DNA damage and repair in Helicobacter pylori-infected gastric mucosa cells

Helicobacter pylori is a common human pathogen and its infection is believed to contribute to gastric cancer. Impaired DNA repair may fuel up cancer transformation by the accumulation of mutation and increased susceptibility to exogenous carcinogens. To evaluate the role of infection of H. pylori in DNA damage and repair we determined: (1) the level of endogenous basal, oxidative and alkylative DNA damage, and (2) the efficacy of removal of DNA damage induced by hydrogen peroxide and the antibiotic amoxicillin in the H. pylori-infected and non-infected GMCs. DNA damage and the efficacy of DNA repair were evaluated by the alkaline single cell gel electrophoresis (comet assay). Specific damage to the DNA bases were assayed with the DNA repair enzymes formamidopyrimidine-DNA glycosylase (Fpg) recognizing oxidized DNA bases and 3-methyladenine-DNA glycosylase II (AlkA) recognizing alkylated bases. The level of basal and oxidative DNA in the infected GMCs was higher than non-infected cells. H. pylori-infected GMCs displayed enhanced susceptibility to hydrogen peroxide than control cells. There was no difference between the efficacy of DNA repair in the infected and non-infected cells after treatment with hydrogen peroxide and amoxicillin. Our results indicate that H. pylori infection may be correlated with oxidative DNA damage in GMCs. Therefore, these features can be considered as a risk marker for gastric cancer associated with H. pylori infection and the comet assay may be applied to evaluate this marker.     

Increase of antigen-presenting cells in the gastric mucosa of Helicobacter pylori-infected children

BACKGROUND: Infection with Helicobacter pylori leads to an increase of T cells in the gastric mucosa of children. In contrast to peripheral blood, where monocytes are the most abundant antigen-presenting cells, CD14+ macrophages are very rare in infected gastric mucosa. We postulated that other types of antigen-presenting cells must be present in infected gastric mucosa. MATERIAL AND METHODS: Antral biopsies were obtained from 56 children. The cellular expression of major histocompatibility complex (MHC) class II molecules, CD1a/b, and CD23, which are involved in antigen presentation were analyzed by immunohistochemistry. In addition, T cells (CD4, CD8, CD25, and gamma/delta-TCR), B cells (anti-IgM), macrophages (CD14) and granulocytes (CD15) were quantified. RESULTS: Twenty-eight children were H. pylori-infected. Thirteen children were healthy, 15 had other gastric pathologies. T cells (p<.0001), B cells (p<.0001), CD23+ (p<.0001), and CD1a/b+ (p<.005) cells were significantly increased in the lamina propria of H. pylori-infected children, whereas macrophages were rare without significant differences among the groups. Within the epithelium, CD8+ T lymphocytes predominated clearly over CD4+ cells. H. pylori-negative children had only few MHC class II-positive cells within the gastric epithelium, whereas MHC class II antigens were strongly expressed on epithelial cells (p<.0001) of all H. pylori-infected children. CONCLUSION: Helicobacter pylori infection leads to an enhanced expression of antigen-presenting molecules together with a parallel rise of T cells in the lamina propria. This may represent an effort of the immune system to optimize local immune responses against H. pylori. We speculate that the epithelium participates in the initiation of a local immune response against H. pylori.     

IL-21 is highly produced in Helicobacter pylori-infected gastric mucosa and promotes gelatinases synthesis

Helicobacter pylori (Hp) infection is associated with gastric inflammation and ulceration. The pathways of tissue damage in Hp-infected subjects are complex, but evidence indicates that T cell-derived cytokines enhance the synthesis of matrix metalloproteinases (MMP) that contribute to mucosal ulceration and epithelial damage. In this study, we have examined the role of the T cell cytokine IL-21 in Hp-infected gastric mucosa and evaluated whether IL-21 regulates MMP production by gastric epithelial cells. We show that IL-21 is constitutively expressed in gastric mucosa and is more abundant in biopsy specimens and purified mucosal CD3(+) T cells from Hp-infected patients compared with normal patients and disease controls. We also demonstrate that IL-21R is expressed by primary gastric epithelial cells, as well as by the gastric epithelial cell lines AGS and MKN28. Consistently, AGS cells respond to IL-21 by increasing production of MMP-2 and MMP-9, but not MMP-1, MMP-3, MMP-7, or tissue inhibitors of MMP. Analysis of signaling pathways leading to MMP production reveals that IL-21 enhances NF-kappaB but not MAPK activation, and inhibition of NF-kappaB activation reduces IL-21-induced MMP-2 and MMP-9 production. Finally, we show that treatment of Hp-infected gastric explants with anti-IL-21 reduces epithelial cell-derived MMP-2 and MMP-9 production. These data indicate that IL-21 is overexpressed in Hp-infected gastric mucosa where it could contribute to increased epithelial gelatinase production.     

