Effects of supplementing coated methionine in a high plant-protein diet on growth, antioxidant capacity, digestive enzymes activity and expression of TOR signaling pathway associated genes in gibel carp, Carassius auratus gibelio.

This study explored the impacts of supplementation of different levels of coated methionine (Met) in a high-plant protein diet on growth, blood biochemistry, antioxidant capacity, digestive enzymes activity and expression of genes related to TOR signaling pathway in gibel carp (Carassius auratus gibeilo). A high-plant protein diet was formulated and used as a basal diet and supplemented with five different levels of coated Met at 0.15, 0.30, 0.45, 0.60 and 0.75%, corresponding to final analyzed Met levels of 0.34, 0.49, 0.64, 0.76, 0.92 and 1.06%. Three replicate groups of fish (initial mean weight, 11.37 ± 0.02 g) (20 fish per replicate) were fed the test diets over a 10-week feeding period. The results indicated that with the increase of coated Met level, the final weight, weight gain (WG) and specific growth rate initially boosted and then suppressed, peaking at 0.76% Met level (P< 0.05). Increasing dietary Met level led to significantly increased muscle crude protein content (P< 0.05) and reduced serum alanine aminotransferase activity (P< 0.05). Using appropriate dietary Met level led to reduced malondialdehyde concentration in hepatopancreas (P< 0.05), improved superoxide dismutase activity (P< 0.05), and enhanced intestinal amylase and protease activities (P< 0.05). The expression levels of genes associated with muscle protein synthesis such as insulin-like growth factor-1, protein kinase B, target of rapamycin and eukaryotic initiation factor 4E binding protein-1 mRNA were significantly regulated, peaking at Met level of 0.76% (P< 0.05). In conclusion, supplementing optimal level of coated Met improved on fish growth, antioxidant capacity, and the expression of TOR pathway related genes in muscle. The optimal dietary Met level was determined to be 0.71% of the diet based on quadratic regression analysis of WG.

Comprehensive study on the effect of dietary leucine supplementation on intestinal physiology, TOR signaling and microbiota in juvenile turbot (Scophthalmus maximus L.).

Intestinal damage and inflammation are major health and welfare issues in aquaculture. Considerable efforts have been devoted to enhancing intestinal health, with a specific emphasis on dietary additives. Branch chain amino acids, particularly leucine, have been reported to enhance growth performance in various studies. However, few studies have focused on the effect of leucine on the intestinal function and its underlying molecular mechanism is far from fully illuminated. In the present study, we comprehensively evaluated the effect of dietary leucine supplementation on intestinal physiology, signaling transduction and microbiota in fish. Juvenile turbot (Scophthalmus maximus L.) (10.13 ± 0.01g) were fed with control diet (Con diet) and leucine supplementation diet (Leu diet) for 10 weeks. The findings revealed significant improvements in intestinal morphology and function in the turbot fed with Leu diet. Leucine supplementation also resulted in a significant increase in mRNA expression levels of mucosal barrier genes, indicating enhanced intestinal integrity. The transcriptional levels of pro-inflammatory factors il-1ß, tnf-α and irf-1 was decreased in response to leucine supplementation. Conversely, the level of anti-inflammatory factors tgf-ß, il-10 and nf-κb were up-regulated by leucine supplementation. Dietary leucine supplementation led to an increase in intestinal complement (C3 and C4) and immunoglobulin M (IgM) levels, along with elevated antioxidant activity. Moreover, dietary leucine supplementation significantly enhanced the postprandial phosphorylation level of the target of rapamycin (TOR) signaling pathway in the intestine. Finally, intestinal bacterial richness and diversity were modified and intestinal bacterial composition was re-shaped by leucine supplementation. Overall, these results provide new insights into the beneficial role of leucine supplementation in promoting intestinal health in turbot, offering potential implications for the use of leucine as a nutritional supplement in aquaculture practices.

Effects of dietary tryptophan on the antioxidant capacity and immune response associated with TOR and TLRs/MyD88/NF-κB signaling pathways in northern snakehead, Channa argus (Cantor, 1842).

