Metabolic regulator ERRγ governs gastric stem cell differentiation into acid-secreting parietal cells.

Parietal cells (PCs) produce gastric acid to kill pathogens and aid digestion. Dysregulated PC census is common in disease, yet how PCs differentiate is unclear. Here, we identify the PC progenitors arising from isthmal stem cells, using mouse models and human gastric cells, and show that they preferentially express cell-metabolism regulator and orphan nuclear receptor Estrogen-related receptor gamma (Esrrg, encoding ERRγ). Esrrg expression facilitated the tracking of stepwise molecular, cellular, and ultrastructural stages of PC differentiation. EsrrgP2ACreERT2 lineage tracing revealed that Esrrg expression commits progenitors to differentiate into mature PCs. scRNA-seq indicated the earliest Esrrg+ PC progenitors preferentially express SMAD4 and SP1 transcriptional targets and the GTPases regulating acid-secretion signal transduction. As progenitors matured, ERRγ-dependent metabolic transcripts predominated. Organoid and mouse studies validated the requirement of ERRγ for PC differentiation. Our work chronicles stem cell differentiation along a single lineage in vivo and suggests ERRγ as a therapeutic target for PC-related disorders.

Gastric Serotonin Biosynthesis and Its Functional Role in L-Arginine-Induced Gastric Proton Secretion.

Among mammals, serotonin is predominantly found in the gastrointestinal tract, where it has been shown to participate in pathway-regulating satiation. For the stomach, vascular serotonin release induced by gastric distension is thought to chiefly contribute to satiation after food intake. However, little information is available on the capability of gastric cells to synthesize, release and respond to serotonin by functional changes of mechanisms regulating gastric acid secretion. We investigated whether human gastric cells are capable of serotonin synthesis and release. First, HGT-1 cells, derived from a human adenocarcinoma of the stomach, and human stomach specimens were immunostained positive for serotonin. In HGT-1 cells, incubation with the tryptophan hydroxylase inhibitor p-chlorophenylalanine reduced the mean serotonin-induced fluorescence signal intensity by 27%. Serotonin release of 147 ± 18%, compared to control HGT-1 cells (set to 100%) was demonstrated after treatment with 30 mM of the satiating amino acid L-Arg. Granisetron, a 5-HT3 receptor antagonist, reduced this L-Arg-induced serotonin release, as well as L-Arg-induced proton secretion. Similarly to the in vitro experiment, human antrum samples released serotonin upon incubation with 10 mM L-Arg. Overall, our data suggest that human parietal cells in culture, as well as from the gastric antrum, synthesize serotonin and release it after treatment with L-Arg via an HTR3-related mechanism. Moreover, we suggest not only gastric distension but also gastric acid secretion to result in peripheral serotonin release.

Molecular mechanisms of Wischnewski spot development on gastric mucosa in fatal hypothermia: an experimental study in rats.

Numerous dark-brown-coloured small spots called "Wischnewski spots" are often observed in the gastric mucosa in the patients dying of hypothermia, but the molecular mechanisms through which they develop remain unclear. We hypothesised that hypothermia may activate the secretion of gastric acid and pepsin, leading to the development of the spots. To investigate this, we performed experiments using organotypic rat gastric tissue slices cultured at 37 °C (control) or 32 °C (cold). Cold loading for 6 h lowered the extracellular pH in the culture medium. The mRNA expression of gastrin, which regulates gastric acid secretion, increased after cold loading for 3 h. Cold loading increased the expression of gastric H+,K+-ATPase pump protein in the apical canalicular membrane and resulted in dynamic morphological changes in parietal cells. Cold loading resulted in an increased abundance of pepsin C protein and an elevated mRNA expression of its precursor progastricsin. Collectively, our findings clarified that cold stress induces acidification by activating gastric H+,K+-ATPase pumps and promoting pepsin C release through inducing progastricsin expression on the gastric mucosa, leading to tiny haemorrhages or erosions of the gastric mucosa that manifest as Wischnewski spots in fatal hypothermia.

Cystine/Glutamate Antiporter (xCT) Is Required for Chief Cell Plasticity After Gastric Injury.

