c-Jun N-terminal kinase 2 (JNK2) depletion prevents gastrointestinal acute radiation syndrome (GI-ARS) induced epithelial barrier dysfunction and gut microbiota dysbiosis

Background: Exposure to high-energy ionizing radiation in a mass casualty event involving nuclear radiation is a major threat. Radiation exposure beyond 6 Grays (Gy) results in Gastrointestinal Acute Radiation Syndrome (GI-ARS), in which death is imminent without interventions, yet no such approved treatment is available. This is due to a lack of molecular-level understanding of GI-ARS pathogenesis in the gut, which involves Reactive Oxygen Species (ROS) production, intestinal epithelial tight junction (TJ) disruption, barrier dysfunction, microbiota dysbiosis, and endotoxemia. ROS-activated c-Jun N-terminal kinase-2 (JNK2) plays a role in multiple tissue injuries. Here, we investigated the role of JNK2 in radiation-induced intestinal injury. Methods: Wild-type (WT) and JNK2 knockout (KO) mice were subjected to total body irradiation (TBI; 9 Gy) or partial body irradiation with 5% bone marrow shielded (PBI). At 24 and 48h post-irradiation, we assessed intestinal mucosal permeability to FITC-inulin, TJ integrity by immuno-fluorescence confocal microscopy, mucosal injury by histopathology, inflammation by cytokine expression, and endotoxemia by plasma lipopolysaccharide (LPS). Paneth cell function was assessed by RT-PCR, confocal microscopy, and proteomics of antimicrobial proteins (AMP). The gut microbiota was analyzed using 16S rRNA sequencing and metagenomics. The role of JNK2 in radiation-induced TJ disruption was determined in vitro using Caco-2 cell monolayers and mouse enteroids (WT & KO), and assessing epithelial permeability, actin cytoskeleton remodeling, and TJ integrity. Results: TBI and PBI caused intestinal epithelial TJ disruption, barrier dysfunction, mucosal damage, and inflammation in WT but not KO mice. TBI and PBI-induced elevation of plasma LPS was absent in KO mice. Radiation reduced Paneth cell gene expression in WT but not KO mouse ileum, as confirmed by reduced DEFA4 and Lyz1 expression by immunofluorescence staining. The intestinal luminal proteome showed distinct radiation-, strain-, and sex-dependent differences. Radiation depleted AMPs, such as defensins, in WT but not KO mice. Metagenomic analyses revealed distinct taxonomic compositions of the gut microbiota in sham and irradiated WT and KO mice. Radiation increased the abundance of pathogenic bacteria, such as E. coli, and reduced the abundance of probiotics, such as Akkermansia and Lactobacillus, in WT but not KO mice. SP600125, a pan-JNK isoform inhibitor, blocked radiation-induced epithelial permeability and TJ disruption in Caco-2 cell monolayers. Radiation caused actin cytoskeleton remodeling and TJ disruption in WT but not KO enteroids. Conclusion: These findings indicate that JNK2 activation plays a key role in radiation-induced gut microbiota dysbiosis and epithelial TJ disruption, thereby serving as a key mechanism in GI-ARS pathogenesis. These results suggest that JNK2 inhibitors can be explored as potential therapeutics for GI-ARS. Supported by NIH/NIAID - U01AI170019, U01AI172991. 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.