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.

Effects of selenoprotein extracts from Cardamine hupingshanensis on growth, selenium metabolism, antioxidant capacity, immunity and intestinal health in largemouth bass Micropterus salmoides.

This study aimed to assess the impact of dietary selenoprotein extracts from Cardamine hupingshanensis (SePCH) on the growth, hematological parameters, selenium metabolism, immune responses, antioxidant capacities, inflammatory reactions and intestinal barrier functions in juvenile largemouth bass (Micropterus salmoides). The base diet was supplemented with four different concentrations of SePCH: 0.00, 0.30, 0.60 and 1.20 g/Kg (actual selenium contents: 0.37, 0.59, 0.84 and 1.30 mg/kg). These concentrations were used to formulate four isonitrogenous and isoenergetic diets for juvenile largemouth bass during a 60-day culture period. Adequate dietary SePCH (0.60 and 1.20 g/Kg) significantly increased weight gain and daily growth rate compared to the control groups (0.00 g/Kg). Furthermore, 0.60 and 1.20 g/Kg SePCH significantly enhanced amounts of white blood cells, red blood cells, platelets, lymphocytes and monocytes, and levels of hemoglobin, mean corpuscular volume and mean corpuscular hemoglobin in the hemocytes. In addition, 0.60 and 1.20 g/Kg SePCH increased the mRNA expression levels of selenocysteine lyase, selenophosphate synthase 1, 15 kDa selenoprotein, selenoprotein T2, selenoprotein H, selenoprotein P and selenoprotein K in the fish liver and intestine compared to the controls. Adequate SePCH not only significantly elevated the activities of antioxidant enzymes (Total superoxide dismutase, catalase, glutathione reductase, glutathione peroxidase), the levels of total antioxidant capacity and glutathione, while increased mRNA transcription levels of NF-E2-related factor 2, Cu/Zn-superoxide dismutase, catalase, glutathione reductase and glutathione peroxidase. However, adequate SePCH significantly decreased levels of malondialdehyde and H2O2 and the mRNA expression levels of kelch-like ECH-associated protein 1a and kelch-like ECH-associated protein 1b in the fish liver and intestine compared to the controls. Meanwhile, adequate SePCH markedly enhanced the levels of immune factors (alkaline phosphatase, acid phosphatase, lysozyme, complement component 3, complement component 4 and immunoglobulin M) and innate immune-related genes (lysozyme, hepcidin, liver-expressed antimicrobial peptide 2, complement component 3 and complement component 4) in the fish liver and intestine compared to the controls. Adequate SePCH reduced the levels of pro-inflammatory cytokines (tumour necrosis factor-α, interleukin 8, interleukin 1ß and interferon γ), while increasing transforming growth factor ß1 levels at both transcriptional and protein levels in the liver and intestine. The mRNA expression levels of mitogen-activated protein kinase 13 (MAPK 13), MAPK14 and nuclear factor kappa B p65 were significantly reduced in the liver and intestine of fish fed with 0.60 and 1.20 g/Kg SePCH compared to the controls. Histological sections also demonstrated that 0.60 and 1.20 g/Kg SePCH significantly increased intestinal villus height and villus width compared to the controls. Furthermore, the mRNA expression levels of tight junction proteins (zonula occludens-1, zonula occludens-3, Claudin-1, Claudin-3, Claudin-5, Claudin-11, Claudin-23 and Claudin-34) and Mucin-17 were significantly upregulated in the intestinal epithelial cells of 0.60 and 1.20 g/Kg SePCH groups compared to the controls. In conclusion, these results found that 0.60 and 1.20 g/Kg dietary SePCH can not only improve growth, hematological parameters, selenium metabolism, antioxidant capacities, enhance immune responses and intestinal functions, but also alleviate inflammatory responses. This information can serve as a useful reference for formulating feeds for largemouth bass.

Selenium yeast improve growth, serum biochemical indices, metabolic ability, antioxidant capacity and immunity in black carp Mylopharyngodnpiceus.

