Chronic Psychological Stress Disrupted the Composition of the Murine Colonic Microbiota and Accelerated a Murine Model of Inflammatory Bowel Disease

<div><p>The effect of psychological stress on the gastrointestinal microbiota is widely recognized. Chronic psychological stress may be associated with increased disease activity in inflammatory bowel disease, but the relationships among psychological stress, the gastrointestinal microbiota, and the severity of colitis is not yet fully understood. Here, we examined the impact of 12-week repeated water-avoidance stress on the microbiota of two inbred strains of T cell receptor alpha chain gene knockout mouse (background, BALB/c and C57BL/6) by means of next-generation sequencing of bacterial 16S rRNA genes. In both mouse strains, knockout of the T cell receptor alpha chain gene caused a loss of gastrointestinal microbial diversity and stability. Chronic exposure to repeated water-avoidance stress markedly altered the composition of the colonic microbiota of C57BL/6 mice, but not of BALB/c mice. In C57BL/6 mice, the relative abundance of genus <i>Clostridium</i>, some members of which produce the toxin phospholipase C, was increased, which was weakly positively associated with colitis severity, suggesting that expansion of specific populations of indigenous pathogens may be involved in the exacerbation of colitis. However, we also found that colitis was not exacerbated in mice with a relatively diverse microbiota even if their colonic microbiota contained an expanded phospholipase C-producing <i>Clostridium</i> population. Exposure to chronic stress also altered the concentration of free immunoglobulin A in colonic contents, which may be related to both the loss of bacterial diversity in the colonic microbiota and the severity of the colitis exacerbation. Together, these results suggest that long-term exposure to psychological stress induces dysbiosis in the immunodeficient mouse in a strain-specific manner and also that alteration of microbial diversity, which may be related to an altered pattern of immunoglobulin secretion in the gastrointestinal tract, might play a crucial role in the development of chronic stress-induced colitis.</p></div

Composition of the colonic microbiota of BALB/c <i>Tcra</i>^−/− mice after exposure to 12-week repeated water-avoidance stress.

<p>(a) Bacterial taxonomic profiling of the colonic microbiota at the genus level. (b) Bacterial richness as represented by observed species rarefaction measured based on 1 to 75 000 sequences. Mean ± standard deviation. (c) Average unweighted UniFrac distance. (d) Principal-coordinates analysis plot based on the unweighted UniFrac distance. HET, <i>Tcra</i>^−/+ mice not exposed to repeated water-avoidance stress (rWAS); CON, <i>Tcra</i>^{<i>−/−</i>} mice not exposed to rWAS; LFW, <i>Tcra</i>^−/− mice exposed to low-frequency (1 day) rWAS; HFW, <i>Tcra</i>^−/− mice exposed to high-frequency (5 days) rWAS. PC1 and PC2 are the first two principal coordinates. * <i>P</i> < 0.05; n.s., not significant (<i>P</i> > 0.05).</p

Correlation between histological score and concentration of <i>Clostridium perfringens</i> or phylogenetic diversity.

<p>(a) <i>C</i>. <i>perfringens</i> was quantified in colonic contents by means of quantitative PCR. The concentration of <i>C</i>. <i>perfringens</i> was positively correlated with histological score; however, this was not statistically significant (<i>P</i> = 0.17). (b) Phylogenetic diversity was calculated based on 75 000 reads per sample. The phylogenetic diversity of colonic microbiota was significantly negatively correlated with the histological score (<i>P</i> < 0.001). RES, <i>Tcra</i>^−/− mice resistant to rWAS-induced colitis; SEN, <i>Tcra</i>^−/− mice sensitive to rWAS-induced colitis.</p

Detection ratio and mean abundance of operational taxonomic units (OTUs) significantly increased in <i>Tcra</i>^−/− mice exposed to repeated water-avoidance stress.

