Gut microbiota isolated from Paneth cell TCF4-deficient mouse renders protection against radiation-induced intestinal injury

Background: Ionizing radiation exposure induces intestinal injury associated with epithelial tight junction (TJ) disruption, mucosal barrier dysfunction, microbiota dysbiosis, and endotoxemia. Our recent study showed that radiation-induced microbiota dysbiosis is associated with downregulation of Wnt signaling and Paneth cell dysfunction. To determine the role of Paneth cell Wnt signaling downregulation and the microbiota dysbiosis in radiation injury, we generated Paneth cell-specific TCF4 knockout (TCF4ΔPC) mice. In this study, we evaluated the effects of Paneth cell TCF4 deficiency and the gut microbiota isolated from TCF4ΔPC mice on radiation-induced intestinal injury. Methods: TCF4ΔPC (KO) and corresponding TCF4Flox (WT) mice were subjected to partial body irradiation with 5% bone marrow protection (IR group) or sham-treated (SHAM group). Gut microbiota was harvested from the colon of SHAM and IR-treated WT and KO mice, generating four different microbiota donors (WT-SHAM, WT-IR, KO-SHAM, KO-IR). Pseudo-germ-free (PGF) mice were created by antibiotic treatment of wild-type mice (Jackson Lab). PGF mice were transplanted with colonic microbiota from WT-SHAM, WT-IR, KO-SHAM, and KO-IR donors and subjected to PBI-BM5 or SHAM treatment. Intestinal injury was assessed at 48 hours after irradiation. Body weight changes, colon lengths, intestinal mucosal permeability to FITC-inulin, confocal immunofluorescence microscopy of epithelial TJ, intestinal histopathology, and mucosal cytokine/chemokine expression were evaluated. Results: KO mice showed a significant reduction of Paneth cell-specific gene expression, including α-defensins. PBI-BM5 caused more severe disruption of intestinal epithelial TJ, mucosal barrier dysfunction, mucosal inflammation, and endotoxemia in KO mice compared with WT mice. In PGF mice that received WT-SHAM or KO-SHAM microbiota, radiation reduced body weight and colon length; there was no significant difference in the radiation effects between WT-SHAM- and KO-SHAM-transplanted mice. Similar results were obtained when PGF mice received WT-IR and KO-IR transplants. However, the radiation-induced change in body weight was significantly attenuated in mice that received KO-IR transplant, suggesting protection by the KO-IR microbiota. Mucosal permeability in the ileum and colon was decreased considerably by radiation in PGF mice that received KO-SHAM, but not WT-SHAM transplants, again suggesting protection by the KO-SHAM microbiota. Immunofluorescence confocal microscopy of TJ proteins also revealed radioprotective effects by KO-SHAM and KO-IR microbiota. WT-SHAM microbiota showed no apparent effect on TJ integrity in the ileum of SHAM or IR-treated PGF mice. But WT-IR microbiota by itself disrupted TJ in the ileum. On the other hand, KO-IR microbiota did not affect TJ integrity in SHAM-treated PGF mice; however, it attenuated radiation-induced TJ disruption, indicating that KO-IR microbiota is protective against radiation injury. The protective effects of KO-IR microbiota were further supported by histopathology and mucosal cytokine/chemokine expression. Conclusion: This study shows that Paneth cell TCF4 deletion and alterations in downstream Paneth cell functions modify the gut microbiota composition, which is protective against radiation injury. Paneth cell dysfunction due to TCF4 deficiency may have triggered a mucosal defense response, leading to a protective microbiota composition Funding Support: NIH/NIAID- U01-AI170019, U01-AI172991 This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.