Cellular stress-related protein expression in Helicobacter pylori-infected gastric epithelial AGS cells

Helicobacter pylori infection leads to gastroduodenal inflammation, peptic ulceration, and gastric carcinoma. Moreover, H. pylori may induce disease-specific protein expression in gastric epithelial cells. The present study was aimed at determining differentially expressed proteins in H. pylori-infected gastric epithelial AGS cells. AGS cells were treated with H. pylori at a bacterium/cell ratio of 300:1 for 12 h. Altered protein patterns as separated by two-dimensional electrophoresis using pH gradients of 4-7 were conclusively identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) analysis of the peptide digests. Four differentially expressed proteins, whose expression levels were increased by more than two-fold in H. pylori-infected cells, were analyzed. These proteins (14-3-3 protein alpha/beta, cullin homolog 3, alpha-enolase, ezrin) are known to be related to cell proliferation, cell adhesion, and carcinogenesis, and may be mediated by cellular stress, such as reactive oxygen species. In conclusion, the identification of these differentially expressed proteins provide valuable information for the understanding of the pathophysiologic mechanisms of H. pylori-induced gastric diseases, and may be useful as prognostic indices of H. pylori-related gastric disorders.     

Trace element levels in serum and gastric mucosa in patients with Helicobacter pylori positive and negative gastritis

BackgroundMost trace elements are inhibited by Helicobacter pylori-infection, and variations in specific element levels are linked to the development of stomach cancer. This is the first study to show the relationship between serum and tissue concentrations of twenty-five trace elements and H. pylori infection status. This study purposed to define serum and tissue trace element levels of 25 healthy individuals with Helicobacter pylori-positive gastritis and Helicobacter pylori-negative gastritis and to reveal their relationship with the disease.MethodsStudy groups consisted of sixty-two patients with Helicobacter pylori-positive, thirty-seven patients with Helicobacter pylori-negative, and thirty healthy individuals. Serum and tissue concentrations of twenty-five elements (aluminum, boron, arsenic, barium, calcium, beryllium, copper, cadmium, iron, chromium, mercury, lithium, potassium, magnesium, sodium, manganese, nickel, phosphorus, lead, scandium, strontium, selenium, tellurium, titanium, zinc) were defined by inductively coupled plasma optical emission spectrometry.ResultsExcept for copper, lithium, and strontium elements in serum samples, other trace elements differed significantly between the groups (p < 0.05). The serum chromium (p = 0.002), mercury (p = 0.001), boron (p < 0.001), and cadmium (p < 0.001) levels of H. pylori-negative gastritis and H. pylori-positive gastritis participants were significantly different, and their serum concentrations were less than 0.5 µ/l. Boron, barium, beryllium, chromium, lithium, phosphorus and strontium elements in tissue samples did not differ significantly between the groups (p > 0.05). Manganese, nickel, tellurium and titanium elements were not detected in tissue and serum samples. The mean concentrations of calcium, beryllium, chromium, iron, potassium, lithium, magnesium, scandium, and selenium were higher in the tissues of patients with H. pylori gastritis compared to healthy control tissues. Also, cadmium could not be detected in tissue samples. There was a significant difference between H. pylori-infected tissue and serum chromium levels (p = 0.001), with lower levels detected in tissue samples.ConclusionThis is the first study that we are knowledgeable of that reports the concentrations of twenty five elements in both serum and tissue samples, as well as the relationship between trace elements and Helicobacter pylori-infection status. Dietary adjustment is indicated as an adjunct to medical therapy to stabilize trace elements because Helicobacter pylori bacteria cause inflammation and impair element absorption in gastritis patients. We also think that this study will shed light on studies on the relationship between Helicobacter pylori-trace elements and serum-tissue/healthy serum-tissue trace element levels of patients with Helicobacter pylori gastritis.     

HLA-DR expression on CD8 lymphocytes from gastric mucosa in urease-positive and urease-negative gastritis

Abstract We isolated lymphocytes from chronically inflamed gastric mucosa. We analysed the expression of IL-2 receptors (CD25), transferin receptors (CD71) and HLA-DR molecules on T lymphocytes by flow cytometric analysis in 16 patients with urease-positive and in 7 patients with urease-negative chronic gastritis. In G0, G1 and G2 histological type (Sydney classification) of gastritis the number of lymphocytes obtained from the gastric mucosa biopsies was too low for the flow cytometric analysis. However, in G3 histological type of chronic gastritis we obtained enough cells for the flow cytometric analysis in 75 %. We demonstrated a significant increase in HLA-DR expression on CD8 cells from patients with urease-positive gastritis compared to urease-negative gastritis. We also observed a statistically non-significant increase in HLA-DR expression on CD3 cells, and in CD71 expression on both CD3 and CD8 cells in urease-positive gastritis. However, no difference in CD25 expression was found between the two types of gastritis.     

