fusion between tubulovesicles and apical membranes. We observed, for the first time, hypergastrinemia and foveolar hyperplasia in the gastric fundic region of the knockdown mice. Dilation of fundic glands was observed, the percentage of parietal and chief cells was reduced, and that of mucous-secreting cells was increased. The parietal cells of knockdown mice contained dilated tubulovesicles and abnormal mitochondria, and subsets of these cells contained abnormal vacuoles and multilamellar structures. Therefore, lack of ezrin not only causes achlorhydria and hypergastrinemia but also changes the structure of gastric glands, with severe perturbation of the secretory membranes of parietal cells.
Assuntos
Proteínas do Citoesqueleto/metabolismo , Epitélio/fisiologia , Mucosa Gástrica/fisiologia , Regulação da Expressão Gênica/fisiologia , Células Parietais Gástricas/metabolismo , Acloridria/metabolismo , Animais , Anticorpos , Proteínas do Citoesqueleto/genética , Mucosa Gástrica/citologia , Gastrinas/sangue , Técnicas de Silenciamento de Genes , Lectinas , Camundongos , Camundongos Transgênicos , Microscopia de Fluorescência , Mucina-6/genética , Mucina-6/metabolismo , Mucinas/genética , Mucinas/metabolismo , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Células Parietais Gástricas/ultraestrutura , Peptídeos/genética , Peptídeos/metabolismo , RNA/genética , RNA/metabolismo , Fator Trefoil-2 , Regulação para CimaRESUMO
The present studies were designed to examine the effects of ClC-2 ablation on cellular morphology, parietal cell abundance, H/K ATPase expression, parietal cell ultrastructure and acid secretion using WT and ClC-2-/- mouse stomachs. Cellular histology, morphology and proteins were examined using imaging techniques, electron microscopy and western blot. The effect of histamine on the pH of gastric contents was measured. Acid secretion was also measured using methods and secretagogues previously established to give maximal acid secretion and morphological change. Compared to WT, ClC-2-/- gastric mucosal histological organization appeared disrupted, including dilation of gastric glands, shortening of the gastric gland region and disorganization of all cell layers. Parietal cell numbers and H/K ATPase expression were significantly reduced by 34% (P<0.05) and 53% (P<0.001) respectively and cytoplasmic tubulovesicles appeared markedly reduced on electron microscopic evaluation without evidence of canalicular expansion. In WT parietal cells, ClC-2 was apparent in a similar cellular location as the H/K ATPase by immunofluorescence and appeared associated with tubulovesicles by immunogold electron microscopy. Histamine-stimulated [H+] of the gastric contents was significantly (P<0.025) lower by 9.4 fold (89%) in the ClC-2-/- mouse compared to WT. Histamine/carbachol stimulated gastric acid secretion was significantly reduced (range 84-95%, P<0.005) in ClC-2-/- compared to WT, while pepsinogen secretion was unaffected. Genetic ablation of ClC-2 resulted in reduced gastric gland region, reduced parietal cell number, reduced H/K ATPase, reduced tubulovesicles and reduced stimulated acid secretion.
Assuntos
Canais de Cloreto/genética , Digestão/fisiologia , Ácido Gástrico/metabolismo , ATPase Trocadora de Hidrogênio-Potássio/biossíntese , Células Parietais Gástricas/metabolismo , Animais , Transporte Biológico , Canais de Cloro CLC-2 , Contagem de Células , Vesículas Citoplasmáticas/metabolismo , Digestão/genética , Imunofluorescência , ATPase Trocadora de Hidrogênio-Potássio/metabolismo , Camundongos , Camundongos Knockout , Microscopia Eletrônica , Células Parietais Gástricas/ultraestrutura , Pepsinogênio A/metabolismoRESUMO
OBJECTIVE: To investigate whether the ultrastructure and hydrogen potassium adenosine triphosphate (H+ /K+ ATPase) expression of human parietal cells were associated with aging. METHODS: In all, 50 participants who underwent gastroscopy due to dyspepsia were divided into two age groups, with 19 in the younger group (YG, aged 20-59 years) and 31 in the elder group (EG, aged ≥60 years). The ultrastructure of their parietal cell was determined by electron microscopy (EM), and the expressions of H+ /K+ ATPase α-subunit mRNA and ß-unit protein were detected. Furthermore, 24-h esophageal pH monitoring was performed in the two groups. RESULTS: EM images showed no distinct difference in the morphology and distribution of parietal cells or the acid secretion-related organelle between the two groups. There were no differences between YG and EG in the proportion of mitochondria and the tubulovesicular system area. The expressions of H+ /K+ ATPase α-subunit mRNA and ß-subunit protein showed no age-related alteration between YG and EG. The expression of H+ /K+ ATPase α-subunit mRNA in EG was higher than that in YG, whereas the expression of ß-subunit protein was significantly higher in those aged ≥80 years than in the YG. No significant difference was found in the 24-h esophageal pH monitoring between YG and EG. CONCLUSION: Acid secretion-related organelles in parietal cells do not degenerate with aging, the expression of H+ /K+ ATPase even shows a trend to increase, indicating the existence of intact molecular biological basis for acid secretion in healthy elderly individuals.
