Protective roles of arginine, threonine, and tryptophan against oxidative stress in chicken intestinal epithelial cells

Chicken intestinal epithelial cells (cIECs) play a crucial role in nutrient absorption and maintaining barrier integrity but are highly susceptible to oxidative stress. This study investigated the efficacy of arginine (Arg), tryptophan (Trp), and threonine (Thr) in protecting cIECs from hydrogen peroxide (H₂O₂) induced oxidative stress. The experimental design consisted of 8 treatment groups: a positive control (PC, cells incubated with fresh medium), a negative control (NC, cells exposed to 1 mM H₂O₂), and 6 additional groups pretreated with either 250 or 500 mM Arg, Trp, or Thr for 24 h (6 replicates per treatment) prior to 6 h incubation with 1 mM H₂O₂. The results demonstrated that all amino acid treatments significantly improved (<italic>p </italic>< 0.05) cell viability compared to NC. Total antioxidant capacity (TAC) was markedly lower (<italic>p </italic>< 0.05) in NC than in PC; however, 250 μM Arg and 250 μM Trp significantly enhanced (<italic>p </italic>< 0.05) TAC compared to NC. Paracellular permeability was significantly lower (<italic>p </italic>< 0.05) in PC compared with NC. Treatments with 500 μM Arg, 250 μM Trp, and both Thr levels reduced (<italic>p </italic>< 0.05) paracellular permeability compared to NC and other amino acid groups. At all time points, transepithelial electrical resistance was consistently greater (<italic>p </italic>< 0.05) in PC than in NC, with notable effects observed for 500 μM Arg and Thr treatments, which exhibited greater (<italic>p </italic>< 0.05) values than the other amino acid treatments. Expression of tight junction-related genes, including <italic>zonula occludens-1</italic>, <italic>claudin</italic>, <italic>occludin</italic>, and <italic>junctional adhesion molecule 2</italic>, was greater (<italic>p </italic>< 0.05) in PC than in NC. All amino acid treatments increased (<italic>p </italic>< 0.05) the expression of <italic>claudin</italic> and <italic>occludin</italic> compared with NC. In conclusion, Arg, Trp, and Thr attenuate oxidative stress in cIECs by enhancing cell viability, antioxidant capacity, and barrier function, primarily through upregulation of tight junction-related gene expression.