This experiment was conducted to investigate the impacts of dietary selenium yeast (SeY) on the growth performance, fish body composition, metabolic ability, antioxidant capability, immunity and inflammatory responses in juvenile black carp (Mylopharyngodn piceus). The base diet was supplemented with 0.00, 0.30 and 0.60 g/kg SeY (0.04, 0.59 and 1.15 mg/kg of selenium) to form three isonitrogenous and isoenergetic diets for juvenile black carp with a 60-day. Adequate dietary SeY (0.30 and 0.60 g/kg) could significantly increase the weight gain (WG), special growth rate (SGR) compared to the SeY deficient groups (0.00 g/kg) (P < 0.05). Meanwhile, 0.30 and 0.60 g/kg SeY elevated the mRNA levels of selenoprotein T2 (SEPT2), selenoprotein H (SEPH), selenoprotein S (SEPS) and selenoprotein M (SEPM) in the liver and intestine compared with the SeY deficient groups (P < 0.05). Adequate dietary SeY could promote glucose catabolism and utilization through activating glucose transport (GLUT2), glycolysis (GCK, HK, PFK, PK, PDH), tricarboxylic acid cycle (ICDH and MDH), glycogen synthesis (LG, GCS and GBE) and IRS/PI3K/AKT signal pathway molecules (IRS2b, PI3Kc and AKT1) compared with the SeY deficient groups (P < 0.05). Similarly, adequate dietary SeY could improve lipid transport and triglycerides (TG) synthesis through increasing transcription amounts of CD36, GK, DGAT, ACC and FAS in the fish liver compared with the SeY deficient groups (P < 0.05). In addition, adequate SeY could markedly elevate activities of antioxidant enzymes (T-SOD, CAT, GR, GPX) and contents of T-AOC and GSH, while increased transcription amounts of Nrf2, Cu/Zn-SOD, CAT, and GPX in fish liver and intestine (P < 0.05). However, adequate SeY notably decreased contents of MDA, and the mRNA transcription levels of Keap1 in the intestine compared with the SeY deficient groups (P < 0.05). Adequate SeY markedly increased amounts or levels of the immune factors (ALP, ACP, LZM, C3, C4 and IgM) and the transcription levels of innate immune-related functional genes in the liver and intestine (LZM, C3 and C9) compared to the SeY deficient groups (P < 0.05). Moreover, adequate SeY could notably reduce levels of IL-8, IL-1ß, and IFN-γ and elevate TGF-1ß levels in fish intestine (P < 0.05). The transcription levels of MAPK13, MAPK14 and NF-κB p65 were notably reduced in fish intestine treated with 0.30 and 0.60 g/kg SeY (P < 0.05). In conclusion, these results suggested that 0.30 and 0.60 g/kg SeY could not only improve growth performance, increase Se, glucose and lipid metabolic abilities, enhance antioxidant capabilities and immune responses, but also alleviate inflammation, thereby supplying useful reference for producing artificial feeds in black carp.

Effects of dietary berberine on growth performance, lipid metabolism, antioxidant capacity and lipometabolism-related genes expression of AMPK signaling pathway in juvenile black carp (Mylopharyngodon piceus) fed high-fat diets.

This study aimed to investigate the effects of high-fat diet (HFD) supplemented with berberine on growth, lipid metabolism, antioxidant capacity and lipometabolism-related genes expression of AMPK signaling pathway in juvenile black carp (Mylopharyngodon piceus). Five hundred and forty healthy fish (4.04 ± 0.01 g) were randomly distributed into six groups, and fed six experimental diets: normal-fat diet (NFD, 5% fat), HFD (15% fat), and four HFDs supplemented with graded levels of berberine, respectively. The results showed that, compared with fish fed NFD, HFD had no effects on the growth of fish except for reducing survival rate, whereas HFD caused extensive lipid accumulation, oxidative stress injury and hepatic abnormalities. However, compared with the HFD group, fish fed HFD containing an appropriate berberine (98.26 or 196.21 mg/kg) improved the growth performance, increased hepatic lipid metabolism and antioxidant enzymes activities, and up-regulated the mRNA expression levels of ampk subunits and lipolysis genes such as pparα, cpt-1, acox, atgl and hsl (P < 0.05). Meanwhile, HFD supplemented with an appropriate berberine reduced crude lipid contents in liver and whole-body, decreased serum lipid contents, and ALT and AST activities, and down-regulated the mRNA expression levels of lipogenesis genes such as srebp-1, acc1, gpat, fas and pparγ, and lipid transporter genes such as fatp, fabp and fat/cd36 (P < 0.05). Thus, HFD supplemented with an appropriate berberine could improve growth of black carp, promote lipid metabolism and enhance antioxidant capacity. The lipid-lowering mechanism of berberine might be mediated by activating AMPK pathway, up-regulating lipolysis genes expression, and down-regulating lipogenesis and transport genes expression.

