The role of vitamin E in polyunsaturated fatty acid synthesis and alleviating endoplasmic reticulum stress in sub-adult grass carp (Ctenopharyngodon idella).

Vitamin E (VE) is an essential lipid-soluble vitamin that improves the fish flesh quality. However, the underlying molecular mechanisms remain unclear. This study aimed to investigate the effects of VE on growth performance and flesh quality in sub-adult grass carp (Ctenopharyngodon idella). A total of 450 fish (713.53 ± 1.50 g) were randomly divided into six treatment groups (three replicates per treatment) and fed for nine weeks with different experimental diets (dietary lipid 47.8 g/kg) that contained different levels of VE (5.44, 52.07, 96.85, 141.71, 185.66, and 230.12 mg/kg diet, supplemented as dl-α-tocopherol acetate). Notably, the treatment groups that were fed with dietary VE ranging from 52.07 to 230.12 mg/kg diet showed improvement in the percent weight gain, special growth rate, and feed efficiency of grass carp. Moreover, the treatment groups supplemented with dietary VE level of 141.71, 185.66, and 230.12 mg/kg diet showed enhancement in crude protein, lipid, and α-tocopherol contents in the muscle, and the dietary levels of VE ranging from 52.07 to 141.71 mg/kg diet improved muscle pH24h and shear force but reduced muscle cooking loss in grass carp. Furthermore, appropriate levels of VE (52.07 to 96.85 mg/kg diet) increased the muscle polyunsaturated fatty acid content in grass carp. Dietary VE also increased the mRNA levels of fatty acid synthesis-related genes, including fas, scd-1, fad, elovl, srebp1, pparγ, and lxrα, and up-regulated the expression of SREBP-1 protein. However, dietary VE decreased the expression of fatty acid decomposition-related genes, including hsl, cpt1, acox1, and pparα, and endoplasmic reticulum stress-related genes, including perk, ire1, atf6, eif2α, atf4, xbp1, chop, and grp78, and down-regulated the expression of p-PERK, p-IRE1, ATF6, and GRP78 proteins. In conclusion, dietary VE increased muscle fatty acid synthesis, which may be partly associated with the alleviation of endoplasmic reticulum stress, and ultimately improves fish flesh quality. Moreover, the VE requirements for sub-adult grass carp (713.53 to 1590.40 g) were estimated to be 124.9 and 122.73 mg/kg diet based on percentage weight gain and muscle shear force, respectively.

Emerging role of vitamin D3 in alleviating intestinal structure injury caused by Aeromonas hydrophila in grass carp (Ctenopharyngodon idella).

Bacterial pathogens destroy the structural integrity of functional organs in fish, leading to severe challenges in the aquaculture industry. Vitamin D3 (VD3) prevents bacterial infections and strengthens immune system function via vitamin D receptor (VDR). However, the correlation between VD3/VDR and the structural integrity of functional organs remains unclarified. This study aimed to investigate the influence of VD3 supplementation on histological characteristics, apoptosis, and tight junction characteristics in fish intestine during pathogen infection. A total of 540 healthy grass carp (257.24 ± 0.63 g) were fed different levels of VD3 (15.2, 364.3, 782.5, 1,167.9, 1,573.8, and 1,980.1 IU/kg) for 70 d. Subsequently, fish were challenged with Aeromonas hydrophila, a pathogen that causes intestinal inflammation. Our present study demonstrated that optimal supplementation with VD3 (1) alleviated intestinal structural damage, and inhibited oxidative damage by reducing levels of oxidative stress biomarkers; (2) attenuated excessive apoptosis-related death receptor and mitochondrial pathway processes in relation to p38 mitogen-activated protein kinase signaling (P < 0.05); (3) enhanced tight junction protein expression by inhibiting myosin light chain kinase signaling (P < 0.05); and (4) elevated VDR isoform expression in fish intestine (P < 0.05). Overall, the results demonstrated that VD3 alleviates oxidative injury, apoptosis, and the destruction of tight junction protein under pathogenic infection, thereby strengthening pathogen defenses in the intestine. This finding supports the rationale for VD3 intervention as an essential practice in sustainable aquaculture.

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.

Tea polyphenols protect against Flavobacterium columnare-induced gill injury via suppression of oxidative stress, inflammation, and apoptosis in grass carp.