Introduction: Dietary tryptophan (Trp) has been shown to influence fish feed intake, growth, immunity and inflammatory responses. The purpose of this study was to investigate the effect and mechanism of Trp on immune system of juvenile northern snakehead (Channa argus Cantor, 1842). Methods: A total of 540 fish (10.21 ± 0.11 g) were fed six experimental diets containing graded levels of Trp at 1.9, 3.0, 3.9, 4.8, 5.9 and 6.8 g/kg diet for 70 days, respectively. Results and Discussion: The results showed that supplementation of 1.9-4.8 g/kg Trp in diets had no effect on the hepatosomatic index (HSI) and renal index (RI), while dietary 3.9 and 4.8 g/kg Trp significantly increased spleen index (SI) of fish. Dietary 3.9, 4.8, 5.9 and 6.8 g/kg Trp enhanced the total hemocyte count (THC), the activities of total antioxidant capacity (T-AOC) and superoxide dismutase (SOD). Malondinaldehyde (MDA) levels in the blood were significantly decreased by consuming 3.9 and 4.8 g/kg Trp. Fish fed with 3.0 and 3.9 g/kg Trp diets up-regulated interleukin 6 (il-6) and interleukin 8 (il-8) mRNA levels. The expression of tumor necrosis factor α (tnf-α) was highest in fish fed with 3.0 g/kg Trp diet, and the expression of interleukin 1ß (il-1ß) was highest in fish fed with 3.9 g/kg Trp diet. Dietary 4.8, 5.9 and 6.8 g/kg Trp significantly decreased il-6 and tnf-α mRNA levels in the intestine. Moreover, Trp supplementation was also beneficial to the mRNA expression of interleukin 22 (il-22). Additionally, the mRNA expression levels of target of rapamycin (tor), toll-like receptor-2 (tlr2), toll-like receptor-4 (tlr4), toll-like receptor-5 (tlr5) and myeloid differentiation primary response 88 (myd88) of intestine were significantly up-regulated in fish fed 1.9, 3.0 and 3.9 g/kg Trp diets, and down-regulated in fish fed 4.8, 5.9 and 6.8 g/kg Trp diets. Dietary 4.8 and 5.9 g/kg Trp significantly increased the expression of inhibitor of nuclear factor kappa B kinase beta subunit (ikkß) and decreased the expression of inhibitor of kappa B (iκbα), but inhibited nuclear transcription factor kappa B (nf-κb) mRNA level. Collectively, these results indicated that dietary 4.8 g/kg Trp could improve antioxidant capacity and alleviate intestinal inflammation associated with TOR and TLRs/MyD88/NF-κB signaling pathways.

Influence of dietary replacement of fish meal with fish soluble meal on growth and TOR signaling pathway in juvenile black sea bream (Acanthopagrus schlegelii).

An 8-week feeding trial was conducted to evaluate the effect of replacement of fish meal (FM) with fish soluble meal (FSM) on growth performance, feed utilization and expression of genes involved in TOR signaling pathway for juvenile black sea bream (Acanthopagrus schlegelii). Six isonitrogenous (41%) and isolipidic diets were prepared to contain graded levels of FSM which replaced 0% (control diet), 10%, 20%, 30%, 40% and 60% protein from FM. Triplicate groups of 20 fish with initial weight 0.51 ± 0.01 g were fed with experimental diets twice daily to apparent satiation. The results showed significant differences in growth performance and feed utilization among all treatments, final body weight (FBW), percent weight gain (PWG), specific growth rate (SGR) and protein efficiency ratio (PER) significantly increased with dietary replacement levels of FM with FSM increasing from 0% to 40% (P < 0.05), PWG, SGR and PER were significantly reduced when replacement of FM with FSM further increased from 40% to 60%. Based on PWG against replacement levels of FM with FSM, A two-slope broken-line model analysis indicated that the optimal replacement of FM with FSM is to be 42.59%. Moreover, the lowest feed conversion ratio (FCR) was observed in fish fed the 40% FSM replacement diet. Muscle amino acid profile in muscle revealed that total essential amino acids, arginine and threonine were significantly influenced by replacement levels of FSM, while there was no significant difference in NEAA among all treatments. The hematological indices were not affected by the replacement levels of FM with FSM. The relative expression levels of irs-1, pi3k, akt, igf-1, s6k1 and tor were up-regulated when replacement levels of FM with FSM increased from 0% to 40%, and higher values were observed in fish fed with 40% FSM replacement diet compared to those fed the other diets. However, relative expression of 4e-bp2 was down-regulated when replacement levels of FM with FSM increased from 0% to 40% (P < 0.05). In summary, the results of present study indicated that FSM could be a viable alternative protein source for black sea bream, dietary FSM supplementation could improve growth and up-regulate the relative expression of irs-1, pi3k, akt, igf-1, s6k1 genes related to TOR signaling pathway in liver of juvenile black sea bream.