The influence of Lactobacillus brevis on ornithine decarboxylase activity and polyamine profiles in Helicobacter pylori-infected gastric mucosa

Background. Functional probiotics may prevent Helicobacter pylori infection, and some evidence suggests that they also possess antitumor properties. Lactobacillus brevis (CD2) is a functional Lactobacillus strain with peculiar biochemical features, essentially related to the activity of arginine deiminase. This enzyme catalyzes the catabolism of arginine and affects the biosynthesis of polyamines (putrescine, spermidine, and spermine). Polyamines are polycations found in high concentrations in both normal and neoplastic cells. Our aims were: 1, to assess whether oral administration of L. brevis (CD2) affects H. pylori survival in the human gastric mucosa; 2, to evaluate the effects of L. brevis (CD2) on polyamine biosynthesis in gastric biopsies from H. pylori‐positive patients. Materials and Methods. For 3 weeks before endoscopy, 22 H. pylori‐positive dyspeptic patients randomly received (ratio 1 : 1) high oral doses of L. brevis (CD2) or placebo. Before and after treatment, H. pylori infection was determined by urea breath test (UBT). In gastric biopsies, ornithine decarboxylase activity and polyamine levels were, respectively, evaluated by a radiometric technique and high‐pressure liquid chromatography (HPLC). Results. L. brevis (CD2) treatment did not eradicate H. pylori. However, a reduction in the UBT delta values occurred, suggesting a decrease in intragastric bacterial load. Significantly, L. brevis (CD2) induced a decrease in gastric ornithine decarboxylase activity and polyamine levels. Conclusions. Our data support the hypothesis that L. brevis (CD2) treatment decreases H. pylori colonization, thus reducing polyamine biosynthesis. Alternatively, the arginine deiminase activity following L. brevis (CD2) administration might cause arginine deficiency, preventing polyamine generation from gastric cells.     

Oxidative damage of the gastric mucosa in Helicobacter pylori positive chronic atrophic and nonatrophic gastritis, before and after eradication

Background. Helicobacter pylori is the main cause of gastritis and a primary carcinogen. The aim of this study was to assess oxidative damage in mucosal compartments of gastric mucosa in H. pylori positive and negative atrophic and nonatrophic gastritis. Materials and methods. Five groups of 10 patients each were identified according to H. pylori positive or negative chronic atrophic (Hp‐CAG and CAG, respectively) and nonatrophic gastritis (Hp‐CG and CG, respectively), and H. pylori negative normal mucosa (controls). Oxidative damage was evaluated by nitrotyrosine immunohistochemistry in the whole mucosa and in each compartment at baseline and at 2 and 12 months after eradication. Types of intestinal metaplasia were classified by histochemistry. Results. Total nitrotyrosine levels appeared significantly higher in H. pylori positive than in negative patients, and in Hp‐CAG than in Hp‐CG (p < .001); no differences were found between H. pylori negative gastritis and normal mucosa. Nitrotyrosine were found in foveolae and intestinal metaplasia only in Hp‐CAG. At 12 months after H. pylori eradication, total nitrotyrosine levels showed a trend toward a decrease in Hp‐CG and decreased significantly in Hp‐CAG (p = .002), disappearing from the foveolae (p = .002), but remaining unchanged in intestinal metaplasia. Type I and II of intestinal metaplasia were present with the same prevalence in Hp‐CAG and CAG, and did not change after H. pylori eradication. Conclusions. Oxidative damage of the gastric mucosa increases from Hp‐CG to Hp‐CAG, involving the foveolae and intestinal metaplasia. H. pylori eradication induces a complete healing of foveolae but not of intestinal metaplasia, reducing the overall oxidative damage in the mucosa.     

Level of group E prostaglandins in gastric mucosa of patients with various forms of chronic gastritis

There was studied prostaglandins E concentration in bioptats of gastric mucosa in 158 patients with various forms and localization of chronic gastritis using a radioimmunological method. Chronic gastritis without atrophy and with mild atrophy showed to have decreased levels of prostaglandins E in comparison with the control. In the patients with atrophic gastritis with "reconstruction" of epithelium there was revealed an increase of prostaglandins concentration, which was maximal in the patients with atrophic-hyperplastic forms of the disease.     

Metabolic alterations in patients with Helicobacter pylori-related gastritis: The H. pylori-gut microbiota-metabolism axis in progression of the chronic inflammation in the gastric mucosa

Purpose

To characterize the serum metabolism in patients with Helicobacter pylori-positive and H. pylori-negative gastritis.