Optimal dietary curcumin improved growth performance, and modulated innate immunity, antioxidant capacity and related genes expression of NF-κB and Nrf2 signaling pathways in grass carp (Ctenopharyngodon idella) after infection with Aeromonas hydrophila.

This study investigated the effects of dietary curcumin on growth performance, non-specific immunity, antioxidant capacity and related genes expression of NF-κB and Nrf2 signaling pathways in grass carp (Ctenopharyngodon idella). A total of 525 juvenile grass carps with mean initial body weight of (5.30 ± 0.10) g were randomly distributed into five groups with three replicates each, fed five diets containing graded levels of curcumin (0, 196.11, 393.67, 591.46 and 788.52 mg/kg diet) for 60 days. After feeding trial, fifteen fish per tank were challenged with Aeromonas hydrophila and the mortalities were recorded for 7 days. The results showed that optimal dietary curcumin (393.67 mg/kg diet) improved the weight gain (WG) and specific growth rate (SGR) of juvenile grass carp, reduced feed conversion ratio (FCR) and the mortalities after challenge (P < 0.05). Moreover, optimal dietary curcumin increased the activities of lysozyme (LYZ) and acid phosphatase (ACP), and complement 3 (C3) and C4 levels, decreased alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities in serum of grass carp after injection with A. hydrophila (P < 0.05). Meanwhile, optimal dietary curcumin up-regulated the mRNA levels of LYZ, C3 and antimicrobial peptides [hepcidin, liver-expressed antimicrobial peptide-2 (LEAP-2), ß-defensin], and anti-inflammatory cytokines of interleukin-10 (IL-10) and transforming growth factor ß1 (TGF-ß1), and inhibitor of κBα (IκBα), whereas down-regulated pro-inflammatory cytokines of tumor necrosis factor-α (TNF-α), IL-1ß, IL-6 and IL-8, and nuclear factor kappa B p65 (NF-κB p65), IκB kinases (IKKα, IKKß and IKKγ) mRNA levels in the liver and blood of grass carp after injection with A. hydrophila (P < 0.05). In addition, optimal dietary curcumin increased the reduced glutathione (GSH) content and activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione S-transferase (GST) and glutathione reductase (GR), reduced reactive oxygen species (ROS) and malondialdehyde (MDA) levels in the liver of grass carp after injection with A. hydrophila (P < 0.05). Meanwhile, optimal dietary curcumin up-regulated the mRNA levels of these antioxidant enzymes and nuclear factor erythroid 2-related factor 2 (Nrf2), whereas down-regulated Kelch-like ECH-associated protein (Keap) 1a and Keap 1b mRNA levels (P < 0.05) in the liver and blood of grass carp after injection with A. hydrophila. Thus, optimal dietary curcumin supplementation could promote growth of juvenile grass carp, reduce FCR, and enhance disease resistance, innate immunity and antioxidant capacity of fish, attenuating inflammatory response. However, dietary excessive curcumin had negative effect on fish. Based on second-order regression analysis between dietary curcumin contents and weight gain, the optimum requirement of dietary curcumin in juvenile grass carp was determined to be 438.20 mg/kg diet.

Growth, antioxidant capacity, intestinal morphology, and metabolomic responses of juvenile Oriental river prawn (Macrobrachium nipponense) to chronic lead exposure.

Understanding the mechanisms of metal toxicity to organisms farmed for food may suggest mitigation strategies. We determined the 24-, 48-, 72-, and 96-h median lethal concentrations of lead in juvenile oriental river prawn (Macrobrachium nipponense). The prawns were then exposed to sub-lethal concentrations (13.13 and 26.26 µg/L) of lead for 60 days and growth, antioxidant enzyme activity, intestinal morphology, and metabolite profiles were assessed. Prawns exposed to 26.26 µg/L but not to 13.13 µg/L lead exhibited lower weight gain than controls. The lead burden in muscle was 0.067 and 0.25 µg/g of dry weight exposed to 13.13 and 26.26 µg/L, respectively. Levels of glutamic oxaloacetic transaminase and glutamic-pyruvic transaminase were not altered following exposure. Exposure increased malondialdehyde activity in the hepatopancreas and decreased superoxide dismutase and glutathione peroxidase activities. Catalase activity first increased and then decreased as lead concentrations increased. Some intestinal epithelial cells disassociated from the basement membrane in prawns exposed to 13.13 µg/L lead. Intestinal epithelial cells in prawns exposed to 26.26 µg/L lead separated completely from the basement membrane. Gas chromatography-mass spectrometry metabolomics assays showed the 13.13-µg/L exposure did not elicit significant metabolic alterations. Exposure to 26.26 µg/L lead differentially up-regulated 58 metabolites and down-regulated 21 metabolites. The metabolites identified were involved in galactose, purine, glutathione, and carbon metabolism, biosynthesis of amino acids and steroids, and neuroactive ligand-receptor interaction. These data indicate that chronic lead exposure can adversely affect growth, increase accumulation in muscle, impair intestinal morphology, and induce oxidant stress or neurotoxicity-related effects in M. nipponense.

