Patrón EPIYA en cepas de Helicobacter pylori CagA positivas en pacientes del Hospital Regional de Talca / EPIYA pattern in Helicobacter pylori CagA positive strains in patients of the Regional Hospital of Talca

BACKGROUND: The clinical outcome of Helicobacter pylori (H. pylori) infection has been related to the presence of CagA protein. This protein is highly polymorphic and its oncogenic ability depends on the number and type of tyrosine phosphorylation sites in the EPIYAs repeat sequences (A, B, C and D). AIM: To determine the EPIYA patterns of the CagA gene in H. pylori strains and its relationship with gastrointestinal pathology in infected patients of the Regional Hospital of Talca. MATERIAL AND METHODS: The strains were isolated from gastric biopsies and characterized by bacteriological and molecular methods. Gastrointestinal pathology was characterized by histopathological analysis. For the determination of the presence of the cagA gene and the EPIYAs standards, the conventional PCR technique was used. RESULTS: 138 DNA samples from H. pylori strains were analyzed. 92.0% (127/138) of the isolates carried the cagA gene, of which 66 (52.0%) corresponded to the EPIYA-ABC pattern, 43 (33.8%) to the EPIYA-ABCC pattern and 21 16.5%) to the EPIYA-ABCCC phosphorylation pattern. 50.4% (64/127) of cagA positive strains isolated from dyspeptic patients in the Maule region have more than two C sites of phosphorylation. The number of EPIYAs C motifs was associated with the presence of more severe histopathological damage in the gastric mucosa.

Correlations between the CagA Antigen and Serum Levels of Anti-Helicobacter pylori IgG and IgA in Children

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.

An Inverse Relationship between the Expression of the Gastric Tumor Suppressor RUNX3 and Infection with Helicobacter pylori in Gastric Epithelial Dysplasia

BACKGROUND/AIMS: This study was performed to determine the association between RUNX3 expression and Helicobacter pylori infection in premalignant gastric lesions. METHODS: We examined 107 patients with gastric epithelial dysplasia who had undergone endoscopic mucosal resection or submucosal dissection. All tissue samples were evaluated by RUNX3 staining and subclassified by immunophenotype. H. pylori infection in dysplastic lesions and the normal surrounding tissue was examined by silver staining, and cagA status was assessed by polymerase chain reaction. RESULTS: The loss of RUNX3 expression was observed in 62 cases (57.9%), and an association with H. pylori infection was found in 54 cases (50.5%). The infection rate with the cagA-positive H. pylori strain was 63.0%. In RUNX3-negative lesions, the rate of H. pylori infection (p=0.03) and the frequency of category 4 lesions (according to the revised Vienna classification) were high (p=0.02). In addition, the gastric mucin phenotype was predominant. In RUNX3-negative category 4 lesions, the rate of cagA-positive H. pylori infection rate was high but not significantly increased (p=0.08). CONCLUSIONS: Infection with H. pylori is associated with inactivation of RUNX3 in early gastric carcinogenesis. This mechanism was prominent in gastric cancer with a gastric mucin phenotype.

Gastrokine 1 Expression in the Human Gastric Mucosa Is Closely Associated with the Degree of Gastritis and DNA Methylation

PURPOSE: Gastrokine 1 plays an important role in gastric mucosal defense. Additionally, the Gastrokine 1-miR-185-DNMT1 axis has been shown to suppress gastric carcinogenesis through regulation of epigenetic alteration. Here, we investigated the effects of Gastrokine 1 on DNA methylation and gastritis. MATERIALS AND METHODS: Expression of Gastrokine 1, DNMT1, EZH2, and c-Myc proteins, and the presence of Helicobacter pylori CagA protein were determined in 55 non-neoplastic gastric mucosal tissue samples by western blot analysis. The CpG island methylation phenotype was also examined using six markers (p16, hMLH1, CDH1, MINT1, MINT2 and MINT31) by methylation-specific polymerase chain reaction. Histological gastritis was assessed according to the updated Sydney classification system. RESULTS: Reduced Gastrokine 1 expression was found in 20 of the 55 (36.4%) gastric mucosal tissue samples and was closely associated with miR-185 expression. The Gastrokine 1 expression level was inversely correlated with that of DNMT1, EZH2, and c-Myc, and closely associated with the degree of gastritis. The H. pylori CagA protein was detected in 26 of the 55 (47.3%) gastric mucosal tissues and was positively associated with the expression of DNMT1, EZH2, and c-Myc. In addition, 30 (54.5%) and 23 (41.9%) of the gastric mucosal tissues could be classified as CpG island methylation phenotype-low and CpG island methylation phenotype-high, respectively. Reduced expression of Gastrokine 1 and miR-185, and increased expression of DNMT1, EZH2, and c-Myc were detected in the CpG island methylation phenotype-high gastric mucosa. CONCLUSIONS: Gastrokine 1 has a crucial role in gastric inflammation and DNA methylation in gastric mucosa.

