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Health benefits, flavour quality indicators and physical properties were analysed after feeding grass carp graded concentrations of soybean isoflavones (SIF) (0, 25, 50, 75, 100 and 125 mg/kg) for 60 days. The results demonstrated that optimal dietary SIF supplementation improved the protein and total PUFA content, especially healthcare n-3 PUFA (C18: 3n-3, EPA and DHA), and increased the flavour-related free amino acid [especially umami amino acid] and 5'-inosine monophosphate content, improving the health benefits and flavour quality indicators in the muscle of grass carp. In addition, optimal dietary SIF supplementation (25 or 50 mg SIF/kg diet) enhanced some physical properties [water-holding capacity and tenderness] and increased the collagen content; however, it reduced cathepsin activity and apoptosis. SIF supplementation enhanced the glutathione content and the activity of antioxidant enzymes (except CuZnSOD) by regulating their gene expression. The gene expression could be regulated by NF-E2-related factor 2 (Nrf2) signalling in the muscle of grass carp. We demonstrated that optimal dietary SIF supplementation elevated the health benefits, flavour quality indicators and physical properties of fish muscle. [ABSTRACT FROM AUTHOR]

This study was conducted to examine the effects of dietary valine on the physical and flavor characteristics, fatty acid (FA) profile, antioxidant status and Nrf2-dependent antioxidant enzyme gene expression in the muscle of young grass carp (Ctenopharyngodon idella) fed increasing levels of valine (4.3, 8.0, 10.6, 13.1, 16.9 and 19.1 g/kg) for 8 weeks. Compared with the control group, the group fed valine showed improved physical characteristics of fish fillets (increased relative shear force, hydroxyproline, protein and lipid levels and decreased cathepsin B and L activities, as well as cooking loss, were observed). Moreover, valine improved the flavor of young grass carp fillets by increasing the amino acid (AA) concentration in fish muscle (increased aspartic acid, threonine, glutamine, cystine, methionine, leucine, tyrosine, phenylalanine, lysine, histidine, arginine and valine concentrations were observed). Additionally, optimal valine supplementation increased the potential health benefits to humans by decreasing the saturated FA (C15:0 and C16:0) concentration and increasing the unsaturated FA (monounsaturated FAs (MUFAs), such as C16:1, C18:1c+t and C20:1, and polyunsaturated FAs (PUFAs), such as C18:3n-3, C20:2 and C22:6) concentration. In addition, the reduced glutathione (GSH) content and the activities of Cu/Zn superoxide dismutase (SOD1), catalase (CAT) and Selenium-dependent glutathione peroxydase (Se-GPx) increased under valine supplementation (P < 0.05). Furthermore, the SOD1, CAT and Se-GPx mRNA levels increased with dietary valine levels, possibly due to the up-regulation of NF-E2-related factor 2 (Nrf2), target of rapamycin (TOR) and ribosomal protein S6 kinase 1 (S6K1) and the down-regulation of Kelch-like-ECH-associated protein 1 (Keap1) in muscle (P < 0.05). In conclusion, valine improved the physical and flavor characteristics, FA profile, and antioxidant status and regulated the expression of the antioxidant enzyme genes Nrf2, Keap1, TOR and S6K1 in fish fillets. [ABSTRACT FROM AUTHOR]

The liver and intestine are susceptible to the oxidative damage which could result in several diseases. Choline deficiency induced oxidative damage in rat liver cells. Thus, this study aimed to investigate the potential molecular mechanisms responsible for choline deficiency-induced oxidative damage. Juvenile Jian carp were fed diets differing in choline content [165 (deficient group), 310, 607, 896, 1167 and 1820 mg/kg diet] respectively for 65 days. Oxidative damage, antioxidant enzyme activities and related gene expressions in the hepatopancreas and intestine were measured. Choline deficiency decreased choline and phosphatidylcholine contents, and induced oxidative damage in both organs, as evidenced by increased levels of oxidative-stress markers (malondialdehyde, protein carbonyl and 8-hydroxydeoxyguanosine), coupled with decreased activities of antioxidant enzymes [Copper-zinc superoxide dismutase (CuZnSOD), manganese superoxide dismutase (MnSOD), glutathione peroxidase (GPx) and glutathione-S-transferase (GST)]. However, choline deficiency increased glutathione contents in the hepatopancreas and intestine. Furthermore, dietary choline deficiency downregulated mRNA levels of MnSOD, GPx1b, GST-rho, mGST3 and Kelch-like ECH associating protein 1 (Keap1b) in the hepatopancreas, MnSOD, GPx1b, GPx4a, GPx4b, GST-rho, GST-theta, GST-mu, GST-alpha, GST-pi and GST-kappa in the intestine, as well as intestinal Nrf2 protein levels. In contrast, choline deficiency upregulated the mRNA levels of GPx4a, GPx4b, mGST1, mGST2, GST-theta, GST-mu, Keap1a and PKC in the hepatopancreas, mGST3, nuclear factor erythoid 2-related factor 2 (Nrf2) and Keap1a in the intestine, as well as hepatopancreatic Nrf2 protein levels. This study provides new evidence that choline deficiency-induced oxidative damage is associated with changes in the transcription of antioxidant enzyme and Nrf2/Keap1 signaling molecules in the hepatopancreas and intestine. Additionally, this study firstly indicated that choline deficiency induced varied change patterns of different GPx and GST isoforms. Meanwhile, the changes of some GPx and GST isoforms caused by choline deficiency in the intestine were contrary to those in the hepatopancreas. [ABSTRACT FROM AUTHOR]

