Ligilactobacillus salivarius 8-2 attenuates intestinal mucosal injury in chicks via the mitochondrial fision-glycometabolism reprogramming axis

The intestinal mucosa is crucial for nutrient absorption and barrier function. In this study, a newly identified strain of Ligilactobacillus salivarius (L. sa. 8-2) exhibited a mucosal protective effect in chick intestinal injury models. However, the underlying mechanisms need to be clarified. In vivo, chicks were pre-fed with L. sa. 8-2 supernatant or its precipitate and then challenged with lipopolysaccharide (LPS) or Salmonella Typhimurium (STm). The results showed that L. sa. 8-2 and its supernatant significantly reduced intestinal epithelial apoptosis, increased goblet cell density and Muc2 mRNA abundance, and upregulated Claudin1 protein levels. Specifically, compared with the LPS-challenged group, L. sa. 8-2 supernatant accelerated epithelial renewal, as shown by elevated protein levels of proliferating cell nuclear antigen (PCNA) and increased mRNA abundance of cell cycle regulators Ccnd1 and Cdk2. Moreover, the L. sa. 8-2 supernatant markedly increased protein levels of both the active intestinal stem cell (aISC) marker LGR5 and the reserve ISC (rISC) marker HOPX in the intestinal crypt. In vitro, the enteroids were pre-treated with L. sa. 8-2 supernatant and then challenged with LPS. It was shown that the L. sa. 8-2 supernatant elevated the protein levels of LGR5 and HOPX compared with the injured group. Moreover, co-staining of LGR5 and HOPX revealed that more HOPX⁺ cells were co-localized with LGR5⁺ cells in the L. sa. 8-2 supernatant pretreatment group, suggesting the activation of rISCs. Additionally, compared with the injured groups, L. sa. 8-2 supernatant preserved mitochondrial morphology and upregulated the fission-related protein FIS1. At both the mRNA and protein levels, L. sa. 8-2 supernatant upregulated the pyruvate metabolism-related enzyme LDHA and LDHB, while decreasing mRNA levels of Sdha (an oxidative phosphorylation-related enzyme), suggesting a metabolic shift from oxidative phosphorylation toward glycolysis and an enhancement of pyruvate utilization. Taken together, L. sa. 8-2 alleviates intestinal mucosal injury and promotes epithelial renewal by activating ISCs, which is mediated by promoting mitochondrial fission and subsequent enhancement of pyruvate utilization in the intestinal crypt.