Characterizing and identifying of miRNAs involved in berberine modulating glucose metabolism of Megalobrama amblycephala.

The present study, as one part of a larger project that aimed to investigate the effects of dietary berberine (BBR) on fish growth and glucose regulation, mainly focused on whether miRNAs involve in BBR's modulation of glucose metabolism in fish. Blunt snout bream Megalobrama amblycephala (average weight of 20.36 ± 1.44 g) were exposed to the control diet (NCD, 30% carbohydrate), the high-carbohydrate diet (HCD, 43% carbohydrate) and the berberine diet (HCB, HCD supplemented with 50 mg/kg BBR). After 10 weeks' feeding trial, intraperitoneal injection of glucose was conducted, and then, the plasma and liver were sampled at 0 h, 1 h, 2 h, 6 h, and 12 h. The results showed the plasma glucose levels in all groups rose sharply and peaked at 1 h after glucose injection. Unlike the NCD and HCB groups, the plasma glucose in the HCD group did not decrease after 1 h, while remained high level until at 2 h. The NCD group significantly increased liver glycogen content at times 0-2 h compared to the other two groups and then liver glycogen decreased sharply until at times 6-12 h. To investigate the role of BBR that may cause the changes in plasma glucose and liver glycogen, miRNA high-throughput sequencing was performed on three groups of liver tissues at 2 h time point. Eventually, 20 and 12 differentially expressed miRNAs (DEMs) were obtained in HCD vs NCD and HCB vs HCD, respectively. Through function analyzing, we found that HCD may affect liver metabolism under glucose loading through the NF-κB pathway; and miRNAs regulated by BBR mainly play roles in adipocyte lipolysis, niacin and nicotinamide metabolism, and amino acid transmembrane transport. In the functional exploration of newly discovered novel:Chr12_18892, we found its target gene, adenylate cyclase 3 (adcy3), was widely involved in lipid decomposition, amino acid metabolism, and other pathways. Furthermore, a targeting relationship of novel:Chr12_18892 and adcy3 was confirmed by double luciferase assay. Thus, BBR may promote novel:Chr12_18892 to regulate the expression of adcy3 and participate in glucose metabolism.

Carrier Transport Regulation of Pixel Graphene Transparent Electrodes for Active-Matrix Organic Light-Emitting Diode Display.

Integrating a graphene transparent electrode (TE) matrix with driving circuits is essential for the practical use of graphene in optoelectronics such as active-matrix organic light-emitting diode (OLED) display, however it is disabled by the transport of carriers between graphene pixels after deposition of a semiconductor functional layer caused by the atomic thickness of graphene. Here, the carrier transport regulation of a graphene TE matrix by using an insulating polyethyleneimine (PEIE) layer is reported. The PEIE forms an ultrathin uniform film (≤10 nm) to fill the gap of the graphene matrix, blocking horizontal electron transport between graphene pixels. Meanwhile, it can reduce the work function of graphene, improving the vertical electron injection through electron tunneling. This enables the fabrication of inverted OLED pixels with record high current and power efficiencies of 90.7 cd A-1 and 89.1 lm W-1 , respectively. By integrating these inverted OLED pixels with a carbon nanotube-based thin-film transistor (CNT-TFT)-driven circuit, an inch-size flexible active-matrix OLED display is demonstrated, in which all OLED pixels are independently controlled by CNT-TFTs. This research paves a way for the application of graphene-like atomically thin TE pixels in flexible optoelectronics such as displays, smart wearables, and free-form surface lighting.

Pushing the conductance and transparency limit of monolayer graphene electrodes for flexible organic light-emitting diodes.

Graphene has emerged as an attractive candidate for flexible transparent electrode (FTE) for a new generation of flexible optoelectronics. Despite tremendous potential and broad earlier interest, the promise of graphene FTE has been plagued by the intrinsic trade-off between electrical conductance and transparency with a figure of merit (σDC/σOp) considerably lower than that of the state-of-the-art ITO electrodes (σDC/σOp <123 for graphene vs. ∼240 for ITO). Here we report a synergistic electrical/optical modulation strategy to simultaneously boost the conductance and transparency. We show that a tetrakis(pentafluorophenyl)boric acid (HTB) coating can function as highly effective hole doping layer to increase the conductance of monolayer graphene by sevenfold and at the same time as an anti-reflective layer to boost the visible transmittance to 98.8%. Such simultaneous improvement in conductance and transparency breaks previous limit in graphene FTEs and yields an unprecedented figure of merit (σDC/σOp ∼323) that rivals the best commercial ITO electrode. Using the tailored monolayer graphene as the flexible anode, we further demonstrate high-performance green organic light-emitting diodes (OLEDs) with the maximum current, power and external quantum efficiencies (111.4 cd A-1, 124.9 lm W-1 and 29.7%) outperforming all comparable flexible OLEDs and surpassing that with standard rigid ITO by 43%. This study defines a straightforward pathway to tailor optoelectronic properties of monolayer graphene and to fully capture their potential as a generational FTE for flexible optoelectronics.