Assuntos
Envelhecimento/metabolismo , Ácido Gástrico/metabolismo , ATPase Trocadora de Hidrogênio-Potássio/biossíntese , Células Parietais Gástricas/enzimologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Envelhecimento/genética , Envelhecimento/patologia , Dispepsia/enzimologia , Dispepsia/patologia , Monitoramento do pH Esofágico , Mucosa Gástrica/enzimologia , Gastroscopia , Regulação Enzimológica da Expressão Gênica/fisiologia , ATPase Trocadora de Hidrogênio-Potássio/genética , Humanos , Microscopia Eletrônica , Pessoa de Meia-Idade , Células Parietais Gástricas/ultraestrutura , RNA Mensageiro/genética , Adulto JovemRESUMO
EAC in its early stages, when it can potentially be cured, is rarely symptomatic and is associated with high mortality rates because in part of late-stage diagnosis. Given that DNA repair is an important contributory factor of early-stage malignancy, our study focused on the expression of the base excision repair enzyme N-methylpurine DNA glycosylase (MPG) in EAC disease onset. MPG messenger RNA (mRNA) expression levels were determined using quantitative reverse transcriptase polymerase chain reaction from a maximum of 72 patient samples. Immunohistochemistry was further utilized for the detection of MPG protein, and semiquantitative analysis performed using an H-score approach was carried out on a total of 130 archival tissue samples of different esophageal pathologies. Nuclear localized MPG protein was detected in all nonmalignant tissues derived from the enterohepatic system, with H-score values of 3.9-5.5 ± 0.4-1.0. In cancerous tissues derived from the enterohepatic system, a 9.5-fold increase in the level of MPG mRNA expression was specifically observed in the malignant regions located within the esophagus region. Further analysis revealed a 9- and 14-fold increase in MPG mRNA expression in EAC tumor, node, metastasis stages II and III, respectively, suggesting MPG expression to correlate with EAC disease progression. Immunohistochemistry analysis further showed a sevenfold significant increase in MPG protein expression in EAC tissues. Intriguingly, there was a fivefold significant decrease in nuclear localized MPG protein expression in tissues derived from Barrett's esophagus and low-grade dysplasia. Such findings highlight a complex regulatory pattern governing DNA glycosylase base excision repair initiation, as normal tissue undergoes Barrett's metaplasia and later dedifferentiates to EAC. Indeed, disease-stage-specific alterations in the expression of MPG may highlight a potential role for MPG in determining EAC onset and thus potentially be of clinical relevance for early disease detection and increased patient survival.
