FUNCTIONAL ROLE OF GASTRIC-COMMENSAL BACTERIA IN THE DEVELOPMENT OF CD44V9-POSITIVE CANCER STEM-LIKE CELLS IN <i>H. PYLORI</i>-INFECTED GASTRIC MUCOSA

Introduction. Our previous study reported a significant increase in metachronous recurrence of early gastric cancer after endoscopic submucosal dissection due to CD44v9-positive cancer stem-like cells in H. pylori-infected stomach tissue (Br. J. Cancer 109:379, 2013). Additionally, we demonstrated development of CD44v9-positive cells due to overexpression of capping actin protein of muscle Z-line alpha subunit 1 (CAPZA1) by cells that accumulated H. pylori-derived CagA oncoprotein (Autophagy 15:242, 2019; Cell. Mol. Gastroenterol. Hepatol. 8:319, 2019); CAPZA1 expression is significantly increased by histone deacetylase (HDAC) inhibitors (Autophagy 15:242, 2019). Short-chain fatty acids (SCFAs), a subset of key gut-commensal metabolites, inhibit HDAC (J. Biol. Chem. 253:3364, 1978) and may induce CAPZA1 overexpression in H. pylori-infected gastric mucosa, resulting in the development of CD44v9-positive cells. To test this, we examined whether SCFAs mediate production of CD44v9-positive cells via upregulation of CAPZA1 expression in H. pylori infection, and searched for SCFA-producing bacteria in gastric cancer patients by a Taq amplified (TA) cloning and sequencing technique using a 16S rRNA library of gastric commensal bacteria.
Methods. A 16S rRNA library derived from gastric juice was constructed by TA cloning and sequenced by the Sanger method. H. pylori G27 and G27 cagPAI-deletion mutant strains were used. Gastric epithelial monolayers (mucosoids) were made from gastric organoids (Gut 68:400, 2019).
Results. Butyrate enhanced CAPZA1 expression in AGS cells and mucosoids via enhanced histone acetylation at the CAPZA1 promoter region. CD44v9 expression was induced in CAPZA1-overexpressing cells accumulating CagA secreted by H. pylori. CD44v9-positive cells expressed LGR5, KLF5, and SALL4, suggesting that butyrate imparts stem-cell properties to CD44v9-expressing cells. Specific SCFA-producing bacteria, Streptococcus spp., in stomachs of gastric cancer patients were identified as non-Helicobacter bacteria and their 16S rRNA levels were significantly higher in gastric cancer patients than in controls (AUROC = 0.83 [95% CI, 0.65–1.00]). Butyrate synthesizing gene butCoAT expression in Streptococcus spp. was significantly greater by 2-fold compared with that in E. coli, indicating that Streptococcus spp. have a superior ability to produce butyrate. Culture supernatant of Streptococcus spp. increased expression of CAPZA1 in AGS cells, suggesting that co-infection by Streptococcus spp. and H. pylori increases the risk of CD44v9-positive cancer-stem cell production.
Conclusion. Butyrate produced by Streptococcus spp. fosters development of CD44v9-positive cancer stem-like cells in H. pylori-infected stomach tissue through the enhancement of CAPZA1 expression.