Flavobacterium columnare (F. columnare) is one of the deadliest fish pathogens causing bacterial gill rot disease in various freshwater fish species globally. Tea polyphenols (TPs) are an inexpensive product extracted from tea that have received much attention as a feed additive in aquaculture. The current study was designed to investigate the underlying mechanisms and protective effects of dietary TPs against F. columnare-induced gill injury via suppression of oxidative stress, apoptosis, and inflammation in grass carp. TPs were not supplemented to the diet (control) and were supplemented at 40, 80, 120, 160 or 200 mg/kg diet. The feeding experiment was carried out for 60 days, followed by a 3-Day F. columnare challenge test. The results showed that 120 mg/kg TPs in the diet exerted the following five protective effects in fish gill: (1) control gill-rot disease and improved histopathology, (2) inhibit excessive apoptosis, (3) enhance the activity of antioxidant enzymes and upregulate related gene expression via the Nrf2/Keap1 pathway, (4) increase the activity of immune enzymes, And (5) mediate inflammatory cytokine gene expression via the JAK/STAT3 pathway. Taken together, dietary supplementation with TPs is a compelling approach to protect the gill function of fish against F. columnare.

Implications of vitamin D for flesh quality of grass carp (Ctenopharyngodon idella): antioxidant ability, nutritional value, sensory quality, and myofiber characteristics.

BACKGROUND: Muscle represents a unique and complex system with many components and comprises the major edible part of animals. Vitamin D is a critical nutrient for animals and is known to enhance calcium absorption and immune response. In recent years, dietary vitamin D supplementation in livestock has received increased attention due to biological responses including improving shear force in mammalian meat. However, the vitamin D acquisition and myofiber development processes in fish differ from those in mammals, and the effect of vitamin D on fish flesh quality is poorly understood. Here, the influence of dietary vitamin D on fillet quality, antioxidant ability, and myofiber development was examined in grass carp (Ctenopharyngodon idella). METHODS: A total of 540 healthy grass carp, with an initial average body weight of 257.24 ± 0.63 g, were allotted in 6 experimental groups with 3 replicates each, and respectively fed corresponding diets with 15.2, 364.3, 782.5, 1,167.9, 1,573.8, and 1,980.1 IU/kg vitamin D for 70 d. RESULTS: Supplementation with 1,167.9 IU/kg vitamin D significantly improved nutritional value and sensory quality of fillets, enhancing crude protein, free amino acid, lipid, and collagen contents; maintaining an ideal pH; and reducing lactate content, shear force, and cooking loss relative to respective values in the control (15.2 IU/kg) group. Average myofiber diameter and the frequency of myofibers > 50 µm in diameter increased under supplementation with 782.5-1,167.9 IU/kg vitamin D. Levels of oxidative damage biomarkers decreased, and the expression of antioxidant enzymes and nuclear factor erythroid 2-related factor 2 signaling molecules was upregulated in the 1,167.9 IU/kg vitamin D treatment compared to respective values in the control group. Furthermore, vitamin D supplementation activated cell differentiation by enhancing the expression of myogenic regulatory factors and myocyte enhancer factors compared to that in the control group. In addition, supplementation with 1,167.9 IU/kg vitamin D improved protein deposition associated with protein synthesis molecule (target of rapamycin) signaling and vitamin D receptor paralogs, along with inhibition of protein degradation (forkhead box protein 1) signaling. CONCLUSIONS: Overall, the results demonstrated that vitamin D strengthened antioxidant ability and myofiber development, thereby enhancing nutritional value and sensory quality of fish flesh. These findings suggest that dietary vitamin D supplementation is conducive to the production of nutrient-rich, high quality aquaculture products.

Aflatoxin B1 damaged structural barrier through Keap1a/Nrf2/ MLCK signaling pathways and immune barrier through NF-κB/ TOR signaling pathways in gill of grass carp (Ctenopharyngodon idella).

