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The risk of gastric cancer (GC) declines after Helicobacter pylori (H. pylori) eradication and long-term aspirin use. We evaluated the effects of H. pylori eradication (Cohort 1) and aspirin use (Cohort 2) on the methylation of microRNAs (miRNAs), such as miR-34c, miR-124a-3, miR-129-2, and miR-137, in the gastric mucosa with and without GC, i.e., in atrophic mucosal glands without intestinal metaplasia (non-IM) and intestinal metaplastic glands (IM). DNA was isolated from non-IM and IM separately using laser caption microdissection. In Cohort 1, H. pylori eradication was associated with a significant reduction of miR-124a-3 methylation only in non-IM, but not in IM. miR-129-2 methylation in non-IM may be a surrogate marker of GC in H. pylori-infected patients. In Cohort 2, aspirin did not reverse miRNA methylation in either non-IM or IM, irrespective of H. pylori infection. miR-129-2 methylation in non-IM was an independent predictive marker of GC in H. pylori-infected but not -eradicated patients. These results indicate that H. pylori eradication and aspirin use were less effective for improving methylation in IM than in non-IM; thus, these interventions are recommended at an early stage prior to the development of IM to prevent GC development. In addition, the effects of the interventions were not uniform for each miRNA gene.

The risk of gastric cancer (GC) remains even after H. pylori eradication; thus, other combination treatments, such as chemopreventive drugs, are needed. We evaluated the effects of aspirin on genetic/epigenetic alterations in precancerous conditions, i.e., atrophic mucosa (AM) and intestinal metaplasia (IM), in patients with chronic gastritis who had taken aspirin for more than 3 years. A total of 221 biopsy specimens from 74 patients, including atrophic gastritis (AG) cases without aspirin use (control), AG cases with aspirin use (AG group), and GC cases with aspirin use (GC group), were analyzed. Aspirin use was associated with a significant reduction of CDH1 methylation in AM (OR: 0.15, 95% CI: 0.06-0.41, p = 0.0002), but was less effective in reversing the methylation that occurred in IM. Frequent hypermethylation including that of CDH1 in AM increased in the GC group compared to the AG group, and CDH1 methylation was an independent predictive marker of GC (OR: 8.50, 95% CI: 2.64-25.33, p = 0.0003). In patients with long-term aspirin use, the changes of molecular events in AM but not IM may be an important factor in the reduction of cancer incidence. In addition, methylation of the CDH1 gene in AM may be a surrogate of GC.

Background: Helicobacter pylori induces chronic inflammation and intestinal metaplasia (IM) through genetic and epigenetic changes and activation of intracellular signaling pathways that contribute to gastric carcinogenesis. However, the precise mechanism of IM in gastric carcinogenesis has not been fully elucidated. We previously found that intestine-specific homeobox (ISX) mRNA expression increased in organoids cultured from Helicobacter-infected mouse mucosa. In this study, we elucidate the role of ISX in the development of IM and gastric carcinogenesis., Methods: ISX expression was assessed in Helicobacter-infected mouse and human gastric mucosa. MKN45 gastric cancer cells were co-cultured with H. pylori to determine whether Helicobacter infection induced ISX expression. We established stable MKN45 transfected cells expressing ISX (Stable-ISX MKN45) and performed a spheroid colony formation assay and a xenograft model. We performed ISX immunohistochemistry in cancer and adjacent gastric tissues., Results: ISX expression was increased in mouse and human gastric mucosa infected with Helicobacter. The presence of IM and H. pylori infection in human stomach was correlated with ISX expression. H. pylori induced ISX mRNA and protein expression. CDX1/2, cyclinD1, and MUC2 were upregulated in Stable-ISX MKN45, whereas MUC5AC was downregulated. Stable-ISX MKN45 cells formed more spheroid colonies, and had high tumorigenic ability. ISX expression in gastric cancer and adjacent mucosa were correlated., Conclusions: ISX expression induced by H. pylori infection may lead to IM and hyperproliferation of gastric mucosa through CDX1/2 and cyclinD1 expression, contributing to gastric carcinogenesis.

