The MUC5AC glycoprotein is the primary receptor for Helicobacter pylori in the human stomach

Background and objectives. Helicobacter pylori shows a characteristic tropism for the mucus‐producing gastric epithelium. In infected patients, H. pylori colocalizes in situ with the gastric secretory mucin MUC5AC. The carbohydrate blood‐group antigen Lewis B (LeB) was deemed responsible for the adherence of H. pylori to the gastric surface epithelium. We sought to determine if MUC5AC is the carrier of LeB, and thus if MUC5AC is the underlying gene product functioning as the main receptor for H. pylori in the stomach. Methods. We studied three types of human tissue producing MUC5AC: Barrett's esophagus (BE), normal gastric tissue, and gastric metaplasia of the duodenum (GMD). Tissue sections were immuno‐fluorescently stained for MUC5AC or LeB, and subsequently incubated with one of three strains of Texas red‐labeled H. pylori, one of which was unable to bind to LeB. We determined the colocalization of MUC5AC or LeB with adherent H. pylori. Results. The binding patterns for the two LeB‐binding strains to all tissues were similar, whereas the strain unable to bind to LeB did not bind to any of the tissues. In normal gastric tissue, the LeB‐binding strains always bound to MUC5AC‐ and LeB‐positive epithelial cells. In four nonsecretor patients, colocalization of the LeB‐binding strains was found to MUC5AC‐positive gastric epithelial cells. In BE, the LeB‐binding H. pylori strains colocalized very specifically to MUC5AC‐positive cells. MUC5AC‐producing cells in GMD contained LeB. Yet, LeB‐binding H. pylori not only colocalized to MUC5AC or LeB present in GMD, but also bound to the LeB‐positive brush border of normal duodenal epithelium. Conclusions. Mucin MUC5AC is the most important carrier of the LeB carbohydrate structure in normal gastric tissue and forms the major receptor for H. pylori.     

Helicobacter pylori infection up-regulates gland mucous cell-type mucins in gastric pyloric mucosa

Background. Two types of mucous cell are present in gastric mucosa: surface mucous cells (SMCs) and gland mucous cells (GMCs), which consist of cardiac gland cells, mucous neck cells, and pyloric gland cells. We have previously reported that the patterns of glycosylation of SMC mucins are reversibly altered by Helicobacter pylori infection. In this study, we evaluated the effects of H. pylori infection on the expression of GMC mucins in pyloric gland cells. Methods. Gastric biopsy specimens from the antrums of 30 H. pylori‐infected patients before and after eradication of H. pylori and 10 normal uninfected volunteers were examined by immunostaining for MUC6 (a core protein of GMC mucins), α1,4‐N‐acetyl‐glucosaminyl transferase (α4GnT) (the glycosyltransferase which forms GlcNAcα1‐4Galβ‐R), and GlcNAcα1‐4Galβ‐R (a GMC mucin‐specific glycan). Results. MUC6, α4GnT, and HIK1083‐reactive glycan were expressed in the cytoplasm, supranuclear region, and secretory granules in pyloric gland cells, respectively. The immunoreactivity of MUC6 and α4GnT, but not of GlcNAcα1‐4Galβ‐R, in the pyloric gland increased in H. pylori‐associated gastritis, and after the eradication of H. pylori, the increased expression of MUC6 and α4GnT in the gastric mucosa of H. pylori‐infected patients decreased to almost normal levels. This up‐regulation was correlated with the degree of inflammation. Conclusions. In addition to the synthesis of GMC mucins increasing reversibly, their metabolism or release may also increase reversibly in H. pylori‐associated gastritis. The up‐regulation of the expression of gastric GMC mucins may be involved in defense against H. pylori infection in the gastric surface mucous gel layer and on the gastric mucosa.     