Aquafeeds are susceptible to contamination caused by aflatoxin B1 (AFB1). The gill of fish is an important respiratory organ. However, few studies have investigated the effects of dietary AFB1 exposure on gill. This study aimed to discuss the effects of AFB1 on the structural and immune barrier of grass carp gill. Dietary AFB1 increased reactive oxygen species (ROS) levels, protein carbonyl (PC) and malondialdehyde (MDA) contents, which consequently caused oxidative damage. In contrast, dietary AFB1 decreased antioxidant enzymes activities, relative genes expression (except MnSOD) and the contents of glutathione (GSH) (P < 0.05), which are partly regulated by NF-E2-related factor 2 (Nrf2/Keap1a). Moreover, dietary AFB1 caused DNA fragmentation. The relative genes of apoptosis (except Bcl-2, McL-1 and IAP) were significantly upregulated (P < 0.05), and apoptosis was likely upregulated through p38 mitogen-activated protein kinase (p38MAPK). The relative expressions of genes associated with tight junction complexes (TJs) (except ZO-1 and claudin-12) were significantly decreased (P < 0.05), and TJs were likely regulated by myosin light chain kinase (MLCK). Overall, dietary AFB1 disrupted the structural barrier of gill. Furthermore, AFB1 increased gill sensitivity to F. columnare, increased Columnaris disease and decreased the production of antimicrobial substances (P < 0.05) in grass carp gill, and upregulated the expression of genes involved with pro-inflammatory factors (except TNF-α and IL-8) and the pro-inflammatory response partly attributed to the regulation by nuclear factor κB (NF-κB). Meanwhile, the anti-inflammatory factors were downregulated (P < 0.05) in grass carp gill after challenge with F. columnare, which was partly attributed to the target of rapamycin (TOR). These results suggested that AFB1 aggravated the disruption of the immune barrier of grass carp gill after being challenge with F. columnare. Finally, the upper limit of safety of AFB1 for grass carp, based on Columnaris disease, was 31.10 µg/kg diet.

Cellular antioxidant mechanism of mannan-oligosaccharides involving in enhancing flesh quality in grass carp (Ctenopharyngodon idella).

BACKGROUND: Deterioration of flesh quality has bad effects on consumer satisfaction. Therefore, effects of safe mannan-oligosaccharides (MOS) on flesh quality of grass carp (Ctenopharyngodon idella) muscle were studied. A total of 540 healthy fish (215.85 ± 0.30 g) were randomly divided into six groups and fed six separate diets with graded levels of MOS (0, 200, 400, 600, 800 and 1000 mg kg-1 ) for 60 days. This study aimed at investigating the benefits of dietary MOS on flesh quality (fatty acids, amino acids and physicochemical properties) and the protection mechanism regarding antioxidant status. RESULTS: Optimal MOS could improve tenderness (27.4%), pH (5.5%) while decreasing cooking loss (16.6%) to enhance flesh quality. Meanwhile, optimal MOS improved flavor inosine 5'-monophosphate (IMP) of 11.8%, sweetness and umami-associated amino acid, healthy unsaturated fatty acid (UFA) of 14.9% and n-3 polyunsaturated fatty acids (n-3 PUFAs) especially C20:5n-3 (15.8%) and C22:6n-3 (38.3%). Furthermore, the mechanism that MOS affected pH, tenderness and cooking loss could be partly explained by the reduced lactate, cathepsin and oxidation, respectively. The enhanced flesh quality was also associated with enhanced antioxidant ability concerning improving antioxidant enzymes activities and the corresponding transcriptional levels, which were regulated through NF-E2-related factor 2 (Nrf2) and target of rapamycin (TOR) signaling. Based on pH24h , cooking loss, shear force and DHA (docosahexaenoic acid, C22:6n-3), optimal MOS levels for grass carp were estimated to be 442.75, 539.53, 594.73 and 539.53 mg kg-1 , respectively. CONCLUSION: Dietary MOS is a promising alternative strategy to improve flesh quality of fish muscle. © 2022 Society of Chemical Industry.

Methionine played a positive role in improving the intestinal digestion capacity, anti-inflammatory reaction and oxidation resistance of grass carp, Ctenopharyngodon idella, fry.

A study was carried out to appraisal the function of methionine on intestinal digestion and the health of grass carp (Ctenopharyngodon idella) fry (initial weight 0.36 ± 0.01 g). The fry were fed graded dietary methionine levels (0.33%-1.20% dry matter) in 18 recirculatory tanks (180 L). After an 8-week breeding experiment, the results revealed that 0.71%-1.20% dietary methionine levels markedly upregulated the mRNA levels of intestinal digestion including trypsin, amylase, chymotrypsin and AKP, and 0.71%-0.87% dietary methionine level significantly increased intestinal trypsin activities compared with the 0.33% dietary methionine level. For inflammation, 0.71%-1.20% dietary methionine levels downregulated the mRNA levels of NF-κBp65, IL-1ß, IL-6, IL-8, IL-15 and IL-17D, whereas upregulated the mRNA levels of anti-inflammatory cytokines, including IL-4/13B, IL-10 and IL-11. In terms of antioxidants, although dietary methionine levels had no significant effect on the expression of most core genes of the Nrf2/ARE signaling pathway, such as Nrf2, Keap 1, GPx4, CAT, Cu/Zn-SOD. Furthermore, dietary methionine levels had no significant effect on the expression of p38MAPK, IL-12p35, TGF-ß2 and IL-4/13A. 0.71%-1.20% dietary methionine levels still increased the mRNA levels of GPx1α, GSTR and GSTP1. Furthermore, higher intestinal catalase activity and glutathione contents were also observed in fry fed 0.71%-1.20% diets. In summary, 0.71%-1.20% dietary methionine levels played a positive role in improving the intestinal digestion capacity of digestion, anti-inflammatory reaction and oxidation resistance of grass carp fry. This study provided a theoretical basis for improving the survival rate and growth of grass carp fry.

