A new insight on alleviating the inhibitory effect of aflatoxin B1 on muscle development in grass carp (Ctenopharyngodon idella): the effect of 4-methylesculetin in vivo and in vitro

Abstract

Aflatoxin B1 (AFB1), an important fungal toxin, exists mainly in plant feed ingredients and animals consuming feed contaminated with AFB1 will have reduced growth and impaired health condition mainly due to oxidative stress and reduced immunity. Our previous study found that AFB1 caused oxidative damage and inhibited muscle development of zebrafish. 4-methylesculetin (4-ME), a coumarin derivative, is now used in biochemistry and medicine widely because of its antioxidant function. Whether 4-ME could alleviate the inhibition of muscle development in grass carp induced by AFB1 has not been reported. In this experiment, 720 healthy grass carp (11.40 ± 0.01 g) were randomly divided into 4 groups with 3 replicates of 60 fish each, including control group, AFB1 group (60 μg/kg diet AFB1), 4-ME group (10 mg/kg diet 4-ME), and AFB1+4-ME group (60 μg/kg diet AFB1 + 10 mg/kg 4-ME diet), for a 60 d growth experiment. In vitro, we also set up 4 treatment groups in grass carp primary myoblast, including control group, AFB1 group (15 μmol/L AFB1), 4-ME group (0.5 μmol/L 4-ME) and AFB1+4-ME group (15 μmol/L AFB1+0.5 μmol/L 4-ME). The results showed that dietary AFB1 decreased growth performance of grass carp, damaged the ultrastructure and induced oxidative damage in grass carp muscle, and significantly decreased the mRNA and protein expression levels of myogenin (MyoG), myogenic differentiation (MyoD), myosin heavy chain (MYHC), as well as the protein expression levels of laminin β1, fibronectin and collagen Ⅰ (P < 0.05), significantly activated the protein expression levels of urokinase-type plasminogen activator (uPA), matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9) and phosphorylate-38mitogen-activated protein kinase (p38 MAPK) both in grass carp muscle and grass carp primary myoblast (P < 0.05). Supplementation of AFB1 with 4-ME significantly improved the growth performance inhibition and alleviated the muscle fiber development inhibition and extracellular matrix (ECM) degradation in grass carp induced by AFB1. The present results revealed that supplementation of AFB1 contaminated feed with 4-ME reduced the inhibition of growth and muscle development by alleviating AFB1-induced ECM degradation in grass carp, which might be related to the p38 MAPK/uPA/MMP/ECM pathway. The results implied that 4-ME could be used as a valuable mycotoxin scavenger in animal feed.