We examined the development of gastric cancer risk screening, from rat pepsinogen studies in an experimental rat gastric carcinogenesis model induced with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and human pepsinogen studies in the 1970s and 1980s to the recent "ABC method" for human gastric cancer risk screening. First, decreased expression or absence of a major pepsinogen isozyme, PG1, was observed in the rat gastric mucosa from the early stages of gastric carcinogenesis to adenocarcinomas following treatment with MNNG. In the 1980s, decreases in PGI in the human gastric mucosa and serum were identified as markers of atrophic gastritis. In the 1990s, other researchers revealed that chronic infection with Helicobacter pylori (Hp) causes atrophic gastritis and later gastric cancer. In the 2000s, a gastric cancer risk screening method combining assays to detect serum anti-Hp IgG antibody and serum PGI and PGII levels, the "ABC method", was established. Eradication of Hp and endoscopic follow-up examination after the ABC method are recommended to prevent gastric cancer.

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.

The gastric barrier contains several lines of defence which protect the epithelium from harmful microbes and toxins. Pre-mucosal defence mechanisms include secreted acid (HCl 0.1 mmol/L) and pepsin, which are capable of denaturing tissue. A tightly adherent mucous layer provides the next line of defence, and physically separates any potentially hazardous substance in the lumen from the mucosal surface. Apical secretion of HCO3(-) maintains a non-acidic microenvironment at the mucosal surface. Membrane-bound phospholipids repel soluble toxins, and sulphydryls scavenge reactive oxygen species. However, when noxious agents overwhelm these mechanisms, the epithelium is damaged. Herein, we discuss the pathological and physiological basis for several disease states which are associated with a breakdown in one or more components of the gastric barrier, including: Helicobacter pylori-associated gastritis, atrophic gastritis, stress-related mucosal disease, age-related gastropathy and portal hypertensive gastropathy. The effect of non-steroidal anti-inflammatory drugs and proton pump inhibitors on the gastric mucosa, is explored. Finally, we outline the alterations in mucosal defence caused by alcohol, caffeine, minerals and vitamins.

Based on our previous report on gastric cancer which documented ATP4A and ATP4B mRNA down- regulation in gastric tumors relative to normal gastric tissues, we hypothesized that epigenetic mechanisms could be responsible. ATP4A and ATP4B mRNA expression in gastric cancer cell lines AGS, SNU638 and NUGC-3 was examined using reverse transcriptase PCR (RT-PCR). AGS cells were treated with TSA or 5'-AzaDC and methylation specific PCR (MSP) and bisulfite sequencing PCR (BSP) analysis were performed. MSP analysis was on DNA from paraffin embedded tissues sections and plasma. Expression analysis revealed downregulation of ATP4A and ATP4B genes in gastric cancer cell lines relative to normal gastric tissue, while treatment with 5'-AzaDC re-activated expression of both. Search for CpG islands in their putative promoter regions did not indicate CpG islands (CGI) but only further downstream in the bodies of the genes. Methylation specific PCR (MSP) in the exon1 of the ATP4B gene and exon7 in ATP4A indicated methylation in all the gastric cancer cell lines tested. MSP analysis in tumor tissue samples revealed methylation in the majority of tumor samples, 15/19, for ATP4B and 8/8 for ATP4A. There was concordance between ATP4B and ATP4A down-regulation and methylation status in the tumour samples tested. ATP4B methylation was detectable in cell free DNA from gastric cancer patient's plasma samples. Thus ATP4A and ATP4B down-regulation involves DNA methylation and methylated ATP4B DNA in plasma is a potential biomarker for gastric cancer.