Preventive effects of Cladosiphon fucoidan against Helicobacter pylori infection in Mongolian gerbils

Background. Recently, the acquisition by Helicobacter pylori of resistance to antibiotics has become a serious problem. Therefore, nonantibiotic substances are required to diminish H. pylori‐induced gastric lesions. In the present study, the effects of Cladosiphon fucoidan were examined in terms of H. pylori attachment to porcine gastric mucin in vitro and Helicobacter pylori‐induced gastritis in vivo. Methods. The inhibitory effect of Cladosiphon fucoidan and other polysaccharides on H. pylori attachment to porcine gastric mucin was assayed in vitro with mucin‐coated microtiter plates. The effect of Cladosiphon fucoidan on H. pylori‐induced gastritis was examined in vivo using Mongolian gerbils. H. pylori‐inoculated gerbils were given fucoidan in drinking water. Six weeks after H. pylori‐inoculation, gerbils were sacrificed for macroscopic and microscopic examination of gastric lesions and counting of viable H. pylori in the gastric mucosa. Results. Cladosiphon fucoidan inhibited the H. pylori attachment to porcine gastric mucin at pH 2.0 and 4.0. Two other sulfated polysaccharides, Fucus fucoidan and dextran sulfate sodium, also inhibited the attachment but only at pH 2.0. Inhibitory effects of these three sulfated polysaccharides were not observed at pH 7.2 and nonsulfated polysaccharides, such as mannan and dextran, exerted no influence at any pH. In the in vivo experiment, the H. pylori‐induced gastritis and the prevalence of H. pylori infected animals were markedly reduced by fucoidan in a dose‐dependent manner, at doses of 0.05 and 0.5% in the drinking water. Conclusion. Cladosiphon fucoidan may deserve particular attention as a safe agent that can prevent H. pylori infection and reduce the risk of associated gastric cancer.     

Profiling and identification of eubacteria in the stomach of Mongolian gerbils with and without Helicobacter pylori infection

Background. Mongolian gerbils are frequently used to study Helicobacter pylori‐induced gastritis and its consequences. The presence of an indigenous bacterial flora with suppressive effect on H. pylori may cause difficulties with establishing this experimental model. Aim. The aim of the present study was to determine bacterial profiles in the stomach of Mongolian gerbils with and without (controls) H. pylori infection. Methods. Gastric tissue from H. pylori ATCC 43504 and CCUG 17874 infected and control animals were subjected to microbial culturing and histology. In addition, gastric mucosal samples from H. pylori ATCC 43504 infected and control animals were analyzed for bacterial profiling by temporal temperature gradient gel electrophoresis (TTGE), cloning and pyrosequencing of 16S rDNA variable V3 region derived PCR amplicons. Results. Oral administration of H. pylori ATCC 43504, but not CCUG 17874, induced colonization and gastric inflammation in the stomach of Mongolian gerbils. Temporal temperature gradient gel electrophoresis (TTGE) and partial 16S rDNA pyrosequencing revealed the presence of DNA representing a mixed bacterial flora in the stomach of both H. pylori ATCC 43504 infected and control animals. In both cases, lactobacilli appeared to be dominant. Conclusion. These findings suggest that indigenous bacteria, particularly lactobacilli, may have an impact on the colonization and growth of H. pylori strains in the stomach of Mongolian gerbils.     

A Potential Association between Helicobacter pylori CagA EPIYA and Multimerization Motifs with Cytokeratin 18 Cleavage Rate during Early Apoptosis