Copper (Cu) is a common heavy metal pollutant in aquatic environments that originates from natural as well as anthropogenic sources. The present study investigated whether Cu causes oxidative damage and induces changes in the expression of genes that encode tight junction (TJ) proteins, cytokines and antioxidant-related genes in the intestine of the grass carp (Ctenopharyngodon idella). We demonstrated that Cu decreases the survival rate of fish and increases oxidative damage as measured by increases in malondialdehyde and protein carbonyl contents. Cu exposure significantly decreased the expression of genes that encode the tight junction proteins, namely, claudin (CLDN)-c, -3 and -15 as well as occludin and zonula occludens-1, in the intestine of fish. In addition, Cu exposure increases the mRNA levels of the pro-inflammatory cytokines, specifically, IL-8, TNF-α and its related signalling factor (nuclear factor kappa B, NF-κB), which was partly correlated to the decreased mRNA levels of NF-κB inhibitor protein (IκB). These changes were associated with Cu-induced oxidative stress detected by corresponding decreases in glutathione (GSH) content, as well as decreases in the copper, zinc-superoxide dismutase (SOD1) and glutathione peroxidase (GPx) activities and mRNA levels, which were associated with the down-regulated antioxidant signalling factor NF-E2-related factor-2 (Nrf2) mRNA levels, and the Kelch-like-ECH-associated protein1 (Keap1) mRNA levels in the intestine of fish. Histidine supplementation in diets (3.7 up to 12.2 g/kg) blocked Cu-induced changes. These results indicated that Cu-induced decreases in intestinal TJ proteins and cytokine mRNA levels might be partially mediated by oxidative stress and are prevented by histidine supplementation in fish diet. [ABSTRACT FROM AUTHOR]

The aim of the work was primarily to explore the protective activity pathways of lysine against oxidative damage in fish in vivo and in enterocytes in vitro. First, grass carp were fed diets containing six graded levels of lysine (7.1–19.6 g kg-1 diet) for 56 days. Second, the enterocytes were treated with different concentrations of lysine (0–300 mg/L in media) prior to (pre-treatment), along with (co-treatment) or following (post-treatment) with 6 mg/L of Cu for 24 h. The results indicated that lysine improved grass carp growth performance. Meanwhile, lysine ameliorated lipid and protein oxidation by elevating the gene expression and activity of antioxidant enzymes (superoxide dismutase (SOD), glutathioneperoxidase (GPx), glutathione-S-transferase (GST) and reductase (GR)), and nuclear factor erythroid 2-related factor 2 (Nrf2) mRNA levels in fish intestine. The in vitro studies showed that co- and post-treatment with lysine conferred significant protection against Cu-induced oxidative damage in fish primary enterocytes as measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) OD values, along with alkaline phosphatase (ALP) and lactate dehydrogenase activities, and the depletion of protein carbonyl (PC), malondialdehyde (MDA) and 8-hydroxydeoxyguanosine contents. Moreover, lysine co-treatment decreased the activities and mRNA level of cellular SOD, GPx, GST and GR compared with the Cu-only exposed group. Gene expression of the signalling molecule Nrf2 showed the same pattern as that of SOD activity, whereas Kelch-like ECH-associated protein 1b (Keap1b) followed the opposite trend, indicating that co-treatment with lysine induced antioxidant enzymes that protected against oxidative stress through Nrf2 pathway. In addition, post-treatment with lysine increased proteasomal activity and blocked the Cu-stimulated increase in mRNA levels of GST and associated catalase (CAT) and GST activities (P<0.01 and P<0.001). GR activity and gene expression, and glutathione (GSH) content followed an opposite trend to GST activity (P<0.05). Thus, post-treatment of lysine elevated protein and DNA repair abilities and ameliorated the cellular redox state of enterocytes. The overall results suggest that lysine plays a significant role in the protection of fish intestine in vivo and in vitro through the induction of key antioxidant protection. [ABSTRACT FROM AUTHOR]