Effects of dietary tryptophan levels on antioxidant status and immunity for juvenile blunt snout bream (Megalobrama amblycephala) involved in Nrf2 and TOR signaling pathway.

Dietary administration of tryptophan has been proved improving growth performance of fish. An 8-week feeding trial was conducted to investigate the effects of dietary tryptophan level on antioxidant capacity and immune response through Nrf2 and TOR signaling pathway. The results showed that, 0.08% tryptophan level significantly increased plasma aspartate aminotransferase (AST), while immunoglobulin M (IgM) and alkaline phosphatase (ALP) were strikingly increased by 0.40% level. The level of plasma complement component 3 (C3), alanine aminotransferase (ALT) and albumin (ALB) were independent of tryptophan supplementation. Total superoxide dismutase (T-SOD), catalase (CAT), total antioxidant capacity (T-AOC) and glutathione (GSH) activity were increased with increasing dietary tryptophan level until 0.40% and then decreased, while the level of malondialdehyde (MDA) showed a reverse trend. 0.19% and 0.28% tryptophan level significantly improved the glutathione peroxidase 1 (GPx-1) activity. Compared with 0.08% dietary tryptophan level, 0.40% level significantly improved nuclear factor erythroid 2-related factor 2 (Nrf2), GPx, manganese superoxide dismutase (Mn-SOD), CAT and transforming growth factor-ß (TGF-ß) mRNA level, while Kelch-like ECH-associated protein 1 (Keap1) and interleukin 1ß (IL-1ß) mRNA level were significantly decreased. The relative expression of copper zinc superoxide dismutase (Cu/Zn-SOD), heme oxygenase-1 (HO-1), target of rapamycin (TOR), phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K), protein kinase B (Akt) and interleukin 10 (IL-10) were significantly improved by 0.28% diet, while the mRNA level of tumor necrosis factor-α (TNF-α) and nuclear factor-kappa B (NF-κB) were increased by 0.08% diet. Interleukin 8 (IL-8) mRNA level was not significantly affected by dietary tryptophan. Based on MDA and T-SOD value, the optimal dietary tryptophan level of juvenile blunt snout bream was determined to be 0.33% (1.03% of dietary protein) and 0.36% (1.13% of dietary protein), respectively, using quadratic regression analysis.

Leucine promotes the growth of squabs by increasing crop milk protein synthesis through the TOR signaling pathway in the domestic pigeon (Columba livia).

Leucine (Leu) plays a critical regulatory role in protein synthesis, however, the effects and molecular mechanisms of Leu on crop milk protein in the domestic pigeons (Columba livia) are still unknown. Therefore, the study aimed to investigate the effects of dietary Leu supplementation on crop milk protein synthesis and the growth performance of squabs and the possible underlying mechanism. A total of 240 pairs of breeding pigeons (1102.3 ± 9.5 g/pair) were randomly assigned to 1 of 5 treatments, including a positive control (PC) diet that had adequate crude protein (crude protein, CP = 18%; Leu = 1.30%), a negative control (NC) diet that was low in CP (CP = 16%, Leu = 1.30%), and NC diets supplemented with Leu at 0.15%, 0.45%, or 1.05%. Compared with the NC diet, 0.15 to 0.45% Leu supplementation decreased BW loss and increased relative crop weight, crop thickness, and protein levels in the crop tissue and milk of breeding pigeons. However, dietary supplementation with 1.05% Leu inhibited ADFI in breeding pigeons. Dietary supplementation with 0.15 to 0.45% Leu decreased the mortality rate and increased the BW, eviscerated yield, and breast muscle yield of young squabs. The protein expression levels of the target of rapamycin (TOR), ribosomal protein S6 kinase 1 (S6K1), ribosomal protein S6 kinase (S6), eukaryotic initiation factor 4E binding protein 1 (4EBP1), and eukaryotic translation initiation factor 4E (eIF4E) were upregulated in the crop tissue of breeding pigeons in PC, 0.15% and 0.45% Leu-supplemented groups. Collectively, these results indicated that 0.15 to 0.45% Leu supplementation could decrease BW loss, increase milk protein synthesis in the crop of breeding pigeons, and enhance the survival rate and growth performance of young squabs through the TOR signaling pathway.