Abstract
Animal intestines are extremely susceptible to the adverse effects of feed and external conditions within intensive production systems. 4-Methylesculetin (4-ME) is a natural coumarin with a catechol structure and has potent antioxidant and anti-inflammatory properties. This study was investigating the effects of 4-ME as a functional substance added to the diet on the growth performance, intestine growth, and intestinal integrity of juvenile grass carp (Ctenopharyngodon idella). A total of 1080 healthy grass carp (11.40 ± 0.01 g) were randomly assigned to 18 net cages. The experiment was divided into 6 treatment groups with 3 replicates in each group and 60 fish in each replicate. Different levels of 4-ME (0.00, 2.46, 5.14, 10.14, 19.88, and 39.86 mg/kg) were added to the basal diet. The experimental period of 60 days. Dietary supplementation with 10.14 mg/kg 4-ME increased specific growth rate (SGR), feed efficiency (FE), and whole body composition (crude protein and crude fat) (P < 0.05), but there is no significant effect on crude ash (P > 0.05). Dietary supplementation with 10.14 mg/kg 4-ME increased the content of UDP-glycosyltransferase (UGT) enzymes in hepatopancreas (P < 0.001), and digestive and absorptive enzyme activity in the intestine (P < 0.05). Dietary supplementation with 10.14 mg/kg 4-ME promoted intestine somatic index and fold height by increasing intestinal proliferation in intestine (P < 0.05), possibly through the WNT/β-catenin pathway. Dietary supplementation with 10.14 mg/kg 4-ME improved the structural integrity of intestinal apical junctional complex (AJC), including mRNA expression levels of zo-2b, claudin (-b, c, f, 3c, 11, 12) and increased the protein expression levels of ZO-1, occludin and β-catenin (P < 0.05). Dietary supplementation with 10.14 mg/kg 4-ME were decreased mRNA expression levels of claudin-15a, claudin-15b, ROCK, RhoA, MLCK and the protein expression levels of MLCK and ROCK1 (P < 0.05), and its mechanism may have been mediated through the RhoA/ROCK pathway. In addition, according to SGR, diamine oxidase (DAO) and UGT, the appropriate addition of 4-ME in juvenile grass carp were determined to be 9.62, 9.25, and 10.65 mg/kg, respectively. In this experiment, 4-ME was incorporated into the diet as a functional substance to investigate its effects on the growth performance and intestinal health of fish, indicating that 4-ME has potential for development as a functional feed additive in aquaculture production.