Assuntos
Adenocarcinoma/diagnóstico , Biomarcadores Tumorais/análise , DNA Glicosilases/análise , Reparo do DNA , Neoplasias Esofágicas/diagnóstico , Adenocarcinoma/patologia , Adenocarcinoma/secundário , Esôfago de Barrett/patologia , Estudos de Casos e Controles , Núcleo Celular/ultraestrutura , Transformação Celular Neoplásica/patologia , DNA Glicosilases/genética , Progressão da Doença , Detecção Precoce de Câncer , Neoplasias Esofágicas/patologia , Humanos , Mucosa Intestinal/citologia , Túbulos Renais/citologia , Linfonodos/patologia , Células Mesangiais/ultraestrutura , Metaplasia , Estadiamento de Neoplasias , Células Parietais Gástricas/ultraestrutura , Projetos Piloto , Lesões Pré-Cancerosas/patologia , RNA Mensageiro/análise , Estudos RetrospectivosRESUMO
The subcellular behavior of aluminum and indium, used in medical and industrial fields, was studied in the gastric mucosa and the liver after their intragastric administration to rats, using, two of the most sensitive methods of observation and microanalysis, the transmission electron microscopy, and the secondary ion mass spectrometry. The ultrastructural study showed the presence of electron dense deposits, in the lysosomes of parietal and principal gastric mucosa cells but no loaded lysosomes were observed in the different studied hepatic territories. The microanalytical study allowed the identification of the chemical species present in those deposits as aluminum or indium isotopes and the cartography of their distribution. No modification was observed in control rats tissues. In comparison to previous studies describing the mechanism of aluminum concentration in the gastric mucosa and showing that this element was concentrated in the lysosomes of fundic and antral human gastric mucosa, our study provided additional informations about the types of cells involved in the phenomenon of concentration of aluminum and indium, which are the parietal and the principal cells of the gastric mucosa. Our study demonstrated that these cells have the ability to concentrate selectively aluminum and indium in their lysosomes, as a defensive reaction against intoxication by foreign elements.
Assuntos
Compostos de Alumínio/farmacocinética , Mucosa Gástrica/efeitos dos fármacos , Índio/farmacocinética , Nitratos/farmacocinética , Células Parietais Gástricas/metabolismo , Compostos de Alumínio/administração & dosagem , Animais , Microanálise por Sonda Eletrônica , Mucosa Gástrica/anatomia & histologia , Mucosa Gástrica/metabolismo , Índio/administração & dosagem , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/ultraestrutura , Lisossomos/efeitos dos fármacos , Lisossomos/ultraestrutura , Masculino , Microscopia Eletrônica de Transmissão , Nitratos/administração & dosagem , Células Parietais Gástricas/efeitos dos fármacos , Células Parietais Gástricas/ultraestrutura , Ratos , Ratos Wistar , Espectrometria de Massa de Íon SecundárioAssuntos
Coristoma/diagnóstico , Doenças do Esôfago/diagnóstico , Mucosa Gástrica , Gastroscopia , Microscopia Confocal/métodos , Coristoma/patologia , Meios de Contraste , Doenças do Esôfago/patologia , Esofagite/etiologia , Feminino , Fluoresceína , Ácido Gástrico/metabolismo , Gastroscópios , Humanos , Microscopia Confocal/instrumentação , Células Parietais Gástricas/metabolismo , Células Parietais Gástricas/ultraestrutura , Inibidores da Bomba de Prótons/uso terapêutico , Adulto JovemRESUMO
BACKGROUND & AIMS: Mutations in TRPML1, a lysosomal Ca(2+)-permeable TRP channel, lead to mucolipidosis type IV, a neurodegenerative lysosomal storage disease. An unusual feature of mucolipidosis type IV is constitutive achlorhydria. We produced Trpml1(-/-) (null) mice to investigate the requirement for this protein in gastric acid secretion. METHODS: Trpml1-null mice were generated by gene targeting. The expression of Trpml1 and its role in acid secretion by gastric parietal cells were analyzed using biochemical, histologic, and ultrastructural approaches. RESULTS: Trpml1 is expressed by parietal cells and localizes predominantly to the lysosomes; it was dynamically palmitoylated and dephosphorylated in vivo following histamine stimulation of acid secretion. Trpml1-null mice had significant impairments in basal and histamine-stimulated gastric acid secretion and markedly reduced levels of the gastric proton pump. Histologic and ultrastructural analyses revealed that Trpml1(-/-) parietal cells were enlarged, had multivesicular and multi-lamellated lysosomes, and maintained an abnormal intracellular canalicular membrane. The intralysosomal Ca(2+) content and receptor-mediated Ca(2+) signaling were, however, unaffected in Trpml1(-/-) gastric glands, indicating that Trpml1 does not function in the regulation of lysosomal Ca(2+). CONCLUSIONS: Loss of Trpml1 causes reduced levels and mislocalization of the gastric proton pump and alters the secretory canaliculi, causing hypochlorhydria and hypergastrinemia. The lysosomal enlargement and defective intracellular canaliculi formation observed in Trpml1(-/-) parietal cells indicate that Trpml1 functions in the formation and trafficking of the tubulovesicles. This study provides direct evidence for the regulation of gastric acid secretion by a TRP channel; TRPML1 is an important protein in parietal cell apical membrane trafficking.