New Insight on the Immune Modulation and Physical Barrier Protection Caused by Vitamin A in Fish Gills Infected With Flavobacterium columnare.

In this study, we have investigated the influence of vitamin A on gill barrier function of grass carp (Ctenopharyngodon idella) infected with Flavobacterium columnare. The fish were fed different concentrations of vitamin A diets for 10 weeks and then infected with F. columnare by immersion. We observed that optimal vitamin A significantly prevented gill rot morbidity in fish infected with F. columnare. Further investigations revealed that vitamin A boosted the gill immunity by increasing the contents of complements (C3 and C4), activities of acid phosphatase (ACP) and lysozyme, mRNAs of ß-defensin-1, liver-expressed antimicrobial peptide 2A and 2B (LEAP-2A and LEAP-2B), hepcidin, and anti-inflammatory cytokines like transforming growth factor ß1 (TGF-ß1), TGF-ß2, interleukin-10 (IL-10), and IL-11. It also enhanced the levels of various related signaling molecules including inhibitor protein κBα (IκBα), target of rapamycin (TOR), and ribosome protein S6 kinase 1 (S6K1) but downregulated the expression of pro-inflammatory cytokines including IL-1ß, IL-8, tumor necrosis factor α (TNF-α), and interferon γ2 (IFN-γ2) and related signaling molecules including nuclear factor κB p65 (NF-κB p65) (rather than NF-κB p52), IκB kinase ß (IKKß), IKKγ (rather than IKKα), eIF4E-binding protein 1 (4E-BP1), and 4E-BP2 mRNA levels in fish gills. In addition, dietary vitamin A markedly lowered the concentrations of reactive oxygen species (ROS), malondialdehyde (MDA), and protein carbonyl (PC), increased both the activities and mRNAs of copper/zinc superoxide dismutase (Cu/ZnSOD), MnSOD, glutathione transferases (GSTs), glutathione peroxidase (GPx), and glutathione reductase (GR) associated with upregulation of NF-E2-related factor 2 (Nrf2) mRNAs and downregulation of Kelch-like-ECH-associated protein (Keap1a) and Keap1b mRNAs. Moreover, vitamin A decreased the mRNAs of different apoptotic mediators [caspases 8, 9, 3 (rather than 7)] associated with downregulation of signaling molecule p38 mitogen-activated protein kinase (p38MAPK) mRNAs in fish gills. Besides, vitamin A promoted tight junction (TJ) complex mRNAs [including claudin-b, -c, -3, -7, -12, occludin, and zonula occludens-1 (ZO-1)] that have been linked to the downregulation of myosin light chain kinase (MLCK) signaling. Taken together, the current study demonstrated for the first time that vitamin A markedly enhanced gill health associated with immune modulation and physical barrier protection. Based on protecting fish against gill rot morbidity, ACP activity, and against lipid peroxidation, optimum vitamin A concentrations in on-growing grass carp (262-997 g) were found to be 1,991, 2,188, and 2,934 IU/kg diet, respectively.

A study of the potential effect of yellow mealworm (Tenebrio molitor) substitution for fish meal on growth, immune and antioxidant capacity in juvenile largemouth bass (Micropterus salmoides).