Polimorfismos genéticos de interleucinas IL-1B-511, IL-1RN, IL-10, factor de necrosis tumoral alfa-308 e infección por Helicobacter pylori CagA positivo en cáncer gástrico y úlcera duodenal en diferentes poblaciones en Colombia / Genetic polymorphisms of IL-1B-511, IL-1RN, IL-10 interleukins, tumor necrosis alfa-308 and Positive Helicobacter pylori CagA infection in gastric cancer and duodenal ulcer in different populations in Colombia

Objetivo: Determinar la asociación entre los polimorfismos IL-1B-511, IL-1RN, TNF-α-308, IL-10-819 e IL-101082 y la infección por Helicobacter pylori CagA positivo en un grupo de pacientes con cáncer gástrico y úlcera duodenal en diferentes poblaciones en Colombia. Métodos: Estudio de casos y controles con 341 pacientes: con gastritis no atrófica, 194; con cáncer gástrico, 58; úlcera duodenal con lesiones preneoplásicas, 54; y con úlcera duodenal, 35. La genotipificación de los polimorfismos se hizo por discriminación alélica usando PCR en tiempo real, y la del IL-1RN, por PCR convencional y electroforesis en agarosa. La infección por Helicobacter pylori CagA se determinó mediante ELISA. Se utilizó la regresión logística en el análisis estadístico. Resultados: Ser portador del genotipo IL-1B-511TT (OR=4,69; IC 95% 1,22-18,09) y tener una infección por Helicobacter pylori CagA positivo (OR=4,43; IC 95% 1,72-11,4) se asociaron a cáncer gástrico. Tener una infección por Helicobacter pylori CagA positivo (OR=4,39; IC95% 1,82-10,59) se asoció a la presencia de úlcera duodenal con lesiones preneoplásicas, y ser portador del genotipo IL-1B-511CT se asoció a úlcera duodenal (OR=0,30; IC 95% 0,10-0,91). Conclusión: Los resultados sugieren que la respuesta pro-inflamatoria y la genética virulenta de la bacteria son factores relacionados con los diferentes desenlaces ocasionados por la infección por Helicobacter pylori en la población estudiada; así, el polimorfismo IL-1B-511 es un factor relacionado con cáncer gástrico y úlcera duodenal, y la infección por Helicobacter pylori CagA positivo es un factor asociado a cáncer gástrico y úlcera duodenal con lesiones preneoplásicas.

Objective: To determine the association between the IL-1B-511, IL-1RN, TNF-α-308, IL-10-819 and IL-101082 polymorphisms and positive Heliocobacter pylori CagA infection in a group of patients with gastric cancer and duodenal ulcer in different populations in Colombia. Methods: A case-control study was performed on 341 patients: those with non-atrophic gastritis, 194; with gastric cancer, 58; duodenal ulcer with preneoplastic lesion, 54; and with duodenal ulcer, 35. The genotyping of polymorphisms was done with allelic discrimination using PCR in real time, and that for IL-1RN with conventional PCR and agarose electrophoresis. Helicobacter pylori CagA infection was ascertained with ELISA. Logistic regression was used in statistical analysis. Results: Being a carrier of genotype IL-1B-511TT (OR=4.69; CI 95% 1.22-18.09) and being positive for Helicobacter pylori CagA infection (OR=4.43; CI 95% 1.72-11.4) are associated with gastric cancer. Positive Helicobacter pylori CagA infection (OR=4.39; CI 95% 1.82-10.59) is associated with the presence of duodenal ulcer with preneoplastic lesions, being a carrier of genotype IL-1B-511CT is associated with duodenal ulcer (OR=0.30; CI 95% 0.10-0.91). Conclusion: The results suggest that pro-inflammatory response and virulent bacterial genetics are factors related to the different outcomes brought about by Helicobacter pylori infection in the population studied; that is, the IL-1B-511 polymorphism is a factor related to gastric cancer and duodenal ulcer, and positive Helicobacter pylori CagA infection is a factor associated with gastric cancer and duodenal ulcer with preneoplastic lesions.

Relationship between caga-positive Helicobacter pylori infection and risk of gastric cancer: a case control study in Porto Alegre, RS, Brazil / Relação entre infecção por Helicobacter pylori linhagem cagA-positiva e risco de câncer gástrico: um estudo caso-controle em Porto Alegre, Brasil

CONTEXT: Gastric cancer is the second most common cause of cancer related death worldwide. Although Helicobacter pylori has been classified as a class I carcinogen, the presence of infection is not a factor that alone is able to lead to gastric cancer, and one of the possible explanations for this is the existence of different strains of H. pylori with different degrees of virulence. OBJECTIVES: To investigate the association between cagA-positive H. pylori and gastric cancer, using polymerase chain reaction (PCR) for the detection of this bacterial strain. METHODS: Twenty-nine patients with gastric cancer were matched by sex and age (± 5 years) with 58 patients without gastric cancer, submitted to upper gastrointestinal endoscopy. All patients were evaluated for the status of infection by H. pylori (through urease test, histological analysis and PCR for the genes ureA and 16SrRNA) and by cagA-positive strain (through PCR for cagA gene). RESULTS: Evaluating the presence of infection by cagA-positive H. pylori, it was verified that the rate of infection was significantly higher in the group with gastric cancer when compared with the matched controls, occurring in 62.1 percent and 29.3 percent, respectively (OR = 3.95; CI 95 percent 1.543-10.096). CONCLUSIONS: There is an association between cagA-positive H. pylori strain and risk of gastric cancer.

