4-Methylesculetin alleviated aflatoxin B1-induced liver injury and ferritinophagy through AMPK- TOR-Ulk axis in grass carp (Ctenopharyngodon idella)

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 supplementation 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.

4-Methylesculetin; Aflatoxin B1; Ferritinophagy; Grass carp; Injury; Liver.