This study aimed to evaluate the effects of partial replacement of fish meal (FM) with yellow mealworm (Tenebrio molitor, TM) on the growth performance, food utilization and intestinal immune response of juvenile largemouth bass (Micropterus salmoides). Seven diets containing increasing levels of TM (FM substitution) were designed (approximately 0% (0%), 4% (11.1%), 8.1% (22.2%), 12.2% (33.3%), 16.3% (44.4%), 20.4% (55.5%), and 24.5% (66.6%), designated TM0, TM11, TM22, TM33, TM44, TM55, and TM66, respectively). 420 fish were randomly selected and placed in 21 cages (1 m*1 m*1 m, 7 treatments for triplicate, 20 fish per cage). Fish (initial weight 6.25 ± 0.03 g) were fed seven isonitrogenous (47%) and isocaloric (19 MJ kg-1) diets to satiety twice daily for 8 weeks. Compared to the control group (TM0), TM11 showed no significant difference in the weight gain rate (WGR), specific growth rate (SGR) or feed conversion ratio (FCR), while all other TM inclusion groups presented different degrees of decline. There was no significant difference in the whole-body composition among all groups (P > 0.05). Plasma total protein (TP), triglyceride (TG) and albumin (ALB) contents were significantly decreased in TM55 and TM66 (P < 0.05). The highest plasma aspartate transaminase (AST) activity was observed in TM66 (P < 0.05). TM33, TM44 and TM55 showed the lowest activities of plasma alanine amiotransferase (ALT) and alkaline phosphatase (ALP) (P < 0.05). Moreover, increased mRNA levels of superoxide dismutase (SOD) and catalase (CAT) were measured in the TM11 to TM55 groups, while intestinal SOD activity peaked in TM11 (P < 0.05). With the exception of TM11, the other TM inclusion groups showed significant inhibition of the relative expression of RelA, C3 and TNF-α (P < 0.05). All experimental groups exhibited lower expression of IL-10 than TM0 (P < 0.05). The TM11 group showed significantly upregulated expression of IL-1ß and TGF-ß (P < 0.05). In addition, TLR2 expression was increased in TM11 and TM22 (P < 0.05). Considering enzyme activities and immune-related gene expression, TM supplementation levels should not exceed 4% (TM11).

The role of dietary chromium supplementation in relieving heat stress of juvenile blunt snout bream Megalobrama amblycephala.

The present study assessed the role of dietary chromium (Cr) supplementation in relieving heat stress (HS) of juvenile blunt snout bream Megalobrama amblycephala. The supplemented Cr contents by chromium picolinate (Cr-Pic) was 0 mg/kg (control group), 0.4 mg/kg, 1.6 mg/kg and 12.0 mg/kg, respectively. The fish continued to be fed four diets at suitable temperatures (26 °C) for 2 weeks, and then the temperature was then heated up to 33 °C through thermo-regulated system. The results showed that Cr supplementation had no significant effect on the immune indices and antioxidant indices before HS (P > 0.05). However, Cr supplementation played an important role in relieving HS. After HS, compared with the control group, 1.6 mg/kg and 12.0 mg/kg Cr supplementation groups significantly lowered the plasma glucose level and aspartate transaminase (AST) activity (P < 0.05), and 0.4 mg/kg and 1.6 mg/kg Cr supplementation groups significantly lowered alanine aminotransferase (ALT) activity (P < 0.05). 0.4 mg/kg and 1.6 mg/kg supplementation groups significantly improved hepatic total superoxide dismutase (T-SOD) activity (P < 0.05). Furthermore, 0.4mg/kg-12.0 mg/kg Cr supplementation groups significantly improved the activities of hepatic glutathione peroxidase (GPx) and catalase (CAT) and lowered hepatic malondialdehyde (MDA) level in liver (P < 0.05). The mRNA levels of hepatic copper zinc superoxide dismutase (Cu/Zn-SOD), CAT and GPx were significantly improved in 0.4mg/kg-12.0 mg/kg supplementation Cr groups (P < 0.05), however, there was no significant variation of hepatic manganese superoxide dismutase (Mn-SOD) mRNA levels under different levels of supplementation (P > 0.05). Significantly lower mRNA levels of hepatic pro-inflammatory cytokines observed in 0.4mg/kg-12.0 mg/kg Cr supplementation groups including tumour necrosis factor-α (TNF-α), interleukin 1ß (IL-1ß) and interleukin 8 (IL-8) (P < 0.05), and 0.4mg/kg-12.0 mg/kg Cr supplementation significantly improved the relative expressions of hepatic heat shock protein 70 (HSP70) and heat shock protein 90 (HSP90) (P < 0.05). The present study indicated that dietary Cr supplementation might have no significant effect on immune capacity and antioxidant capacity under normal physiological conditions, whereas it played an important role in relieving HS.

Dietary berberine can ameliorate glucose metabolism disorder of Megalobrama amblycephala exposed to a high-carbohydrate diet.

