To investigate the mechanism of long non-coding RNA plasmacytoma variant translocation 1 (PVT1) in gastric cancer caused by (HP) infection. The expression of PVT1 was detected by quantitative real-time polymerase chain reaction in HP-infected normal gastric epithelial cells GES-1. Gastric cancer cell line SGC-7901 was transfected with PVT1 small interfering RNA and co-cultured with HP,and then the inflammatory cytokines such as tumor necrosis factor-α (TNF-α),interleukin (IL) -1β,IL-6 and IL-8 were detected. After PVT1 was knocked down,the effects of PVT1 on the proliferation and migration of gastric cancer cells were examined by cell scratch assay. RNA-pulldown combined with mass spectrometry was used to detect the protein binding to PVT1,and the result of mass spectrometry was verified by RNA-pulldown combined with Western blot. In HP-infected normal gastric epithelial cells GES-1,quantitative real-time polymerase chain reaction showed that PVT1 was significantly up-regulated (=7.160,=0.019). PVT1 was knocked down in gastric cancer cells,and then infected with HP. The expressions of inflammatory factors including TNF-α (=3.899,=0.011),IL-1β (=14.610,=0.000),and IL-8 (=6.557,=0.001) were significantly inhibited. Although PVT1 knockdown had no significant effect on the proliferation ability of gastric cancer cells,it inhibited the migration of cells. PVT1 might interact with RPS8 protein. PVT1 may act as a pro-inflammatory factor and regulate gastric cancer caused by HP infection.
Cell Line, Tumor ; Cell Movement ; Cytokines ; metabolism ; Epithelial Cells ; cytology ; microbiology ; Gene Knockdown Techniques ; Helicobacter Infections ; pathology ; Helicobacter pylori ; Humans ; Inflammation ; RNA, Long Noncoding ; genetics

We tested correlations between anti-Helicobacter pylori IgG and IgA levels and the urease test, anti-CagA protein antibody, degree of gastritis, and age. In total, 509 children (0-15 years) were enrolled. Subjects were stratified as 0-4 years (n = 132), 5-9 years (n = 274), and 10-15 years (n = 103) and subjected to the urease test, histopathology, ELISA, and western blot using whole-cell lysates of H. pylori strain 51. The positivity rate in the urease test (P = 0.003), the degree of chronic gastritis (P = 0.021), and H. pylori infiltration (P < 0.001) increased with age. The median titer for anti-H. pylori IgG was 732.5 IU/mL at 0-4 years, 689.0 IU/mL at 5-9 years, and 966.0 IU/mL at 10-15 years (P < 0.001); the median titer for anti-H. pylori IgA was 61.0 IU/mL at 0-4 years, 63.5 IU/mL at 5-9 years, and 75.0 IU/mL at 10-15 years (P < 0.001). The CagA-positivity rate was 26.5% at 0-4 years, 36.5% at 5-9 years, and 46.6% at 10-15 years for IgG (P = 0.036), and 11.3% at 0-4 years, 18.6% at 5-9 years, and 23.3% at 10-15 years for IgA (P < 0.001). Anti-H. pylori IgG and IgA titers increased with the urease test grade, chronic gastritis degree, active gastritis, and H. pylori infiltration. Presence of CagA-positivity is well correlated with a high urease test grade and high anti-H. pylori IgG/IgA levels.
Adolescent ; Antibodies, Bacterial/*blood ; Antigens, Bacterial/*analysis/immunology ; Bacterial Proteins/*analysis/immunology/metabolism ; Blotting, Western ; Child ; Child, Preschool ; Chronic Disease ; Enzyme-Linked Immunosorbent Assay ; Female ; Gastritis/pathology ; Helicobacter Infections/blood/microbiology/*pathology ; Helicobacter pylori/isolation & purification/*metabolism ; Humans ; Immunoglobulin A/*blood ; Immunoglobulin G/*blood ; Infant ; Infant, Newborn ; Male ; Severity of Illness Index ; Urease/metabolism

OBJECTIVETo understand the junction protein expression of gastric mucosa including occlusal proteins (occludin), closed protein-4 (claudin-4), zonula occluden-1(ZO-1), epithelial cadherin (E-cadherin), and β ring protein (β-catenin) and the clinical significance in children with Helicobacter pylori (Hp) infection.