Assuntos
Ácido Gástrico/metabolismo , Mucolipidoses/metabolismo , Células Parietais Gástricas/metabolismo , Canais de Cátion TRPM/deficiência , Acloridria/genética , Acloridria/metabolismo , Acloridria/fisiopatologia , Animais , Cálcio/metabolismo , Modelos Animais de Doenças , Gastrinas/sangue , ATPase Trocadora de Hidrogênio-Potássio/metabolismo , Histamina/metabolismo , Hipertrofia , Lipoilação , Lisossomos/metabolismo , Lisossomos/ultraestrutura , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mucolipidoses/genética , Mucolipidoses/patologia , Mucolipidoses/fisiopatologia , Células Parietais Gástricas/ultraestrutura , Fosforilação , Transporte Proteico , Canais de Cátion TRPM/genética , Fatores de Tempo , Canais de Potencial de Receptor TransitórioRESUMO
Ultrastructural characteristics of cell populations in the gastric tube mucosa were studied after plastic surgery for benign esophageal disease. The foveolar epithelium mainly consisted of secretory active mucocytes with small changes in cytoplasmic organelles. Functionally immature parietal cells and gland cells in the epithelium of fundal glands were characterized by atrophy and degeneration. Ultrastructural features of the epithelial compartment attest to adaptive cellular hypersecretion of the mucoid against the background of impaired acid production and enzyme secretion. The stromal compartment was characterized by low pinocytotic activity of endothelial cells in blood vessels and polymorphism of smooth muscle cells. Hypertrophic leiomyocytes were shown to dominate in the population of these cells. Some cells had signs of biosynthesis and reduction of filaments. Single leiomyocytes were degenerated. The observed ultrastructural modification of epithelial and connective cells can be interpreted as structural adaptation of the gastric transplant.
Assuntos
Epitélio/ultraestrutura , Esofagoplastia , Mucosa Gástrica/citologia , Mucosa Gástrica/ultraestrutura , Adulto , Células Endoteliais/citologia , Células Endoteliais/ultraestrutura , Esôfago/citologia , Esôfago/patologia , Esôfago/cirurgia , Feminino , Humanos , Masculino , Microscopia Eletrônica , Pessoa de Meia-Idade , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/ultraestrutura , Células Parietais Gástricas/ultraestruturaRESUMO
Epsin is a ubiquitin-binding endocytic adaptor, which is highly concentrated at clathrin-coated pits and coordinates acquisition of bilayer curvature with coat recruitment and cargo selection. Epsin is encoded by three distinct genes in mammals. Epsin 1 and 2 have broad tissue distribution with high-level expression in the brain. In contrast, epsin 3 was reported to be expressed primarily in immature keratinocytes. Here, we show that epsin 3 is selectively expressed at high levels in the stomach (including the majority of gastric cancers), where it is concentrated in parietal cells. In these cells, epsin 3 is enriched and colocalized with clathrin around apical canaliculi, the sites that control acidification of the stomach lumen via the exo-endocytosis of vesicles containing the H/K ATPase. Deletion of the epsin 3 gene in mice did not result in obvious pathological phenotypes in either the stomach or other organs, possibly because of overlapping functions of the other two epsins. However, levels of EHD1 and EHD2, two membrane tubulating proteins with a role in endocytic recycling, were elevated in epsin 3 knock-out stomachs, pointing to a functional interplay of epsin 3 with EHD proteins in the endocytic pathway of parietal cells. We suggest that epsin 3 cooperates with other bilayer binding proteins with curvature sensing/generating properties in the specialized traffic and membrane remodeling processes typical of gastric parietal cells.