Dietary T-2 toxin induces transcriptomic changes in hepatopancreas of Chinese mitten crab (Eriocheir sinensis) via nutrition metabolism and apoptosis-related pathways.

Long-term feed route exposure to T-2 toxin was proved to elicit growth retarding effects and induction of oxidative stress and apoptosis in Chinese mitten crab (Eriocheir sinensis). However, no study with a holistic perspective has been conducted to date to further describe the in-depth toxicological mechanism of T-2 toxin in E.sinensis. In this study, an RNA-Sequencing (RNA-seq) was used in this study to investigate the effects of feed supplementation with 0 mg/kg and 4 mg/kg T-2 toxin on the hepatopancreas transcriptome of E.sinensis and establish a hepatopancreas transcriptome library of T-2 toxin chronically exposed crabs after five weeks, where 14 differentially expressed genes (DEGs) were screened out across antioxidant, apoptosis, autophagy, glucolipid metabolism and protein synthesis. The actual expression of all the DEGs (Caspase, ATG4, PERK, ACSL, CAT, BIRC2, HADHA, HADHB, ACOX, PFK, eEFe1, eIF4ɑ, RPL13Ae) was also analyzed by real-time quantitative PCR (RT-qPCR). It was demonstrated that long-term intake of large amounts of T-2 toxin could impair antioxidant enzyme activity, promote apoptosis and protective autophagy, disrupt lipid metabolism and inhibit protein synthesis in the hepatopancreas of E.sinensis. In conclusion, this study explored the toxicity mechanism of T-2 toxin on the hepatopancreas of E.sinensis at the mRNA level, which lays the foundation for further investigation of the molecular toxicity mechanism of T-2 toxin in aquatic crustaceans.

Acute exposure to microcystin-LR induces hepatopancreas toxicity in the Chinese mitten crab (Eriocheir sinensis).

The Chinese mitten crab is an important economic species in the Chinese aquaculture industry due to its rich nutritional value and distinct flavor. The hepatopancreas is a popular edible part of the Chinese mitten crab, and therefore, hepatopancreatic health directly determines its quality. However, a large-scale outbreak of hepatopancreatic necrosis syndrome ("Shuibiezi" disease in Chinese), which is caused by abiotic agents correlated with cyanobacteria bloom outbreaks, adversely affects the Chinese mitten crab breeding industry. Cyanobacterial blooms that occur in high-density farming ponds can produce microcystin-LR (MC-LR), which is hepatotoxic in fish and mammals. Hepatopancreas toxicity of MC-LR (0, 25, 50 and 75 µg/kg) was investigated after 48 h of exposure. The MC-LR can cause hepatopancreatic injury by inducing hepatopancreatic structural damage, subcellular structural changes, and cell apoptosis, followed by enhanced lipid peroxidase, reactive oxygen species, and apoptosis-related enzyme (Caspase 3, 8, and 9) activities. These in turn promote gene and protein expression of apoptosis-associated proteases (Caspase 3, 7, and 8, Bcl-2, and Bax), and alter antioxidant system responses (superoxide dismutase, glutathione S-transferase, glutathione peroxidase, glutathione reductase activities, and glutathione content). The present study is the first report on MC-LR hepatotoxicity in the Chinese mitten crab and confirms hepatopancreas toxicity, providing a theoretical basis for enhancing MCs resistance and developing preventive and curative measures against hepatopancreatic disease in the Chinese mitten crab breeding industry.

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.

Protective effects of dietary arginine against oxidative damage and hepatopancreas immune responses induced by T-2 toxin in Chinese mitten crab (Eriocheir sinensis).

T-2 toxin is a secondary metabolite produced by Fusarium spp. that is a major cereal and animal feed contaminant. T-2 toxin has numerous adverse effects on animals, including hepatotoxicity. Arginine (Arg) is closely associated with the regulation of immune responses and antioxidant activity in tissues. The objective of the present study was to evaluate the protective effects of dietary Arg against oxidative damage and immune responses of the hepatopancreas induced by T-2 toxin in Chinese mitten crab. According to the results, 3.17% Arg in the diet decreased alanine aminotransferase, aspartate aminotransferase, and alkaline phosphatase activity in the haemolymph significantly, when compared with the levels of activity in the T-2 toxin group. Arg supplementation also increased superoxide dismutase and glutathione peroxidase activity, while decreasing malondialdehyde concentrations in the hepatopancreas, when compared with the levels in the T-2 toxin group. In addition, 3.17% Arg in the diet increased acid phosphatase and alkaline phosphatase activity in the hepatopancreas, as well as albumin concentrations in the haemolymph, when compared with the T-2 toxin group. Dietary Arg also regulated the expression of antioxidant enzyme-related genes (mitochondrial manganese superoxide dismutase, cytosolic manganese superoxide dismutase, and catalase) and immune related genes (prophenoloxidase, NF-κB-like transcription factor Relish, and lipopolysaccharide-induced TNF-α factor) to alleviate the damage associated with the T-2 toxin. Furthermore, Arg ameliorated damage to the hepatopancreas microstructure in the crabs. The results of the present study indicate that dietary Arg could enhance the antioxidant and immune capacity of Chinese mitten crab against oxidative damage and immune injury to the hepatopancreas induced by T-2 toxin.