The gastrointestinal mucosal barrier plays an essential role in the separation of the inside of the body from the outside environment. Tight junctions (TJs) are the most important component for construction of a constitutive barrier of epithelial cells, and they regulate the permeability of the barrier by tightly sealing the cell-cell junctions. TJ proteins are represented by claudins, occludin, junctional adhesion molecules, and scaffold protein zonula occludens. Among these TJ proteins, claudins are the major components of TJs and are responsible for the barrier and the polarity of the epithelial cells. Gastrointestinal diseases including reflux esophagitis, inflammatory bowel disease, functional gastrointestinal disorders, and cancers may be regulated by these molecules, and disruption of their functions leads to chronic inflammatory conditions and chronic or progressive disease. Therefore, regulation of the barrier function of epithelial cells by regulating the expression and localization of TJ proteins is a potential new target for the treatment of these diseases. Treatment strategies for these diseases might thus be largely altered if symptom generation and/or immune dysfunction could be regulated through improvement of mucosal barrier function. Since TJ proteins may also modify tumor infiltration and metastasis, other important goals include finding a good TJ biomarker of cancer progression and patient prognosis, and developing TJ protein-targeted therapies that can modify patient prognosis. This review summarizes current understanding of gastrointestinal barrier function, TJ protein expression, and the mechanisms underlying epithelial barrier dysregulation in gastrointestinal diseases.

Background/aims: Junctional adhesion molecules (JAMs) are known as integral constituents of cellular tight junctions. However, the functions of JAMs in cancer tissues are controversial and the function of JAM-A in gastric cancer is unclear. Acordingly, we investigated the function of JAM-A in gastric epithelial and gastric cancer cell proliferation, invasion and apoptosis., Methodology: A normal rat gastric mucosa-derived cell line (RGM1), a rat gastric cancer-like cell line established from RGM1 (RGK1), and a human gastric cancer cell line (NCI-N87) were used in this study. To examine the expression of junctional proteins, immunoblotting and immunofluorescent staining were performed with specific antibodies (JAM-A, claudins, occludin and ZO-1). JAM-A was knocked down by small interfering RNA., Results: RGM1 and RGK1 expressed JAM-A, occludin and ZO-1 but not claudins. RGK1 were significantly more invasive than RGM1. JAM-A knock-down significantly decreased the proliferation and the invasion of RGK1 but not of RGM1. JAM-A knock-down significantly decreased the proliferation of NCI-N87 cells and significantly decreased expression of the anti-apoptotic protein Bcl-xL but not the expression of AKT or Mcl-1., Conclusions: JAM-A promotes proliferation and inhibits apoptosis of gastric cancer, suggesting that it has a pivotal role in gastric cancer progression.

Background: Claudins are tight junction (TJ) proteins, and the relationship between the level of expression and localization of TJ protein, and tumor aggressiveness in early gastric cancer (GC) is still far from clear., Aims: To investigate the expression of claudins and Ki-67 in early GC cells and surrounding normal gastric mucosa., Methods: A total of 53 early GC lesions removed via endoscopic mucosal resection or endoscopic submucosal resection were evaluated. All of the GCs were characterized as well to moderately differentiated adenocarcinoma. The labeling index (LI) of Ki-67 was calculated for each sample. To assess the prevalence of epithelial TJs, immunofluorescent staining for claudin-3, claudin-4, and claudin-7 was performed. The immunoreactivity was graded according to the percentage of stained cells., Results: Claudin-3, claudin-4, and claudin-7 expression at TJs in GC and intestinal metaplasia were significantly higher than that in gastric mucosa with no intestinal metaplasia. The Ki-67 LI of GC specimens was inversely correlated with claudin-3 expression, but not with claudin-4 or claudin-7 expression. Claudin-3 expression was significantly lower at the submucosal invasive front of GCs., Conclusions: The down-regulation of claudin-3 was associated with the proliferative potential of GC cells, indicating that claudins may have a pivotal role in the progression of GC.

