BACKGROUND: Helicobacter pylori (H. pylori) has been considered a definitive carcinogen in gastric cancer. Telomerase is activated in gastric cancer and some premalignant gastric lesions, including intestinal metaplasia (IM). In this study, we evaluated the relationships of both H. pylori infection and telomerase activity with endoscopic and histologic features in IM. The effects of H. pylori eradication on endoscopic, histologic and biochemical changes were evaluated. METHODS: Endoscopic biopsies were obtained from 43 patients with IM for rapid urease, histologic and telomerase tests. The endoscopic and histologic features, H. pylori infection and telomerase were assessed. After H. pylori eradication, 15 patients were re-evaluated and compared after 4 months. RESULTS: Thirty-four (79.1%) patients were infected with H. pylori. The incidence of H. pylori infection was borderline correlated to the severity of IM (p=0.076). Telomerase was elevated in eight (18.6%) patients. Telomerase tends to be high in subtype III and endoscopic grade III of IM. After H. pylori eradication, endoscopic extent (p=0.039) and histologic severity (p=0.074) showed improvements, and telomerase decreased significantly (p=0.0001). CONCLUSION: Our data suggest that telomerase is associated with the severity and extent of IM and that H. pylori eradication improves the endoscopic and histologic features in IM, and decreases telomerase activity. H. pylori eradication can be considered one of the methods to prevent gastric cancer in patients with H. pylori-infected IM. Further long-term and large-scaled study will be needed.
Female ; Helicobacter Infections/*enzymology ; *Helicobacter pylori ; Human ; Intestinal Mucosa/enzymology/microbiology/*pathology ; Male ; Metaplasia/enzymology/microbiology ; Middle Aged ; Precancerous Conditions/enzymology/microbiology ; Stomach Neoplasms/*enzymology/microbiology ; Telomerase/*metabolism

OBJECTIVETo study COX-2 expression in H. pylori infected gastric mucosal epithelia and its significance in the carcinogenesis of the stomach.

METHODSRapid urease test and histological examination with basic magnenta staining were used to assess the status of H. pylori infection in the stomach. COX-2 was detected immunohistochemically.

RESULTSCOX-2 immunostaining was positive in 1 out of 12 cases with H. pylori-negative gastric mucosa and also in 1 out of 10 cases with H. pylori-positive gastric mucosa without macroscopic alterations, while COX-2 expression was found to be positive in 5 out of 9 cases with H. pylori related superficial gastritis with mucosal erosions. COX-2 expression was detected in 5 out of 10 cases with H. pylori-positive mild atrophic gastritis, 8 out of 10 cases with H. pylori-positive moderate-severe atrophic gastritis and intestinal metaplasia, and 6 out of 8 cases with H. pylori-positive moderate-severe dysplasia. COX-2 expression was positive in 22 out of 32 cases of gastric cancer.

CONCLUSIONH. pylori may induce COX-2 expression of gastric mucosal epithelia in chronic superficial gastritis, which is related to the development of mucosal injury. According to gastric mucosal carcinogenesis pattern up-regulation of COX-2 expression is associated with gastric mucosal carcinogenesis, and involved in the early development of premalignant lesions.

Adult ; Aged ; Cyclooxygenase 2 ; biosynthesis ; genetics ; Female ; Gastric Mucosa ; enzymology ; Gastritis ; enzymology ; microbiology ; Helicobacter Infections ; enzymology ; Helicobacter pylori ; Humans ; Male ; Middle Aged