METHODSeventy patients in whom gastric endoscopy was performed because of nausea, vomiting, abdominal pain, bloating, acid reflux, melena, and other gastrointestinal symptoms were enrolled in this study from Dec. 2010 to Apr. 2013 in our hospital. Informed consent was signed by their parents, and the study was in accordance with the principles of medical ethics. Hp positivity was confirmed if both respiratory urea test (RUT) and Hp were positive by gastric mucosal pathology. Gastric mucosal samples from 70 patients were enrolled in this study, 23 of them were Hp negative, 47 of them were Hp positive (24 cases without peptic ulcer, 23 cases with peptic ulcer). The mRNA levels and protein expression of tight junction protein of gastric mucosa were measured by RT-PCR and Western blot respectively. The location and semi quantitative content of E-cadherin and β-catenin in gastric mucosa were detected by immunohistochemical staining method.

RESULTThe mRNA level of E-cadherin, β-catenin, ZO-1 in the Hp positive group regardless of peptic ulcer was significantly lower than that in the Hp negative group. Hp positive without peptic ulcer group were 0.0008, 0.0040, 0.0014, respectively; Hp positive with peptic ulcer group were 0.0010, 0.0090, 0.0013, respectively; Hp negative group were 0.0137, 0.0423, 0.0198, respectively (F values were 36.956, 39.893, 38.962, respectively, all P<0.05). The expression of claudin-4 mRNA in Hp positive group with peptic ulcer increased significantly, the difference among Hp positive group with peptic ulcer, Hp positive group without peptic ulcer and Hp negative group was statistically significant (0.1438 vs. 0.0926 vs. 0.0789) (F value was 11.964, P<0.05), while the difference of occludin mRNA levels among the three groups was not statistically significant.Immunohistochemistry results showed that the score of E-cadherin, β-catenin positive cell in the Hp positive patients were also significantly lower than that in the Hp negative group (t values were 3.981 and 2.340, all P<0.05, respectively). Western blot results showed that the protein levels of β-catenin in Hp positive group with peptic ulcer were significantly lower than that in Hp negative group, while the protein levels of E-cadherin in Hp positive patients regardless of peptic ulcer were decreased significantly in Hp negative group.

CONCLUSIONOur results revealed that the tight junction protein E-cadherin, β-catenin, ZO-1 expression of gastric mucosa were decreased in children with Hp infection, while claudin-4 expression was increased in Hp positive patients with peptic ulcer, suggesting that damage to gastric epithelial barrier function may be the main pathogenesis of Hp associated gastric diseases in children.

Blotting, Western ; Cadherins ; metabolism ; Child ; Claudin-4 ; metabolism ; Gastric Mucosa ; metabolism ; pathology ; Helicobacter Infections ; metabolism ; Helicobacter pylori ; Humans ; Immunohistochemistry ; Occludin ; metabolism ; Peptic Ulcer ; metabolism ; microbiology ; RNA, Messenger ; Tight Junction Proteins ; metabolism ; Tight Junctions ; metabolism ; Zonula Occludens-1 Protein ; metabolism ; beta Catenin ; metabolism