BACKGROUND & AIMS: Many differentiated epithelial cell types are able to reprogram in response to tissue damage. Although reprogramming represents an important physiological response to injury, the regulation of cellular plasticity is not well understood. Damage to the gastric epithelium initiates reprogramming of zymogenic chief cells into a metaplastic cell lineage known as spasmolytic polypeptide-expressing metaplasia (SPEM). The present study seeks to identify the role of xCT, a cystine/glutamate antiporter, in chief cell reprogramming after gastric injury. We hypothesize that xCT-dependent reactive oxygen species (ROS) detoxification is required for the reprogramming of chief cells into SPEM. METHODS: Sulfasalazine (an xCT inhibitor) and small interfering RNA knockdown were used to target xCT on metaplastic cells in vitro. Sulfasalazine-treated wild-type mice and xCT knockout mice were analyzed. L635 or DMP-777 treatment was used to chemically induce acute gastric damage. The anti-inflammatory metabolites of sulfasalazine (sulfapyridine and mesalazine) were used as controls. Normal gastric lineages, metaplastic markers, autophagy, proliferation, xCT activity, ROS, and apoptosis were assessed. RESULTS: xCT was up-regulated early as chief cells transitioned into SPEM. Inhibition of xCT or small interfering RNA knockdown blocked cystine uptake and decreased glutathione production by metaplastic cells and prevented ROS detoxification and proliferation. Moreover, xCT activity was required for chief cell reprogramming into SPEM after gastric injury in vivo. Chief cells from xCT-deficient mice showed decreased autophagy, mucus granule formation and proliferation, as well as increased levels of ROS and apoptosis compared with wild-type mice. On the other hand, the anti-inflammatory metabolites of sulfasalazine did not affect SPEM development. CONCLUSIONS: The results presented here suggest that maintaining redox balance is crucial for progression through the reprogramming process and that xCT-mediated cystine uptake is required for chief cell plasticity and ROS detoxification.

A human mode of intestinal type gastric carcinoma.

Gastric cancer is a malignant tumor originating from the gastric mucosa epithelium. Intestinal type gastric cancer is frequently taken on elderly men, and there are many high incidence areas around the world. Intestinal type gastric cancer often is accompanied by gastric mucosal atrophy, intestinal metaplasia. The clinical manifestation involves hypergastrinemia, low stomach acid, PG I/II progressive decreasing, anemia, and protein energy malnutrition. The neck cells of gastric glands act as tissue stem cells to regenerate the gastric glands. In addition to secreting gastric acid and intrinsic factor, the parietal cells also have the function of inducing differentiation of themselves and gastric epithelial cells. When the function of parietal cells is normal, the neck cells differentiate into mature cells, and the glands regenerate intact. When the function of parietal cells is defective, the neck cells maybe differentiate into mature intestinal cells, and the gastric glands will regenerate in form of the intestinal metaplasia. When the function of parietal cells is lost, the neck cells can not differentiate into mature cells successfully, and the accumulation of immature cells in gastric mucosal tissue forms atypical hyperplasia of different degrees and cancers of various differentiation grades. Any factors that can reduce the function of parietal cell could result in intestinal type gastric carcinogenesis. Adrenal cortical hypofunction can make the parietal cell hypofunction, hypohematopoiesis, protein synthesis rates reducing and protein degradation rates increasing. The patients develop gastric cancer, and come with lack of gastric acid and intrinsic factor, anemia, protein energy malnutrition. Autoimmune gastritis can produce parietal cell antibodies to damage parietal cells. Patients with autoimmune gastritis gastric exhibit hypergastrinemia, lack of gastric acid and internal factor, higher incidence of gastric cancer. H. pylori can damage gastric parietal cells directly and indirectly. When declining in quantity of parietal cells, the patients exhibit hypergastrinemia, low gastric acid, mucosa atrophy, intestinal metaplasia and gastric cancer. Medicine that inhibits the function of parietal cells also could increase the risk of gastric cancer development. The distribution of mucosa atrophy, intestinal metaplasia and intestinal type gastric cancer is opposite with the distribution of parietal cells in stomach. With age the quantity of parietal cells decreases, the atrophy area of gastric mucosa extends upward from antrum to body and downward from cardia to body along lesser curvature, and the location of distal gastric cancer moves upward and the gastric cardiac cancer increase.

A signalling cascade of IL-33 to IL-13 regulates metaplasia in the mouse stomach.