BACKGROUND/AIMS: In this study, we analysed the changes of matrix metalloproteinase-9 (MMP-9) expression in the gastric antral epithelium in respect to H. pylori eradication. METHODS: Twenty patients with H. pylori-positive chronic gastritis or peptic ulcer were studied. The expression of MMP-9 in the gastric antral biopsy specimens were compared before and after H. pylori eradication using immunohistochemical study. The positive rates and intensity of MMP-9 staining were evaluated at surface mucous cells and pyloric gland cells. RESULTS: The positive rate of MMP-9 staining in antral mucosal epithelial cells of H. pylori chronic gastritis is 63.8%. The positive rates of MMP-9 staining in the surface mucous cells and pyloric gland cells were 75.5% and 52.0% before H. pylori eradication, respectively. On the contrary, the rates were 85.5% and 82.0% after eradication. The MMP-9 overexpression in the pyloric gland cells were noticeably increased after H. pylori eradication. Strong positive staining of MMP-9 was increased significantly after H. pylori eradication in the pyloric gland cells. CONCLUSIONS: These results suggest that MMP-9 over-expression is associated with H. pylori infection as a host inflammatory response. The increased expression after H. pylori eradication indicates that MMP-9 may have a important role in remodeling or early tissue repairing process of gastric mucosa.
Adult ; Aged ; English Abstract ; Female ; Gastric Mucosa/*enzymology ; Gastritis/drug therapy/enzymology/microbiology ; Gelatinase B/*metabolism ; Helicobacter Infections/drug therapy/*enzymology/microbiology ; *Helicobacter pylori ; Humans ; Immunohistochemistry ; Male ; Middle Aged ; Peptic Ulcer/drug therapy/enzymology/microbiology ; Pyloric Antrum

BACKGROUND/AIMS: The cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS), the proteins that have the role in the gastric carcinogenesis, are stimulated by H. pylori infection in the gastric mucosa. The aim of this study was to evaluate the expression of COX-2 and iNOS proteins one year after the eradication of H. pylori. METHODS: Gastric antral mucosa from fifty eight patients with chronic gastritis who were all infected with H. pylori was examined for the expression of COX-2 and iNOS proteins before and one year after the eradication of H. pylori by immunohistochemical stain. RESULTS: COX-2 and iNOS proteins were expressed in the epithelial cells and interstitial inflammatory cells of gastric mucosa. Percent expressions of COX-2 and iNOS were significantly decreased one year after the eradication in the patients with cured infection, but not in those having persistent H. pylori. COX-2 and iNOS expressions were well correlated with H. pylori density, acute and chronic inflammation of gastric mucosa. CONCLUSIONS: The eradication of H. pylori can decrease the expression of COX-2 and iNOS in the gastric mucosa in long-term period. This seems to be due to the removal of H. pylori itself and related regression of gastric inflammation.
Cyclooxygenase 2/immunology/*metabolism ; Drug Therapy, Combination ; Gastric Mucosa/*enzymology ; Helicobacter Infections/drug therapy ; *Helicobacter pylori ; Humans ; Nitric Oxide Synthase Type II/immunology/*metabolism ; Time Factors

The gene encoding urease subunit A (ureA) of Helicobacter pylori (H. pylori) was cloned from H. pylori isolate by polymerase chain reaction (PCR). Sterile distilled water instead of DNA served as negative control. The nucleotide sequence of the amplified product was determined. Homologous analysis of the ureA against that reported by Clayton CL and the GenBank and SwissProt databases were performed with the BLAST program at the Genome Net through the Internet. 0.8 kb PCR product was amplified from all H. pylori clinical isolators. The nucleotide sequence of the ureA was determined. The nucleotide sequence of the ureA began with ATG as the initiation codon and terminated in TAA as stop codon. The coding regions had a 44% G + C content. The DNA sequence was 98% homologous to that reported by Clayton CL (688 out of 702 residues were identical). The derived amino-acid sequences of the ureA were 99% homologous to that reported by Clayton CL (232 out of 234 residues were identical). The nucleotide sequence and the predicted protein showed significant homology to ureA of H. pylori in the NCBI Entrez database.
Base Sequence ; Cloning, Molecular ; DNA, Bacterial/chemistry ; DNA, Bacterial/genetics ; *Genes, Bacterial ; Genetic Code ; Helicobacter Infections/microbiology ; Helicobacter pylori/enzymology ; Helicobacter pylori/*genetics ; Helicobacter pylori/isolation & purification ; Molecular Sequence Data ; Polymerase Chain Reaction ; Sequence Analysis, DNA ; Transcription, Genetic ; Urease/*genetics ; Urease/metabolism