BACKGROUND/AIMS: Helicobacter pylori cytotoxin-associated gene A (CagA) has been suggested to be involved in the inactivation of Runt-related transcription factor 3 (RUNX3), a known gastric carcinoma tumor suppressor gene. It remains unclear how H. pylori CagA initiates or maintains RUNX3 promoter methylation and inactivates its protein expression in gastric carcinoma. METHODS: RUNX3 promoter methylation status, RUNX3 expression, and H. pylori CagA were investigated in 76 sample pairs of gastric carcinoma tissue. The patients' medical records were reviewed. The association between RUNX3 methylation or loss of RUNX3 expression and clinicopathologic variables according to H. pylori CagA status were investigated. RESULTS: In gastric carcinoma patients with H. pylori CagA-positive infection, RUNX3 methylation did not show association with lymphatic invasion, venous invasion, and TNM stages. However RUNX3 methylation was observed more frequently in poorly differentiated adenocarcinoma and signet ring cell carcinoma (77.8% vs. 20.0%, p=0.023) in early stage. In gastric carcinoma patients with H. pylori CagA-positive infection, loss of RUNX3 expression did not show association with lymphatic invasion, venous invasion, and TNM stages. However loss of RUNX3 expression was observed more frequently in early gastric carcinoma than in advanced gastric carcinoma (84.2% vs. 75.0%, p=0.51), but this difference was not significant. CONCLUSIONS: In gastric carcinoma patients with H. pylori CagA-positive infection, RUNX3 methylation or loss of RUNX3 expression did not show correlation with lymphovascular invasion and TNM stages. In early gastric carcinoma patients with H. pylori CagA-positive infection, RUNX3 methylation was observed more in poorly differentiated adenocarcinoma and signet ring cell carcinoma.
Adult ; Aged ; Aged, 80 and over ; Antigens, Bacterial/*metabolism ; Bacterial Proteins/*metabolism ; Cell Line, Tumor ; Core Binding Factor Alpha 3 Subunit/genetics/*metabolism ; Female ; *Gene Expression Regulation, Neoplastic ; Helicobacter Infections/complications/microbiology ; Helicobacter pylori/isolation & purification/*metabolism ; Humans ; Immunohistochemistry ; Lymphatic Metastasis ; Male ; Methylation ; Middle Aged ; Neoplasm Staging ; Promoter Regions, Genetic ; Retrospective Studies ; Stomach Neoplasms/complications/microbiology/*pathology

OBJECTIVETo investigate the correlation between infection with L-form of Helicobacter pylori (Hp-L) and the expressions of macrophage migration inhibition factor (MIF), matrix metalloproteinase 9 (MMP9), and vascular endothelial growth factor (VEGF) in gastric cancer.

METHODSHp-L was examined in 80 gastric carcinoma and 50 adjacent normal tissues by Gram staining and immunohistochemical staining, and the expressions of MIF, MMP9 and VEGF were detected by immunohistochemical staining; the expression of MIF mRNA was detected by RT-PCR and the expression of MIF, MMP9 and VEGF proteins were detected by Western blotting in 30 fresh gastric cancer tissues and the corresponding adjacent tissues.

RESULTSOf the 80 gastric carcinoma tissues, 57 (71.25%) showed Hp-L positivity detected by both Gram staining and immunohistochemical staining, as compared with a rate of only 14% in the adjacent normal tissues (P<0.05). The gastric carcinoma tissues showed higher expression levels of MIF, MMP9 and VEGF proteins than the corresponding adjacent normal mucosa; the positivity MIF, MMP-9 and VEGF proteins were significantly higher in Hp-L-positive gastric carcinoma than in Hp-L-negative cases (P<0.05). Positive correlations were found between Hp-L positivity and the expressions of MIF, MMP-9 and VEGF (r=0.598, 0.292, 0.341, respectively, P<0.05). The 30 fresh gastric cancer tissues showed also significantly higher MIF mRNA expression and MIF, MMP-9 and VEGF protein expressions than the adjacent tissues (t=3.729, P<0.01). The expressions of MIF and MMP-9 were also related to the clinicopathological factors including lymph node metastasis and depth of invasion (P<0.05).

CONCLUSIONInfection with L-form of Hp-L can be an important factor that contributes to the invasion and metastasis of gastric carcinoma, the mechanism of which involves up-regulated expressions of MIF, MMP-9 and VEGF.

Adult ; Aged ; Aged, 80 and over ; Female ; Helicobacter Infections ; metabolism ; pathology ; Helicobacter pylori ; Humans ; L Forms ; Lymphatic Metastasis ; Macrophage Migration-Inhibitory Factors ; metabolism ; Male ; Matrix Metalloproteinase 9 ; metabolism ; Middle Aged ; Stomach Neoplasms ; metabolism ; microbiology ; pathology ; Vascular Endothelial Growth Factor A ; metabolism