Background: Gastroesophageal reflux is the most favored etiology for globus; however, recent studies suggest that a cervical inlet patch (heterotopic gastric mucosa in the cervical esophagus) involves globus. This study aimed to reveal whether these two conditions were associated with globus., Methods: A prospective cross-sectional survey was conducted for 2116 consecutive patients who underwent esophagogastroduodenoscopy in our hospital. Patients were interviewed about globus sensations and reflux symptoms prior to endoscopy. Non-erosive reflux disease was considered with reflux symptoms without reflux esophagitis. Inlet patch was detected using narrow-band imaging., Results: Sixty-three patients were excluded, and of the remaining 2053 patients (1117 male, 61 years old on average), 120 (5.8%) and 284 (13.8%) had globus and inlet patch, respectively. Multivariate analysis (n = 1584) following the exclusion of proton pump inhibitor users revealed that female gender, younger age (<60), inlet patch, and reflux symptoms were independent risk factors for the development of globus; however, reflux esophagitis was an independent factor for reducing the development of globus. In a reanalysis of non-erosive reflux disease instead of reflux symptoms, inlet patch and non-erosive reflux disease were independent risk factors for the development of globus (odds ratio, 2.9 and 12.1, respectively)., Conclusions: Non-erosive reflux disease was more closely associated with globus than an inlet patch. Our controversial results indicating a strong association of globus with non-erosive reflux disease and an inverse association of globus with reflux esophagitis may suggest that mechanisms other than gastroesophageal reflux as the etiology, such as an esophageal visceral hypersensitivity, involve globus.

Afferent fibers innervating the gastrointestinal tract have major roles in consciously evoked sensations including pain. However, little is known about the molecules involved in mechanonociception from the upper gastrointestinal tract. We recently reported that activation of extracellular signal-regulated kinase 1/2 (ERK1/2), a member of the mitogen-activated protein kinase cascade in primary afferent neurons, was induced by noxious gastric distention in the rat, and that the activation of ERK1/2 in dorsal root ganglion (DRG) neurons can be implicated in acute visceral pain. Transient receptor potential (TRP) A1, a member of the TRP family of cation channels, was expressed in both DRG and nodose ganglion (NG) neurons innervating the stomach and in nerve fibers in the gastric wall. TRPA1 was coexpressed with ERK1/2 in gastric primary afferent neurons, and attenuation of TRPA1 activation using antisense peptides and a specific blocker led to suppression of both ERK1/2 activation and visceromotor responses. TRPA1 also significantly colocalized with substance P (SP) and calcitonin gene-related peptide (CGRP) in the thoracolumbar DRG, NG and stomach. These data indicate that SP and CGRP may also be released by TRPA1 activation in primary afferent neurons to elicit neurogenic inflammation and promote visceral hyperalgesia., (Copyright 2010 S. Karger AG, Basel.)

Although reactive oxygen species have been implicated as mediators of gastrointestinal injury, their influence on the function of gastric epithelial tight junctions (TJs), which create a paracellular permeability barrier, needs to be fully investigated. H2O2 exposure to MKN28 gastric epithelial monolayers caused a significant decrease in trans-epithelial electrical resistance (TEER) and a significant increase in dextran permeability. Oxidant-mediated gastric epithelial permeability was significantly attenuated by a radical scavenger, rebamipide. H2O2 decreased the amount of claudin-3 protein but not claudin-4, -7, and JAM-A. Rebamipide significantly attenuated H2O2-induced decrease in claudin-3 protein. Small interfering RNA (siRNA) against claudin-3 treatment specifically decreased claudin-3 as seen by immunoblotting and immunofluorescent staining. Gastric TEER was significantly decreased with the treatment of siRNA against claudin-3. This is the first study to demonstrate that claudin-3 is involved in the barrier function of gastric epithelial cells and that rebamipide abolishes the H2O2-induced decrease in claudin-3 protein.

