Helicobacter pylori evolution during progression from chronic atrophic gastritis to gastric cancer and its impact on gastric stem cells

Abstract

We have characterized the adaptations of Helicobacter pylori to a rarely captured event in the evolution of its impact on host biology-the transition from chronic atrophic gastritis (ChAG) to gastric adenocarcinoma-and defined the impact of these adaptations on an intriguing but poorly characterized interaction between this bacterium and gastric epithelial stem cells. Bacterial isolates were obtained from a single human host colonized with a single dominant strain before and after his progression from ChAG to gastric adenocarcinoma during a 4-year interval. Draft genome assemblies were generated from two isolates, one ChAG-associated, the other cancer-associated. The cancer-associated strain was less fit in a gnotobiotic transgenic mouse model of human ChAG and better able to establish itself within a mouse gastric epithelial progenitor-derived cell line (mGEP) that supports bacterial attachment. GeneChip-based comparisons of the transcriptomes of mGEPs and a control mouse gastric epithelial cell line revealed that, upon infection, the cancer-associated strain regulates expression of GEP-associated signaling and metabolic pathways, and tumor suppressor genes associated with development of gastric cancer in humans, in a manner distinct from the ChAG-associated isolate. The effects on GEP metabolic pathways, some of which were confirmed in gnotobiotic mice, together with observed changes in the bacterial transcriptome are predicted to support aspects of an endosymbiosis between this microbe and gastric stem cells. These results provide insights about how H. pylori may adapt to and influence stem cell biology and how its intracellular residency could contribute to gastric tumorigenesis.

Fig. 1.

Association of H. pylori with gastric epithelial progenitors. In this multilabel immunohistochemical analysis, bacteria (red) are seen in association with BrdU-labeled GEPs (green) in the epithelium of a gnotobiotic transgenic tox176 mouse colonized with the cancer-associated Kx2 isolate. The epithelial marker, E-cadherin, appears white, and nuclei are stained blue with DAPI. Dashed lines indicate Kx2-associated progenitor cells that were targeted by n-LCM for transcriptional assays (see SI Text for details). (Scale bar: 25 μm.)

Fig. 2.

Flow-chart showing the approach used to define isolate-specific and progenitor-specific mGEP transcriptional responses to infection with the Kx1 and Kx2 isolates.

Fig. 3.

Kx2-associated up-regulation of antizyme inhibitor 1 expression in gastric epithelial progenitors in vitro and in vivo. (A) Mouse GeneChip analysis showing mGEP-specific up-regulation of antizyme inhibitor 1 (Azin1) after infection with the cancer-associated Kx2 strain. Three independent experiments for each condition were performed. Numbers at the bottom indicate standard deviations above (red) or below (green) the mean level of expression (black) of the Azin1 gene. (B) qRT-PCR of mGEP responses to Kx1 and Kx2 infection. *, P < 0.05; **, P < 0.01 in a two-sample two-tailed Student's t test with three biological replicates per condition for each mGEP transcript. (C) Up-regulation of Azin1 and Odc1 in gastric epithelial progenitor cells recovered by n-LCM from a tox176 mouse that had been colonized for 6 months with the Kx2 strain (mean values ± 1 SD for two to three technical replicates are plotted). *, P < 0.05.