Dietary reduced glutathione supplementation can improve growth, antioxidant capacity, and immunity on Chinese mitten crab, Eriocheir sinensis.

Eriocheir sinensis is an important aquaculture species in China, and its yield and quality are threatened by oxidative stress caused by deteriorating water conditions. Reduced glutathione (GSH) is an endogenous antioxidant, but whether dietary GSH can increase the resistance of E. sinensis to environmental stress remains unclear. Therefore, in this study, crabs were fed with dietary GSH (0, 300, 600, 900, and 1200 mg/kg diet weight) for up to 10 weeks to determine the effects of different dietary GSH concentrations on growth, antioxidant capacity, and immunity of E. sinensis. The results showed that the weight gain rate and survival rate increased significantly as dietary GSH levels increased from 0 to 900 mg/kg, but decreased at 1200 mg/kg. Compared with the control group, the diet supplemented with 900 mg/kg GSH not only increased the concentration of GSH in the haemolymph and hepatopancreas, but also enhanced the activity of glutathione peroxidase (GSH-Px) (p < 0.05). Diets supplemented with 600 or 900 mg/kg GSH significantly increased the enzymes activities of superoxide dismutase (SOD), lysozyme (LZM), alkaline phosphatase, and acid phosphatase, and significantly decreased the content of malondialdehyde. To understand the changes in the activity of these enzymes further, the expression of related genes was detected. Diets supplemented with 600 or 900 mg/kg GSH significantly upregulated the genes expressions of cytosolic manganese SOD, mitochondrial manganese SOD, copper, zinc-SOD, GSH-Px, LZM, and prophenoloxidase activating factor, and significantly down regulated the expression of Toll-like receptor 1, Toll-like receptor 2, Dorsal, and the myeloid differentiation factor 88. However, a diet supplemented with 1200 mg/kg GSH decreased those positive indicators. Overall, our results demonstrated that an appropriate diet supplemented with 600 or 900 mg/kg GSH enhances antioxidant capacity and immunity, which will enhance the general health of E. sinensis.

Dietary glutathione supplementation enhances antioxidant activity and protects against lipopolysaccharide-induced acute hepatopancreatic injury and cell apoptosis in Chinese mitten crab, Eriocheir sinensis.

Eriocheir sinensis (E. sinensis) is an important aquaculture species in China. However, deteriorating water environments lead to oxidative stress in these crabs, which subsequently reduces their quality and yield. Glutathione (GSH) is an endogenous antioxidant that is used to mitigate oxidative stress. However, whether dietary GSH can enhance the resistance of E. sinensis to oxidative stress remains unclear. Herein, crabs were fed dietary GSH (the basal diet was supplemented with 0, 300, 600, 900, and 1200 mg/kg diet weight of GSH) for up to 3 weeks and, then, challenged with lipopolysaccharide (LPS; 400 µg/kg body weight). After 6 h, their hepatopancreas were sampled. Diet supplementation with 600 and 900 mg/kg diet weight GSH not only increased the content of GSH in the hepatopancreas, but also enhanced the activities and mRNA expressions of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and glutathione-S-transferase (GST) (P < 0.05), compared to that in control crabs challenged with LPS alone. Diet supplementation with 600 or 900 mg/kg GSH also significantly increased the enzyme activities of GSH reductase and γ-glutamyl cysteine synthetase (γ-GCS) in LPS-treated crabs. Haematoxylin-eosin (HE) staining, electron microscopy, and flow cytometry were used to examine the structure and subcellular structure of and apoptosis in the hepatopancreas. The histopathology and sub-microstructure analysis results also showed that diet supplementation with 600 or 900 mg/kg GSH significantly alleviated damage in crabs challenged with LPS and decreased reactive oxygen species (ROS) levels and cell apoptosis ratios in the hepatopancreas, compared to the LPS-treated crabs. To further understand the effect of dietary GSH on LPS-induced apoptosis, the activities and gene or protein expressions of apoptosis-related factors were evaluated. As a result, diet supplementation with 600 or 900 mg/kg GSH significantly decreased the activities of caspases-3, -8, and -9 and inhibited the relative expression of caspase-3 and -8 but increased the expression of B-cell lymphoma-2 (bcl-2) and B-cell lymphoma-2-associated X inhibitor (bax inhibitor) in crabs challenged with LPS. This treatment further significantly downregulated the relative protein levels of caspase-3, -8, -9 and Bax and upregulated those of Bcl-2 in crabs challenged with LPS. However, treatment with 1200 mg/kg GSH caused the opposite effects. Overall, our results reveal that appropriate diets supplemented with 600 or 900 mg/kg GSH could enhance the antioxidant capacity and anti-apoptotic mechanisms in crabs after LPS injection, thereby providing a theoretical basis for the application of dietary GSH in E. sinensis.