Proton pump inhibitors (PPIs) are now commonly used for the treatment of acid related diseases such as peptic ulcer and reflux esophagitis. Because of their ability to produce direct inhibition of the proton pump, PPIs provide more sustained increase of the gastric pH than H(2)-receptor (H(2)R) antagonists. Diverse reports have been published on gastric epithelial cell modality associated with PPI treatment both in animal models and clinical settings. The present review summarizes the recent accumulated evidence on gastric epithelial cell modality associated with PPI treatment, including the formation of gastric carcinoid tumors and fundic gland polyps, and the development of gastric mucosal atrophy. Long-term PPI treatment has been reported to cause enlargement of the parietal cells and enterochromaffin-like cells, and to decrease the number of chief cells without affecting A-like cell. Although the development of gastric carcinoid tumors after chronic PPI treatment has been reported in animal studies, no such occurrences have been demonstrated in humans. The effect of PPIs on the formation of fundic gland polyps and the development of atrophic gastritis should be investigated in future studies.

Alterations in the E-cadherin gene are associated with sporadic and hereditary diffuse-type gastric cancer. To determine how the loss of function of E-cadherin affects gastric epithelial cell phenotypes, we generated transgenic mice using the Cre-loxP system in which the E-cadherin gene is specifically knocked out in the parietal cell lineage. In the transgenic mice, expression of E-cadherin was lost or reduced in proton pump-expressing parietal cells, which became round in shape and were pushed out of the glands to accumulate in the stromal area. Additionally, gastric mucosa exhibited hyperplasia from 3 months in the mice, some cells of which later became positive for trefoil factor 2, a marker of spasmolytic polypeptide-expressing metaplasia. From 6 months, E-cadherin-negative/proton pump-negative cells appeared from the parietal cell lineage, which increased in number to form cell clusters. Moreover, signet ring-like cells, which are morphologically similar to signet ring carcinoma cells, were found in the cell clusters from 12 months. However, no invasive gastric adenocarcinomas were found in the E-cadherin-deficient mice, even at 24 months or later. These data indicate that the loss of E-cadherin induces possible pre-cancerous lesions in the gastric mucosa but may not be sufficient for its malignant conversion.

Shh (Sonic Hedgehog) is a morphogen involved in gastric fundic gland differentiation in the adult. Shh expression is reduced in Helicobacter pylori-associated intestinal metaplastic change of the gastric epithelium and mice that lack Shh show intestinal transformation of the gastric mucosa. Similarly, in the stomach of Cdx2 (caudal-type homeobox 2)-transgenic mice, the gastric mucosa is replaced by intestinal metaplastic mucosa. The aim of the present study was to use Cdx2-transgenic mice to investigate: (i) Shh expression in the intestinal metaplastic mucosa of the Cdx2-transgenic mouse stomach; and (ii) the relationship between Shh and Cdx2. We determined that Shh mRNA levels were dramatically reduced in the intestinal metaplastic mucosa of the Cdx2-transgenic mouse stomach compared with the normal (wild-type) mouse stomach. This was not due to hypermethylation of the Shh promoter, but instead we showed that Cdx2 directly bound to the TATA box region of the Shh promoter. Cdx2 also down-regulated transcription of the Shh gene in the human gastric carcinoma cell lines AGS, MKN45 and MKN74. In conclusion, Cdx2 reduced Shh expression by binding to the unmethylated Shh promoter in the intestinal metaplastic mucosa of Cdx2-transgenic mouse stomach.

Gastric pathology is a common complication in diabetes mellitus. The aim of the study was to evaluate the functions and morphological changes of the parietal cells of the rat stomach after streptozotocin-induced diabetes. Diabetes mellitus was induced in Wistar rats by a single intraperitoneal injection of streptozotocin (60 mg/kg body weight). The rats were weighed weekly and sacrificed after 6 months. The glandular portion of the stomach was removed and processed for H(+)-K(+)-ATPase immunohistochemistry and light and electron microscopy studies. Acid secretion was measured in vivo. After 6 months of diabetes, the mean weight of the rats was significantly lower (P < 0.001) compared to control. The mean weight of the stomach to body weight percentage increased significantly (P < 0.001) compared to control. The blood glucose level in diabetic rats was significantly higher (P < 0.001) than in normal control. Diabetic rats showed significant (P < 0.001) decrease in basal and stimulated acid secretion when compared to control. Electron micrographs of the parietal cells of glandular stomach of diabetic rats revealed significant (P < 0.0002) reduction in the number of mitochondria and a small though not significant increase in the number of canaliculi in the parietal cells compared with normal. Immunohistochemistry showed reduced H(+)-K(+)-ATPase (P < 0.00001) compared to control. Long-term diabetes induces morphological as well as functional changes in gastric parietal cells. The decrease in the number of mitochondria accompanied by reduced in H(+)-K(+)-ATPase in parietal cells may explain the reduced acid secretion observed in diabetics.