BACKGROUND/AIMS: Helicobacter pylori infection is linked to the development of gastric cancer. H. pylori-associated gastric inflammation is considered to be the first important step in the histogenesis of such neoplasia. However, studies that compare proteome of gastric mucosa infected with or without H. pylori are lacking. METHODS: We employed proteomics analysis on the endoscopic biopsy specimens of gastric mucosa obtained from two groups (30 cases): healthy subjects without H. pylori infection (15 cases), and gastritis patients with H. pylori infection (15 cases). The pooled proteins obtained from gastric mucosa infected with or without H. pylori were separated by two-dimensional gel electrophoresis and analyzed by a computer-aided program. The altered protein expressions were then identified by mass spectrometry and validated by Western blotting and immunohistochemistry. RESULTS: On mass spectrometry using MALDI TOF(TM) Analyzer, the up-regulation of Keratin 1, ezrin, adenosine triphosphate (ATP) synthase subunit alpha mitochondrial isoform c, Keratin type I cytoskeletal 19, and Keratin type I cytoskeletal 9 were identified; in contrast, 71 kd heat shock cognate protein, ATP synthase subunit alpha mitochondrial precursor, and annexin IV were down-regulated. Among them, membrane cytoskeleton linker ezrin was validated using Western blot and immunohistochemistry. CONCLUSIONS: Expression of ezrin was significantly different between the gastric mucosa with and without H. pylori infection. Therefore, ezrin could be considered a promising potential molecular marker for detecting H. pylori infection in gastric mucosa.
Blotting, Western ; Cytoskeletal Proteins/metabolism ; Down-Regulation ; Electrophoresis, Gel, Two-Dimensional ; Female ; Gastric Mucosa/*metabolism/microbiology ; Gastritis/complications/metabolism/pathology ; Gastroscopy ; Helicobacter Infections/complications/metabolism/*pathology ; *Helicobacter pylori ; Humans ; Immunohistochemistry ; Male ; Proteome/*analysis ; *Proteomics ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Up-Regulation

OBJECTIVETo establish an animal model of gastric cancer by long-term infection of Helicobacter pylori (H.pylori) and to elucidate the pathogenesis by proteomics analysis.

METHODSFifty male Mongolian gerbils (4-5 week-old and weighted 60-100 g) were infected with H.pylori and the gastric tissues were obtained after the infection at 3, 6, 12 and 24 months. Histological changes were evaluated by H-E staining of the gastric tissue sections. Detection of H.pylori was performed by in-vitro culture of fresh gastric tissue samples, PCR amplification of H.pylori 16s rRNA and localization by silver staining. In addition, proteins extracted from gastric tissue samples were subjected to two-dimensional electrophoresis (2-DE) at various infection time points. Protein spots with increased quantity over the course of H.pylori infection were selected and analyzed by LC-MS/MS. Finally, differentially expressed proteins between human gastric cancer tissue samples and lymph nodes were analyzed by real-time RT-PCR.

RESULTSColonization of H.pylori was observed in gastric tissue of gerbils as early as 3 months after H.pylori infection, and persisted till 24 months. Pathological examination of infected animals showed various histological changes including acute gastritis, atrophic gastritis, intestinal metaplasia and gastric carcinoma. Seventy-eight differentially expressed proteins were identified by proteomics analysis, among which 36 proteins were up-regulated and 42 were down-regulated. Analyzed by LC-MS/MS, ten proteins were identified, including lactate dehydrogenase, ATP synthase, fatty acid-binding protein, COX5B, peroxiredoxin-4, peroxide reductase, transgelin, succinyl-CoA ligase, keratin and protein disulfide-isomerase A2, among which transgelin, ATP synthase and lactate dehydrogenase were highly expressed in human gastric carcinoma and lymph nodes.

CONCLUSIONSH.pylori infection induces the expression of transgelin, ATP synthase and lactate dehydrogenase, implying possible roles in the pathogenesis of gastric diseases including cancer.