Background and Aims: Helicobacter pylori is a highly diverse pathogen, which encounters epithelial cells as the initial defense barrier during its lifelong infection. The structure of epithelial cells can be disrupted through cleavage of microfilaments. Cytokeratin 18 (CK18) is an intermediate filament, the cleavage of which is considered an early event during apoptosis following activation of effector caspases. Methods: Helicobacter pylori strains were isolated from 76 dyspeptic patients. cagA 3’ variable region and CagA protein status were analyzed by PCR and western blotting, respectively. Eight hours post‐co‐culture of AGS cells with different H. pylori strains, flow cytometric analysis was performed using M30 monoclonal antibody specific to CK18 cleavage‐induced neo‐epitope. Results: Higher rates of CK18 cleavage were detected during co‐culture of AGS cells with H. pylori strains bearing greater numbers of cagA EPIYA‐C and multimerization (CM) motifs. On the other hand, H. pylori strains with greater numbers of EPIYA‐B relative to EPIYA‐C demonstrated a decrease in CK18 cleavage rate. Thus, H. pylori‐mediated cleavage of CK18 appeared proportional to the number of CagA EPIYA‐C and CM motifs, which seemed to be downplayed in the presence of EPIYA‐B motifs. Conclusions: Our observation associating the heterogeneity of cagA variants with the potential of H. pylori strains in the induction of CK18 cleavage as an early indication of apoptosis in gastric epithelial cells supports the fact that apoptosis may be a type‐specific trait. However, additional cagA‐targeted experiments are required to clearly identify the role of EPIYA and CM motifs in apoptosis and/or the responsible effector molecules.     

Gastric and duodenum microflora analysis after long-term Helicobacter pylori infection in Mongolian Gerbils

Background: Long‐term Helicobacter pylori infection leads to chronic gastritis, peptic ulcer, and gastric malignancies. Indigenous microflora in alimentary tract maintains a colonization barrier against pathogenic microorganisms. This study is aimed to observe the gastric and duodenum microflora alteration after H. pylori infection in Mongolian Gerbils model. Materials and Methods: A total of 18 Mongolian gerbils were randomly divided into two groups: control group and H. pylori group that were given H. pylori NCTC J99 strain intragastrically. After 12 weeks, H. pylori colonization was identified by rapid urease tests and bacterial culture. Indigenous microorganisms in stomach and duodenum were analyzed by culture method. Histopathologic examination of gastric and duodenum mucosa was also performed. Results: Three of eight gerbils had positive H. pylori colonization. After H. pylori infection, Enterococcus spp. and Staphylococcus aureus showed occurrences in stomach and duodenum. Lactobacillus spp. showed a down trend in stomach. The levels and localizations of Bifidobacterium spp., Bacteroides spp., and total aerobes were also modified. Bacteroides spp. significantly increased in H. pylori positive gerbils. No Enterobacteriaceae were detected. Positive colonization gerbils showed a higher histopathologic score of gastritis and a similar score of duodenitis. Conclusions: Long‐term H. pylori colonization affected the distribution and numbers of indigenous microflora in stomach and duodenum. Successful colonization caused a more severe gastritis. Gastric microenvironment may be unfit for lactobacilli fertility after long‐term H. pylori infection, while enterococci, S. aureus, bifidobacteria, and bacteroides showed their adaptations.     

CagA tyrosine phosphorylation in gastric epithelial cells caused by Helicobacter pylori in patients with gastric adenocarcinoma

Background. Tyrosine phosphorylation of Helicobacter pylori cytotoxin‐associated protein of in gastric epithelial cells is reported. The goals of this study are first to examine the occurrence of CagA tyrosine phosphorylation in H. pylori strains isolated from patients with gastric adenocarcinoma and gastritis, and second to clarify the relationship between the diversity of tyrosine phosphorylation motifs and the presence of CagA tyrosine phosphorylation. Methods. Fifty‐eight clinical isolates of H. pylori from patients with gastric adenocarcinoma (29 cases) and gastritis (29 cases) were studied for CagA tyrosine phosphorylation by Western blotting. Sequence diversity of tyrosine phosphorylation motifs was analysed among positive‐ or negative‐CagA tyrosine phosphorylation isolates. Results. Positive CagA tyrosine phosphorylation was found in 93.1% (27 of 29) of strains from gastric adenocarcinoma patients and 51.7% (15 of 29) of strains from gastritis patients (p < 0.001). Intact motifs were found in H. pylori isolates with CagA tyrosine phosphorylation. Of the 16 negative CagA tyrosine phosphorylation isolates, intact tyrosine phosphorylation motifs were found in 15 isolates. Conclusions. CagA tyrosine phosphorylation, which is significantly greater in strains from gastric adenocarcinoma patients, may play a role in gastric carcinogenesis, and could be a better marker of more virulent strains than the cag pathogenicity island in Asia, where the cag pathogenicity island is present in nearly all H. pylori strains. Sequence diversity of tyrosine phosphorylation motifs on CagA was not related to the presence of tyrosine phosphorylation. The absence of tyrosine phosphorylation motif might result in negative tyrosine phosphorylation phenotypes, but such motifs are not the sole factors associated with CagA tyrosine phosphorylation.     