Assuntos
Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Polaridade Celular , Endocitose/fisiologia , Mucosa Gástrica/metabolismo , Células Parietais Gástricas/fisiologia , Proteínas Adaptadoras de Transporte Vesicular/genética , Animais , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , ATPase Trocadora de Hidrogênio-Potássio/metabolismo , Humanos , Camundongos , Camundongos Knockout , Células Parietais Gástricas/ultraestrutura , Estômago/anatomia & histologia , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Distribuição Tecidual , Proteínas de Transporte Vesicular/genética , Proteínas de Transporte Vesicular/metabolismoRESUMO
Chronic gastritis was morphologically verified in 119 patients. The count of parietal cells per 100 epitheliocytes of fundic glands was estimated. The sizes of parietal cells were measured by the morphometric computer program Videotest. For the diagnosis of autoimmune chronic gastritis (ACG), the level of antibodies to H+K+ATPase of gastric parietal cells in the patients' serum was determined by enzyme immunoassay. Childhood ACG has been found to be characterized by hyperplasia and structural changes of parietal cells with their preserved total count. The specific features of adult ACG include a significant reduction in the number of parietal cells, their sizes depending on the level of antiparietal autoantibodies, and marked ultrastructural changes as impairments in the differentiation and destruction of cytoplasm, organelles, and cell membrane.
Assuntos
Autoanticorpos/sangue , Doenças Autoimunes/sangue , Doenças Autoimunes/patologia , Gastrite/sangue , Gastrite/patologia , ATPase Trocadora de Hidrogênio-Potássio/metabolismo , Células Parietais Gástricas/metabolismo , Células Parietais Gástricas/ultraestrutura , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Autoanticorpos/imunologia , Doenças Autoimunes/enzimologia , Doenças Autoimunes/imunologia , Membrana Celular/enzimologia , Membrana Celular/imunologia , Membrana Celular/ultraestrutura , Criança , Doença Crônica , Citoplasma/enzimologia , Citoplasma/imunologia , Citoplasma/ultraestrutura , Feminino , Gastrite/enzimologia , Gastrite/imunologia , ATPase Trocadora de Hidrogênio-Potássio/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Organelas/enzimologia , Organelas/imunologia , Organelas/ultraestruturaRESUMO
In the mammalian gastrointestinal tract, the cell fate decisions that specify the development of multiple, diverse lineages are governed in large part by interactions of stem and early lineage progenitor cells with their microenvironment, or niche. Here, we show that the gastric parietal cell (PC) is a key cellular component of the previously undescribed niche for the gastric epithelial neck cell, the progenitor of the digestive enzyme secreting zymogenic (chief) cell (ZC). Genetic ablation of PCs led to failed patterning of the entire zymogenic lineage: progenitors showed premature expression of differentiated cell markers, and fully differentiated ZCs failed to develop. We developed a separate mouse model in which PCs localized not only to the progenitor niche, but also ectopically to the gastric unit base, which is normally occupied by terminally differentiated ZCs. Surprisingly, these mislocalized PCs did not maintain adjacent zymogenic lineage cells in the progenitor state, demonstrating that PCs, though necessary, are not sufficient to define the progenitor niche. We induced this PC mislocalization by knocking out the cytoskeleton-regulating gene Cd2ap in Mist1(-/-) mice, which led to aberrant E-cadherin localization in ZCs, irregular ZC-ZC junctions, and disruption of the ZC monolayer by PCs. Thus, the characteristic histology of the gastric unit, with PCs in the middle and ZCs in the base, may depend on establishment of an ordered adherens junction network in ZCs as they migrate into the base.