PURPOSE: The role of the Ferric Uptake Regulator (FUR) in the acid resistance of Helicobacter pylori (H. pylori) has been thought to be independent of urease. However, we demonstrated in this study that Fur influences urease activity. MATERIALS AND METHODS: A fur knockout mutant of H. pylori was constructed by replacing the Fur gene with a kanamycin resistant marker gene. The wild-type H. pylori and fur mutant were compared for survival. The integrity of the inner membrane of the bacteria was evaluated by confocal microscopy using membrane-permeant and -impermeant fluorescent DNA probes. Urease activity of intact H. pylori was measured between pH 3 and 8. Real time PCR of both strains was performed for urease genes including ureI, ureE, ureF, ureG, and ureH. RESULTS: The fur deletion affected the survival of H. pylori at pH 4. The urease activity curve of the intact fur mutant showed the same shape as the wild-type but was 3-fold lower than the wild-type at a pH of less than 5. Real time PCR revealed that the expression of all genes was consistently down-regulated in the fur mutant. CONCLUSION: The results of this study showed that fur appears to be involved in acid resistant H. pylori urease activity.
Bacterial Proteins/genetics/*physiology ; Helicobacter pylori/*enzymology/genetics ; Hydrogen-Ion Concentration ; Microscopy, Confocal ; Models, Biological ; Mutation ; Repressor Proteins/genetics/*physiology ; Urease/*metabolism

OBJECTIVETo clone UreB gene of Helicobacter pylori (H. pylori) isolated from children to pGEX-4T-1 expression plasmid, and do sequence analysis.

METHODSA pair of specific primer was designed according to H. pylori UreB gene in the GenBank. Using H. pylori strains isolated from children as a template, a UreB gene was obtained by PCR. After EcoR I and Not I digestion, the PCR production was linked with pGEX-4T-1 which was digested with the same enzymes. The recombinant plasmid was transformed into E.coli BL21 and identified by double enzyme digestion and sequence analysis. The sequence results were compared with the gene sequence in the GenBank.

RESULTSA UreB gene was successfully amplified from children's H. pylori strain GZCH1. It was 1710 bp in size. The objective band was identified by double enzyme digestion. DNA sequence showed that UreB was in the correct open reading frame. The sequence comparison analysis showed that DNA and amino acid sequence identities of UreB gene with other strains were 98%. The sequence of UreB of H. pylori strain GZCH1 was submitted to GenBank (accession number:FJ455126).

CONCLUSIONSUreB of H. pylori strain GZCH1 is successfully cloned to pGEX-4T-1, which provides a basis for research of oral H. pylori vaccine.

Amino Acid Sequence ; Bacterial Vaccines ; immunology ; Child ; Cloning, Molecular ; Helicobacter pylori ; enzymology ; immunology ; Humans ; Male ; Molecular Sequence Data ; Urease ; chemistry ; genetics ; immunology

OBJECTIVETo clone Helicobacter pylori ureB gene, to construct prokaryotic expression system of the gene and to identify immunogenicity of the fusion protein.

METHODSThe ureB gene from a clinical isolate Y06 of H.pylori was amplified by high fidelity PCR. The nucleotide sequence of the target DNA amplification fragment was sequenced after T-A cloning. The expression vector pET32a with inserted ureB gene was constructed. ureB fusion protein was expressed in E.coli strain BL21DE3 induced by IPTG at different dosages. Western blot using antibody against whole cell of H.pylori as well as immunodiffusion assay using antiserum of rabbit against the fusion protein was applied to determine immunogenicity of the fusion protein.

RESULTSIn comparison with the reported corresponding sequences, the homology of nucleotide sequence of the cloned ureB gene was from 96.88% approximate, equals 97.82%, while the homology of its putative amino acid sequence was as high as 99.65% approximate, equals 99.82%. The expression output of UreB protein in pET32a-ureB-BL21DE3 system was approximately 40%of the total bacterial proteins. UreB protein was able to combine with antibody against whole cell of H.pylori and induce rabbit to produce high titer antibody after the animal was immunized with the protein.