Role of Mucin Lewis Status in Resistance to Helicobacter pylori Infection in Pediatric Patients

Background: Helicobacter pylori causes gastritis, peptic ulcer and is a risk factor for adenocarcinoma and lymphoma of the stomach. Gastric mucins, carrying highly diverse carbohydrate structures, present functional binding sites for H. pylori and may play a role in pathogenesis. However, little information is available regarding gastric mucin in children with and without stomach diseases. Materials and Methods: Expression of mucins and glycosylation was studied by immunohistochemistry on gastric biopsies from 51 children with and without H. pylori infection and/or peptic ulcer disease. Results: In all children, MUC5AC was present in the surface epithelium and MUC6 in the glands. No MUC6 in the surface epithelium or MUC2 was detected in any section. The Le^b and Le^a blood group antigens were present in the surface epithelium of 80% and 29% of children, respectively. H. pylori load was higher in Le^b negative children than in Le^b positive individuals (mean ± SEM 17.8 ± 3.5 vs 10.8 ± 1.5; p < 0.05), but there was no correlation between Le^a or Le^b status and gastritis, nodularity, and gastric or duodenal ulcer (DU). Expression of sialyl‐Le^x was associated with H. pylori infection, and DU. Conclusions: Mucin expression and glycosylation is similar in children and adults. However, in contrast to adults, pediatric H. pylori infection is not accompanied by aberrant expression of MUC6 or MUC2. Furthermore, the lower H. pylori density in Le^b positive children indicates that H. pylori is suppressed in the presence of gastric mucins decorated with Le^b, the binding site of the H. pylori BabA adhesin.     

Seroreactivity to 19.5-kDa antigen in combination with absence of seroreactivity to 35-kDa antigen is associated with an increased risk of gastric adenocarcinoma

Background. Only a minority of those infected with Helicobacter pylori will develop gastric cancer. Stratification of H. pylori strains based on carcinogenic potential will provide a basis for selective surveillance and eradication therapy. We studied the anti‐H. pylori antibody profile in Asian patients with gastric adenocarcinoma to identify any H. pylori antigen that may be associated with an increased or decreased risk of gastric carcinoma. Patients and Methods. A case‐control study comparing the seroprevalence of antibodies with various H. pylori antigens in Singaporeans with gastric adenocarcinoma and the normal Singaporean population was carried out using both conventional immunoglobulin (Ig) G enzyme‐linked immunosorbent assay (ELISA) and Western blot immunoassay. Results. The seroprevalence among 44 gastric adenocarcinoma cases (70.5% males, mean age 66.7 ± 13.5 years) and 261 controls (49.8% males, mean age 61.5 ± 4.1 years) was 90.9% vs. 50.2% by IgG ELISA. In the H. pylori‐positive male population, those suffering from gastric adenocarcinoma had significantly lower seroreactivity to the 35‐kDa antigen compared with asymptomatic controls (p = .0198, OR = 3.79, 95% CI 1.24–11.61). Seropositivity to the 19.5 kDa antigen was also found to be associated with the presence of gastric adenocarcinoma in Singaporean males (p = .022, OR = 4.17, 95% CI 1.22–14.28). A ‘high‐risk’ phenotype consisting of absence of a band at 35‐kDa in combination with the presence of a band at 19.5‐kDa was significantly associated with the presence of gastric adenocarcinoma (p = .002, OR = 3.7, 95% CI 1.6–8.6). Conclusions. Stratification of H. pylori strains based on their potential for carcinogenesis, such as those strains that are seropositive for the 19.5 kDa antigen and seronegative for the 35‐kDa antigen, may provide a basis for selective eradication of H. pylori infection and future vaccine development.     

