Effects of 30 d of dietary treatments on (a) body weight and (b) body length of juvenile zebrafish. Values are means ± SEM, n 3. *Significantly different (P < 0·05, two-way (dietary Se × dietary VE or MHY1485) ANOVA followed by Bonferroni–Dunn multiple comparison). ASe, the basal Se-adequate diet; DSe, the basal Se-deficient diet; VE, DL-α-tocopherol acetate, antioxidant; MHY1485, activator of the target of rapamycin.

Effects of 30 d of dietary treatments on Se status in skeletal muscle of juvenile zebrafish. (a) Total Se concentration. (b) Gpx activity. (c) Relative mRNA levels of selenoprotein genes. Values are means ± SEM, n 3. *Significantly different (P < 0·05, two-way (dietary Se × dietary VE or MHY1485) ANOVA followed by Bonferroni–Dunn multiple comparison). ASe, the basal Se-adequate diet; DSe, the basal Se-deficient diet; VE, DL-α-tocopherol acetate, antioxidant; MHY1485, activator of the target of rapamycin; Gpx, glutathione peroxidase; dio, iodothyronine deiodinase; seleno, selenoprotein.

Effects of 30 d of dietary treatments on the levels of (a) ROS and (b) oxidative status biomarkers in skeletal muscle of juvenile zebrafish. (a) Three different ROS fluorescent probes (DCFH-DA, DHE and Amplex red) were applied to detect non-specific ROS, superoxide anion and hydrogen peroxide, respectively. (b) The levels of protein carbonyl and malondialdehyde were detected to evaluate oxidative status. Values are means ± SEM, n 3. *Significantly different, ns represents not significantly different (P < 0·05, two-way (dietary Se × dietary VE or MHY1485) ANOVA followed by Bonferroni-Dunn multiple comparison). ASe, the basal Se-adequate diet; DSe, the basal Se-deficient diet; VE, DL-α-tocopherol acetate, antioxidant; MHY1485, activator of the target of rapamycin; ROS, reactive oxygen species; DCFH-DA, 2’,7’-dichlorodihydrofluorescein diacetate; DHE, dihydroethidium; MDA, malondialdehyde.

Effects of 30 d of dietary treatments on Akt-TORC1 pathway in skeletal muscle of juvenile zebrafish. Data were obtained 2 h after a meal by (a) Western blotting assay, and (b) the relative phosphorylation levels of proteins in Akt-TORC1 pathway were calculated. Values are means ± SEM, n 3. *Significantly different (P < 0·05, two-way (dietary Se × dietary VE or MHY1485) ANOVA followed by Bonferroni–Dunn multiple comparison). ASe, the basal Se-adequate diet; DSe, the basal Se-deficient diet; VE, DL-α-tocopherol acetate, antioxidant; MHY1485, activator of the target of rapamycin; Akt, protein kinase B; TOR, the target of rapamycin; TORC1, the target of rapamycin complex 1; S6K1, 70 kDa ribosomal protein S6 kinase; S6, ribosomal protein S6; eEF2, eukaryotic translation elongation factor 2; 4E-BP1, eukaryotic translation initiation factor 4E binding protein.

Effects of 30 d of dietary treatments on protein synthesis in skeletal muscle of juvenile zebrafish. Muscle protein synthesis rate was detected 2 h after a meal using the surface sensing of translation method and quantified by measuring the incorporation of exogenous puromycin into nascent peptides. Puromycin-labelled peptides were detected by (a) Western blotting assay using an antibody against puromycin, and (b) the results are normalised to α-Tubulin. Values are means ± SEMs, n 3. *Significantly different (P < 0·05, two-way (dietary Se × dietary VE or MHY1485) ANOVA followed by Bonferroni–Dunn multiple comparison). ASe, the basal Se-adequate diet; DSe, the basal Se-deficient diet; VE, DL-α-tocopherol acetate, antioxidant. MHY1485, activator of the target of rapamycin.

Effects of 30 d of dietary treatments on histological characteristics of skeletal muscle in juvenile zebrafish. (a) Zebrafish whole-body transverse sections were obtained at the vent level and stained with wheat germ agglutinin (green, for plasma membrane). (b) Representative images of whole-body transverse sections, and the corresponding zoom-in images for white muscle fibres. The area of white muscle within the whole-body transverse section is presented in the image obtained from zebrafish fed the ASe. WM, white muscle; SC, spinal cord; NC, notochord; V, vent. Scale bar for whole-body transverse section, 500 μm; scale bar for white muscle fibre, 100 μm. Images were subjected to a morphological survey to calculate (c) the total cross-sectional area of white muscle and (d) the mean diameter of white muscle fibres. Values are means ± SEM, n 3. *Significantly different (P < 0·05, two-way (dietary Se× dietary VE or MHY1485) ANOVA followed by Bonferroni–Dunn multiple comparison). ASe, the basal Se-adequate diet; DSe, the basal Se-deficient diet; VE, DL-α-tocopherol acetate, antioxidant. MHY1485, activator of the target of rapamycin.

A schematic diagram for the regulatory mechanism of Se in skeletal muscle fibre hypertrophy in zebrafish. (a) Under Se adequate status, zebrafish skeletal muscle exhibits a low concentration of ROS, an active PI3K-Akt-TORC1 signalling and an efficient protein synthesis, which attribute to maintain the normal hypertrophy process of skeletal muscle fibres in zebrafish. (b) Under Se-deficient status, the down-regulation of antioxidant selenoproteins lead to a high concentration of ROS. The high concentration of ROS dephosphorylates and inactivate Akt, thereby inhibiting TORC1 pathway-mediated protein synthesis and leading to the suppressed hypertrophy of skeletal muscle fibres. IGF1, insulin-like growth factor 1; IRSs, insulin receptor substrates; PI3K, phosphatidylinositol-3-kinase; PIP2, phosphoinositide-4,5-biphosphate; PIP3, phosphoinositide-3,4,5-triphosphate; PDK1, phosphoinositide-dependent kinase 1; Akt, protein kinase B; TORC1, the target of rapamycin complex 1; TORC2, the target of rapamycin complex 2; ROS, reactive oxygen species.