<p>Detection ratio and mean abundance of operational taxonomic units (OTUs) significantly increased in <i>Tcra</i>^−/− mice exposed to repeated water-avoidance stress.</p

Histological score criteria.

<p>Histological score criteria.</p

Concentrations of luminal IgA and IgG in <i>Tcra</i>^−/− mice exposed to repeated water-avoidance stress (rWAS).

<p>Concentrations of total IgA and IgG in the colonic content of each mouse were measured by using a sandwich enzyme-linked immunosorbent assay. Data are presented as mean + standard deviation. CON, <i>Tcra</i>^−/− mice not exposed to rWAS; LFW, <i>Tcra</i>^−/− mice exposed to low-frequency (1 day) rWAS; HFW, <i>Tcra</i>^−/− mice exposed to high-frequency (5 days) rWAS; RES, <i>Tcra</i>^−/− mice resistant to rWAS-induced colitis; SEN, <i>Tcra</i>^−/− mice sensitive to rWAS-induced colitis.</p

Relative abundances of operational taxonomic units (OTUs).

<p>Heatmap showing the relative abundances of 70 OTUs and their taxonomic assignment acquired by using the Ribosomal Database Project classifier. Taxa names in brackets are annotations supplied by the Greengenes database and are not officially accepted by the Society for General Microbiology. HET, <i>Tcra</i>^−/+ mice not exposed to repeated water-avoidance stress (rWAS); CON, <i>Tcra</i>^−/− mice not exposed to rWAS; RES, <i>Tcra</i>^−/− mice resistant to rWAS-induced colitis, SEN, <i>Tcra</i>^−/− mice sensitive to rWAS-induced colitis.</p

Disturbance in the Mucosa-Associated Commensal Bacteria Is Associated with the Exacerbation of Chronic Colitis by Repeated Psychological Stress; Is That the New Target of Probiotics?

<div><p>Psychological stress can exacerbate inflammatory bowel disease. However, the mechanisms underlying how psychological stress affects gut inflammation remain unclear. Here, we focused on the relationship between changes in the microbial community of mucosa-associated commensal bacteria (MACB) and mucosal immune responses induced by chronic psychological stress in a murine model of ulcerative colitis. Furthermore, we examined the effect of probiotic treatment on exacerbated colitis and MACB composition changes induced by chronic psychological stress. Repeated water avoidance stress (rWAS) in B6-<i>Tcra</i>^-/- mice severely exacerbated colitis, which was evaluated by both colorectal tissue weight and histological score of colitis. rWAS treatment increased mRNA expression of <i>UCN2</i> and <i>IFN-γ</i> in large intestinal lamina propria mononuclear cells (LI-LPMC). Interestingly, exacerbated colitis was associated with changes in the microbial community of MACB, specifically loss of bacterial species diversity and an increase in the component ratio of <i>Clostridium</i>, revealed by 16S rRNA gene amplicon analysis. Finally, the oral administration of a probiotic <i>Lactobacillus</i> strain was protective against the exacerbation of colitis and was associated with a change in the bacterial community of MACB in rWAS-exposed <i>Tcra</i>^-/- mice. Taken together, these results suggested that loss of species diversity in MACB might play a key role in exacerbated colitis induced by chronic psychological stress. In addition, probiotic treatment may be used as a tool to preserve the diversity of bacterial species in MACB and alleviate gut inflammation induced by psychological stress.</p></div

Detection ratio and mean abundance of operational taxonomic units (OTUs) changed in rWAS-exposed <i>Tcra</i>^-/- mice.

<p>Detection ratio and mean abundance of operational taxonomic units (OTUs) changed in rWAS-exposed <i>Tcra</i>^-/- mice.</p

Effect of LcS on the OTUs significantly changed in MACB of rWAS-exposed <i>Tcra</i>^−/− mice.

<p>Effect of LcS on the OTUs significantly changed in MACB of rWAS-exposed <i>Tcra</i>^−/− mice.</p