Florsheim, Esther Borges

Cholera toxin (CTx) is a potent enterotoxin and widely used mucosal adjuvant, yet the acute host response to CTx itself, in the absence of a co-administered antigen, remains vague. In this study, we integrated physiological and metabolic phenotyping with intestinal transcriptomics, epithelial imaging, immune profiling, hepatic gene-expression analysis, and paired microbiome profiling to characterize the response to a single oral CTx exposure in mice. Within 24 h, CTx remodeled immune-cell composition in both epithelial and lamina propria compartments, with responses shaped by sex and genetic background. Recurrent changes included increased epithelial group 3 innate lymphoid cells, reduced lamina propria RORγt+ regulatory T cells, and decreased epithelial ILC1/NK cells in males of both strains. CTx also increased villus DCLK1+ tuft cells and reduced MUC2 immunoreactivity. Jejunal transcriptomics revealed induction of inflammatory defense, glycolytic, autophagy, and lipid-utilization programs together with suppression of proliferative and mitochondrial programs, consistent with coordinated stress adaptation. These local changes were accompanied by rapid restructuring of the fecal microbial community and by systemic hypometabolism, despite markedly different hypothermic responses across strains. CTx also elicited a selective hepatic stress and metabolic program, including conserved suppression of Cyp7a1 across sexes and strains. Together, these findings show that CTx alone rapidly reorganizes the intestinal immune, epithelial, metabolic, and microbial environment while suppressing whole-body metabolism. This work provides an integrated framework for understanding the acute host response to CTx and for interpreting its widespread use as a mucosal adjuvant.

ABSTRACT Maternal obesity is associated with increased risk of sporadic colorectal cancer (CRC) in offspring, suggesting that early-life environmental exposures durably shape disease susceptibility. Intestinal stem cells (ISCs), long-lived drivers of epithelial renewal and tumor initiation, are well poised to mediate this effect; however, how maternal obesity influences ISC programming during development remains poorly understood. Using mouse models of diet-induced obesity, we show that exposure to a maternal high-fat Western diet (mHFD) during pre- and postnatal development stably programs colonic ISCs. Offspring exhibit increased ISC proliferation, enhanced self-renewal, a hypermetabolic state, and altered epithelial lineage composition that persists into adulthood despite dietary normalization. These changes are accompanied by increased tumor burden following loss of Apc heterozygosity. Mechanistically, we identify the pro-inflammatory cytokine IL-17A as a key extrinsic driver and PPARd/a nuclear receptors as intrinsic mediators of the mHFD phenotype, revealing an immune-epithelial axis that programs ISC function during early life. Together, our findings demonstrate that maternal metabolic environments durably enhance stem cell fitness, providing a mechanistic link between developmental exposure and adult disease risk.

Abstract Description Cholera toxin (CT), an exotoxin produced by Vibrio cholerae, causes severe diarrheal disease and contributes to the third leading cause of death in children under 5. Beyond its pathogenicity, CT possesses potent mucosal adjuvant properties, inducing type 2 immunity in the gastrointestinal tract. However, the immune response to acute oral CT exposure remains poorly characterized. To address this, we intragastrically administered a single nonlethal dose of CT to mice and assessed physiological and immune responses over 24 hours. In C57BL/6 mice, CT induced severe hypothermia, while BALB/c mice exhibited milder effects, with males of both strains showing heightened sensitivity. All treated animals developed diarrhea, and C57BL/6 mice experienced intestinal edema lasting up to 24 hours. Transcriptomic analysis of the small intestine revealed acute upregulation of inflammatory mediators such as IL-1β and IL-33. Immune profiling by full-spectrum flow cytometry showed CT-induced changes in the lamina propria compartment, including increased ILC3, reduced Treg RORγT+ and CD11b+CD103+ dendritic cells. In the epithelial layer, CT slightly elevated neutrophils, eosinophils, monocytes, and ILC3 while reducing CD3+ T lymphocytes and ILC1/ NK cells. These findings emphasize the broader action of CT in pathogenesis and immunity, shedding light on how immune sensing of bacterial toxins can drive protective defenses and adjuvant activity. Funding Sources Supported by Hypothesis Fund Topic Categories Mucosal and Regional Immunology (MUC)