Assuntos
Diferenciação Celular , Linhagem da Célula , Celulas Principais Gástricas/citologia , Células Epiteliais/citologia , Nicho de Células-Tronco/citologia , Células-Tronco/citologia , Proteínas Adaptadoras de Transdução de Sinal/deficiência , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/deficiência , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Caderinas/metabolismo , Polaridade Celular , Proliferação de Células , Celulas Principais Gástricas/metabolismo , Celulas Principais Gástricas/ultraestrutura , Proteínas do Citoesqueleto/deficiência , Proteínas do Citoesqueleto/metabolismo , Células Epiteliais/metabolismo , Células Epiteliais/ultraestrutura , Junções Intercelulares/metabolismo , Junções Intercelulares/ultraestrutura , Camundongos , Modelos Biológicos , Células Parietais Gástricas/citologia , Células Parietais Gástricas/metabolismo , Células Parietais Gástricas/ultraestrutura , Ligação Proteica , Transporte Proteico , Nicho de Células-Tronco/metabolismo , Nicho de Células-Tronco/ultraestrutura , Células-Tronco/metabolismo , Células-Tronco/ultraestruturaRESUMO
Slc26a9 is a recently identified anion transporter that is abundantly expressed in gastric epithelial cells. To study its role in stomach physiology, gene targeting was used to prepare mice lacking Slc26a9. Homozygous mutant (Slc26a9(-/-)) mice appeared healthy and displayed normal growth. Slc26a9 deletion resulted in the loss of gastric acid secretion and a moderate reduction in the number of parietal cells in mutant mice at 5 weeks of age. Immunofluorescence labeling detected the H-K-ATPase exclusively on the apical pole of gastric parietal cells in Slc26a9(-/-) mice, in contrast to the predominant cytoplasmic localization in Slc26a9(+/+) mice. Light microscopy indicated that gastric glands were dilated, and electron micrographs displayed a distinct and striking absence of tubulovesicles in parietal cells and reductions in the numbers of parietal and zymogen cells in Slc26a9(-/-) stomach. Expression studies indicated that Slc26a9 can function as a chloride conductive pathway in oocytes as well as a Cl(-)/HCO(3)(-) exchanger in cultured cells, and localization studies in parietal cells detected its presence in tubulovesicles. We propose that Slc26a9 plays an essential role in gastric acid secretion via effects on the viability of tubulovesicles/secretory canaliculi and by regulating chloride secretion in parietal cells.
Assuntos
Antiporters/deficiência , Membrana Celular/patologia , Ácido Gástrico/metabolismo , Deleção de Genes , Células Parietais Gástricas/metabolismo , Células Parietais Gástricas/patologia , Animais , Animais Recém-Nascidos , Proteínas de Transporte de Ânions/metabolismo , Antiporters/metabolismo , Biomarcadores/metabolismo , Células COS , Chlorocebus aethiops , Imunofluorescência , ATPase Trocadora de Hidrogênio-Potássio/metabolismo , Concentração de Íons de Hidrogênio , Immunoblotting , Camundongos , Camundongos Knockout , Células Parietais Gástricas/enzimologia , Células Parietais Gástricas/ultraestrutura , Proteínas SLC4A , Transportadores de Sulfato , Titulometria , XenopusRESUMO
Helicobacter pylori persistently colonize the human stomach and have been linked to atrophic gastritis and gastric carcinoma. Although it is well known that H. pylori infection can result in hypochlorhydria, the molecular mechanisms underlying this phenomenon remain poorly understood. Here we show that VacA permeabilizes the apical membrane of gastric parietal cells and induces hypochlorhydria. The functional consequences of VacA infection on parietal cell physiology were studied using freshly isolated rabbit gastric glands and cultured parietal cells. Secretory activity of parietal cells was judged by an aminopyrine uptake assay and confocal microscopic examination. VacA permeabilization induces an influx of extracellular calcium, followed by activation of calpain and subsequent proteolysis of ezrin at Met(469)-Thr(470), which results in the liberation of ezrin from the apical membrane of the parietal cells. VacA treatment inhibits acid secretion by preventing the recruitment of H,K-ATPase-containing tubulovesicles to the apical membrane of gastric parietal cells. Electron microscopic examination revealed that VacA treatment disrupts the radial arrangement of actin filaments in apical microvilli due to the loss of ezrin integrity in parietal cells. Significantly, expression of calpain-resistant ezrin restored the functional activity of parietal cells in the presence of VacA. Proteolysis of ezrin in VacA-infected parietal cells is a novel mechanism underlying H. pylori-induced inhibition of acid secretion. Our results indicate that VacA disrupts the apical membrane-cytoskeletal interactions in gastric parietal cells and thereby causes hypochlorhydria.
