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Randomized Controlled Trial
. 2025 Jun 3;32(1):55.
doi: 10.1186/s12929-025-01149-3.

Probiotics ameliorate H. pylori-associated gastric β-catenin and COX-2 carcinogenesis signaling by regulating miR-185

Yao-Jong Yang  1   2 , Chung-Tai Wu  3   4 , Hsiu-Chi Cheng  3   4 , Wei-Ying Chen  4 , Joseph T Tseng  5 , Wei-Lun Chang  3   4 , Bor-Shyang Sheu  6   7
Affiliations
Randomized Controlled Trial

Probiotics ameliorate H. pylori-associated gastric β-catenin and COX-2 carcinogenesis signaling by regulating miR-185

Yao-Jong Yang et al. J Biomed Sci. .

Abstract

Background: This study aimed to investigate whether probiotics can ameliorate the H. pylori-induced Wnt/β-catenin-related COX-2 carcinogenesis signaling pathway by regulating the expression of microRNAs (miRNAs).

Methods: An H. pylori isolate and GES-1 cells were used to establish a COX-2-associated carcinogenesis axis. Western blot analysis was conducted to investigate Wnt/β-catenin and COX-2 signaling. Next-generation sequencing and DIANA Tools identified significant differences in miRNA expressions. The probiotics Lactobacillus acidophilus and Bifidobacterium lactis were used to study anti-carcinogenesis effects in GES-1 and miRNA-transfected GES-1 cells. The H. pylori-infected patients with intestinal metaplasia (IM) were randomly allocated into probiotic treatment or not after successful eradication, the IM regression was assessed by the 2nd esophagogastroduodenoscopy one year after treatment.

Results: Pretreatment with probiotics significantly reduced H. pylori-induced nuclear β-catenin phosphorylation and COX-2 levels in GES-1 cells. Among 9 significantly altered miRNAs, miR-185 was the only miRNA targeting the Wnt/β-catenin signaling pathway. H. pylori increased miR-185 expression and upregulated COX-2 carcinogenesis through the Wnt/β-catenin pathway, but not the JAK2/STAT3 pathway. B. lactis ameliorated H. pylori-induced miR-185 expression and nuclear β-catenin/COX-2 signaling in a dose-dependent manner. In the 6-month probiotic-treated patients had a significantly higher IM regression rate than controls (intention-to-treat: 37.5 vs 11.5%, OR: 4.60, 95% CI: 1.134-18.65, p = 0.025; per-protocol: 46.2 vs 17.6%, OR: 4.00, 95% CI: 0.923-17.33, p = 0.055). Patients without IM regression had significantly higher miR-185 levels in follow-up biopsies (p < 0.01).

Conclusions: Pretreatment with B. lactis ameliorated the H. pylori-induced COX-2 carcinogenesis pathway by reducing miR-185 expression, which targets Wnt/β-catenin signaling. (ClinicalTrials.gov, NCT05544396).

Keywords: H. pylori; COX-2; Carcinogenesis; MiR-185; Probiotics; Wnt/β-catenin.

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Conflict of interest statement

Declarations. Ethics approval and consent to participate: The study were approved by the Institutional Review Board, National Cheng Kung University Hospital, Taiwan. (A-BR-106-085). Consent for publication: Not applicable. The name of Randomized Clinical Trials registry and the registration number: Name: Study on the Probiotics Regulating miRNA in H. pylori-induced Wnt/β-catenin Gastric Carcinogenesis. ClinicalTrials.gov ID: NCT05544396. Date of registered this trial: March 20, 2024. https://clinicaltrials.gov/study/NCT05544396 . Competing interests: The authors declare no competing interests.

Figures

Fig. 1
Fig. 1
The levels of IL-6, IL-11, and IL-8 after H. pylori infection in both cancerous (AGS, MKN45, and SUN1) and primary GES-1 cells. The cells were seeded in one layer on the plates. H. pylori and PBS (control) were co-incubated with the cells for 24 h, and the supernatant was collected and evaluated for IL-6 (a and d), IL-11 (b and e), and IL-8 (c) levels by ELISA
Fig. 2
Fig. 2
H. pylori infection induced β-catenin/COX-2 expressions in GES-1 and AGS cells. A The nuclear and cytoplasmic protein levels (fold) of GES-1 cells incubated with H. pylori or PBS (control) for 24 h. B The nuclear and cytoplasmic protein levels (fold) of AGS cells incubated with H. pylori or PBS (control) for 24 h. C The protein levels (fold) of cytoplasmic JAK2 and nuclear p-STAT3 in both GES-1 and AGS cells after co-culture with H. pylori or PBS for 24 h. D The protein levels (fold) of Wnt3α and p-GSK3β of GES-1 and AGS cells co-cultured with H. pylori or PBS for 24 h
Fig. 3
Fig. 3
Pretreatment with probiotics (A. L. acidophilus, B. B. lactis, A + B. mixture of A and B) reduced nuclear phosphorylated β-catenin and COX-2 which were upregulated by H. pylori. The cells were pretreated with probiotics (MOI 100) for 4 h. The cells were washed with PBS 3 times and then co-cultured with H. pylori for 24 h. The nuclear protein was harvested for A COX-2 and β-catenin and B Wnt3α/p-GSK3β and JAK2/nuclear p-STAT3
Fig. 4
Fig. 4
The miR-185 transcription level in GES-1 and miR-185-overexpressing GES-1 (miR-185 mimic) cells after H. pylori infection. The relative quantity of miR-185 was measured by real-time PCR at A 4 h and B 24 h. The protein levels of nuclear p-β-catenin C, p-STAT3 E, and COX-2 D in GES-1 and miR-185-overexpressing GES-1 (miR-185 mimic) cells after H. pylori infection
Fig. 5
Fig. 5
Effect of pretreatment with. probiotics (A. L. acidophilus, B. B. lactis, A + B. mixture of A and B) on the relative quantity of miR-185 in A GES-1 and B miR-185-overexpressing GES-1 (miR-185 mimic) cells after H. pylori infection. The pretreatment with differential doses of B. lactis (B50: MOI 50, B100: MOI 100, and B200: MOI200) on the relative quantity of miR-185 in C GES-1 and D miR-185 mimic cells and nuclear p-β-catenin and COX-2 in E GES-1 and F miR-185 mimic cells after H. pylori infection
Fig. 6
Fig. 6
The mean relative transcription levels of miR-185 in antrum biopsies between A IM and non-IM patients without H. pylori infection. B The mean miR-185 expression in antrum biopsies of H. pylori-infected patients with IM who completed a 6-month course of probiotic treatment 1 year after eradication therapy. C The mean miR-185 transcription levels stratified by regression and non-regression of IM at the second EGD. EGD, esophagogastroduodenoscopy; IM, intestinal metaplasia
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