Removal of mucus for ultrastructural observation of the surface of human gastric epithelium using pronase

Background. Helicobacter pylori adhering to the human gastric epithelium causes gastric diseases such as ulcer, carcinoma and lymphoma. It is thus important to observe in detail both the surface of the epithelial cells and the H. pylori that adhered to it for the elucidation of H. pylori‐induced diseases by scanning electron microscopy (SEM). Since the thick mucus layer blocks the observation of the cell surface and the bacteria, it is generally eliminated during the processing for SEM by roughly mechanical methods, but these treatments also demolish the ultrastructure of the cells. We studied the nonmechanical method for removal of mucus layer of gastric epithelium using pronase. Materials and Methods. To determine the optimal concentration of pronase, mucin was used as a substrate for inhibition of the viscosity. Pronase was added in 2% mucin at the concentration of 10, 50, 100, 500, 1000, 2000 or 5000 unit/ml and the flowing time of the mixture was measured. Based on the digestion experiment, biopsied specimens from 24 patients with dyspepsic symptoms were fixed in glutaraldehyde and then washed in rolling with different concentration of pronase. After the pretreatment by pronase, the specimens were treated according to the standard process for SEM. Results. We succeeded in removing the mucus layer on the surface of epithelial cells from the biopsied specimens fixed in glutaraldehyde by rinsing with 2000 unit/ml pronase for 24 hours. Conclusions. Using our digestive method without destroying the ultrastructure, the earliest stage which H. pylori has adhered onto the human gastric epithelium can be observed for the investigation of H. pylori‐induced gastric disorders by SEM.     

Human gastric mucins differently regulate Helicobacter pylori proliferation, gene expression and interactions with host cells

Helicobacter pylori colonizes the mucus niche of the gastric mucosa and is a risk factor for gastritis, ulcers and cancer. The main components of the mucus layer are heavily glycosylated mucins, to which H. pylori can adhere. Mucin glycosylation differs between individuals and changes during disease. Here we have examined the H. pylori response to purified mucins from a range of tumor and normal human gastric tissue samples. Our results demonstrate that mucins from different individuals differ in how they modulate both proliferation and gene expression of H. pylori. The mucin effect on proliferation varied significantly between samples, and ranged from stimulatory to inhibitory, depending on the type of mucins and the ability of the mucins to bind to H. pylori. Tumor-derived mucins and mucins from the surface mucosa had potential to stimulate proliferation, while gland-derived mucins tended to inhibit proliferation and mucins from healthy uninfected individuals showed little effect. Artificial glycoconjugates containing H. pylori ligands also modulated H. pylori proliferation, albeit to a lesser degree than human mucins. Expression of genes important for the pathogenicity of H. pylori (babA, sabA, cagA, flaA and ureA) appeared co-regulated in response to mucins. The addition of mucins to co-cultures of H. pylori and gastric epithelial cells protected the viability of the cells and modulated the cytokine production in a manner that differed between individuals, was partially dependent of adhesion of H. pylori to the gastric cells, but also revealed that other mucin factors in addition to adhesion are important for H. pylori-induced host signaling. The combined data reveal host-specific effects on proliferation, gene expression and virulence of H. pylori due to the gastric mucin environment, demonstrating a dynamic interplay between the bacterium and its host.     

