Abstract: 【Objective】 To investigate the effect of aflatoxin B₁ (AFB₁) on the apoptosis of porcine jejunum epithelial cells (IPEC-J2) by inducing intracellular reactive oxygen species (ROS) production, providing a theoretical basis for the pathogenic mechanism of AFB1. 【Methods】 Cells were stimulated with AFB₁ at concentrations of 0, 20, 40, 60, 80 and 100 μg/mL for 12 h.CCK-8 assay kit was used to detect cell viability and determine the toxic effect of AFB₁ on IPEC-J2 cells, and then immunofluorescence assay were used to detect intracellular ROS level after the cells were stimulated with 0, 10, 20 and 30 μg/mL AFB_1.At the same time, Real-time quantitative PCR and Western blotting were used to detect the expression of key factors related to apoptotic pathways in cells.ROS inhibitor N-acetyl-L-cysteine (NAC) was used to stimulate cells, and then the effect of AFB₁ on the expression of apoptotic factors in cells was detected. 【Result】 The results showed that AFB₁ at concentrations of 0-20 μg/mL had no significant effect on cell viability (P＞0.05), and as the concentration of AFB₁ increased, the viability of IPEC-J2 cells showed a significant or extremely significant decrease trend compared with blank control group (P＜0.05 or P＜0.01).ROS immunofluorescence analysis showed that ROS level was highly dependent on AFB₁ concentration, and as the concentration of AFB₁ increased, the intracellular ROS level was significantly or extremely significantly increased compared with blank control group (P＜0.05 or P＜0.01).Real-time quantitative PCR and Western blotting results showed that when the concentration of AFB₁ was 20 μg/mL, the expression of Caspase-3 mRNA and protein were significantly higher than that of blank control group (P＜0.05).When the concentration of AFB₁ was 30 μg/mL, the expression of Bax and Bax/Bcl-2 mRNA and protein were significantly higher than that of blank control group (P＜0.05), while the protein expressions of Bax and Bcl-2 were significantly lower than those of blank control group(P＜0.05).Pretreatment of cells with ROS inhibitor NAC followed by AFB₁ addition resulted in a decrease in apoptotic factors to varying degrees compared with the AFB₁-only treatment group (P＜0.05). 【Conclusion】 AFB₁ could promote the upregulation of apoptotic factors Bax, Bcl-2, and Caspase-3 in IPEC-J2 cells, and this process might be related to the accumulation of intracellular reactive oxygen species.

Key words: aflatoxins B₁; reactive oxygen species; cell apoptosis; porcine jejunum epithelial cells