Fecal microbiota transplantation alleviates DSS-induced colitis: increased fecal butyrate, reduced colonic p65 phosphorylation, and altered Th17/Treg ratios in the spleen and mesenteric lymph nodes.

BACKGROUND: Ulcerative colitis (UC) development and progression are associated with intestinal dysbiosis, altered short-chain fatty acid (SCFA) metabolism, and immune dysregulation. Although fecal microbiota transplantation (FMT) has therapeutic potential, its key effector metabolites and regulatory pathways remain unclear. METHODS: Male BALB/c mice with 2.5% dextran sulfate sodium (DSS)-induced colitis received FMT, 5-aminosalicylic acid (5-ASA), sodium butyrate (NaB), or pyrrolidine dithiocarbamate (PDTC). Colitis severity was evaluated using body weight, disease activity index, colon length, and hematoxylin and eosin staining. Cytokines in colonic tissue and serum were quantified by enzyme-linked immunosorbent assay. Colonic p-p65/p65 and p-IκBα/IκBα ratios and tight-junction proteins were measured by Western blotting. Th17/Treg proportions in the spleen and mesenteric lymph nodes were assessed by flow cytometry. Gut microbiota composition and fecal SCFA profiles were examined by 16S rRNA sequencing and targeted metabolomics, respectively. RESULTS: Compared with DSS, FMT markedly attenuated weight loss, disease activity, colon shortening, and histopathological injury. In colonic tissue, FMT reduced IL-17A and IL-21 and increased TGF-β1. In serum, it increased IL-10 and TGF-β1 and reduced IL-21. FMT increased colonic Claudin-1, Occludin, and zonula occludens-1 expression and reduced the colonic p-p65/p65 ratio. It also reduced Th17 proportions and increased Treg proportions in the spleen and mesenteric lymph nodes. Microbiota analysis indicated improved diversity and reshaping of DSS-induced dysbiosis, including recovery of Firmicutes at the phylum level and Lachnospiraceae at the family level. FMT was associated with higher endpoint fecal acetate and butyrate concentrations than DSS, with butyrate also higher than in the 5-ASA group, supporting its evaluation as an FMT-associated readout. In a separate experiment, NaB, PDTC, and combined NaB/PDTC treatments were each associated with changes in selected outcomes relative to DSS. CONCLUSION: FMT alleviates DSS-induced colitis and is associated with microbiota shifts, increased endpoint fecal butyrate concentration, and a reduced colonic p-p65/p65 ratio. It is also associated with increased colonic tight-junction protein expression and altered Th17/Treg proportions in the spleen and mesenteric lymph nodes. These findings represent parallel associations and do not establish causal relationships among the measured outcomes.