Tight junctions create a paracellular permeability barrier that is breached when nonsteroidal anti-inflammatory drugs cause gastrointestinal injury, including increased gastrointestinal permeability. However, the mechanism by which aspirin affects the function of gastric epithelial tight junctions is unknown. Thus, we examined the effect of aspirin on gastric mucosal barrier properties and tight junction organization using MKN28, a human gastric epithelial cell line that expresses claudin-3, claudin-4, claudin-7, zonula occludens (ZO)-1, and occludin, but not claudin-2 or claudin-5, as determined by immunoblot analysis and immunofluorescent staining. Aspirin (5 mM) treatment of MKN28 gastric epithelial monolayers significantly decreased transepithelial electrical resistance and increased dextran permeability. Both aspirin-mediated permeability and phosphorylation of p38 MAPK were significantly attenuated by SB-203580 (a p38 MAPK inhibitor) but not by U-0126 (a MEK1 inhibitor) or SP-600125 (a JNK inhibitor). Aspirin significantly decreased the quantity of claudin-7 protein produced by MKN28 cells but not the quantity of claudin-3, claudin-4, ZO-1, or occludin. The aspirin-induced decrease in claudin-7 protein was completely abolished by SB-203580 pretreatment. These results demonstrate, for the first time, that claudin-7 protein is important in aspirin-induced gastric barrier loss and that p38 MAPK activity mediates this epithelial barrier dysfunction.

Background: It is still disputed whether gastric atrophy or intestinal metaplasia improves after the cure of Helicobacter pylori infection., Aim: To clarify the histological changes after the cure of H. pylori infection through a literature survey., Methods: Fifty-one selected reports from 1066 relevant articles were reviewed. The extracted data were pooled according to histological parameters of gastritis based on the (updated) Sydney system., Results: Activity improved more rapidly than inflammation. Eleven of 25 reports described significant improvement of atrophy. Atrophy was not improved in one of four studies with a large sample size (> 100 samples) and in two of five studies with a long follow-up period (> 12 months), suggesting that disagreement between the studies was not totally due to sample size or follow-up period. Methodological flaws, such as patient selection, and statistical analysis based on the assumption that atrophy improves continuously and generally in all patients might be responsible for the inconsistent results. Four of 28 studies described significant improvement of intestinal metaplasia [corrected]., Conclusions: Activity and inflammation were improved after the cure of H. pylori infection. Atrophy did not improve generally among all patients, but improved in certain patients. Improvement of intestinal metaplasia was difficult to analyse due to methodological problems including statistical power.

Background: Cyclooxygenase-2 (COX-2) is one of the rate-limiting enzymes for prostaglandin synthesis from arachidonic acid. Although it is known that inhibition of cyclooxygenase activity delays ulcer healing, the regulatory relationship between COX-2 and its metabolites in gastric epithelial cell proliferation is not well known., Aim: To investigate whether COX-2 has an effect on gastric mucosal cell proliferation and further studied whether such effect is mediated only by prostaglandin E2 (PGE2), a representative metabolite of arachidonates in the gastric mucosa., Methods: Artificial wounds of defined area size were created on complete monolayer cell sheets of isolated rat gastric epithelial cells and rat gastric cell line RGM1 under the addition of arachidonic acid or a COX-2 selective inhibitor, JTE522. Repair of wounds was assessed by monitoring wound size, with cell proliferation detected using 5-bromodeoxyuridine staining. Quantity of secreted PGE2 was measured by enzyme immunoassay., Results: Stimulation of foetal calf serum increased the expression of COX-2 protein and inhibition of COX-2 retarded wound healing with reduction of cell proliferation. Arachidonic acid increased PGE2 production and accelerated restoration. Combination of JTE522 and arachidonic acid resulted in a marked retardation of wound healing compared to the control, but JTE522 did not completely suppress the increase in cellular PGE2 content following the addition of arachidonate., Conclusions: The difference in the effects of JTE522 on PGE2 production and on wound healing suggest that the involvement of COX-2 in gastric epithelial cell proliferation is not mediated solely by PGE2.