OBJECTIVE: Alternatively activated macrophages (M2) are associated with the progression of spasmolytic polypeptide-expressing metaplasia (SPEM) in the stomach. However, the precise mechanism(s) and critical mediators that induce SPEM are unknown. DESIGN: To determine candidate genes important in these processes, macrophages from the stomach corpus of mice with SPEM (DMP-777-treated) or advanced SPEM (L635-treated) were isolated and RNA sequenced. Effects on metaplasia development after acute parietal cell loss induced by L635 were evaluated in interleukin (IL)-33, IL-33 receptor (ST2) and IL-13 knockout (KO) mice. RESULTS: Profiling of metaplasia-associated macrophages in the stomach identified an M2a-polarised macrophage population. Expression of IL-33 was significantly upregulated in macrophages associated with advanced SPEM. L635 induced metaplasia in the stomachs of wild-type mice, but not in the stomachs of IL-33 and ST2 KO mice. While IL-5 and IL-9 were not required for metaplasia induction, IL-13 KO mice did not develop metaplasia in response to L635. Administration of IL-13 to ST2 KO mice re-established the induction of metaplasia following acute parietal cell loss. CONCLUSIONS: Metaplasia induction and macrophage polarisation after parietal cell loss is coordinated through a cytokine signalling network of IL-33 and IL-13, linking a combined response to injury by both intrinsic mucosal mechanisms and infiltrating M2 macrophages.

Activation of Secretagogue Independent Gastric Acid Secretion via Endothelial Nitric Oxide Synthase Stimulation in Rats.

BACKGROUND/AIMS: L-arginine is an important mediator of cell division, wound healing, and immune function. It can be transformed by the nitric oxide synthase (NOS) to nitric oxide (NO), an important cell signaling molecule. Recent studies from our laboratory demonstrate specific effects of L-arginine (10mM) exposure on gastric acid secretion in rat parietal cells. METHODS: Studies were performed with isolated gastric glands and the pH sensitive dye BCECF-AM +/- L-arginine to examine its effects on acid secretion. The direct NO-donor diethylamine NONOate sodium salt hydrate, was also used while monitoring intracellular pH. The specific inhibitor of the intracellular NO signal cascade ODQ was also used. RESULTS: We found that gastric proton extrusion was activated with application of L-arginine (10mM), in a separate series when L-arginine (10mM) + L-NAME (30µM) were added there was no acid secretion. Addition of the NO-donor diethylamine NONOate sodium salt hydrate (10µM) also induced acid secretion. When the selective sGC-inhibitor ODQ was added with NONOate we did not observe acid secretion. CONCLUSION: We conclude that L-arginine is a novel secretagogue, which can mediate gastric acid secretion. Furthermore, the intake of L-arginine causes direct activation of the H+, K+ ATPase and increased proton extrusion from parietal cells resulting in the increased risk for acid-related diseases. The NO/sGC/cGMP pathway has never been described as a possible intracellular mechanism for H+, K+ ATPase activation before and presents a completely new scientific finding. Moreover, our studies demonstrate a novel role for L-NAME to effectively eliminate NOS induced acid secretion and thereby reducing the risk for L-arginine inducible ulcer disease.

MST4 kinase phosphorylates ACAP4 protein to orchestrate apical membrane remodeling during gastric acid secretion.

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.

Characterization by Lectin Histochemistry of Two Subpopulations of Parietal Cells in the Rat Gastric Glands.

Parietal cells undergo a differentiation process while they move from the isthmus toward the pits and the base region of the gastric gland. The aim of this work was to analyze the rat gastric glands by lectin histochemistry to show the glycans expressed by upper (young) and lower (old) parietal cells. We used lectins recognizing the most frequent sugar moieties in mammals. Each lectin was assayed alone and in combination with several deglycosylation pretreatments: (1) ß-elimination, which removes O-linked oligosaccharides; (2) incubation with Peptide-N-glycosidase F, to remove N-linked glycans; (3) acid hydrolysis, which removes terminal sialic acid moieties; (4) methylation-saponification, to remove sulfate groups from sugar residues; and (5) glucose oxidase, a technique carried out with the lectin concanavalin A to convert glucose into gluconic acid. The lectins from Helix pomatia, Dolichos biflorus (DBA), Glycine max (soybean), Maclura pomifera, Arachis hypogaea (peanut), Bandeiraea simplicifolia (lectin I-B4), and Datura stramonium showed a different glycan expression in the parietal cells throughout the gastric gland. This difference supports that parietal cells undergo a maturation/degeneration process while the cells descend along the gland. The role of DBA as a marker of parietal cells previously reported should be taken with caution because these cells showed different reactivity for the lectin, ranging from negative to strong labeling.

A single transcription factor is sufficient to induce and maintain secretory cell architecture.

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."

Wnt/ß-catenin promotes gastric fundus specification in mice and humans.

