Leucine Improved Growth Performance, Muscle Growth, and Muscle Protein Deposition Through AKT/TOR and AKT/FOXO3a Signaling Pathways in Hybrid Catfish Pelteobagrus v achelli × Leiocassis longirostris

Abstract

(1) Background: l-leucine (Leu) plays a positive role in regulating protein turnover in skeletal muscle in mammal. However, the molecular mechanism for the effects of Leu on muscle growth and protein deposition is not clearly demonstrated in fish. This study investigated the effects of dietary Leu on growth performance and muscle growth, protein synthesis, and degradation-related signaling pathways of hybrid catfish (Pelteobagrus vachelli♀ × Leiocassis longirostris♂). (2) Methods: A total of 630 hybrid catfish (23.19 ± 0.20 g) were fed 6 different experimental diets containing graded levels of Leu at 10.0 (control), 15.0, 20.0, 25.0, 30.0, 35.0, and 40.0 g Leu kg^-1 for 8 weeks. (3) Results: Results showed that dietary Leu increased percent weight gain (PWG), specific growth rate (SGR), FI (feed intake), feed efficiency (FE), protein efficiency ratio (PER), muscle fibers diameter, and muscle fibers density; up-regulated insulin-like growth factor I (IGF-I), insulin-like growth factor I receptor (IGF-IR), proliferating cell nuclear antigen (PCNA), myogenic regulation factors (MyoD, Myf5, MyoG, and Mrf4), and MyHC mRNA levels; increased muscle protein synthesis via regulating the AKT/TOR signaling pathway; and attenuated protein degradation via regulating the AKT/FOXO3a signaling pathway. (4) Conclusions: These results suggest that Leu has potential role to improve muscle growth and protein deposition in fish, which might be due to the regulation of IGF mRNA expression, muscle growth related gene, and protein synthesis and degradation-related signaling pathways. Based on the broken-line model, the Leu requirement of hybrid catfish (23.19-54.55 g) for PWG was estimated to be 28.10 g kg^-1 of the diet (73.04 g kg^-1 of dietary protein). These results will improve our understanding of the mechanisms responsible for muscle growth and protein deposition effects of Leu in fish.

Keywords: hybrid bagrid catfish; leucine; muscle growth; protein degradation; protein synthesis.

Figure 1

Broken-line analysis of PWG for hybrid catfish fed diets containing graded levels of Leu for 8 weeks.

Figure 2

Microstructure of white muscle in cross-section (bars represent 50 μm; (a), 10.0 g Leu kg⁻¹ diet group; (b), 25.0 g Leu kg⁻¹ diet group; (c), 40.0 g Leu kg⁻¹ diet group).

Figure 3

Effect of dietary Leu on protein content (mg protein·g tissue⁻¹ (A)) and RNA/protein ratio (mg RNA·g protein⁻¹ (B)) in hybrid catfish muscle. Data represent means ± SEM of three replicates, with six fish in each replicate. Values having different letters are significantly different (P < 0.05).

Figure 4

Effects of dietary Leu on IGF-I and IGF-IR gene expressions in muscle of hybrid catfish. Values are means ± SEM of three replicates, with six fish in each replicate, and different letters denote significant differences (P < 0.05).

Figure 5

Effects of dietary Leu on muscle growth related gene mRNA expressions in muscle of hybrid catfish. Values are means ± SEM of three replicates, with six fish in each replicate, and different letters denote significant differences (P < 0.05).

Figure 6

Effect of dietary Leu on the PI3K/AKT/TOR signaling pathway in hybrid catfish muscle. (A) Effect of dietary Leu on PI3K, AKT, TOR, S6K1, and 4E-BP1 mRNA expressions in muscle of hybrid catfish. (B) Effect of dietary Leu on the protein expressions of P-AKT (Ser473), AKT, P-TOR (Ser2448), TOR, P-S6K1 (Thr421/Ser424), and S6K1 in muscle of hybrid catfish. Results were expressed as the ratio of P-AKT and AKT (C), P-TOR and TOR (D), and P-S6K1 and S6K1 (E) protein levels. Data represent means ± SEM of three replicates, with six fish in each replicate. Values having different letters are significantly different (P < 0.05).

Figure 7

Effects of dietary Leu on the AKT/FOXO3a signaling pathway in hybrid catfish muscle. (A) Effect of dietary Leu on FOXO3a, MURF-1, and MAFBX mRNA expressions in muscle of hybrid catfish. (B) Effect of dietary Leu on the protein expressions of P-FOXO3a (Ser253), FOXO3a, MURF-1, and MAFBX in muscle of hybrid catfish. Results were expressed as the ratio of P-FOXO3a and FOXO3a (C), MURF-1 (D), and MAFBX (E) protein levels. Data represent means ± SEM of three replicates, with six fish in each replicate. Values having different letters are significantly different (P < 0.05).