Background: Aspirin is known to cause adverse effects, including gastric mucosal injury, and to retard gastric wound healing. Growth factors including hepatocyte growth factor (HGF), epidermal growth factor (EGF) and insulin-like growth factor-I (IGF-I) have been shown to play an important role in the repair of gastric mucosal injury., Aim: To employ the cultured gastric epithelial cell model to elucidate the effects of aspirin, as well as several growth factors (HGF, EGF and IGF-I), on gastric wound repair., Methods: Isolated rabbit gastric epithelial cells (92% mucous cells) were cultured in F-12 medium and formed a complete monolayer cell sheet in 48 h. A wound with a cell-free area of constant size (2 mm2) was then created and the wound repair process was monitored by measuring wound size every 12 h. Proliferating cells were detected by BrdU staining. Effects of aspirin (8 mM), HGF (10 ng/mL), EGF (10 ng/mL) and IGF-I (30 ng/mL) were assessed., Results: Aspirin significantly retarded wound healing, but simultaneous addition of growth factors significantly accelerated wound repair compared with aspirin alone. Growth factors reversed the aspirin-induced inhibition of cell proliferation., Conclusion: Growth factors, including HGF, EGF and IGF-I, reversed the aspirin-induced inhibition of wound repair through their cytoprotective effects on gastric epithelial cells.

Background: Some of mucosal defensive agents have anti-Helicobacter pylori activities. However, their effectiveness in eradicating H. pylori infection has not been evaluated., Aim: To assess the additive effect of mucosal defensive agents in eradication regimens using statistical analysis., Methods: Pertinent studies were retrieved using the Medline and the Igaku-chuo-zasshi databases in Japan, reference and congress abstract lists. Studies in which regimens consisted of dual or triple therapy with mucosal defensive agents and without them, were selected from the retrieved studies. Eradication rates were extracted from studies according to intention-to-treat analysis. We evaluated the efficacies of mucosal defensive agents by pooled relative risk of eradication rates and its 95% confidence intervals (95% CI), which were calculated by Mantel-Haenszel method. Heterogeneity among the studies in treatment effect was evaluated by a chi2-test., Results: In dual therapy regimens, mucosal defensive agents demonstrated significant additive effects (pooled relative risk 1.41; 95% CI: 1.24-1.61). In triple therapy regimens, these agents did not provide significant additive effect. The clinical usefulness of specific agents could not be established, when each agent was analysed independently., Conclusions: Mucosal defensive agents improve the cure rate when used with existing dual therapy regimens for eradicating H. pylori infection.

In this review article we discuss the role of growth factors in gastric ulcer healing using an in vitro wound repair model with gastric epithelial and mesenchymal cells. Several growth factors accelerate gastric epithelial and mesenchymal wound healing in vitro with acceleration of cell migration and proliferation. Epidermal growth factor, transforming growth factor-alpha (TGFalpha), hepatocyte growth factor, and insulin accelerate predominantly gastric epithelial wound healing; and TGFbeta and basic fibroblast growth factor predominantly accelerate gastric mesenchymal wound healing. Platelet-derived growth factor-betabeta and insulin-like growth factor-1 (IGF-1) accelerate both significantly. Among these growth factors, IGF-1 produced from fibroblasts plays a key role in the gastric epithelial-mesenchymal interaction during the process of gastric wound healing.

Background: Although the detailed mechanism is unclear, zinc and its derivative, polaprezinc, have been reported to accelerate gastric ulcer healing in vivo., Aim: To investigate the detailed cellular mechanism of polaprezinc on gastric epithelial cells and fibroblasts with special attention to insulin-like growth factor I (IGF-I)., Methods: Isolated rabbit gastric epithelial cells formed a complete monolayer, from which a circular artificial wound with constant size was made. The restoration process was monitored by measuring wound size up to 48 h. Either polaprezinc, IGF-I, fibroblast conditioned medium or neutralized medium conditioned by anti-IGF-I antibody was added at the time of wounding. The expression of mRNA of IGF-I, hepatocyte growth factor (HGF) and transforming growth factor alpha (TGF-alpha) in fibroblasts with or without polaprezinc treatment was tested using reverse transcription polymerase chain reaction (RT-PCR). Gastric epithelial cell proliferation was also examined by bromodeoxyuridine (BrdU) staining., Results: IGF-I and fibroblast conditioned medium treatment accelerated gastric epithelial restoration which included cell migration and proliferation. However, polaprezinc and neutralized conditioned medium treatment did not accelerate epithelial repair. RT-PCR for growth factor mRNA revealed the IGF-I mRNA expression in fibroblasts was increased after treatment with polaprezinc., Conclusion: Polaprezinc induced IGF-I production from mesenchymal cells, resulting in stimulation of epithelial cell restoration through a paracrine pathway. IGF-I may play an important role in gastric wound repair.

