Protective role of phenylalanine on the ROS-induced oxidative damage, apoptosis and tight junction damage via Nrf2, TOR and NF-κB signalling molecules in the gill of fish

Abstract

This study explored the possible preventive effects of dietary phenylalanine (Phe) on antioxidant responses, apoptosis and tight junction protein transcription in the gills of young grass carp (Ctenopharyngodon idella). Fish were fed six different experimental diets containing graded levels of Phe (3.4–16.8 g kg−1) for 8 weeks. The results showed that Phe deficiency induced protein oxidation and lipid peroxidation by decreasing the glutathione content and the activities and mRNA levels of Cu/Zn superoxide dismutase (SOD1), catalase (CAT), glutathione peroxidase (GPx), glutathione reductase (GR) and glutathione-S-transferase (GST) in fish gill (P < 0.05). These results may be ascribed to the downregulation of NF-E2-related factor 2 (Nrf2), target of rapamycin (TOR) and ribosomal protein S6 kinase 1 (S6K1), and the upregulation of Kelch-like-ECH-associated protein 1 a (Keap1a) expression in grass carp gills (P < 0.05). Additionally, Phe deficiency induced DNA fragmentation via the up-regulation of Caspase 3, Caspase 8 and Caspase 9 mRNA expression (P < 0.05). These results may be ascribed to the improvement in reactive oxygen species (ROS) levels in the fish gills (P < 0.05). Furthermore, the results indicated that Phe deficiency decreased Claudin b, Claudin 3, Occludin and ZO-1 transcription and increased Claudin 15 expression in the fish gills (P < 0.05). These effects were partly due to the downregulation of interleukin 10 (IL-10), transforming growth factor β (TGF-β) and inhibitor factor κBα (iκBα) and the upregulation of relative mRNA expression of interleukin 1β (IL-1β), interleukin 8 (IL-8), tumour necrosis factor-α (TNF-α) and nuclear transcription factor-κB p65 (NF-κB p65) (P < 0.05). Taken together, the results showed that Phe deficiency impaired the structural integrity of fish gills by regulating the expression of tight junction proteins, cytokines, antioxidant enzymes, NF-κB p65, iκBα, TOR, Nrf2, Keap1 and apoptosis-related genes in the fish gills.