Influences of glycyrrhetinic acid (GA) dietary supplementation on growth, feed utilization, and expression of lipid metabolism genes in channel catfish (Ictalurus punctatus) fed a high-fat diet.

An 8-week feeding trial was performed to test the effects of glycyrrhetinic acid (GA) on growth and some gene expression of hepatic lipid metabolism in channel catfish (initial body weight, 3.5 ± 0.02 g) fed high-fat diets. Fish were fed the control diet, high-fat diet (HFD), and HFD supplemented with 0.4, 0.8, and 1.2 mg/kg GA in 15 tanks at a stocking density of 21 fish/tank. Fish fed HFD were significantly lower in body weight gain and specific growth rate but higher in feed intake and feed conversion ratio in comparison to the control. Supplement of GA at 1.2 mg/kg remarkably improved these parameters as compared to the control diet. High levels of cholesterol (TC), triglycerides (TG), low-density lipoprotein (LDL), and low levels of high-density lipoprotein (HDL) in plasma were observed in fish fed HFD; the opposite was observed for fish fed HFD supplemented with GA. The transcription of fatty acid synthase (FAS), sterol regulatory element-binding protein-1 (SREBP1), liver X receptor alpha (LXRα), and hormone-sensitive lipase (HSL) was upregulated, while that of carnitine palmitoyltransferase I (CPT1), peroxisome proliferator-activated receptors alpha (PPARα), acyl-CoA oxidase (ACO), and microsomal triglyceride transfer protein (MTTP) mRNA expression were downregulated in fish fed HFD. The opposite was observed in fish fed HFD supplemented with GA as well as the control group. In conclusion, supplementing the HFD with GA at 1.2 mg/kg could improve the growth performance and lipid metabolism of channel catfish consuming HFD.

Dietary fructooligosaccharide can mitigate the negative effects of immunity on Chinese mitten crab fed a high level of plant protein diet.

An 8-week feeding trial was carried out under controlled condition to evaluate the effect of dietary fructooligosaccharide (FOS) on growth performance, whole body composition, antioxidant status and immunity of crabs fed high levels of plant protein diets. Thus, six experimental diets were formulated (designated as F0P50, F0P60, F0P70, F0.2P50, F0.2P60 and F0.2P70), which contain two FOS levels (0 or 0.2%) and three plant protein levels (50, 60, or 70%) according to a 2 × 3 factorial design. The results showed that weight gain increased significantly as dietary plant protein level decreased from 70% to 50%. At 50% plant protein level, the addition of 0.2% FOS can significantly elevate weight gain (WG) (P < 0.05). The highest value in survival rate was observed in crabs fed F0.2P50 and F0.2P60 diet. Crabs fed F0.2P50 diet showed significantly higher crude protein content (P < 0.05) compared with those in other groups, but there were no significant differences in the contents of moisture, crude lipid and ash among all groups (P > 0.05). Catalase (CAT) activity in crabs fed F0.2P50 increased significantly (P < 0.05) compared with crabs fed F0P60, F0P70, F0.2P60 and F0.2P70, but malondialdehyde (MDA) concentrations decreased significantly (P < 0.05). Meanwhile, nitric oxide (NO) concentration, acid phosphatase (ACP) and alkaline phosphatase (AKP) activities of crabs fed 0.2% FOS diets increased significantly (P < 0.05) compared with crabs fed 0% FOS diets. The expressions of prophenoloxidase (propo) was significantly (P < 0.05) affected only by dietary plant protein levels with the highest values observed in 50% plant protein diet, whereas the opposite was true for Myeloid differentiation factor 88 (myd88). The mRNA expressions of mitochondrial manganese superoxide dismutase (mtmnsod), lipopolysaccharide-induced TNF-α factor (litaf) and toll like receptors (tlrs) were significantly affected (P < 0.05) by both FOS and plant protein levels. The cytosolic manganese superoxide dismutase (cytmnsod) mRNA expressions in F0.2P50 and F0.2P60 groups were significantly higher than those in F0P70 and F0.2P70 groups. The results in this study indicated that supplementation with 0.2% FOS can enhance growth performance in crabs fed lower plant protein diets and as well improve immunity in those fed with higher plant protein diets.