Despite their remarkable safety profile and lack of clinical side effects, proton pump inhibitors (PPIs) induce a transmucosal gastric leak to non-electrolyte probes of various sizes. The ex vivo addition of PPIs to isolated rat gastric corpus increases transmucosal permeability in a dose-dependent manner, which corresponds with PPIs' dose-dependent inhibition of acid secretion. Upon the addition of omeprazole, lansoprazole, or esomeprazole, a small decrease in transepithelial resistance and the concomitant stimulation of short circuit current was observed. Additionally, transepithelial flux of (14)C-[D]-mannitol (MW 182.17) across the gastric mucosa increased by a mean of 68% immediately following the addition of 200 microM omeprazole. This flux increase was bidirectional. Omeprazole also increased the paracellular permeability to larger radiolabeled probes, including (14)C-sucrose (MW 342.3) and (14)C-polyethylene glycol (MW 4,000) by 118% and 350%, respectively. However, the flux of still larger probes, 10,000 and 70,000 MW dextrans, was not increased. Because PPIs are so widely used and are assumed to be innocuous, this transmucosal gastric leak must be further investigated, as it may carry considerable biomedical implications.

Proton pump inhibitors are the second most commonly prescribed drug class in the United States. The increased utilization of PPIs parallels the rising incidence of reflux disease. Owing to their clinical efficacy and relative lack of tachyphylaxis, PPIs have largely displaced H-2 receptor antagonists in the treatment of acid peptic disorders. The elevation of intragastric pH and subsequent alterations of gastric physiology induced by PPIs may yield undesired effects within the upper GI tract. The ubiquity of the various types of H(+), K(+)-ATPase could also contribute to non-gastric effects. PPIs may influence physiology in other ways, such as inducing transepithelial leak.

There is considerable debate whether peroxisome proliferator-activated receptor beta/delta (PPARbeta/delta) ligands potentiate or suppress colon carcinogenesis. Whereas administration of a PPARbeta ligand causes increased small intestinal tumorigenesis in Apc(min/+) mice, PPARbeta-null (Pparb-/-) mice exhibit increased colon polyp multiplicity in colon cancer bioassays, suggesting that ligand activation of this receptor will inhibit colon carcinogenesis. This hypothesis was examined by treating wild-type (Pparb+/+) and Pparb-/- with azoxymethane, coupled with a highly specific PPARbeta ligand, GW0742. Ligand activation of PPARbeta in Pparb+/+ mice caused an increase in the expression of mRNA encoding adipocyte differentiation-related protein, fatty acid-binding protein, and cathepsin E. These findings are indicative of colonocyte differentiation, which was confirmed by immunohistochemical analysis. No PPARbeta-dependent differences in replicative DNA synthesis or expression of phosphatase and tensin homologue, phosphoinositide-dependent kinase, integrin-linked kinase, or phospho-Akt were detected in ligand-treated mouse colonic epithelial cells although increased apoptosis was found in GW0742-treated Pparb+/+ mice. Consistent with increased colonocyte differentiation and apoptosis, inhibition of colon polyp multiplicity was also found in ligand-treated Pparb+/+ mice, and all of these effects were not found in Pparb-/- mice. In contrast to previous reports suggesting that activation of PPARbeta potentiates intestinal tumorigenesis, here we show that ligand activation of PPARbeta attenuates chemically induced colon carcinogenesis and that PPARbeta-dependent induction of cathepsin E could explain the reported disparity in the literature about the effect of ligand activation of PPARbeta in the intestine.