Six groups of grass carp (average weight 266.9 ± 0.6 g) were fed diets containing 197, 385, 770, 1082, 1436 and 1795 mg choline/kg, for 8 weeks. Fish growth, and muscle nutrient (protein, fat and amino acid) content of young grass carp were significantly improved by appropriate dietary choline. Furthermore, muscle hydroxyproline concentration, lactate content and shear force were improved by optimum dietary choline supplementation. However, the muscle pH value, cooking loss and cathepsins activities showed an opposite trend. Additionally, optimum dietary choline supplementation attenuated muscle oxidative damage in grass carp. The muscle antioxidant enzyme (catalase and glutathione reductase did not change) activities and glutathione content were enhanced by optimum dietary choline supplementation. Muscle cooking loss was negatively correlated with antioxidant enzyme activities and glutathione content. At the gene level, these antioxidant enzymes, as well as the targets of rapamycin, casein kinase 2 and NF-E2-related factor 2 transcripts in fish muscle were always up-regulated by suitable choline. However, suitable choline significantly decreased Kelch-like ECH-associated protein 1 a (Keap1a) and Kelch-like ECH-associated protein 1 b (Keap1b) mRNA levels in muscle. In conclusion, suitable dietary choline enhanced fish flesh quality, and the decreased cooking loss was due to the elevated antioxidant status that may be regulated by Nrf2 signaling. [ABSTRACT FROM AUTHOR]

The present study explored the impact of dietary isoleucine (Ile) on fish growth and flesh quality and revealed a possible role of muscle antioxidant defense in flesh quality in relation to dietary Ile. Grass carp (weighing 256.8±3.5 g) were fed diets containing six graded levels of Ile (3.8, 6.6, 9.3, 12.5, 15.2 and 18.5 g/kg) for eight weeks. The results indicated that compared with Ile deficiency (3.8 g/kg diets) and excess (18.5 g/kg diets) groups, 9.3–15.2 g Ile/kg diet supplementations promoted fish growth and muscle fat deposition, whereas 6.6–15.2 g Ile/kg diets supplementation enhanced muscle nutrients (protein and total EAAs) deposition. Furthermore, muscle shear force, pH value, and hydroxyproline concentration were improved by 9.3–12.5, 9.3 and 9.3 g Ile/kg diet supplementations, respectively. However, muscle cooking loss, lactate content, and activities of cathepsin B and L were decreased by 6.6–15.2, 9.3–12.5, 9.3–12.5 and 9.3–15.2 g Ile/kg diet supplementations, respectively. Additionally, 6.6–15.2 and 6.6–12.5 g Ile/kg diet supplementations attenuated malondialdehyde and protein carbonyl contents, respectively. The activities of copper/zinc superoxide dismutase (Cu/Zn-SOD) and glutathione peroxidase (GPx), and glutathione content were enhanced by 6.6–9.3, 6.6–12.5 and 6.6–15.2 g Ile/kg diet supplementations, respectively. Moreover, the relative mRNA expressions of antioxidant enzymes, including Cu/Zn-SOD (6.6–12.5 g/kg diets) and GPx (12.5 g/kg diets), as well as antioxidant-related signaling molecules, including NF-E2-related factor 2 (Nrf2) (6.6–12.5 g/kg diets), target of rapamycin (6.6–12.5 g/kg diets), ribosomal S6 protein kinase 1 (9.3–12.5 g/kg diets) and casein kinase 2 (6.6–12.5 g/kg diets), were up-regulated when Ile diet supplementations were administered at these levels, respectively, whereas the relative mRNA expression of Kelch-like ECH-associated protein 1 was down-regulated with 9.3 g Ile/kg diet supplementations. Collectively, the present study indicated that optimum isoleucine improved flesh quality, partly due to the activation of antioxidant defense through the Nrf2 signaling pathway. [ABSTRACT FROM AUTHOR]