Effects of dietary protein level on growth performance, digestive enzyme activity, and gene expressions of the TOR signaling pathway in fingerling Pelteobagrus fulvidraco.

An 8-week feeding trial was conducted to investigate effects of dietary protein levels (37, 40, and 43%) on the growth performance, feed utilization, digestive enzyme activity, and gene expressions of target of rapamycin (TOR) signaling pathway in fingerling yellow catfish. One hundred and eighty fingerlings (average weight 0.77 ± 0.03 g) were equally distributed across four replicate tanks for each of the three treatments, with 15 fish per tank. No difference (P > 0.05) was observed in initial body weight, survival rate (SR), hepatosomatic index (HSI), viscera index (VSI), dressing percentage (DP), and condition factor (CF) among all the treatments. The diet containing 40% protein increased significantly (P < 0.05) final body weight, weight gain rate (WGR), specific growth rate (SGR), protein efficiency ratio (PER), nitrogen retention (NRE), and energy retention (ERE) in fish. The highest protease activity in the stomach and intestine was observed in the P40 group (P < 0.05), while amylase and lipase were not significantly different (P > 0.05). The transcriptional levels of IGF-1, IGF-1R, and Akt were significantly (P < 0.05) higher in fish fed P40 or P43 than those of fish fed P37. TOR and S6K1 mRNA expressions were significantly (P < 0.05) increased in the P40 groups. Hence, the diet containing 40% protein would be suitable for the optimum growth and effective protein utilization of fingerling Pelteobagrus fulvidraco. In vitro, the transcriptional levels of IGF-1, IGF-1R, Akt, TOR, and S6K1 in hepatocyte supplemented with a 40-µM mixed amino acids were significantly (P < 0.05) higher compared to other treatments. No difference (P > 0.05) was observed in eukaryotic translation initiation factor 4E-binding protein 1 in vivo and in vitro among all the treatments. Effects of dietary protein level on growth performance likely are involved in the activation of TOR signaling pathway in fingerling Pelteobagrus fulvidraco.

Selection of reference genes for miRNA quantitative PCR and its application in miR-34a/Sirtuin-1 mediated energy metabolism in Megalobrama amblycephala.

MiRNAs are small, non-coding RNAs that downregulate gene expression at post-transcriptional levels. They have emerged as important regulators involved in metabolism, immunity, and cancer. Real-time quantitative PCR (RT-qPCR) is an effective and main method for quantifying target miRNA. For robust RT-qPCR method, suitable reference genes play crucial roles in data normalization. Blunt snout bream (Megalobrama amblycephala) is an economically important aquaculture species; however, no reference genes dedicated for qPCR method has been identified for this species so far. The objective of this study was to screen stable reference genes for miRNA RT-qPCR and demonstrated its application in energy metabolism in blunt snout bream. The stabilities of ten potential reference genes (miR-21-1-5p, miR-107a-3p, miR-222a-3p, miR-146a-5p, miR-101a-3p, miR-22a-3p, miR-103-3p, miR-456-3p, miR-221-3p, and U6 (RNU6A)) were evaluated in nine tissues (brain, muscle, liver, skin, spleen, heart, gill, intestine, and eye) under normal condition and in three tissues (liver, intestine, and spleen) under four stresses (heat stress, ammonia stress, bacterial challenge, and glycolipid stress). Using GeNorm, NormFinder, and RefFinder softwares, we discovered that different tissues and stresses are both important variability factors for the expression stability of miRNAs. After verifying miR-34a/Sirtuin-1 expressions in high-carbohydrate diet-induced blunt snout bream, we eventually identified that the most stable reference gene in this species was miR-221-3p, and the best combination of reference genes were miR-221-3p and miR-103-3p.

Glucose-6-phosphate dehydrogenase in blunt snout bream Megalobrama amblycephala: molecular characterization, tissue distribution, and the responsiveness to dietary carbohydrate levels.