Identification, characterization, and expression analysis of adiponectin receptors in black carp Mylopharyngodon piceus in response to dietary carbohydrate.

In the present study, three cDNAs of AdipoRs (MpAdipoR1a, MpAdipoR1b, and MpAdipoR2) were identified from juvenile black carp Mylopharyngodon piceus. There were 375, 378, and 356 amino acids in the MpAdipoR1a, MpAdipoR1b, and MpAdipoR2, respectively. BLAST analysis reveals that MpAdipoRs share high identities with other known AdipoRs from zebrafish, rainbow trout, human, etc. And there were all seven transmembrane regions in the amino acid sequences of MpAdipoR1s and MpAdipoR2, respectively. The relative expression levels of MpAdipoR1s were higher in the liver, blood, brain, and eyes in black carp (p < 0.05). Relatively higher expression of MpAdipoR2 was detected in the liver (p < 0.05) and then in the adipose tissues and blood by real-time PCR assays. The relative expression levels of AdipoR mRNA in the liver, muscle, brain, blood, and adipose tissues were detected by real-time PCR in black carp fed with four levels of dietary carbohydrate (CHO) (10.65, 19.43, 28.84, and 37.91%) for 9 weeks, respectively. The expression levels of MpAdipoR1s in the liver, muscle, brain, and blood were induced and reached to the maximum at optimal dietary CHO (19.43 or 28.84%) and then were decreased at 37.91% dietary CHO, although there were no significant differences on the expression levels of MpAdipoR1a in the liver between 19.43, 28.84, and 37.91% dietary CHO. Similarly, MpAdipoR2 were also induced to the maximal levels in the liver, muscle, brain, and blood at optimal dietary CHO (19.43 or 28.84%) compared with that at 10.65% dietary CHO, although no significant differences were observed on the expression levels of MpAdipoR2 in the liver and muscle between groups fed 19.43 and 37.91% dietary CHO (p > 0.05). However, the expression levels of MpAdipoRs in the adipose tissues were significantly downregulated at excessive dietary CHO (37.91%) compared with other groups (p < 0.05). These results indicated that MpAdipoRs were inducible proteins and might enhance the understanding of their vital roles in the regulation of glucose metabolic homeostasis in black carp.

Effect of trichlorfon on hepatic lipid accumulation in crucian carp Carassius auratus gibelio.

This study evaluated the toxic effects of the organophosphate pesticide trichlorfon on hepatic lipid accumulation in crucian carp Carassius auratus gibelio. Seventy-five fish were divided into five groups (each group in triplicate), and then exposed to 0, 0.5, 1.0, 2.0, and 4.0 mg/L of trichlorfon and fed with commercial feed for 30 d. At the end of the experiment, plasma and hepatic lipid metabolic biochemical status were analyzed. Triglyceride contents were significantly (P < 0.05) increased in liver but decreased in plasma after 1.0, 2.0, and 4.0 mg/L trichlorfon treatments. Plasma insulin contents were markedly (P < 0.05) increased when trichlorfon concentrations were 0.5, 1.0, and 4.0 mg/L. There were no significant differences in hepatic hormone-sensitive lipase contents between the trichlorfon-treated fish and the controls. Hepatic cyclic adenosine 3', 5'-monophosphate, very-low-density lipoprotein, and apolipoprotein B100 contents were decreased in the fish when trichlorfon concentration was 2.0 mg/L. Furthermore, electron microscope observations showed rough endoplasmic reticulum dilatation and mitochondrial vacuolization in hepatocytes with trichlorfon exposure. On the basis of morphological and physiological evidence, trichlorfon influenced crucian carp hepatic pathways of lipid metabolism and hepatocellular ultrastructure, which resulted in lipid accumulation in the liver.