Detection of Helicobacteraceae in intestinal biopsies of children with Crohn's disease

Background: Given that members of Helicobacteraceae family colonize the intestinal mucus layer, it has been hypothesized that they may play a role in Crohn’s disease. This study investigated the presence of Helicobacteraceae DNA in biopsies collected from children with Crohn’s disease and controls. Materials and Methods: The presence of Helicobacteraceae DNA was investigated in intestinal biopsies collected from 179 children undergoing colonoscopy (Crohn’s disease n = 77, controls n = 102) using a Helicobacteraceae‐specific PCR. Results: Members of the Helicobacteraceae were detected in 32/77 children with Crohn’s disease (41.5%) and 23/102 controls (22.5%). Statistical analysis showed the prevalence of Helicobacteraceae detected in patients to be significantly higher than that in controls (p = .0062). Analysis of non‐pylori Helicobacteraceae showed that their prevalence was also significantly higher in patients than in controls (p = .04). Helicobacter pylori was detected in 14.0% of the biopsies across all groups. Given that all children tested were negative for gastric H. pylori, this was a surprising finding. Phylogenetic analysis of H. pylori sequences detected in the biopsies showed that the H. pylori strains identified in the patients did not group with gastric H. pylori included in the analysis, but rather with other H. pylori strains detected in the intestine, gall bladder, and liver. Conclusions:  The higher prevalence of Helicobacteraceae DNA in Crohn’s disease patients would suggest that members of this family may be involved in this disease. In addition, phylogenetic analysis of H. pylori strains showed that extragastric sequences clustered together, indicating that different H. pylori strains may adapt to colonize extragastric niches.     

Mechanism of action of low recurrence of gastritis caused by Helicobacter pylori with the type II urease B gene

Background. Low recurrence of gastritis is seen in patients infected with Helicobacter pylori carrying the type II urease B gene, compared with H. pylori carrying types I and III. The underlying mechanism has been studied in terms of the urease activity and interleukin (IL)‐8 production capacity of different strains of H. pylori. Materials and Methods. Forty‐five patients infected with different strains of H. pylori (type I; 15, type II; 15 and type III; 15) were enrolled in the study. H. pylori was isolated from gastric mucosa and cultured in the presence of urea at pH 5.5 to evaluate urease activity. The capacity of different strains of H. pylori to induce IL‐8 mRNA and IL‐8 from a human gastric cancer cell line and human peripheral blood mononuclear cells was evaluated. Results. The urease activity of type II H. pylori[523 ± 228 µg of ammonia/dl/10⁸ colony‐forming units (CFU)/ml] was significantly lower than that of type I (1355 ± 1369 µg of ammonia/dl/10⁸ CFU/ml) and type III (1442 ± 2229 µg of ammonia/dl/10⁸ CFU/ml) (p < .05). Gastric cancer cells cocultured with type II H. pylori produced lower levels of IL‐8 mRNA compared with type I and type III H. pylori. The levels of IL‐8 were also significantly lower in cultures induced by type II H. pylori compared with those induced by type I and type III H. pylori. Peripheral blood mononuclear cells also produced lower levels of IL‐8 when cocultured with type II compared with type I H. pylori. Conclusions. These results indicate that both the lower level of urease activity and the low IL‐8‐inducing capacity of type II H. pylori might underlie the lower recurrence rate of gastritis caused by type II H. pylori.     

Four modes of adhesion are used during Helicobacter pylori binding to human mucins in the oral and gastric niches

Background:  Helicobacter pylori causes peptic ulcer disease and gastric cancer, and the oral cavity is likely to serve as a reservoir for this pathogen. We investigated the binding of H. pylori to the mucins covering the mucosal surfaces in the niches along the oral to gastric infection route and during gastric disease and modeled the outcome of these interactions.
Materials and Methods:  A panel of seven H. pylori strains with defined binding properties was used to identify binding to human mucins from saliva, gastric juice, cardia, corpus, and antrum of healthy stomachs and of stomachs affected by gastritis at pH 7.4 and 3.0 using a microtiter-based method.
Results:  H. pylori binding to mucins differed substantially with the anatomic site, mucin type, pH, gastritis status, and H. pylori strain all having effect on binding. Mucins from saliva and gastric juice displayed the most diverse binding patterns, involving four modes of H. pylori adhesion and the MUC5B, MUC7, and MUC5AC mucins as well as the salivary agglutinin. Binding occurred via the blood-group antigen-binding adhesin (BabA), the sialic acid-binding adhesin (SabA), a charge/low pH-dependent mechanism, and a novel saliva-binding adhesin. In the healthy gastric mucus layer only BabA and acid/charge affect binding to the mucins, whereas in gastritis, the BabA/Le^b-dependent binding to MUC5AC remained, and SabA and low pH binding increased.
Conclusions:  The four H. pylori adhesion modes binding to mucins are likely to play different roles during colonization of the oral to gastric niches and during long-term infection.     