The mechanism of action of gastrocytoprotective agents is not fully understood. We assessed the effects of an anti-ulcer agent, teprenone, and bile acid on epithelial restoration. Teprenone with or without deoxycholic acid was added to a complete confluent cultured gastric epithelial cell sheet after wounding. Restoration was monitored for 48 h, and the wound size was assessed. Migration velocity was measured, and proliferation was detected by sequential staining with bromodeoxyuridine. The labeling index was calculated. Restoration was completed within 48h in controls, whereas deoxycholic acid retarded repair. The migration velocity was suppressed by deoxycholic acid treatment. The number of proliferative cells peaked at 36 h (labeling index, 1.7%) in controls. In the deoxycholic acid group, the maximal labeling index was 0.5% at 48 h. Teprenone abolished the bile acid-induced retardation. Teprenone showed cytoprotective effects against deoxycholic acid-induced inhibition of epithelial cell migration and proliferation.

MS-818 is a novel synthetic pyrimidine compound that stimulates nerve regeneration and promotes synthesis of various growth factors and differentiation of astrocytes. Effects of MS-818 on gastric epithelial cells were assessed using a wound repair model with primary cultured gastric epithelial cells from rabbits. A round wound with a constant cell-free area was created and the process of restoration was monitored by measuring wound size every 12 h. Cell proliferation was monitored by sequential staining with BrdU. As previously reported, EGF (10 ng/ml) accelerated wound repair by promoting cell migration and proliferation. Although MS-818 alone had no effects, MS-818 (10-100 microM) enhanced EGF-induced acceleration of gastric epithelial restoration, including cell migration and proliferation. Although the detailed mechanism of action of this agent is still unclear, MS-818 might have favorable effects on in vivo gastric mucosal repair.

We assessed the influence of epidermal growth factor (EGF) and insulin on gastric epithelial restoration in vitro. Rabbit gastric epithelial cells were cultured and formed a complete monolayer cell sheet in 2 days. We created a wound (1.8 +/- 0.05 mm2) by denuding an area of cells, and EGF (0.1-30 ng/ml) and/or insulin (1 nM-1 microM) was added. The restoration process, which included cell migration and proliferation, was monitored by measuring the cell-free area every 12 h for 2 days. Proliferating cells were detected by sequential staining with bromodeoxyuridine (BrdU). Control cells showed complete repair in 36-48 h and restoration was accelerated dose-dependently by EGF or insulin. EGF plus insulin further accelerated restoration, which was then completed in 12-24 h. EGF and/or insulin increased the number of BrdU- positive cells. The results indicated that EGF and insulin additively accelerated gastric epithelial wound repair by stimulating both the migration and the proliferation of gastric epithelial cells (particularly the former).

Background: Several studies have been reported on the effects of various therapeutic agents in enhancing or suppressing the carcinogenic activity of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). However, it is still unknown whether a mucosal protective agent could suppress its carcinogenic activity., Methods: Twenty-five Wistar male rats were divided into four groups: group 1, MNNG alone; group 2, MNNG + tetraprenylacetone; group 3, control; group 4, tetraprenlacetone alone. MNNG 100 mg/mL, was freely given to groups 1 and 2, and tetraprenylacetone (200 mg/kg intraperitoneal) was additionally administered every other day to the rats in groups 2 and 4. The animals were sacrificed at 10 weeks and the gastric mucosa examined., Results: Atrophic changes were observed in the antrum after 8 weeks of oral administration of MNNG. Furthermore, using immunohistological analysis with 5-bromo-2'-deoxyuridine (BrdU), the proliferative zone was found to be enlarged and shifted upward, although the BrdU labelling index of the proliferative zone was unaltered. Intraperitoneal administration of tetraprenylacetone every other day suppressed the MNNG-induced atrophic change and the alterations proliferative markers. Tetraprenylacetone alone did not have an effect either on morphological or proliferative markers., Conclusion: These observations suggest that gastric mucosal defensive factors may play critical roles in suppressing atrophic change inducing carcinogenesis by an exogenic carcinogen.

