Bacillus coagulans in Combination with Chitooligosaccharides Regulates Gut Microbiota and Ameliorates the DSS-Induced Colitis in Mice

Abstract

Ulcerative colitis (UC) are chronic inflammatory disorders, which may be caused by intestinal barrier dysfunction, immune system disorders and intestinal microbiota dysbiosis. Synbiotic, the combination of probiotics and prebiotics, is thought to be a pragmatic approach in mitigating inflammation in UC. Bacillus coagulans has been recognized as a potential probiotic for treating intestinal diseases because of its favorable industrial and probiotic properties, including sporulation and lactic acid production. In this study, we evaluated the treatment effects of the B. coagulans FCYS01 spores with or without the chitooligosaccharides (COSs) on UC generated using dextran sulfate sodium (DSS) in mice. Supplementation of B. coagulans spores, prebiotic COSs or the synbiotic (the spores + COSs) had a significant positive effect on DSS-induced UC. The disease activity index and histological damage score were significantly reduced after these supplementations. Compared to DSS group, these supplementations also significantly modulated the cytokines IL-4, IL-6, IL-8, IL-10, and C-reactive protein (CRP) levels and significantly maintained expressions of tight junction proteins and mucin protein and promotes recovery of the intestinal barrier. In addition, these supplementations regulate the composition of gut microbiota and improve the production of short-chain fatty acids (SCFAs), through enrichment of SCFA-producing bacteria, such as Akkermansia and Ruminococcus species. In summary, the synbiotic ameliorated the overall inflammatory status of the experimental UC model and showed a better treatment effect than B. coagulans or COSs did alone as revealed by the markers such as, colon length, IL-4 and Occludin levels. IMPORTANCE Probiotic and prebiotic are believed to be useful in alleviating the inflammatory, thereby resolving or preventing the severity of UC. Spore-forming bacteria Bacillus coagulans show advantages of stability and probiotic effects, being suggested as the important probiotics for UC treatment. Here, we demonstrate that administration of B. coagulans spores, chitooligosaccharides (COSs), or the synbiotic attenuates DSS-induced colitis and significantly correlates with altered gut immune responses. The treatment effect of the synbiotic is inferred to be relied on the enrichment of probiotic bacteria, such as Akkermansia and Ruminococcaceae species, which are reported to be crucial important for gut health. Our findings facilitate the development of therapeutic and preventive strategies for UC using spore-forming lactic acid bacteria in combination with COSs.

Keywords: Bacillus coagulans; chitooligosaccharides; microbiota; short-chain fatty acids; synbiotic.

FIG 1

Animal experiment design of supplementation with B. coagulans, COSs, and synbiotic on DSS-induced colitis mice. Colitis was induced by administration of 2.5% DSS in drinking water for the first 7 days. B. coagulans spores (B. co), COSs, or the synbiotic (spores + COSs) were supplemented in the regular diet for the last 7 days. Ctrl: healthy control group fed with regular diet.

FIG 2

Treatment effect of B. coagulans, COSs, and synbiotic on DSS-induced colitis mice. (A) body weight change, (B) disease activity index (DAI), (C) macroscopic images of the colons, (D) colons length, (E) spleen weight/body weight ratio. Data are shown as means ± SEM (n = 10 per group). Data with different superscript letters (a, b and c) are significantly different (P < 0.05) according to two-way ANOVA (A and B) and one-way ANOVA (D and E) followed by Tukey’s test. Ctrl: healthy control; DSS: DSS-treated group; B. co, COSs, and Synbiotic: supplementation of B. coagulans spores, COSs, or spores + COSs, respectively.

FIG 3

Treatment effect of B. coagulans, COSs, and synbiotic on DSS-induced colon injury and inflammation. (A) Microscopic images of proximal colonic tissues stained with H&E. Scale bars represent 100 μm. The right images are the enlarged part of the left images. Arrows indicate the damage of colon. (B) Histological score calculated after microscopic analyses of the colons. (C) Myeloperoxidase (MPO) activity in proximal colonic tissues. Data were shown as means ± SEM. Data with different superscript letters (a and b) were significantly different (P < 0.05) according to one-way ANOVA followed by Tukey’s test. Ctrl: healthy control; DSS: DSS-treated group; B. co, COSs, and Synbiotic: supplementation of B. coagulans spores, COSs, or spores + COSs, respectively.

FIG 4

Effect of B. coagulans, COSs, and synbiotic on regulation of immune markers. Protein levels of cytokines, including (A) IL-6, (B) IL-8, (C) IL-1β, (D) IFN-γ, (E) TNF-α, (F) CRP, (G) IL-4, and IL-10 (H) of proximal colon tissues, were analyzed by ELISA. Data are shown as means ± SEM (n = 5 per group). Data with different superscript letters (a, b and c) are significantly different (P < 0.05) according to one-way ANOVA followed by Tukey’s test. Ctrl: healthy control; DSS: DSS-treated group; B. co, COSs and Synbiotic: supplementation of B. coagulans spores, COSs, or spores + COSs, respectively.

FIG 5

B. coagulans, COSs and synbiotic regulated the composition of gut microbiota. (A-C) Alpha-diversity indices. (D) Principal-component analysis of Bray-Curtis distance (the difference in PC1 and PC2 was 21.8% and 14.4%). (E and F) The relative abundance of fecal bacterial phylum (E), and family (F). (G) Analysis of differences in the microbial taxa by LEfSe in different groups. (H) The relative abundance of B. coagulans. Concentrations of fecal acetate (I), propionate (J), and butyrate (K). Ctrl: healthy control; DSS: DSS-treated group; B. co, COSs and Synbiotic: supplementation of B. coagulans spores, COSs or spores + COSs, respectively.

FIG 6

Effect of B. coagulans, COSs, and synbiotic on the expression of tight junction proteins. (A) Microscopic images of proximal colonic tissues stained with immunohistochemical of Occludin, Claudin, Zo-1, Muc2. Scale bars represent 100 μm. Arrows indicate the IHC positivity. (B) Mean optical densities with Image-pro plus soft of respective group is illustrated in the graph. Data are shown as means ± SEM. Data with different superscript letters are significantly different (P < 0.05) according to one-way ANOVA followed by Tukey’s test. Ctrl: healthy control; DSS: DSS-treated group; B. co, COSs, and Synbiotic: supplementation of B. coagulans spores, COSs, or spores + COSs, respectively.