Methods

Clinical data and serum gastric function parameters, PGI (pepsinogen I), PGII, PGR (PGI/II), and G-17 (gastrin-17) of 117 patients with chronic gastritis were collected, including 57 H. pylori positive and 60 H. pylori negative subjects. Twenty cases in each group were randomly selected to collect intestinal mucosa specimens and serum samples. The gut microbiota profiles were generated by 16S rRNA gene sequencing, and the serum metabolites were analyzed by a targeted metabolomics approach based on liquid chromatography–mass spectrometry (LC–MS) technology.

Results

Altered expression of 20 metabolites, including isovaleric acid, was detected in patients with HPAG. Some taxa of Bacteroides, Fusobacterium, and Prevotella in the gut microbiota showed significant correlations with differentially expressed metabolites between H. pylori positive and H. pylori negative individuals. As a result, an H. pylori-gut microbiota-metabolism (HGM) axis was proposed.

Conclusion

Helicobacter pylori infection may influence the progression of mucosal diseases and the emergence of other complications in the host by altering the gut microbiota, and thus affecting the host serum metabolism.     

P53 protein and Ki-67 expression in chronic gastritis patients with positive Helicobacter pylori infection

The present study was carried out to assess the predictive value and expression of the proliferative activity of Ki-67 and the expression of p53 protein in Helicobacter pylori associated chronic gastritis. This study comprised archival blocks from 20 dyspeptic patients who at National Hepatology and Tropical Medicine Research Institute underwent a diagnostic oesophago-gastroduodenoscopy with multiple gastric antral endoscopic biopsies for histological examination. The blocks were cut at 5 nM thicknesses, stained by hematoxylin and eosin to score the inflammatory grade and subjected to Giemsa stain to assess H. pylori infection, and then immune–histochemical method was done to determine protein P53 and Ki-67. The obtained results indicated that there was no significant association between the expression of Ki67 and P53 in the studied cases. There was no significant association between Ki67 and P53 in the presence of intestinal atrophy, intestinal metaplasia, intestinal activity and intestinal inflammation. While, there was significant association between Ki67 and P53 in intestinal dysplasia, P = 0.015, 0.025, respectively. It could be concluded that the significant association of the proliferative marker Ki-67 and apoptotic marker p53 protein with intestinal dysplasia may be one of the main predictive values in the development of gastric carcinoma in patients with gastritis secondary to H. pylori infection.     

Eradication of Helicobacter pylori and resolution of gastritis in the gastric mucosa of IL-10-deficient mice

BACKGROUND: Helicobacter pylori has been shown to induce pronounced gastric inflammation in the absence of interleukin-10 (IL-10) by 6 weeks post inoculation. The ability of IL-10(-/-) mice to eradicate H. pylori has not been demonstrated, possibly due to early sacrifice. Therefore, the long-term effect of enhanced gastritis on H. pylori colonization was determined in IL-10(-/-) mice. METHODS: C57BL/6 and IL-10(-/-) mice were infected with H. pylori and assessed for the degree of gastritis, bacterial load, and in vitro T-cell recall response at 4 and 16 weeks of infection. RESULTS: Infection of IL-10(-/-) mice resulted in significantly more severe gastritis than wild-type control mice and eradication of H. pylori by 4 weeks post inoculation. By 16 weeks, the level of gastritis in IL-10(-/-) was reduced to the levels observed in wild-type mice. Splenocytes from IL-10(-/-) mice were prone to produce significantly greater amounts of IFN-gamma than wild-type mice when stimulated with bacterial antigens. CONCLUSIONS: These results indicate that the host is capable of spontaneously eradicating H. pylori from the gastric mucosa when inflammation is elevated beyond the chronic inflammation induced in wild-type mice, and that the gastritis dissipates following bacterial eradication. Additionally, these data provide support for a model of gastrointestinal immunity in which naturally occurring IL-10-producing regulatory T cells modulate the host response to gastrointestinal bacteria.     

Epithelial response of the rat gastric mucosa to chronic superficial injury.

Chronic injury to the healthy gastric mucosa with noxious agents such as aspirin or alcohol induces a progressive strengthening of the stomach wall against these insults. The present study examined the histologic response of the rat gastric mucosa to chronic destruction of the superficial mucosa for one month with hypertonic saline. The number, position and morphology of proliferating, parietal, G and D cells were followed during mucosal injury and one month of recovery. The results showed that chronic injury reduced parietal cell numbers by about 30 percent, particularly in the middle of the mucosal thickness where a clear zone was formed by hypertrophy of mucous neck-like cells. G cells were also reduced by about 50 percent, but there were no changes in D cells. Chronic injury induced a marked increase in the number of antral (+112 percent) and fundic (+250 percent) proliferating cells. CONCLUSION: The rat gastric mucosa responds to chronic superficial injury by down-regulation of acid secretory cells and gastrin secreting cells and an up-regulation of proliferating cells. The appearance of a prominent layer of mucous neck-like cells may indicate a new secretory function for these cells.