Effects of aspirin on the development of Helicobacter pylori-induced gastric inflammation and heterotopic proliferative glands in Mongolian gerbils

Background: Helicobacter pylori infection is a major cause of gastritis and gastric carcinoma. Aspirin has anti‐inflammatory and antineoplastic activity. The aim of the present study was to determine the effects of aspirin on H. pylori‐induced gastritis and the development of heterotopic proliferative glands. Methods: H. pylori strain SS1 was inoculated into the stomachs of Mongolian gerbils. Two weeks after inoculation, the animals were fed with the powder diets containing 0 p.p.m. (n = 10), 150 p.p.m. (n = 10), or 500 p.p.m. (n = 10) aspirin. Mongolian gerbils were killed after 36 weeks of infection. Uninfected Mongolian gerbils (n = 10) were used as controls. Histologic changes, epithelial cell proliferation and apoptosis, and prostaglandin E₂ (PGE₂) levels of gastric tissue were determined. Results: H. pylori infection induced gastric inflammation. Administration of aspirin did not change H. pylori‐induced gastritis, but alleviated H. pylori‐induced hyperplasia and the development of heterotopic proliferative glands. Administration of aspirin accelerated H. pylori‐associated apoptosis but decreased H. pylori‐associated cell proliferation. In addition, the increased gastric PGE₂ levels due to H. pylori infection were suppressed by treatment with aspirin, especially at the dose of 500 p.p.m. Conclusions: Aspirin alleviates H. pylori‐induced hyperplasia and the development of heterotopic proliferative glands. Moreover, aspirin increases H. pylori‐induced apoptosis. We demonstrated the antineoplastic activities of aspirin in H. pylori‐related gastric carcinogenesis.     

Helicobacter pylori does not mediate the formation of carcinogenic N-nitrosamines

Background. Both N‐nitroso compounds and colonization with Helicobacter pylori represent known risk‐factors for the development of gastric cancer. Endogenous formation of N‐nitroso compounds is thought to occur predominantly in acidic environments such as the stomach. At neutral pH, bacteria can catalyze the formation of N‐nitroso compounds. Based on experiments with a noncarcinogenic N‐nitroso compound as end product, and using only a single H. pylori strain, it was recently reported that H. pylori only displays a low nitrosation capacity. As H. pylori is a highly diverse bacterial species, it is reasonable to question the generality of this finding. In this study, several genetically distinct H. pylori strains are tested for their capacity to form carcinogenic N‐nitrosamines. Materials and Methods. Bacteria were grown in the presence of 0–1000 µM morpholine and nitrite (in a 1 : 1 molar ratio), at pH 7, 5 and 3. Results. Incubation of Neisseria cinerea (positive control) with 500 µM morpholine and 500 µM nitrite, resulted in a significant increase in formation of N‐nitrosomorpholine, but there was no significant induction of N‐nitrosomorpholine formation by any of the H. pylori strains, at any of the three pH conditions. Conclusion. H. pylori does not induce formation of the carcinogenic N‐nitrosomorpholine in vitro. The previously reported weak nitrosation capacity of H. pylori is not sufficient to nitrosate the more difficultly nitrosatable morpholine. This probably also holds true for other secondary amines. These results imply that the increased incidence of gastric cancer formation that is associated with gastric colonization by H. pylori is unlikely to result from the direct induced formation of carcinogenic nitrosamines by H. pylori. However, this has to be further confirmed in in vivo studies.     