Despite the global prevalence of gastric disease, there are few adequate models in which to study the fundus epithelium of the human stomach. We differentiated human pluripotent stem cells (hPSCs) into gastric organoids containing fundic epithelium by first identifying and then recapitulating key events in embryonic fundus development. We found that disruption of Wnt/ß-catenin signalling in mouse embryos led to conversion of fundic to antral epithelium, and that ß-catenin activation in hPSC-derived foregut progenitors promoted the development of human fundic-type gastric organoids (hFGOs). We then used hFGOs to identify temporally distinct roles for multiple signalling pathways in epithelial morphogenesis and differentiation of fundic cell types, including chief cells and functional parietal cells. hFGOs are a powerful model for studying the development of the human fundus and the molecular bases of human gastric physiology and pathophysiology, and also represent a new platform for drug discovery.

Rapid healing of a patient with dramatic subacute combined degeneration of spinal cord: a case report.

BACKGROUND: Prevalence of cobalamin deficiency is high especially in older patients and an immediate therapy start is necessary to prevent irreversible neurological damages. Unfortunately, the diagnosis of cobalamin deficiency is difficult and at present, there is no consensus for diagnosis of this deficiency. Therefore, we aim to elucidate a meaningful diagnostic pathway by a case report with an initially misleading medical history. CASE PRESENTATION: A 57 year-old Caucasian man suffering from dramatic myelosis of the cervical posterior columns. Apart from associated neurological symptoms (tactile hypaesthesia, reduced vibration sensation, loss of stereognosis and of two-point-discrimination) there were no further complaints; especially no gastrointestinal, haematological or psychiatric disorders were provable. Cobalamin (vitamin B12) serum level was normal. The diagnosis of subacute combined degeneration of spinal cord was confirmed by an elevated methylmalonic acid, and hyperhomocysteinemia. Cobalamin deficiency was caused by asymptomatic chronic atrophic inflammation of the stomach with a lack of intrinsic factor producing gland cells. This was revealed by increased gastrin and parietal cell antibodies and finally confirmed by gastroscopy. Parenteral substitution of cobalamin rapidly initiated regeneration. CONCLUSIONS: This case demonstrates that normal cobalamin serum levels do not rule out a cobalamin deficiency. In contrast, path-breaking results can be achieved by determining homocysteine, holotranscobalamin, and methylmalonic acid.

Unique Cellular Lineage Composition of the First Gland of the Mouse Gastric Corpus.

The glandular stomach has two major zones: the acid secreting corpus and the gastrin cell-containing antrum. Nevertheless, a single gland lies at the transition between the forestomach and corpus in the mouse stomach. We have sought to define the lineages that make up this gland unit at the squamocolumnar junction. The first gland in mice showed a notable absence of characteristic corpus lineages, including parietal cells and chief cells. In contrast, the gland showed strong staining of Griffonia simplicifolia-II (GSII)-lectin-positive mucous cells at the bases of glands, which were also positive for CD44 variant 9 and Clusterin. Prominent numbers of doublecortin-like kinase 1 (DCLK1) positive tuft cells were present in the first gland. The first gland contained Lgr5-expressing putative progenitor cells, and a large proportion of the cells were positive for Sox2. The cells of the first gland stained strongly for MUC4 and EpCAM, but both were absent in the normal corpus mucosa. The present studies indicate that the first gland in the corpus represents a unique anatomic entity. The presence of a concentration of progenitor cells and sensory tuft cells in this gland suggests that it may represent a source of reserve reparative cells for adapting to severe mucosal damage.

An update on the LIM and SH3 domain protein 1 (LASP1): a versatile structural, signaling, and biomarker protein.

The gene encoding the LIM and SH3 domain protein (LASP1) was cloned two decades ago from a cDNA library of breast cancer metastases. As the first protein of a class comprising one N-terminal LIM and one C-terminal SH3 domain, LASP1 founded a new LIM-protein subfamily of the nebulin group. Since its discovery LASP1 proved to be an extremely versatile protein because of its exceptional structure allowing interaction with various binding partners, its ubiquitous expression in normal tissues, albeit with distinct expression patterns, and its ability to transmit signals from the cytoplasm into the nucleus. As a result, LASP1 plays key roles in cell structure, physiological processes, and cell signaling. Furthermore, LASP1 overexpression contributes to cancer aggressiveness hinting to a potential value of LASP1 as a cancer biomarker. In this review we summarize published data on structure, regulation, function, and expression pattern of LASP1, with a focus on its role in human cancer and as a biomarker protein. In addition, we provide a comprehensive transcriptome analysis of published microarrays (n=2,780) that illustrates the expression profile of LASP1 in normal tissues and its overexpression in a broad range of human cancer entities.