Background: Anti-ulcer agents exert various functional effects on gastric epithelial cells., Aim: The effects of a novel gastro-cytoprotective agent (rebamipide) on epithelial restoration following bile acid damage were assessed using primary cultured rabbit gastric epithelial cells., Methods: Rebamipide was added to complete confluent cell sheets with deoxycholic acid just after creating a cell-free wound (2 mm2). The restoration was monitored and analysed by phase contrast microscopy and an image analyser for 48 h. The migration speed was measured during the initial 3 h after wounding. Cell proliferation was detected by staining for bromodeoxyuridine (BrdU) at 12-h intervals. The labelling index was calculated per unit area. The major cytoskeletal protein actin was detected by immunohistochemical staining., Results: In the controls, restoration was completed 48 h following wounding. Deoxycholic acid retarded this process. The addition of rebamipide to deoxycholic acid abolished the bile acid-induced retardation. The migration speed was 26 microns/h in the controls. 15 microns/h in the deoxycholic acid group and 27 microns/h in the deoxycholic acid plus rebamipide group. In the controls, BrdU-positive cells, which were rarely detected in the initial 24 h, were maximal at 36 h (labelling index 1.7%). In the deoxycholic acid group, proliferation was inhibited (peak labeling index; 0.5% at 48 h). Actin-containing stress fibres were detected throughout the cells and the periphery of the lamellipodia in the controls, and were disrupted in the deoxycholic acid-treated group. Rebamipide prevented these effects., Conclusions: Deoxycholic acid significantly retarded restoration by the inhibition of both cell migration and proliferation, potentially through an effect on the cytoskeleton. Rebamipide protected the mucosal cells from bile acid mediated injury.

The purpose of this study was to clarify the effect of ethanol on confluent monolayer gastric epithelial cells that had a round cell-free area in the center of the culture dish. Restoration of such "wounding" was evaluated quantitatively every 12 hr using a computer image analyzer with and without ethanol. Without ethanol, restoration was achieved within 48 hr. Exposure to ethanol retarded cellular restoration significantly. Staining for actin and myosin in the control group revealed the presence of lamellipodia and stress fibers. However, in the ethanol group narrowed lamellipodia and few stress fibers were observed. In conclusion, ethanol retarded the migration and proliferation of cultured gastric mucosal cells after in vitro wounding, possibly by damaging the cytoskeletal system.

This study was conducted to examine the possible relationship among connexin 32 (Cx32) expression, cell proliferation and differentiation in the normal stomach, N-methyl-N'-nitro-nitrosoguanidine (MNNG)-induced atrophic gastritis, and carcinoma in rats. Atrophic gastritis and adenocarcinoma were induced by the administration of MNNG for 8 and 30 weeks, respectively. Cell proliferation was detected by staining with 5-bromo-2'-deoxyuridine (BrdU). The proliferative zone (BrdU-positive zone), located in the lower third of the gastric gland in controls, was elongated in atrophic gastritis. In adenocarcinoma, BrdU-positive cells were distributed diffusely. Cx32 expression was investigated by an indirect immunofluorescence method. In both control and atrophic gastritis specimens, Cx32 fluorescence was abundant in the surface epithelium, but was rarely detected in the glandular portion or the proliferative zone. The length of the Cx32-positive mucosa was significantly less than the control value in atrophic gastritis and no such positive mucosa was visible in adenocarcinoma. The results of this study indicate that the loss of cell-cell communication through the gap junction, associated with elongation of the proliferative cell zone, may be manifested much earlier than carcinoma. We regard this model as useful for investigating the development of atrophic gastritis into gastric carcinoma.

There are many published reports suggesting a close relationship between intestinal metaplasia and gastric carcinogenesis, but there are few studies that examine the cellular kinetics of these tissues in humans. Thus, we sought to characterize the proliferative zone of intestinal metaplasia of the human stomach and correlate this with its known malignant potential. We examined the incorporation of bromodeoxyuridine into 228 human endoscopic biopsy specimens from duodenal mucosa (n=35) and non-intestinalized antral mucosa (n=127) as well as antral mucosa with intestinal metaplasia (n = 66). The proliferative zone in specimens with intestinal metaplasia was deeper when compared to non-intestinalized antral mucosa, but was more superficial than that of duodenal mucosa. Although the labeling index of intestinalized mucosa was similar to that of non-intestinalized antral mucosa, the size of the proliferative zone was significantly increased in intestinal metaplasia. The dislocation of the proliferative zone with an increase in its size in intestinal metaplasia is considered to be a hallmark of gastric intestinal metaplasia.

