Abstract
Inflammatory bowel disease (IBD) remains a global health challenge linked to intestinal barrier disruption, microbiota dysbiosis, and immune dysregulation, though the interplay of these mechanisms remains poorly defined. Here, we investigated the therapeutic potential of Lactobacillus johnsonii N5 in a murine dextran sulfate sodium (DSS)-induced colitis model. Prophylactic N5 administration alleviated colitis symptoms (weight loss, colon shortening), reduced fecal and serum lipocalin-2 levels, and suppressed colonic pro-inflammatory cytokines (IL-1β, IL-6). N5 preserved microbial diversity, enhanced mucus secretion, and reinforced mucosal barrier integrity, preventing colitis onset. Therapeutically, N5 attenuated disease progression by downregulating IL-1β, IL-6, IL-8 expression, restoring Lactobacillus populations, and suppressing Escherichia-Shigella expansion, thereby reducing bacterial translocation and systemic inflammation. N5 promoted Ki67+ epithelial proliferation, accelerating mucosal repair. Mechanistically, N5 targeted neutrophil-mediated gut-liver injury, suppressing coagulation pathways in colon-liver transcriptomes, reducing hepatic lesions, platelet aggregation, CD162+ neutrophil recruitment, and H3cit + neutrophil extracellular trap (NET) formation. N5's effects were partially recapitulated by DNase I in vivo and/or by its metabolites in vitro, suggesting its action involves metabolite-driven NET inhibition alongside DNase-like NET clearance. These findings illuminate N5's dual role in IBD-prophylactic barrier fortification and therapeutic resolution of neutrophilic inflammation, and highlight its potential as a multifaceted probiotic therapy.