Tritrichomonas muris sensitizes the intestinal epithelium to doxorubicin-induced apoptosis

Doxorubicin (DXR) is a widely used chemotherapy drug that can induce severe intestinal mucositis. Although the influence of gut bacteria on DXR-induced damage has been documented, the role of eukaryotic commensals remains unexplored. We discovered Tritrichomonas muris (Tmu) in one of our mouse colonies exhibiting abnormal tuft cell hyperplasia, prompting an investigation into its impact on DXR-induced intestinal injury. Mice from Tmu-colonized and Tmu-excluded facilities were injected with DXR. Tissue morphology and gene expression were evaluated at acute injury (6 h) and regenerative (72 h and 120 h) phases. Changes to crypt and villus morphology were more subtle than previously reported and region-specific, with significantly shorter jejunal villi in Tmu⁺ mice at 72 h post-DXR compared with Tmu^- controls. Most notably, we observed elevated rates of DXR-induced apoptosis, measured by cleaved caspase 3 (CC3) staining, in Tmu⁺ intestinal crypts at 6 h post-DXR. Tmu⁺ mice also exhibited reduced expression of active intestinal stem cell (aISC) marker Lgr5 and facultative ISC (fISC) marker Ly6a at 6 h post-DXR compared with Tmu^- controls. Tmu, but not DXR, was associated with increased inflammation and expression of type 2 cytokines IL-5 and IL-13. Tmu⁺ mice also exhibited a decreased fecal abundance of Lactobacillus, which promotes gut barrier integrity, and reduced claudin expression, indicating potential barrier dysfunction that could explain the increase in DXR-induced apoptosis. These findings highlight the significant influence of commensal microbiota, particularly eukaryotic organisms like Tmu, on intestinal biology and response to chemotherapy, underscoring the complexity of gut microbiota interactions in drug-induced mucositis.NEW & NOTEWORTHY Our study found that the eukaryotic commensal Tritrichomonas muris (Tmu) significantly increases DXR-induced intestinal apoptosis in mice. Tmu also reduces Lgr5 expression post-DXR injury and elevates inflammation and type 2 cytokine expression in the absence of injury. 16S sequencing identifies decreased abundance of protective Lactobacillus in Tmu colonized mice, as well as decreased expression of barrier-forming claudins, which may explain increased apoptosis. These findings emphasize the complex role of microbiota in drug-induced intestinal damage.