CONTEXTO: O câncer gástrico é a segunda causa mais comum de mortes relacionadas à neoplasia em todo o mundo. Embora o Helicobacter pylori seja classificado como um carcinógeno classe I, a presença da infecção não é um fator que isoladamente possa conduzir ao câncer gástrico e, uma das possíveis justificativas, é a existência de diferentes linhagens de H. pylori com diferentes graus de virulência. OBJETIVO: Investigar a associação entre H. pylori cagA-positivo e câncer gástrico, utilizando a reação em cadeia de polimerase (PCR) para a detecção desta linhagem bacteriana. MÉTODOS: Vinte e nove pacientes com câncer gástrico foram pareados por sexo e por idade (± 5 anos) com 58 pacientes sem câncer gástrico, submetidos a endoscopia digestiva alta. Todos os pacientes foram avaliados quanto à presença de infecção pelo H. pylori (com teste da urease, análise histológica e PCR para os genes ureA e 16SrRNA) e pela linhagem cagA desta bactéria (com PCR para o gene cagA). RESULTADOS: Avaliando a presença de infecção por H. pylori cagA-positivo, verificou-se que a taxa da infecção era significativamente mais alta no grupo de pacientes com câncer gástrico, quando comparado com o grupo controle, ocorrendo em 62,1 por cento e em 29,3 por cento, respectivamente (OR = 3,95; CI 95 por cento 1,543-10,096). CONCLUSÕES: Há associação entre H. pylori cagA-positivo e risco de câncer gástrico.

Changes of reactive oxygen species levels in the AGS cells induced by Helicobacter pylori extracts / 中华消化杂志

Objective To investigate the relationship between Helicobacter pylori (H. pylori) infection and concentration of reactive oxygen species (ROS) in AGS cells. Methods AGS cells were cultured with either Hp11638 (CagA~+ , VacA~+ ) extract or Hp11638 mutant (CagA~+ , VacA~-) extract for 48 hours, then the cells and supernatants were collected. The concentration of ROS in AGS cells was measured by flow cytometry. The eytochrome C reduction was detected by spectrophotometer at 550 nm. Results The ROS levels in the AGS cells were correlated with two H. pylori strains in a concentration- and time-dependent manner. The ROS levels in AGS cells treated with Hp11638 extract in different concentrations or times were correspondingly higher than those treated with Hp11638 mutant extract. Similar results were found in examination of cytochrome C reduction. Conclusion The elevation of ROS in AGS cells is related to effects of H. pylori proteins, and the VaeA protein involves in the process.

Caracterización molecular y bioinformática de la proteína CagA de Helicobacter pylori a partir de biopsias gástricas de pacientes colombianos / Molecular and bioinformatic characterization of Helicobacter pylori CagA protein using gastric biopsies of Colombian patients

El gen cagA de Helicobacter pylori codifica para la proteína CagA considerada uno de los factores de virulencia cuya presencia se asocia a un mayor riesgo de padecer enfermedades gástricas severas. El presente estudio planteó como objetivo el diseño de una estrategia molecular y bioinformática útil en la determinación de la presencia de secuencias repetitivas que pueden contener uno o más motivos de fosforilación (EPIYA). Se amplificó y secuenció la región variable de cagA en muestras H. pylori CagA positivas. Se realizó una búsqueda y selección de herramientas bioinformáticas que permitieran establecer las características de los motivos EPIYA. La presencia de motivos tipo EPIYA-A y EPIYA-B, seguido por una a dos repeticiones de EPIYA-C, similares a los reportados para países de Occidente, fueron encontrados. De las aplicaciones bioinformáticas evaluadas, solo un conjunto de herramientas demostró ser útil en la caracterización de las unidades de repetición en la proteína CagA.

Helicobacter pylori CagA protein, the cagA gen product, has been considered as a virulence factor associated with a considerable increase risk for develops severe gastric illness. The purpose of this research was to design a molecular and bioinformatics strategy that allowed the establishment of phosphorylation status of the tyrosine residue of the CagA protein. The amplification and sequencing of the variable fragment region of cagA in the positive CagA samples were used to do the bioinformatics analysis in order to establish the characteristics of the EPIYA motifs. The presence of the EPIYA-A and EPIYA-B motifs, followed by one or two EPIYA-C repetitions, similar to those reported previously for occidental countries were set up. From the different bioinformatics applications that were employed only one group of tools proved to be useful to characterize the repeated units presents in the CagA protein.