Assuntos
Acloridria/metabolismo , Citoesqueleto de Actina/metabolismo , Proteínas de Bactérias/metabolismo , Membrana Celular/metabolismo , Infecções por Helicobacter/metabolismo , Helicobacter pylori/metabolismo , Células Parietais Gástricas/metabolismo , Acloridria/microbiologia , Citoesqueleto de Actina/ultraestrutura , Animais , Cálcio/metabolismo , Calpaína/metabolismo , Membrana Celular/microbiologia , Membrana Celular/ultraestrutura , Permeabilidade da Membrana Celular , Células Cultivadas , Proteínas do Citoesqueleto/metabolismo , Gastrite Atrófica/metabolismo , Gastrite Atrófica/microbiologia , Gastrite Atrófica/patologia , Infecções por Helicobacter/microbiologia , Infecções por Helicobacter/patologia , Humanos , Células Parietais Gástricas/microbiologia , Células Parietais Gástricas/ultraestrutura , Coelhos , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/microbiologia , Neoplasias Gástricas/ultraestruturaRESUMO
It is clinicopathologically important to elucidate the cell kinetics for the maintenance of normal gastric epithelium. In a rat gastric mucosa isolated after stimulation, a number of cells were exfoliated into the gastric lumen of the pit region. The present study was undertaken to clarify the origin of exfoliated cells and their histochemical profiles by taking the advantages of cryotechniques. As results, most of the exfoliated cells were identified as pit-parietal cells labeled with both peanut-lectin and anti-H+/K+-ATPase antibody. Quantitative analysis verified a time-dependent increase in the number of exfoliated cells in the gastric mucosa isolated after stimulation. The exfoliated cells exhibited a diffuse intracellular staining for E-cadherin, suggesting a dissociation of the adhesion molecule prior to the cell exfoliation. It should be noted that most of the exfoliated cells were negative to the apoptotic markers (TUNEL staining and caspase-3). Ultrastructurally, autophagosome-like structures consisting of H+/K+-ATPase positive membranes were frequently seen in the exfoliated pit-parietal cells. In addition, the pit-parietal cell exfoliation was accompanied by sealing of their basal portion with the cytoplasmic processes of adjacent surface mucous cells. The present morphological findings provide a new insight into the cell kinetics in the gastric epithelium in vitro.
Assuntos
Mucosa Gástrica/fisiologia , Células Parietais Gástricas/fisiologia , Animais , Antígenos de Diferenciação/metabolismo , Caderinas/metabolismo , Caspase 3/metabolismo , Polaridade Celular/fisiologia , Substituição ao Congelamento/métodos , Mucosa Gástrica/ultraestrutura , ATPase Trocadora de Hidrogênio-Potássio/metabolismo , Técnicas In Vitro , Masculino , Células Parietais Gástricas/ultraestrutura , Ratos , Ratos WistarRESUMO
OBJECTIVE: To observe the effect of selective cyclooxygenase-2 (COX-2) inhibitor on the healing of experimental gastric ulcer in rats and explore its mechanisms in light of gastric acid secretion. METHODS: Gastric ulcers were induced in rats by an application of acetic acid to the serosal surface of the anterior gastric body. The effects of selective COX-2 inhibitor, celecoxib, on the healing of gastric ulcer, the total acidity of gastric juice, the expressions of H+, K+-ATPase mRNA and protein, and the ultrastructure of the parietal cell were observed in comparison with the effects of normal saline. RESULTS: Nine days after ulcer induction, the ulcer area was 11.9-/+3.1 mm square and 19.7-/+3.8 mm square in rats with normal saline and celecoxib treatments, respectively (P<0.01). The total acidity of gastric juice and the expressions of H+, K+-ATPase mRNA and protein in celecoxib group were significantly higher than that in normal saline group at both 6 and 9 days after ulcer induction, but no significant difference was found between the two groups in the amount of secretary canaliculus and microvillus. CONCLUSION: Selective COX-2 inhibitor can significantly delay the healing of experimental gastric ulcer in rats, the mechanism of which might be associated with enhanced digestive action of gastric acid on the new granulation tissue at the ulcer base as a result of celecoxib-stimulated gastric acid secretion of the parietal cells.