摘要The mycotoxin aflatoxin B1(AFB1),frequently identified in animal feed and raw materials,induces oxidative stress as a primary toxicological consequence.The coumarin compound 4-methylesculetin(4-ME)possesses notable antioxidant properties,leading to its application in medical contexts.Given that the liver is the principal organ targeted by AFB1,this study investigated the potential mechanism through which 4-ME mitigated hepatic injury induced by AFB1 in grass carp.The grass carp(initial body weight of 11.40±0.01 g)were randomly divided into 4 treatment groups(3 replicates/treatment and 60 fish/replicate),which were control group(Control),60 μg/kg AFB1 group(AFB1),10 mg/kg 4-ME group(4-ME)and 60 μg/kg AFB1+10 mg/kg 4-ME group(AFB1+4-ME).The present study found that AFB1 caused liver injury,dietary 4-ME supplementation enhanced the antioxidant capacity through the Nrf2 pathway,decreased the levels of AFB1-DNA adducts(P＜0.001)and total bile acid(TBA)(P＜0.001)in liver,and decreased the protein expression of CYP3A4 in liver(P＜0.001),inhibited the transcriptional levels of endoplasmic reticulum stress(ERS)-related genes(including XBP1,IRE1,ATF6,Chop,EIF2α,PERK,and GRP78)(P＜0.05),autophagy-related genes(including beclin 1,LC3,and ATG12)(P＜0.05)and apoptosis-related genes(including Bax,caspase-8,and caspase-3)(P＜0.05).Dietary 4-ME supple-mentation also decreased the contents of iron(P=0.004),and increased SLC7A11(P=0.001)and GPx4 protein levels(P＜0.001)in liver,and alleviated AFB1-induced elevation of AMPK and Ulk1 genes and proteins expression(P＜0.05),and the decreased of TOR gene and protein expression(P＜0.05)in live.In summary,AFB1 induced oxidative damage,ERS,apoptosis,and autophagy in the liver of grass carp,which are associated with ferroptosis and linked to the activation of the AMPK-TOR-Ulk1 signaling axis.Notably,supplementation with 4-ME mitigated these effects.The findings offer new theoretical insights into the potential of 4-ME to alleviate ferritinophagy-related diseases induced by AFB1 in grass carp.