This study aimed to characterize the full-length cDNA of glucose-6-phosphate dehydrogenase (G6PD) from Megalobrama amblycephala with its responses to dietary carbohydrate levels characterized. The cDNA obtained covered 2768 bp with an open reading frame of 1572 bp. Sequence alignment and phylogenetic analysis revealed a high degree of conservation (77-97%) among most fish and other higher vertebrates. The highest transcription of G6PD was observed in kidney followed by liver, whereas relatively low abundance was detected in eye. Then, the transcriptions and activities of G6PD as well as lipid contents were determined in the liver, muscle, and the adipose tissue of fish fed two dietary carbohydrate levels (30 and 42%) for 12 weeks. Hepatic transcriptions of fatty acid synthetase (FAS), acetyl-CoA carboxylase α (ACCα), sterol regulatory element-binding protein-1 (SREBP1), and peroxisome proliferator-activated receptor γ (PPARγ) were also measured to corroborate the lipogenesis derived from carbohydrates. The G6PD expressions and activities in both liver and the adipose tissue as well as the lipid contents in whole-body, liver, and the adipose tissue all increased significantly after high-carbohydrate feeding. Hepatic transcriptions of FAS, ACCα, SREBP1, and PPARγ were also up-regulated remarkably by the intake of a high-carbohydrate diet. These results indicated that the G6PD of M. amblycephala shared a high similarity with that of other vertebrates. Its expressions and activities in tissues were both highly inducible by high-carbohydrate feeding, as also held true for the transcriptions of other enzymes and/or transcription factors involved in lipogenesis, evidencing an enhanced lipogenesis by high dietary carbohydrate levels.

Fishmeal replacement by rice protein concentrate with xylooligosaccharides supplement benefits the growth performance, antioxidant capability and immune responses against Aeromonas hydrophila in blunt snout bream (Megalobrama amblycephala).

This study aimed to investigate the effects of fishmeal (FM) replacement by rice protein concentrate (RPC) with a xylooligosaccharides supplement on the growth performance, antioxidant capability and immune response against Aeromonas hydrophila in blunt snout bream (Megalobrama amblycephala). Fish (46.85 ±â€¯0.34 g) were randomly assigned to one of 6 diets, namely the control diet (containing FM), the RPC diet (FM replaced by RPC) and RPC diet supplemented with 0.5, 1.5, 2.3 and 3% XOS respectively, for 8 weeks. After the feeding trial, fish were challenged by Aeromonas hydrophila for 96 h with the blood and liver sample obtained at 48 and 96 h respectively. The results showed that the final weight, weight gain and protein efficiency ratio of fish fed RPC diet were significantly (P < 0.05) lower than that of the control group, whereas the opposite was true for FCR. However, the supplement of 1.5% XOS remarkably (P < 0.05) improved these parameters compared to the control diet. Plasma total iron binding capacity of fish fed the RPC diet showed little difference (P > 0.05) with that of the control group, but it enhanced significantly (P < 0.05) with the supplement of 1.5% XOS compared to the control group. After bacterial infection, plasma lysozyme (LYM), complement 3, complement 4, myeloperoxidase (MPO), acid phosphatase (ACP) and alkaline phosphatase activities, as well as immunoglobulin M, levels all increased significantly (P < 0.05) with the maximum value is attained at 48 h, then they decreased significantly (P < 0.05) with further increasing time at 96 h. Similar results were also observed in liver superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX) activities as well as malondialdehyde (MDA) content. Regarding dietary treatment, these parameters of fish fed RPC diet showed little difference (P > 0.05) with those fed the control diet but were significantly enhanced (P < 0.05) when RPC diet was supplemented with 1.5-2.3% XOS, except for hepatic (MDA) content which showed an opposite trend compared to the control group. After 96 h of challenge, the relative percentage survival (RPS) of fish fed XOS was significantly higher (P < 0.05) than that of fish offered the control and RPC diet. In addition, significant (P < 0.05) interactions between dietary XOS and sampling time were also observed in plasma LYM, MPO, ACP, and hepatic SOD, CAT, GPX, MDA, as well as RPS. In conclusion, this study indicated that dietary supplementation of 1.5%XOS could significantly improve the growth performance, antioxidant capability, innate immunity and A. hydrophila resistance of blunt snout bream fed diets with FM replaced by RPC.

Effects of dietary glucose and starch levels on the growth, apparent digestibility, and skin-associated mucosal non-specific immune parameters in juvenile blunt snout bream (Megalobrama amblycephala).