Clinicopathological analysis of early-stage gastric cancers detected after successful eradication of Helicobacter pylori

Background and Aims: The results of a randomized controlled study and meta‐analysis study have recently proved that Helicobacter pylori eradication has a preventive effect against the development of metachronous and primary gastric cancer. However, gastric cancer is sometimes detected after successful eradication. There is a lack of study about gastric cancers in eradicated patients. To clarify the characteristics of gastric cancers detected after H. pylori eradication, we analyzed the clinicopathological features of these cancers. Methods: The subjects were 18 early‐stage gastric cancer specimens resected from 17 patients who had received successful eradication of H. pylori from February 1995 to March 2009. The control group consisted of 36 specimens from noneradicated patients with persistent H. pylori infection who were matched with the subjects in age, sex, and depth of invasion. Clinicopathological features and mucin phenotypes of gastric cancer were clinically and immunohistologically evaluated. Results: The average diameter of gastric cancer was smaller and Ki‐67 index was lower in the eradication group. The morphological distribution of depression types was significantly lower in the control group. Immunohistochemical phenotyping revealed that 72.2% of the lesions in the eradicated group were complete gastric type or gastric predominant mixed type, whereas the percentages of gastric type and intestinal type in the control group were similar. Conclusion: Our findings indicate that the clinicopathological characteristics of gastric cancers detected after H. pylori eradication are different from those of gastric cancers in patients with persistent H. pylori infection. H. pylori eradication may suppress intestinalization during the development of gastric cancer.     

The Interaction of Helicobacter pylori with the Adherent Mucus Gel Layer Secreted by Polarized HT29-MTX-E12 Cells

Helicobacter pylori colonises the gastric mucosa of humans. The majority of organisms live in mucus. These organisms are an important reservoir for infection of the underlying epithelium. Cell culture models for H. pylori infection do not normally possess a mucus layer. The interaction of H. pylori with TFF1, a member of the trefoil factor family found in gastric mucin, is mediated by lipopolysaccharide. To test the hypothesis that the interaction of H. pylori with TFF1 promotes mucus colonization we characterised the interaction of H. pylori with a mucus secreting cell line, HT29-MTX-E12. An isogenic mutant of H. pylori with truncated core oligosaccharides was produced and binding to TFF1 and ability to colonise HT29-MTX-E12 cells determined. The adherent mucus layer of HT29-MTX-E12 cells contained the gastric mucin MUC5AC and trefoil factors, TFF1 and TFF3. H. pylori was found within the mucus layer in discrete clusters and in close association with TFF1. It also interacted with the membrane bound mucin MUC1 and replicated when co-cultured with the cells. An isogenic mutant of H. pylori with a truncated LPS core did not interact with TFF1, and colonization of HT29-MTX-E12 cells was reduced compared to the wild-type strain (p<0.05). Preincubation of cells with wild type LPS but not with truncated LPS resulted in reduced colonization by H. pylori. These results demonstrate that the interaction of TFF1 with H. pylori is important for colonization of gastric mucus and the core oligosaccharide of H. pylori LPS is critical for this interaction to occur. HT29-MTX-E12 cells are a useful system with which to study the interaction of bacteria with mucosal surfaces and the effect of such interactions on mediating colonization.     

Binding of extracellular matrix proteins by lactobacilli

Ten gut and ten vaginalLactobacillus strains were investigated for their ability to bind type I collagen (Cn-I) and four selected gut lactobacilli were investigated for their binding to other extracellular matrix (ECM) molecules. Immobilized Cn-I (100 mg/L) in wells of microtitre plates was bound by all 10 autoaggregating vaginal strains and by 3 strains of gut lactobacilli from piglets in the range ofA ₅₇₀ readings 0.114–1.806.L. acidophilus strain SV31 was much more adherent than the rest of strains. All four gut lactobacilli tested for binding to other ECM molecules displayed good binding to porcine fibronectin and heparin and some of them bound weakly to fetuin and porcine mucin. No binding of these strains was observed to bovine mucin, bovine fibrinogen and bovine lactoferrin.