Atopic dermatitis (AD), a chronic inflammatory skin disease prevalent in Western children, is characterized by an IgE-mediated response and a mixed type 1 and type 2 immune response. While genetic factors contribute, increasing prevalence suggests significant environmental influence. The gut microbiota's role in AD pathogenesis is increasingly recognized, with potential for microbiota modulation to alleviate inflammation. Previous studies showed a strong correlation between gut microbiota composition and inflammatory skin response in oxazolone-induced AD in mice, with a transferable high- and low-responding phenotype. This study aimed to investigate the effect of a prebiotic, XOS, on gut microbiota composition and the severity of oxazolone-induced AD in BALB/c mice, testing the hypothesis that prebiotic intervention would alter gut microbiota composition and consequently influence the severity of the disease. Successful modulation of the gut microbiota could offer a novel therapeutic approach for AD.

Existing literature highlights the complex interplay between the gut microbiota and the development of atopic dermatitis. Studies have demonstrated a correlation between specific gut microbial compositions and the severity of AD, suggesting that modulation of the gut microbiota could be a promising therapeutic strategy. Previous research using fructooligosaccharides (FOS) in murine models has shown a reduction in AD severity. However, the mechanisms underlying these effects are not fully understood. The current understanding is that the gut microbiota influences the immune system, potentially through the production of short-chain fatty acids and other metabolites, which can modulate the inflammatory response. Furthermore, the transferability of AD susceptibility along with the gut microbiota in germ-free mice highlights the critical role of the intestinal environment in shaping immune responses.

Thirty-two four-week-old female BALB/cJBomTac mice were randomly assigned to two groups: a control group fed a standard diet and an XOS group fed a diet supplemented with 47.5 g XOS/kg. Atopic dermatitis was induced by sensitizing and repeatedly challenging the mice with oxazolone solution (0.8% for sensitization, 0.4% for challenges). Ear thickness and clinical skin inflammation scores were assessed. Histological analysis of ear biopsies evaluated epidermal spongiosis, dermal hyperplasia, mast cell infiltration, and fibroplasia. Serum IgE levels were measured using ELISA, and ear tissue cytokine levels were assessed using a multiplex assay. Gut microbiota composition was determined using 16S rRNA gene amplicon sequencing before and after oxazolone treatment. Statistical analysis included unpaired t-tests, Mann-Whitney U-tests, ANOVA, and chi-square tests, as appropriate. Specific details on the methodology used for each assessment such as histological scoring, cytokine measurement, and sequencing are given in detail within the paper.

Oxazolone treatment successfully induced atopic dermatitis in all mice, increasing ear thickness and causing clinical symptoms. The XOS-supplemented diet significantly reduced the number of high responders to oxazolone-induced dermatitis, both clinically and histopathologically. While serum IgE levels and ear tissue cytokine levels did not differ significantly between groups, a large inter-individual variation was observed. Gut microbiota analysis revealed that the XOS-fed mice clustered separately from the control mice based on unweighted PCoA, mainly driven by higher *Prevotella* spp. abundance and decreased *Lactobacillus reuteri*. Importantly, in the control group, *Prevotella* spp. abundance correlated negatively with clinical ear inflammation score, serum IgE levels, and various cytokine levels. In contrast, in the XOS group, no significant correlations were found.

The study's findings demonstrate that dietary XOS can effectively reduce the severity of oxazolone-induced AD in mice. This effect is likely mediated through alterations in the gut microbiota, particularly an increase in *Prevotella* spp. This contrasts with previous studies using FOS, where bifidogenic effects were implicated in the reduction of AD severity. This study, however, focuses on non-bifidogenic effects of XOS in a specific population of mice not harboring *Bifidobacterium* spp, and therefore reveals a distinct mechanism. The negative correlation between *Prevotella* abundance and inflammation markers in the control group strongly supports the protective role of *Prevotella* in AD. The lack of significant differences in immunological parameters between groups may be due to the high inter-individual variation in the control group, potentially masking subtle effects of XOS. The study emphasizes the importance of considering individual gut microbial profiles when evaluating the impact of prebiotics and highlights the need for more research to unravel the complex interactions between specific gut bacteria and immune responses in AD.

This study demonstrates that dietary XOS can mitigate oxazolone-induced atopic dermatitis in mice, specifically by increasing the abundance of *Prevotella* spp. in the gut. This indicates a potential therapeutic role for *Prevotella* spp. in AD, possibly through modulation of the inflammatory response. Future studies should investigate the underlying mechanisms of *Prevotella*'s protective effects and evaluate the potential of *Prevotella*-directed therapies for AD. Further research is needed to explore the impact of prebiotics on AD across different mouse strains and to clarify the role of *Prevotella* in AD.

The study used a murine model of AD, which may not fully recapitulate the complexity of human AD. The large inter-individual variation observed in both groups could have masked more subtle effects of the XOS diet. The study did not investigate the mechanisms through which *Prevotella* spp. might exert its protective effect, which would be a crucial area for future research. Moreover, the absence of *Bifidobacterium* spp in the population tested limits the ability to compare the effects of XOS in those species containing *Bifidobacterium* spp.