Original Article
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World J Gastrointest Oncol. Mar 15, 2013; 5(3): 50-59
Published online Mar 15, 2013. doi: 10.4251/wjgo.v5.i3.50
CagA EPIYA polymorphisms in Colombian Helicobacter pylori strains and their influence on disease-associated cellular responses
Carlos Alberto Fajardo, Andrés Javier Quiroga, Andrea Coronado, Karen Labrador, Nicole Acosta, Pilar Delgado, Carlos Jaramillo, María Mercedes Bravo
Carlos Alberto Fajardo, Andrés Javier Quiroga, Andrea Coronado, Karen Labrador, María Mercedes Bravo, Cancer and Infectious Agents Research Group, National Cancer Institute, Bogotá 110411, Colombia
Nicole Acosta, Pilar Delgado, Carlos Jaramillo, Molecular Diagnostics and Bioinformatics Laboratory, Biological Sciences Department, Los Andes University, Bogotá 111711, Colombia
Author contributions: Bravo MM and Quiroga AJ designed the study; Fajardo CA, Quiroga AJ, Coronado A, Labrador K and Acosta N performed the research; Fajardo CA, Quiroga AJ, Coronado A, Labrador K, Acosta N, Delgado P, Jaramillo C and Bravo MM analyzed the data; Fajardo CA and Bravo MM wrote the paper.
Supported by National Cancer Institute, Bogotá, Colombia, Grant No. 41030310 to Bravo MM and Sciences Faculty, Los Andes University, Bogotá, Colombia
Correspondence to: María Mercedes Bravo, MS, Head, Cancer and Infectious Agents Research Group, National Cancer Institute, Bogotá 110411, Colombia. mbravo@cancer.gov.co
Telephone: +57-1-3341111 Fax: +57-1-3341360
Received: August 28, 2012
Revised: December 22, 2012
Accepted: January 14, 2013
Published online: March 15, 2013

AIM: To investigate the influence of the CagA diversity in Helicobacter pylori (H. pylori) strains from Colombia on the host cell biology.

METHODS: Eighty-four H. pylori-cagA positive strains with different Glu-Pro-Ile-Tyr-Ala (EPIYA) motifs patterns, isolated from patients with gastritis (n = 17), atrophic gastritis (n = 17), duodenal ulcer (n = 16), intestinal metaplasia (n = 16) and gastric cancer (n = 18), were included. To determine the integrity of the cag pathogenicity island (cagPAI) we evaluated the presence of cagA, cagT, cagE, and cag10 genes by polymerase chain reaction. AGS gastric epithelial cells were infected with each strain and assayed for translocation and tyrosine phosphorylation of CagA by western blot, secretion of interleukin-8 (IL-8) by enzyme-linked immuno sorbent assay after taking supernatants from cocultures and cell elongation induction. For cell elongation quantification, coculture photographs were taken and the proportion of “hummingbird” cells (> 15 μm) was determined.

RESULTS: Overall 72% (60/84) of the strains were found to harbor a functional cagPAI. Levels of phosphorylated CagA were significantly higher for isolates from duodenal ulcer than the ones in strains from gastritis, atrophic gastritis, intestinal metaplasia and gastric cancer (49.1% ± 23.1% vs 21.1% ± 19.5%, P < 0.02; 49.1% ± 23.1% vs 26.2% ± 14.8%, P < 0.045; 49.1% ± 23.1% vs 21.5% ± 19.5%, P < 0.043 and 49.1% ± 23.1% vs 29.5% ± 27.1%, P < 0.047 respectively). We observed variable IL-8 expression levels ranging from 0 to 810 pg/mL and from 8.8 to 1442 pg/mL at 6 h and 30 h post-infection, respectively. cagPAI-defective strains did not induce detectable levels of IL-8 at 6 h post-infection. At 30 h post-infection all strains induced IL-8 expression in AGS cells, although cagPAI-defective strains induced significantly lower levels of IL-8 than strains with a functional cagPAI (57.1 ± 56.6 pg/mL vs 513.6 ± 338.6 pg/mL, P < 0.0001). We did not observe differences in the extent of cell elongation induction between strains with a functional or a defective cagPAI in 6 h cocultures. At 24 h post infection strains with functional cagPAI showed high diversity in the extent of hummingbird phenotype induction ranging from 7% to 34%. cagPAI defective strains induced significantly lower levels of elongation than strains with functional cagPAI with one or more than one EPIYA-C motif (15.1% ± 5.2% vs 18.9% ± 4.7%, P < 0.03; and 15.1% ± 5.2% vs 20.0% ± 5.1%, P < 0.003 respectively). No differences were observed in cellular elongation induction or IL-8 expression among H. pylori strains bearing one and more than one EPIYA-C motifs, neither at 6 h nor at 24 h of coculture. There were no associations between the levels of induction of cell elongation or IL-8 expression and number of EPIYA motifs or pathology.

CONCLUSION: The present work describes a lack of association between H. pylori CagA protein EPIYA motifs variations from Colombian isolates and disease-associated cellular responses.

Keywords: Helicobacter pylori, cagA 3’ region, CagA protein, Interleukin 8, Cell elongation, Glu-Pro-Ile-Tyr-Ala