Dietary dimethylglycine alleviates cyclic heat stress-induced jejunal injury in yellow-feathered broilers by inhibiting the TLR4/NF-κB signaling pathway and restoring MUC2 expression

This study aimed to investigate the protective effect and mechanism of dietary dimethylglycine (DMG) on jejunal injury in yellow-feathered broilers subjected to cyclic heat stress (HS). A total of 180 35-day-old male yellow-feathered broilers were randomly allocated into 3 groups: the thermoneutral condition with basal diet group (CON), the cycle heat stress with basal diet group (HS, 32-35°C), and the cycle heat stress with 0.1% DMG in basal diet group (HS_DMG, 32-35°C). The experiment included a 7-day heating period followed by a 14-day cyclic heat stress period. Growth performance, rectal temperature, jejunal morphology and ultrastructure, inflammatory cytokine levels, and protein expression related to the TLR4/NF-κB pathway and MUC2 were evaluated. Additionally, primary chicken jejunal epithelial cells were isolated and exposed to heat stress with or without DMG pretreatment to validate in vivo findings. Results showed that heat stress significantly increased rectal temperature (P < 0.05), had a tendency to reduce body weight (BW, P = 0.058) and average daily gain (ADG, P = 0.050), and increase the feed-to-gain ratio (F/G, P = 0.053). Histologically, HS caused epithelial shedding, reduced villus height and villus height/crypt depth, and induced microvilli damage and mitochondrial swelling, all of which were attenuated by DMG supplementation. Heat stress also elevated jejunal mucosa levels of IL-6, IL-8, IL-1β, and TNF-α (P < 0.05), while DMG reduced IL-6, IL-8, and TNF-α (P < 0.05). Immunohistochemistry and western blotting revealed that HS decreased MUC2 expression and increased TLR4 and p-NF-κB-p65 protein levels (P < 0.05), whereas DMG reversed these changes (P < 0.05). In vitro, DMG pretreatment alleviated HS-induced reduction in inflammatory cytokine release, and alterations in TLR4, NF-κB, and MUC2 protein expression. In conclusion, dietary DMG supplementation alleviates cyclic heat stress-induced jejunal injury in yellow-feathered broilers by suppressing the TLR4/NF-κB signaling pathway and restoring MUC2 expression.