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. 2023 Sep 30;28(3):93-105.
doi: 10.15430/JCP.2023.28.3.93.

The Protective Effect of Roseburia faecis Against Repeated Water Avoidance Stress-induced Irritable Bowel Syndrome in a Wister Rat Model

Soo In Choi  1 , Nayoung Kim  1   2   3 , Ryoung Hee Nam  1 , Jae Young Jang  1   3 , Eun Hye Kim  1 , SungChan Ha  1 , Kisung Kang  4 , Wonseok Lee  4 , HyeLim Choi  5 , Yeon-Ran Kim  5 , Yeong-Jae Seok  5 , Cheol Min Shin  1 , Dong Ho Lee  1   2
Affiliations

The Protective Effect of Roseburia faecis Against Repeated Water Avoidance Stress-induced Irritable Bowel Syndrome in a Wister Rat Model

Soo In Choi et al. J Cancer Prev. .

Erratum in

Abstract

Roseburia faecis, a butyrate-producing, gram-positive anaerobic bacterium, was evaluated for its usefulness against repeated water avoidance stress (WAS)-induced irritable bowel syndrome (IBS) in a rat model, and the underlying mechanism was explored. We divided the subjects into three groups: one without stress exposure, another subjected to daily 1-hour WAS for 10 days, and a third exposed to the same WAS regimen while also receiving two different R. faecis strains (BBH024 or R22-12-24) via oral gavage for the same 10-day duration. Fecal pellet output (FPO), a toluidine blue assay for mast cell infiltration, and fecal microbiota analyses were conducted using 16S rRNA metagenomic sequencing. Predictive functional profiling of microbial communities in metabolism was also conducted. FPO and colonic mucosal mast cell counts were significantly higher in the WAS group than in the control group (male, P = 0.004; female, P = 0.027). The administration of both BBH024 (male, P = 0.015; female, P = 0.022) and R22-12-24 (male, P = 0.003; female, P = 0.040) significantly reduced FPO. Submucosal mast cell infiltration in the colon showed a similar pattern in males. In case of fecal microbiota, the WAS with R. faecis group showed increased abundance of the Roseburia genus compared to WAS alone. Moreover, the expression of a gene encoding a D-methionine transport system substrate-binding protein was significantly elevated in the WAS with R. faecis group compared to that in the WAS (male, P = 0.028; female, P = 0.025) group. These results indicate that R. faecis is a useful probiotic for treating IBS and colonic microinflammation.

Keywords: Irritable bowel syndrome; Probiotics; Rats; Roseburia faecis.

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Conflict of interest statement

CONFLICTS OF INTEREST No potential conflicts of interest were disclosed.

Figures

Figure 1
Figure 1. The fecal pellet output (FPO) and mast cell count from each repeated water avoidance stress (WAS) experiment with two different Roseburia faecis strains as probiotics candidates.
FPO data represent the 10-day sessions with weight calibration for male and female rats in the BBH024 experiments. The difference between male and female body weight was normalized by multiplying female FPO with the ratio of male body weight to female body weight. The cumulative FPO differed significantly between WAS-exposed groups and no-stress group in the later stages of BBH024 experiments (A) male and (B) female, as well as the R22-12-24 experiments (C) male and (D) female. The mean FPO of (E) BBH024 experiment and (F) R22-12-24 experiment indicated a significant reduction in FPO in WAS with probiotics group with statistical significance. The mucosal mast cells were counted from toluidine blue stained colonic tissue slides about each gavaged R. faecis strain, (G) BBH024, and (H) R22-12-24. The mast cell count in the colonic mucosa was divided into the area of the colonic mucosa. CON, control.
Figure 2
Figure 2. Colonic mucosal mRNA expression from each repeated water avoidance stress (WAS) experiment with two different Roseburia faecis strains as probiotics candidates.
Mucosal qPCR analysis of serine protease (PRSS) gene 1 (PRSS1: A, B) and 2 (PRSS2: C, D) revealed increased expression in the stress group compared to that in the control (CON) group in both males and females, although statistical significance was not reached. The mRNA expression of the inflammatory cytokine IL-1β (E, F) was significantly increased in the male stress group compared to that in the control group in R22-12-24 gavaged experiments. Furthermore, the inflammatory cytokine IL-6 (G, H) showed increased gene expression in the male stress group compared to that in the control group and showed a statistically significant decrease in expression in the BBH024 gavaged group.
Figure 3
Figure 3. Butyric acid and bile acid (BA) concentrations in fecal samples from repeated water avoidance stress (WAS) experiments with two different Roseburia faecis strains as probiotic candidates.
(A) The butyric acid concentration in feces after 10 days of WAS sessions. Unexpectedly, there was no statistical significance in both (A) BBH024 and (B) R22-12-24 strains and no correlation with previous cultured media results was noted. (C, D) The ratio of cholic acid (CA, primary BA) to deoxycholic acids (DCAs, secondary BA) in feces after 10 days of WAS sessions. In the case of R22-12-24, females showed significantly decreased levels of CA/DCA ratio (D, P = 0.025). (E, F) The ratio of chenodeoxycholic acid (CDCA, primary BA) to lithocholic acids (LCA, secondary BA) in feces after 10 days of WAS sessions. In the case of BBH024, females showed significantly decreased levels of CDCA/LCA ratio (E, P = 0.027). CON, control.
Figure 4
Figure 4. Fecal microbiome characteristics in rats exposed to repeated water avoidance stress (WAS) with Roseburia faecis R22-12-24 as a potential probiotic.
The α-diversity observed operational taxonomic units (OTUs) (A) and Shannon (B) were decreased in male stress-exposed groups while the female stress-exposed group showed increased α-diversity, but there was no statistically significant. The β-diversity was derived from generalized UniFrac distance and represented through principal coordinates analysis (PCoA) male (C, P = 0.037) and female (D, P = 0.011) separately. Both male and female stress groups exhibited significant separation from the control (CON) group. The taxonomic composition of each group in the phylum level (E) highlighted sex-based differences between males and females subjected to WAS. At the genus level, the orally gavaged genus Roseburia (F) showed slightly increased levels in R22-12-24 gavaged group compared to those in the stress group both in males (P = 0.045) and females (P = 0.054).
Figure 5
Figure 5. Predictive functional profiling of microbial communities, based on the Kyoto Encyclopedia of Genes and Genomes (KEGG) database, regarding metabolism related to carbohydrate, amino acid, lipid, nucleotide, and energy.
Panels (A) and (B) represent male (M) and female (F) data, respectively. Panel (C) illustrates module M00053, which is the module about deoxyribonucleotide biosynthesis (ADP/GDP/CDP/UDP→dATP/dGTP/dCTP/dUTP), the significantly different modules for both sexes are shown in the scatter graph. Panels (D) and (E) depict predictive functional profiling of microbial communities through the KEGG orthology database for males and females, respectively. Panel (F) focuses on K02073, which is the gene encoding D-methionine transport system substrate-binding protein, are shown in the scatter graph with both sexes. WAS, water avoidance stress; CON, control.
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FUNDING This work was supported by the Technology Innovation Program (20018499) funded by the Ministry of Trade, Industry & Energy (MOTIE, Korea).

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