Copper exposure elicits pyroptosis and inflammation in bursa of Fabricius and omega-3 reverses this phenomenon by inhibiting NF-κB-NLRP3-GSDMD axis: In vivo, in vitro, and in silico investigations

The overuse of copper (Cu) in feed additives has become a major issue, disrupting animal and ecosystem balance and impairing animal immunity. Omega-3 (ω-3) PUFAs, with immunomodulatory effects, may counteract the immunotoxicity. This study, via in vivo, in vitro, and in silico methods, explored Cu²⁺ exposure's impact on chicken bursa of Fabricius (BF). Cu²⁺ raised ROS levels in BF and PBL, causing oxidative damage. ω-3 intervention reduced ROS, restored antioxidant Enzymes, and alleviated stress. Moreover, through in vivo, in vitro, and in silico studies, we found ω-3 effectively inhibited Cu²⁺-induced Pyroptosis and inflammation. It modulated the NF-κB-NLRP3-GSDMD signaling pathway, down-regulating the mRNA and protein expression of key genes like NLRP3 and GSDMD, and reducing NF-κB phosphorylation to block the inflammatory cascade. Besides, ω-3 mitigated BF pathological changes, such as lessening inflammatory cell infiltration and trabecular damage. Overall, this research reveals how ω-3 alleviates copper-induced immune dysfunction. The findings can guide safer feed additive strategies, ensuring the safety of animal-derived food and a healthier food chain.