Spatial control of proton pump H,K-ATPase docking at the apical membrane by phosphorylation-coupled ezrin-syntaxin 3 interaction.

The digestive function of the stomach depends on acidification of the gastric lumen. Acid secretion into the lumen is triggered by activation of a cAMP-dependent protein kinase (PKA) cascade, which ultimately results in the insertion of gastric H,K-ATPases into the apical plasma membranes of parietal cells. A coupling protein is ezrin whose phosphorylation at Ser-66 by PKA is required for parietal cell activation. However, little is known regarding the molecular mechanism(s) by which ezrin operates in gastric acid secretion. Here we show that phosphorylation of Ser-66 induces a conformational change of ezrin that enables its association with syntaxin 3 (Stx3) and provides a spatial cue for H,K-ATPase trafficking. This conformation-dependent association is specific for Stx3, and the binding interface is mapped to the N-terminal region. Biochemical analyses show that inhibition of ezrin phosphorylation at Ser-66 prevents ezrin-Stx3 association and insertion of H,K-ATPase into the apical plasma membrane of parietal cells. Using atomic force microscopic analyses, our study revealed that phosphorylation of Ser-66 induces unfolding of ezrin molecule to allow Stx3 binding to its N terminus. Given the essential role of Stx3 in polarized secretion, our study presents the first evidence in which phosphorylation-induced conformational rearrangement of the ezrin molecule provides a spatial cue for polarized membrane trafficking in epithelial cells.

Reduced GLP-1R expression in gastric glands of patients with type 2 diabetes mellitus.

BACKGROUND: The incretin effect is reduced in type 2 diabetes mellitus (T2DM) patients. Whether the impaired function of the enteropancreatic axis in these patients is due to defective GLP-1 receptor (GLP-1R) expression in extrapancreatic target organs is not known. AIMS AND METHODS: To compare the GLP-1R expression and distribution in gastric mucosa biopsies of patients with (n =22) and without (n =22) T2DM referred for routine esophagogastroduodenoscopies. GLP-1R mRNA levels were estimated by real-time PCR. The intensity of GLP-1R immunostaining, frequency, and types of glandular cells bearing GLP-1R and their glandular distribution in different stomach mucosa regions were evaluated by immunohistochemical morphological semiquantitative and quantitative analysis. RESULTS: Mean mRNA GLP-1R levels were significantly reduced in patients with T2DM compared with nondiabetic patients (P < .02). Immunohistochemical analysis revealed that the reduced GLP-1R expression in T2DM patients was due to a decreased intensity of immunostaining (P < .01). The number of glandular GLP-1R-bearing cells in both body and antrum mucosa was decreased in T2DM patients. Most notably, the frequency of GLP-1R immunoreactive acid-secreting parietal cells was reduced in the neck area of the gastric principal glands of T2DM patients (P < .01). No correlation was found between the reduced GLP-1R expression and clinical parameters including body mass index, age, glycosylated hemoglobin, and disease duration. CONCLUSION: This is the first evidence of reduced GLP-1R expression in gastric glands of T2DM patients. These data demonstrate that the defective function of the incretin axis in T2DM may also result from decreased GLP-1R expression in its extrapancreatic target organs.

Basolateral expression of GRP94 in parietal cells of gastric mucosa.

GRP94 is a member of the heat shock protein family normally confined to the endoplasmic reticulum that sometimes escapes the KDEL-mediated retention system. It is overexpressed in some gastric and other gastrointestinal carcinomas, but little is known about the physiological role of GRP94 in gastric mucosa. We investigated the membrane presence of GRP94 in parietal cells, which secrete acid into the gastric lumen, using subcellular fractionation, selective solubilization of membrane proteins, Western blotting, and radio-ligand binding and provided evidence of functional GRP94 expression at the surface of gastric mucosa parietal cells anchored to the basolateral domain. Our results show that GRP94 is not an integral membrane protein since 50 mM Na2CO3 treatment dissociates part of it from the membrane. However, 100 mM Na2CO3 treatment did not extract all GRP94 from the membrane, which indicates that it is strongly associated with it. The presence of GRP94 in isolated plasma membrane was demonstrated by Western blotting and its functionality by radio-ligand binding experiments. Both the K(D) value obtained in saturation experiments with N-ethylcarboxamido-[3H]adenosine at 4°C, at the nanomolar range, and the inhibition constant of its binding by radicicol, the most specific GRP94 inhibitor, indicate that active receptor regions are exposed at the membrane surface. Western blotting of plasma membrane subfractions showed that GRP94 is mainly expressed in the basolateral membrane of gastric parietal cells, while its presence in the apical domain is negligible, thereby inferring a role for GRP94 in processes operating in this membrane domain.