Animals ; Disease Models, Animal ; Gastritis ; microbiology ; pathology ; Gerbillinae ; Helicobacter Infections ; complications ; metabolism ; Helicobacter pylori ; genetics ; Humans ; L-Lactate Dehydrogenase ; metabolism ; Male ; Metaplasia ; Microfilament Proteins ; metabolism ; Muscle Proteins ; metabolism ; Proteomics ; Proton-Translocating ATPases ; metabolism ; RNA, Ribosomal, 16S ; analysis ; Stomach Neoplasms ; metabolism ; microbiology ; Tandem Mass Spectrometry

Gastric cancer is one of the most common malignancies and a leading cause of cancer mortality worldwide. The pathogenesis mechanisms of gastric cancer are still not fully clear. Inactivation of tumor suppressor genes and activation of oncogenes caused by genetic and epigenetic alterations are known to play significant roles in carcinogenesis. Accumulating evidence has shown that epigenetic silencing of the tumor suppressor genes, particularly caused by hypermethylation of CpG islands in promoters, is critical to carcinogenesis and metastasis. Here, we review the recent progress in the study of methylations of tumor suppressor genes involved in the pathogenesis of gastric cancer. We also briefly describe the mechanisms that induce tumor suppressor gene methylation and the status of translating these molecular mechanisms into clinical applications.
Apoptosis ; Cell Adhesion ; Cell Cycle ; CpG Islands ; genetics ; DNA Methylation ; DNA Repair ; Epigenesis, Genetic ; Genes, Tumor Suppressor ; Helicobacter Infections ; genetics ; Helicobacter pylori ; Humans ; Neoplasm Invasiveness ; Promoter Regions, Genetic ; genetics ; Stomach Neoplasms ; genetics ; metabolism ; microbiology ; pathology ; Tumor Suppressor Proteins ; genetics ; metabolism

Helicobacter pylori (H. pylori) has been a major concern as a gastric pathogen with unique features since discovered in the end of the 20th century. Recent data on comparative genome study have revealed that H. pylori has successfully survived with its host though over 58,000 years of evolution and migration from continent to continent. To maintain the symbiotic relationship with human, H. pylori has come up with ways to induce host tolerance as well as exert harmful injuries. Studies about H. pylori have accumulated the knowledge about how the cellular and molecular interactions are controlled and regulated to decide whether the symbiotic relationship is directed to diseases or peaceful mutualism. We reviewed recent literatures and research outcomes about the H. pylori and host interaction in molecular and cellular basis.
Adaptive Immunity ; Epithelial Cells/metabolism/microbiology/pathology ; Helicobacter Infections/immunology/metabolism/*pathology ; Helicobacter pylori/*immunology ; *Host-Pathogen Interactions ; Humans ; Symbiosis ; T-Lymphocytes/immunology/metabolism

OBJECTIVETo evaluate the H. pylori and Epstein-Barr virus infection in cardiac and distal gastric adenocarcinoma tissues in residents in Cixian county, a high risk area of esophageal cancer in Hebei province, and to explore the putative role of H. pylori and Epstein-Barr virus infection in the carcinogenesis of adenocarcinoma at different subsites of stomach.

METHODSH. pylori and Epstein-Barr virus latent membrane protein 1 (EBV-LMP1) immunopositivities were determined by Elivision(TM) plus immunohistochemical staining in 190 gastric adenocarcinoma tissues including 144 cases of cardiac adenocarcinoma and 46 cases of distal gastric adenocarcinoma. The relationship between H. pylori and Epstein-Barr virus infection and the subsite, Laurén type as well as other clinicopathological features of gastric adenocarcinoma were analyzed.

RESULTSNo significant difference was found between the H. pylori detection rates in cardiac and distal gastric adenocarcinomas(56.9% vs. 65.2%, P > 0.05). The detection rate of H. pylori in intestinal type was significantly higher than that in the diffuse type distal gastric adenocarcinomas (71.8% vs. 28.6%, P < 0.05). No positive expression of EBV-LMP1 was found in the gastric adenocarcinomas in this study.

CONCLUSIONSNo significant differences in H. pylori and EBV-LMP1 infections were found between cardiac and distal gastric adenocarcinomas in Cixian county. H. pylori infection is related with the intestinal type of distal gastric adenocarcinoma.

Adenocarcinoma ; microbiology ; pathology ; virology ; Aged ; Cardia ; China ; Epstein-Barr Virus Infections ; pathology ; Female ; Helicobacter Infections ; pathology ; Helicobacter pylori ; isolation & purification ; Humans ; Male ; Middle Aged ; Stomach Neoplasms ; microbiology ; pathology ; virology ; Viral Matrix Proteins ; metabolism