To study the effect of proton pump inhibitor (PPI) treatment on patients with reflux esophagitis and its in vivo effect on apoptosis, p53- and epidermal growth factor receptor (EGFR) expression.

After informed consent was obtained, gastric biopsies of the antrum were taken from patients with reflux oesophagitis prior to and after 6 mo of 20 mg omeprazole (n = 14) or 40 mg esomeprazole (n = 12) therapy. Patients did not take any other medications known to affect the gastric mucosa. All patients were Helicobacter pylori negative as confirmed by rapid urease test and histology, respectively. Cell proliferation, apoptosis, EGFR, and p53 expression were measured by immunohistochemical techniques. At least 600 glandular epithelial cells were encountered and results were expressed as percentage of total cells counted. Was considered statistically significant.

Although there was a trend towards increase of cell proliferation and EGFR expression both in omeprazole and esomeprazole treated group, the difference was not statistically significant. Neither apoptosis nor p53 expression was affected.

Long-term PPI treatment does not significantly increase gastric epithelial cell proliferation and EGFR expression and has no effect on apoptosis and p53 expression.

Sonic hedgehog (Shh) is a morphogen involved in many aspects of patterning of the gut during embryogenesis and in gastric fundic gland homeostasis in the adult. Intestinal metaplastic change of the gastric epithelium is associated with the loss of Shh expression, and mice that lack Shh expression show intestinal transformation of the gastric mucosa. The present study was designed to investigate the alteration of Shh expression in the stomach of an experimental model of Helicobacter pylori (H. pylori) colonization. Male Mongolian gerbils were inoculated with H. pylori and examined 4 and 51 weeks later. The level of Shh mRNA expression was determined by quantitative RT-PCR and in situ hybridization. Shh protein expression was determined by immunoblotting and immunohistochemistry. Shh was expressed in the parietal cells, zymogenic cells, and mucous neck cells of the gastric fundic glands of gerbils. Prolonged colonization by H. pylori led to extension of the inflammation from the antrum to the corpus of the stomach, with loss of Shh expression. Loss of Shh expression correlated with loss of parietal cells, disturbed maturation of the mucous neck cell-zymogenic cell lineage, and increased cellular proliferation. Shh expression is significantly reduced in H. pylori-associated gastritis. These data show for the first time that H. pylori infection leads to down-regulation of the expression of a morphogen with an established role (Shh) in gastric epithelial differentiation.

Chronic gastritis with gastric mucosa atrophy, intestinal metaplasia and endocrine cell hyperplasia are alterations with an increased risk for the development of gastric neoplasias. Immunological studies in autoimmune gastritis, in atrophic Helicobacter pylori gastritis and in studies with transgenic mice point to a central role of the parietal cell in the development of gastric mucosa atrophy. Destruction of gastric epithelial cells alone might not be sufficient for the loss of complete gastric glands. Gastric atrophy, endocrine cell hyperplasia and intestinal and pancreatic metaplasia can be regarded as the result of altered morphogenesis within the gastric mucosa. Impaired expression of the gastric morphogenic factor Sonic Hedgehog by parietal cells and increased expression of the transcriptional activators of intestinal and pancreatic differentiation, namely CDX2 and PDX1, seem to be crucial for the development of gastric atrophy and for intestinal, endocrine and pancreatic transdifferentiation processes. Altered expression of these morphogenic factors is partly caused by changes in the gastric milieu. Further studies concerning the normal and pathological morphogenesis of the gastric mucosa and related tissues might give new insight into the pathogenesis of gastric atrophy and metaplasia.