Cell lineage distribution atlas of the human stomach reveals heterogeneous gland populations in the gastric antrum.

OBJECTIVE: The glands of the stomach body and antral mucosa contain a complex compendium of cell lineages. In lower mammals, the distribution of oxyntic glands and antral glands define the anatomical regions within the stomach. We examined in detail the distribution of the full range of cell lineages within the human stomach. DESIGN: We determined the distribution of gastric gland cell lineages with specific immunocytochemical markers in entire stomach specimens from three non-obese organ donors. RESULTS: The anatomical body and antrum of the human stomach were defined by the presence of ghrelin and gastrin cells, respectively. Concentrations of somatostatin cells were observed in the proximal stomach. Parietal cells were seen in all glands of the body of the stomach as well as in over 50% of antral glands. MIST1 expressing chief cells were predominantly observed in the body although individual glands of the antrum also showed MIST1 expressing chief cells. While classically described antral glands were observed with gastrin cells and deep antral mucous cells without any parietal cells, we also observed a substantial population of mixed type glands containing both parietal cells and G cells throughout the antrum. CONCLUSIONS: Enteroendocrine cells show distinct patterns of localisation in the human stomach. The existence of antral glands with mixed cell lineages indicates that human antral glands may be functionally chimeric with glands assembled from multiple distinct stem cell populations.

Apical vacuole formation by gastric parietal cells in primary culture: effect of low extracellular Ca2+.

In primary culture, the gastric parietal cell's deeply invaginated apical membrane, seen in microscopy by phalloidin binding to F-actin (concentrated in microvilli and a subapical web), is engulfed into the cell, separated from the basolateral membrane (which then becomes the complete plasma membrane), and converted, from a lacy interconnected system of canaliculi, into several separate vacuoles. In this study, vacuolar morphology was achieved by 71% of parietal cells 8 h after typical collagenase digestion of rabbit gastric mucosa, but the tight-junctional protein zonula occludens-1 (ZO-1) was completely delocalized after ∼2 h, when cells were ready for culturing. Use of low-Ca(2+) medium (4 mM EGTA) to release cells quickly from gastric glands yielded parietal cells in which ZO-1 was seen in a small spot or ring, a localization quickly lost if these cells were then cultured in normal Ca(2+) but remaining up to 20 h if they were cultured in low Ca(2+). The cells in low Ca(2+) mostly retained, at 20 h, an intermediate morphology of many bulbous canalicular expansions ("prevacuoles"), seemingly with narrow interconnections. Histamine stimulation of 20-h cells with intermediate morphology caused colocalization of proton-pumping H-K-ATPase with canaliculi and prevacuoles but little swelling of those structures, consistent with a remaining apical pore through which secreted acid could escape. Apparent canalicular interconnections, lack of stimulated swelling, and lingering ZO-1 staining indicate inhibition of membrane fission processes that separate apical from basolateral membrane and vacuoles from each other, suggesting an important role for extracellular Ca(2+) in these, and possibly other, endocytotic processes.

TRIM50 protein regulates vesicular trafficking for acid secretion in gastric parietal cells.

Of the TRIM/RBCC family proteins taking part in a variety of cellular processes, TRIM50 is a stomach-specific member with no defined biological function. Our biochemical data demonstrated that TRIM50 is specifically expressed in gastric parietal cells and is predominantly localized in the tubulovesicular and canalicular membranes. In cultured cells ectopically expressing GFP-TRIM50, confocal microscopic imaging revealed dynamic movement of TRIM50-associated vesicles in a phosphoinositide 3-kinase-dependent manner. A protein overlay assay detected preferential binding of the PRY-SPRY domain from the TRIM50 C-terminal region to phosphatidylinositol species, suggesting that TRIM50 is involved in vesicular dynamics by sensing the phosphorylated state of phosphoinositol lipids. Trim50 knock-out mice retained normal histology in the gastric mucosa but exhibited impaired secretion of gastric acid. In response to histamine, Trim50 knock-out parietal cells generated deranged canaliculi, swollen microvilli lacking actin filaments, and excess multilamellar membrane complexes. Therefore, TRIM50 seems to play an essential role in tubulovesicular dynamics, promoting the formation of sophisticated canaliculi and microvilli during acid secretion in parietal cells.