A 10-week feeding trial was performed to evaluate the effects of different types and levels of carbohydrates in growth performance, apparent digestibility coefficients and skin-associated mucosal non-specific immune parameters in blunt snout bream (Megalobrama amblycephala). Fish were randomly fed four diets containing two carbohydrates (glucose and starch) diets and two carbohydrates levels (330 and 440 g kg-1). High carbohydrate levels remarkably increased the weight gain rate (WGR), apparent digestibility of dry matters, protein and carbohydrates, body crud protein content, plasma levels of aspartate transaminase (AST), and skin-associated mucosal levels of immunoglobulin M (IgM), HDL cholesterol, lysozyme (LZM), advanced the transcriptions of mucin 2 (Muc2), mucin 5b (Muc5b) and apolipoprotein A-I (apoA-I), whereas the opposite was true for feed conversion ratio (FCR), plasma levels of IgM, skin-associated mucosal levels of major histocompatibility complex (MHC) and ß-Defensins, and the transcriptions of heat shock protein 60 (Hsp60). In addition, carbohydrate types of glucose remarkably increased the survival rate, apparent digestibility of dry matters, protein and carbohydartes, body crud ash, plasma levels of total protein (TP), globulin (GLB), immunoglobulin M (IgM), complement C3 and complement C4 and the transcriptions of Muc5b. Whereas the carbohydrate types of starch remarkably increased viscerosomatic index (VSI), hepatosomatic index (HSI), condition factor (CF), abdominal fat percentage (AFP), apparent digestibility of liquid, advanced the transcriptions of Muc2, apoA-I and heat shock protein 70 (Hsp70). Significant interactions between different types and levels of dietary carbohydrates were also observed in WGR, apparent digestibility of dry matters, protein and liquid, body crud ash, plasma levels of TP, albumin (ALB) and AST, skin-associated mucosal levels of major histocompatibility complex (MHC) and ß-Defensins, and the transcriptions of Muc2 and Muc5b. Our results indicate that inclusion of high level of glucose in the diet of blunt snout bream could improve growth performance, nonspecific immunity, and increase the efficiency of protein, which is suggesting that high level of glucose could be used in feed production. However, the proportion of the specific formula of glucose using in feed needs further study.

The mechanism of action of a fat regulator: Glycyrrhetinic acid (GA) stimulating fatty acid transmembrane and intracellular transport in blunt snout bream (Megalobrama amblycephala).

High-fat diets are associated with fatty liver and aberrant hepatic lipid metabolism, and glycyrrhetinic acid (GA) has been shown to exert a beneficial effect on lipolysis and fat deposition in fish. In the present study, we evaluated the effect of GA on the growth performance and expression of hepatic lipid transport related genes in blunt snout bream (Megalobrama amblycephala) fed a high fat diet. Two hundred and sixteen fish (average body weight: 45.57 g ±â€¯0.98 g) were fed three experimental diets (6% fat/L6 group, control, 11% fat/L11 group, and 11% fat with 0.3 mg kg-1 GA/L11GA group) for 8 weeks. Compared to the control group, the weight gain and specific growth rate of high-fat fed group at the end of the trialwere significantly improved (P < .05).However, GA showed no effect on animals' growth performance(P > .05). Dietary supplementation with 0.3 mg kg-1 GA significantly decreased the hepatosomatic index, viscera/body ratio, and intraperitoneal fat ratio (P < .05), and up-regulated the expression levels of fatty acids transport protein (FATP), fatty acids binding protein (FABP), fatty acid translocase (CD36), carnitine palmitoyl transferase I (CPT1) and peroxisome proliferators-activated receptors α (PPARα) compared to both the L6 group and L11 group (P < .05). However, no significant difference was observed in fatty acid synthetase (FAS), acetyl-CoA carboxylase α (ACCα), or lipoprotein lipase (LPL) between groups (P > .05). In conclusion, GA significantly rescued high-fat diet induced hepatic lipid accumulation and metabolism dysfunction in M. amblycephalaby stimulating hepatic fatty acid transport and ß-oxidation. Dietary GA may be used as a promising supplement to alleviate high-fat diet induced side effects on M. amblycephala.

Resveratrol supplementation improves lipid and glucose metabolism in high-fat diet-fed blunt snout bream.

Here, we aimed to investigate whether resveratrol (RSV) can ameliorate high-fat diet (HFD)-induced metabolic disorder in fish. Blunt snout bream (Megalobrama amblycephala) with average weight 27.99 ± 0.56 g were fed a normal fat diet (NFD, 5% fat, w/w), a HFD (11% fat), or a HFD supplemented with 0.04, 0.36, or 1.08% RSV for 10 weeks. As expected, fish fed a HFD developed hepatic steatosis, as shown by elevated hepatic and plasma triglycerides, raised whole body fat, intraperitoneal fat ratio and hepatosomatic index, and increased plasma alanine aminotransferase (ALT) and aspartate aminotransferase (AST). RSV supplementation lessened increases in body mass, whole body fat, and intraperitoneal fat, and alleviated development of hepatic steatosis, elevations of plasma triglyceride and glucose, and abnormalities of ALT and AST in HFD-fed fish. RSV supplementation increased SIRT1 messenger RNA (mRNA) expression and consequently hepatic mRNA expression of adipose triglyceride lipase (ATGL), carnitine palmitoyltransferase (CPT1a), and microsomal triglyceride transfer protein (MTTP), implying upregulation of lipolysis, ß-oxidation, and lipid transport, respectively, in the liver. Conversely, hepatic lipoprotein lipase (LPL), sterol regulatory element-binding protein 1 (SREBP-1c), peroxisome proliferator-activated receptor γ (PPARγ), and ATP citrate lyase (ACLY) mRNA expression were decreased, implying suppression of fatty acid uptake, lipogenesis, and fatty acid synthesis. Additionally, RSV downregulated glucokinase (GCK) and sodium-dependent glucose cotransporter 1 (SGLT1) and upregulated glucose transporter 2 (GLUT2) mRNA expression, thus restoring normal glucose fluxes. Thus, RSV improves lipid and glucose metabolisms in blunt snout bream, which are potentially mediated by activation of SIRT1.

