Fig 2: AQP5 knockout attenuates H. pylori-induced gastritis in mice.A H&E staining and histological score of inflammation in gastric mucosa of mice. B Levels of IL-6 and TNF-α in gastric mucosa measured by ELISA. C Flow cytometry of isolated neutrophils (CD45+CD11b+Ly6C–Ly6G+) and monocytes (CD45+CD11b+Ly6G–Ly6C+). D The contents of immune cells in gastric mucosa of mice detected by flow cytometry. E Quantification of apoptotic cells in gastric mucosa detected by TUNEL assay. *p < 0.05, **p < 0.01, ns not significant. Data were shown as the mean ± standard deviation. n = 6 mice/group. Comparisons of data between two groups were analyzed by independent sample t-test.

Fig 3: H. pylori infection facilitates the activation of Wnt/β-catenin pathway by upregulating AQP5.A The differences in enriched pathways between AQP5 high expression and low expression groups analyzed by GSEA. B β-catenin expression in gastric mucosa of mice detected by immunohistochemistry (n = 6). C β-catenin protein level in gastric mucosa of mice detected by Western blot analysis (n = 6). D AQP5 mRNA level in AGS cells determined by RT-qPCR. E The activation of Wnt/β-catenin pathway in primary GECs of mice detected by TOPFlash luciferase reporter assay. F β-catenin expression in mouse primary GECs detected by immunofluorescence staining. *p < 0.05, **p < 0.01, ns not significant. Data were shown as the mean ± standard deviation. Cell experiments were repeated three times independently. One-way ANOVA with Dunnett’s post hoc test was applied for the comparison of data among multiple groups.

Fig 4: H. pylori augments AQP5 expression by upregulating ASCL1.A Veen map of the transcription factors binding AQP5 predicted by hTFtarget database and AQP5-related genes in microarray data GSE106656. B, ASCL1 expression in 14 gastritis patients and 7 IM patients in microarray data GSE106656. C Correlation between ASCL1 and AQP5 expression in microarray data GSE106656. D ASCL1 mRNA level in mouse gastric mucosa measured by RT-qPCR (n = 6). E, ASCL1 mRNA level in mouse primary GECs measured by RT-qPCR. F Enrichment of ASCL1 on the AQP5 promoter detected by ChIP assay. G The silencing efficiency of si-ASCL1-1 or si-ASCL1-2 as reflected by ASCL1 mRNA level in mouse primary GECs transduced with si-ASCL1-1 or si-ASCL1-2 measured by RT-qPCR. H The target relationship between ASCL1 and AQP5 verified by dual luciferase reporter assay. I ASCL1 mRNA level in H. pylori-infected mouse primary GECs transduced with si-ASCL1 measured by RT-qPCR. *p < 0.05, **p < 0.01. Data were shown as the mean ± standard deviation. Cell experiments were repeated three times independently. Comparisons of data between two groups were analyzed by independent sample t-test. One-way ANOVA with Dunnett’s post hoc test was applied for the comparison of data among multiple groups. The data between groups at different time points were compared by repeated measures ANOVA.

Fig 5: H. pylori infection elevates AQP5 expression in gastric mucosa.A Volcano map of differentially expressed genes in microarray data GSE106656. Red represents upregulated genes and green represents downregulated genes with |log2FC | ≥ 1 and p < 0.05. B, AQP5 expression in 14 gastritis patients and 7 IM patients in microarray data GSE106656. C, H. pylori infection in mice detected by Giemsa staining. D, AQP5 mRNA level in mouse gastric mucosa determined by RT-qPCR (n = 6). E, AQP5 protein level in mouse gastric mucosa determined by immunohistochemistry (n = 3). F, AQP5 protein level in mouse isolated gastric mucosa determined by Western blot analysis (n = 3). G, AQP5 mRNA level in mouse isolated gastric mucosa determined by RT-qPCR (n = 3). *p < 0.05, **p < 0.01. Data were shown as the mean ± standard deviation. Comparisons of data between two groups were analyzed by independent sample t-test. One-way ANOVA was applied for the comparison of data among multiple groups. The data between groups at different time points were compared by repeated measures ANOVA.