Gastric body mucosa atrophy predisposes one to gastric cancer. Disturbances in the gastric differentiation process might play a role in the evolution of gastric atrophy. Sonic hedgehog (Shh) has recently been implicated as a crucial factor in gastric organogenesis and gland differentiation. In this study we investigated the expression of key factors in the Shh pathway, namely Shh and its receptor Patched (Ptc), in normal and pathologic stomach mucosa. Furthermore, the potential role of pH for Shh dysregulation was analyzed. Ten gastric biopsy specimens each from normal gastric mucosa, chronic nonatrophic gastritis, atrophic gastritis, and gastric cancer were included. Expression of Shh and Ptc was analyzed by immunohistochemistry. In normal body mucosa and in nonatrophic body gastritis, Shh was strongly expressed in parietal cells. Ptc was also expressed in gastric chief cells. Shh expression was almost completely lost in atrophic gastritis and in gastric cancer and absent in intestinal metaplasia. Ptc was markedly reduced in atrophy and only weakly positive in intestinal metaplasia and gastric cancer. In in vitro experiments, gastric cancer cell line 23132 was found positive for Shh. In long-term culture as well as in culture conditions with low pH, transcription of Shh in 23132 was significantly increased in quantitative reverse transcription PCR analyses. We concluded that the decreased expression of the Shh pathway in atrophic gastritis and gastric cancer might reflect altered differentiation processes within the gastric unit and contributes to the development of gastric atrophy. The increase of gastric pH might play a role in the development of gastric mucosa atrophy via reduction of Shh transcription.

The role of transcription factor forkhead homologue 6 (Fkh6) gene expressed only in gastrointestinal mesenchymes on the differentiation of gastric epithelia was analyzed by inactivating the gene by targeting disruption. Gastric mucosa exhibited hyperplasias with disordered glandular structures in the absence of gene. Measurement of acid secretion in the isolated whole stomach demonstrated that both basal and stimulated secretions were severely suppressed in the Fkh6-/- stomach, while immunohistochemical studies showed that comparable numbers of parietal cells were differentiated in both wild-type and Fkh6-/- stomachs. Ultrastructurally Fkh6-/- parietal cells were furnished with developed intracellular canaliculi and many mitochondria, but their canaluculi were not enlarged nor fully connected to the luminal surface even when animals were treated with histamine, suggesting that Fkh6-/- parietal cells were far less responsive to acid secretion-inducing stimulations. Some parietal cells contained secretory granules positively stained with anti-pepsinogen antibodies, indicating that they retained characteristics of oxynticopeptic cells found in lower vertebrates. We thus concluded that Fkh6 plays essential roles for the development and differentiation of parietal cells via epithelial-mesenchymal interactions.

The H,K-ATPase of the gastric parietal cell is the most critical component of the ion transport system mediating acid secretion in the stomach. To study the requirement of this enzyme in the development, maintenance, and function of the gastric mucosa, we used gene targeting to prepare mice lacking the alpha-subunit. Homozygous mutant (Atp4a(-/-)) mice appeared healthy and exhibited normal systemic electrolyte and acid-base status but were achlorhydric and hypergastrinemic. Immunocytochemical, histological, and ultrastructural analyses of Atp4a(-/-) stomachs revealed the presence of chief cells, demonstrating that the lack of acid secretion does not interfere with their differentiation. Parietal cells were also present in normal numbers, and despite the absence of alpha-subunit mRNA and protein, the beta-subunit was expressed. However, Atp4a(-/-) parietal cells had dilated canaliculi and lacked typical canalicular microvilli and tubulovesicles, and subsets of these cells contained abnormal mitochondria and/or massive glycogen stores. Stomachs of adult Atp4a(-/-) mice exhibited metaplasia, which included the presence of ciliated cells. We conclude that ablation of the H,K-ATPase alpha-subunit causes achlorhydria and hypergastrinemia, severe perturbations in the secretory membranes of the parietal cell, and metaplasia of the gastric mucosa; however, the absence of the pump appears not to perturb parietal cell viability or chief cell differentiation.