Molecular characterization of AMP-activated protein kinase α2 from herbivorous fish Megalobrama amblycephala and responsiveness to glucose loading and dietary carbohydrate levels.

This study aimed to characterize the full-length cDNA of AMPKα2 in Megalobrama amblycephala, and evaluate its potential role in glucose homeostasis and carbohydrate metabolism. The cDNA obtained covered 1942bp with an open reading frame of 1635bp encoding 545 amino acids. Multiple alignments and phylogenetic analysis revealed a high homology (91-100%) among most fish and higher vertebrates. This AMPKα2 mRNA predominantly expressed in muscle, liver and brain, while little in gill and intestine. Then, the AMPKα2 expressions were determined in the muscle, liver and brain of fish subjected to a glucose load (injected intraperitoneally with 0, 1.67 and 3.34g glucose per kg body weight) and after a 12-week feeding trial (fed two dietary carbohydrate levels: 30% and 43%), respectively. After the glucose load, plasma glycemia peaked at 1h in fish. Thereafter, it decreased significantly to the basal level at 8h. However, AMPKα2 expression in muscle, liver and brain all decreased significantly during the first 2h, then returned to the basal value at 24h. Unlikely, tissue AMPKα2 expression of fish receiving saline solution increased significantly during the whole sampling period. Additionally, high-carbohydrate diet enhanced its expression in liver and muscle, but not that in brain. These findings indicated that the AMPKα2 gene shared a high degree of conservation with that of the other vertebrates. Muscle, liver and brain AMPKα2 expressions were highly induced by glucose administration. Furthermore, high dietary carbohydrate modified its expressions in these tissues.

Effect of dietary betaine on growth performance, antioxidant capacity and lipid metabolism in blunt snout bream fed a high-fat diet.

An 8-week feeding experiment was conducted to determine the effect of dietary betaine levels on the growth performance, antioxidant capacity, and lipid metabolism in high-fat diet-fed blunt snout bream (Megalobrama amblycephala) with initial body weight 4.3 ± 0.1 g [mean ± SEM]. Five practical diets were formulated to contain normal-fat diet (NFD), high-fat diet (HFD), and high-fat diet with betaine addition (HFB) at difference levels (0.6, 1.2, 1.8%), respectively. The results showed that the highest final body weight (FBW), weight gain ratio (WGR), specific growth rate (SGR), condition factor (CF), and feed intake (FI) (P < 0.05) were obtained in fish fed 1.2% betaine supplementation, whereas feed conversion ratio (FCR) was significantly lower in the same group compared to others. Hepatosomatic index (HSI) and abdominal fat rate (AFR) were significantly high in fat group compared to the lowest in NDF and 1.2% betaine supplementation, while VSI and survival rate (SR) were not affected by dietary betaine supplementation. Significantly higher (P < 0.05), plasma total cholesterol (TC), triglycerides (TG), low-density lipoprotein (LDL), aspartate transaminase (AST), alanine transaminase (ALT), cortisol, and lower high-density lipoprotein (HDL) content were observed in HFD but were improved when supplemented with 1.2% betaine. In addition, increase in superoxide dismutase (SOD), catalase (CAT), and reduced glutathione (GSH) in 1.2% betaine inclusion could reverse the increasing malondialdehyde (MDA) level induced by HFD. Based on the second-order polynomial analysis, the optimum growth of blunt snout bream was observed in fish fed HFD supplemented with 1.2% betaine. HFD upregulated fatty acid synthase messenger RNA (mRNA) expression and downregulated carnitine palmitoyltransferase 1, peroxisome proliferator-activated receptor α, and microsomal triglyceride transfer protein mRNA expression; nevertheless, 1.2% betaine supplementation significantly reversed these HFD-induced effects, implying suppression of fatty acid synthesis, ß-oxidation, and lipid transport. This present study indicated that inclusion of betaine (1.2%) can significantly improve growth performance and antioxidant defenses, as well as reduce fatty acid synthesis and enhance mitochondrial ß-oxidation and lipid transportation in high-fat diet-fed blunt snout bream, thus effectively alleviating fat accumulation in the liver by changing lipid metabolism.