Blunt snout bream (Megalobrama amblycephala) were randomly assigned into three diets: normal-carbohydrate diet (NCD, 30% carbohydrate, w/w), high-carbohydrate diet (HCD, 43% carbohydrate), and HCB (HCD supplemented with 50 mg/kg berberine (BBR)). After 10 weeks' feeding trial, the results showed that higher levels of plasma glucose, triglyceride, and total cholesterol were observed in HCD-fed fish than in NCD-fed fish, while HCB feeding significantly ameliorated this effect. Moreover, HCB feeding remarkably reversed HCD-induced hepatic glycogen and lipid contents. In insulin signaling, BBR inclusion restored HCD-induced suppression of insulin receptor substrate mRNA expression and elevation of forkhead transcription factor 1 mRNA expression. In glucose metabolism, upregulated glucose transporter 2 and glycogen synthase mRNA expressions in the HCD group were observed compared to the NCD group. However, BBR adding reduced the mRNA expressions of glycogen synthase, phosphoenolpyruvate carboxykinase, and glucose-6-phosphatase and increased the transcriptional levels of glucose transporter 2 and pyruvate kinase. In lipid metabolism, BBR supplementation could reverse downregulated hepatic carnitine palmitoyl transferase I mRNA expression and upregulated hepatic acetyl-CoA carboxylase and fatty acid synthetase mRNA expressions in the HCD group. Taken together, it demonstrates that BBR could improve glucose metabolism of this species via enhancing liver's glycolysis and insulin signaling, while inhibiting liver's glycogen synthesis and gluconeogenesis. It also indicates that BBR could reduce the metabolic burden of the liver by inhibiting fat synthesis and promoting lipid decomposition, and then enhance fat uptake in peripheral tissues.

The immunoreaction and antioxidant capacity of juvenile blunt snout bream (Megalobrama amblycephala) involves the PI3K/Akt/Nrf2 and NF-κB signal pathways in response to dietary methionine levels.

A 75-day rearing trail was designed to evaluate the immunoreaction and antioxidant capacity of juvenile blunt snout bream in response to dietary methionine levels. Three practical diets were extruded to feed juveniles with graded methionine levels (0.40%, 0.84% and 1.28% dry matter). The data indicated that the plasma concentrations of immunoglobulin M (IgM), complement component 3 (C3) and glutathione (GSH) in the 0.84% methionine diet were markedly upper than those in the 0.40% group (P < 0.05). The activities of plasma antioxidant parameters involving catalase (CAT), total superoxide dismutase (T-SOD), total antioxidant capacity (T-AOC) and glutathione peroxidase (GPx) were significantly increased by the 0.84% diet compared with the 0.40% diet, whereas plasma alanine aminotransferase (ALT) and malondialdehyde (MDA) levels were significantly induced by 0.40% methionine (P < 0.05). Compared with the 0.40% group, 0.84% dietary methionine dramatically upregulated the mRNA expression levels of protein kinase B (Akt), phosphoinositide 3-kinase (PI3K) and nuclear factor erythroid 2-related factor 2 (Nrf2) pathway related genes including CAT, manganese superoxide dismutase (Mn-SOD), heme oxygenase 1 (HO-1) and glutathione peroxidase-1 (GPx-1) in the kidney and liver, and downregulated Kelch-like ECH-associated protein 1 (Keap1) mRNA levels (P < 0.05). Compared with the 0.40% group, the 0.84% dietary methionine strikingly suppressed the mRNA levels of renal and hepatic nuclear factor-kappa B (NF-κB) and pro-inflammatory cytokines (interleukin 1ß (IL-1ß), tumor necrosis factor-α (TNF-α), interleukin 6 (IL-6)), however, improved the mRNA expression levels of anti-inflammatory cytokines involved renal and hepatic transforming growth factor-ß (TGF-ß) and hepatic interleukin 10 (IL-10) (P < 0.05). Renal IL-10 and interleukin 8 (IL-8) mRNA expression levels were not markedly influenced by experimental diets (P > 0.05). Dietary methionine (0.84%) significantly upregulated renal and hepatic heat stress protein 70 (Hsp70), renal B-cell lymphoma-2 (Bcl-2) gene expression levels compared with the 0.40% diet (P < 0.05). In a word, the data represented that 0.84% dietary methionine could enhance the immune and antioxidant capacity of this fish species by inducing PI3K/Akt/Nrf2 pathway and inhibiting NF-κB pathway.