Probiotic Lactobacillus johnsonii Reduces Intestinal Inflammation and Rebalances Splenic Treg/Th17 Responses in Dextran Sulfate Sodium-Induced Colitis

Inflammatory bowel disease (IBD), a chronic inflammatory disorder of the gastrointestinal tract, is frequently complicated by extraintestinal manifestations such as functional hyposplenism. Increasing evidence highlights its pathogenesis as a multifactorial interplay of gut dysbiosis, intestinal barrier dysfunction, and dysregulated immune responses. While probiotics, particularly <i>Lactobacillus</i> spp., have emerged as potential therapeutics for IBD, restoring intestinal homeostasis, their systemic immunomodulatory effects remain underexplored. Here, we investigated the protective role of <i>Lactobacillus johnsonii</i> N5 in DSS-induced colitis, focusing on inflammation inhibition and splenic T cell regulation. Pretreatment with <i>L. johnsonii</i> N5 significantly attenuated colitis severity, as evidenced by preserved body weight, reduced disease activity index, and prevention of colon shortening. N5 suppressed colonic pro-inflammatory factors such as TNF-α, Il-1b, Il-6, and CXCL1, while elevating anti-inflammatory IL-10 at both mRNA and protein levels. Transcriptomic analysis of the spleen revealed that N5 mediated the downregulation of inflammatory pathways, including the IL-17 and TNF signaling pathways, as well as the HIF-1 signaling pathway, and modulated the metabolic pathway of oxidative phosphorylation. Flow cytometry analysis demonstrated that N5 rebalanced splenic Treg/Th17 responses by expanding the Treg population and reducing the production of IL-17A in Th17 cells. Notably, Th17-associated IL-17A positively correlated with intestinal pro-inflammatory mediators, emphasizing the role of Th17 cells in driving colitis. In contrast, splenic Treg abundance positively correlated with colonic IL-10 levels, suggesting a link between systemic immune regulation and intestinal anti-inflammatory responses. Our study underscores the therapeutic potential of targeting gut-immune crosstalk through probiotics, thereby offering valuable insights for developing live bacterial-based interventions for IBD and other inflammatory disorders.