Ethanol exerts damaging effects on gastric mucosa and delays ulcer healing. To investigate the effect of ethanol on the wound repairing process, we used a wound repair model using primary cultured gastric mucosal cells. A confluent monolayer gastric mucosal cell sheet consisting mainly of mucous cells was wounded to make a cell-free area of constant size. Cell-free area was restored with time after wounding and monitored every 12 hr using a computer image analyzer to observe epithelial cell restoration quantitatively in the presence and absence of ethanol (2.0%). It was found that, although the control wound was completely repaired in 36 to 48 hr, the group treated with 2.0% ethanol showed a significant delay of repair. In the control, 5-bromodeoxyuridine-positive cells appeared around the wound in 24 to 36 hr. In contrast, the group treated with 2.0% ethanol showed no 5-bromodeoxyuridine-positive cells during the experiment. In conclusion, 2.0% ethanol retarded the repair of gastric mucosal restoration by inhibiting the initial gastric cell migration, followed by inhibition of proliferation of cells.

Background: Cyclooxygenase-2 (COX-2) is one of the rate-limiting enzymes for prostaglandin synthesis from arachidonic acid. Although it is known that inhibition of cyclooxygenase activity delays ulcer healing, the regulatory relationship between COX-2 and its metabolites in gastric epithelial cell proliferation is not well known. Aim: To investigate whether COX-2 has an effect on gastric mucosal cell proliferation and further studied whether such effect is mediated only by prostaglandin E[sub 2] (PGE[sub 2] ), a representative metabolite of arachidonates in the gastric mucosa. Methods: Artificial wounds of defined area size were created on complete monolayer cell sheets of isolated rat gastric epithelial cells and rat gastric cell line RGM1 under the addition of arachidonic acid or a COX-2 selective inhibitor, JTE522. Repair of wounds was assessed by monitoring wound size, with cell proliferation detected using 5-bromodeoxyuridine staining. Quantity of secreted PGE[sub 2] was measured by enzyme immunoassay. Results: Stimulation of foetal calf serum increased the expression of COX-2 protein and inhibition of COX-2 retarded wound healing with reduction of cell proliferation. Arachidonic acid increased PGE[sub 2] production and accelerated restoration. Combination of JTE522 and arachidonic acid resulted in a marked retardation of wound healing compared to the control, but JTE522 did not completely suppress the increase in cellular PGE[sub 2] content following the addition of arachidonate. Conclusions: The difference in the effects of JTE522 on PGE[sub 2] production and on wound healing suggest that the involvement of COX-2 in gastric epithelial cell proliferation is not mediated solely by PGE[sub 2] . [ABSTRACT FROM AUTHOR]

Aim To conduct a retrospective multicentre study at 31 medical centres in Japan, to investigate the association between the eradication of Helicobacter pylori and the development of new gastric cancer after endoscopic mucosal resection (EMR) for early gastric cancer. Methods Patients included those in whom early gastric cancer had been removed by EMR, and who had received follow-up endoscopic treatment. All patients underwent follow-up endoscopic treatment at least once a year after the initial EMR. The rate of new gastric cancer development was compared among those patients for whom H. pylori had been successfully eradicated and those with persistent H. pylori infection. Results The study included 2835 patients with a median follow-up period of 2 years (range 0.5–12 years). Helicobacter pylori was eradicated in 356 patients (13%). Metachronous gastric cancers developed in eight (2%) patients among those who had been successfully treated for H. pylori, compared with 129 patients (5%) among those with persistent H. pylori infection ( P = 0.021; OR = 0.42; 95% CI = 0.20–0.86). Conclusion Although longer term, prospective randomized controlled trials are needed to better estimate the extent and sustainability of possible benefits, this study suggests that the eradication of H. pylori may help reduce the incidence of metachronous gastric cancer. [ABSTRACT FROM AUTHOR]