CONCLUSIONAn expression system with high efficiency of H.pylori ureB gene has been established successfully. The expressed UreB protein with satisfactory immunogenicity and immunoreactivity can be used as antigen in H.pylori vaccine.

Animals ; Bacterial Vaccines ; immunology ; Base Sequence ; Helicobacter pylori ; enzymology ; immunology ; Humans ; Molecular Sequence Data ; Polymerase Chain Reaction ; Rabbits ; Recombinant Fusion Proteins ; immunology ; Urease ; genetics ; immunology ; Vaccines, Synthetic ; immunology

The gene encoding urease subunit A (ureA) of Helicobacter pylori (H. pylori) was cloned from H. pylori isolate by polymerase chain reaction (PCR). Sterile distilled water instead of DNA served as negative control. The nucleotide sequence of the amplified product was determined. Homologous analysis of the ureA against that reported by Clayton CL and the GenBank and SwissProt databases were performed with the BLAST program at the Genome Net through the Internet. 0.8 kb PCR product was amplified from all H. pylori clinical isolators. The nucleotide sequence of the ureA was determined. The nucleotide sequence of the ureA began with ATG as the initiation codon and terminated in TAA as stop codon. The coding regions had a 44% G + C content. The DNA sequence was 98% homologous to that reported by Clayton CL (688 out of 702 residues were identical). The derived amino-acid sequences of the ureA were 99% homologous to that reported by Clayton CL (232 out of 234 residues were identical). The nucleotide sequence and the predicted protein showed significant homology to ureA of H. pylori in the NCBI Entrez database.
Base Sequence ; Cloning, Molecular ; DNA, Bacterial ; chemistry ; genetics ; Genes, Bacterial ; Genetic Code ; Helicobacter Infections ; microbiology ; Helicobacter pylori ; enzymology ; genetics ; isolation & purification ; Humans ; Molecular Sequence Data ; Polymerase Chain Reaction ; Sequence Analysis, DNA ; Transcription, Genetic ; Urease ; genetics ; metabolism

OBJECTIVETo investigate the effects of the two-valence vaccine consisting of Helicobacter pylori (H. pylori) catalase and urease subunit UreB in preventing H. pyloriinfection in mice.

METHODSC57BL/6 mice were divided into 7 groups and immunized with intragastric administration of catalase and UreB (both 100 microg) plus cholera toxin (CT, 2 microg), catalase (100 microg) plus CT (2 microg), UreB (100 microg) plus CT (2 microg), catalase (100 microg), UreB (100 microg), CT (2 microg), or PBS, respectively, once a week for 4 consecutive weeks. Two weeks after the last immunization, all the mice were challenged by live H. pylori, and sacrificed 4 weeks after the challenge to obtain the gastric mucosa samples for detecting H. pylori using semi-quantitative bacterial culture assay.

RESULTSThe total protection rate in mice immunized with the two-valence vaccine, single-valence vaccine of catalase, and single-valence vaccine of UreB was 83.3% (20/24), 41.7% (10/24) and 54.2% (13/24), respectively, and the rate in the other 4 groups were all 0. The H. pyloricolony density in mice with vaccination was significantly lower than that of other 4 groups (P<0.05). The total protection rate and H. pylori colony density differed significantly between the two-valence vaccination group and the single-valence vaccination groups (P<0.05).

CONCLUSIONThe two-valence vaccine consisting of catalase, UreB and adjuvant has better immunoprotective effects than the single-valence vaccines.

Animals ; Bacterial Proteins ; immunology ; Bacterial Vaccines ; immunology ; Catalase ; immunology ; Helicobacter Infections ; immunology ; prevention & control ; Helicobacter pylori ; enzymology ; immunology ; Immunization ; methods ; Male ; Mice ; Mice, Inbred C57BL ; Random Allocation ; Urease ; immunology