This study aimed to investigate the effects of threonine (Thr) on the digestive and absorptive ability, proliferation and differentiation of enterocytes, and gene expression of juvenile Jian carp (Cyprinus carpio var. Jian). First, seven isonitrogenous diets containing graded levels of Thr (7.4–25.2 g/kg diet) were fed to the fishes for 60 days. Second, enterocyte proliferation and differentiation were assayed by culturing enterocytes with graded levels of Thr (0–275 mg/l) in vitro. Finally, enterocytes were cultured with 0 and 205 mg/l Thr to determine protein synthesis. The percent weight gain (PWG), specific growth rate, feed intake, feed efficiency, protein retention value, activities of trypsin, lipase and amylase, weights and protein contents of hepatopancreas and intestine, folds heights, activities of alkaline phosphatase (AKP), γ- glutamyl transpeptidase and Na+/K+-ATPase in all intestinal segments, glutamate-oxaloacetate transaminase (GOT) and glutamate-pyruvate transaminase (GPT) activities in hepatopancreas, and 4E-BP2 gene expression in muscle, hepatopancreas and intestinal segments were significantly enhanced by Thr (p<0.05). However, the plasma ammonia concentration and TOR gene expression decreased (p<0.05). In vitro, Thr supplement significantly increased cell numbers, protein content, the activities of GOT, GPT, AKP and Na+/K+-ATPase, and protein synthesis rate of enterocytes, and decreased LDH activity and ammonia content in cell medium (p<0.05). In conclusion, Thr improved growth, digestive and absorptive capacity, enterocyte proliferation and differentiation, and protein synthesis and regulated TOR and 4E-BP2 gene expression in juvenile Jian carp. The dietary Thr requirement of juvenile Jian carp was 16.25 g/kg diet (51.3 g/kg protein) based on quadratic regression analysis of PWG. [ABSTRACT FROM AUTHOR]

β-conglycinin has been identified as one of the major feed allergens. However, studies of β-conglycinin on fish are scarce. This study investigated the effects of β-conglycinin on the growth, digestive and absorptive ability, inflammatory response, oxidative status and gene expression of juvenile Jian carp (Cyprinus carpio var. Jian) in vivo and their enterocytes in vitro. The results indicated that the specific growth rate (SGR), feed intake, and feed efficiency were reduced by β-conglycinin. In addition, activities of trypsin, chymotrypsin, lipase, creatine kinase, Na+,K+-ATPase and alkaline phosphatase in the intestine showed similar tendencies. The protein content of the hepatopancreas and intestines, and the weight and length of the intestines were all reduced by β-conglycinin. β-conglycinin increased lipid and protein oxidation in the detected tissues and cells. However, β-conglycinin decreased superoxide dismutase (SOD), catalase (CAT), glutathione-S-transferase (GST), glutathione peroxidase (GPx) and glutathione reductase (GR) activities and glutathione (GSH) content in the intestine and enterocytes. Similar antioxidant activity in the hepatopancreas was observed, except for GST. The expression of target of rapamycin (TOR) gene was reduced by β-conglycinin. Furthermore, mRNA levels of interleukin-8 (IL-8), tumor necrosis factor-α (TNF-α), and transforming growth factor-β (TGF-β) genes were increased by β-conglycinin. However, β-conglycinin increased CuZnSOD, MnSOD, CAT, and GPx1b gene expression. In conclusion, this study indicates that β-conglycinin induces inflammation and oxidation, and causes dysfunction of intestinal digestion and absorption in fish, and finally reduces fish growth. The results of this study provide some information to the mechanism of β-conglycinin-induced negative effects. [ABSTRACT FROM AUTHOR]

This study aimed to investigate the effects of threonine (Thr) on the digestive and absorptive ability, proliferation and differentiation of enterocytes, and gene expression of juvenile Jian carp (Cyprinus carpio var. Jian). First, seven isonitrogenous diets containing graded levels of Thr (7.4-25.2 g/kg diet) were fed to the fishes for 60 days. Second, enterocyte proliferation and differentiation were assayed by culturing enterocytes with graded levels of Thr (0-275 mg/l) in vitro. Finally, enterocytes were cultured with 0 and 205 mg/l Thr to determine protein synthesis. The percent weight gain (PWG), specific growth rate, feed intake, feed efficiency, protein retention value, activities of trypsin, lipase and amylase, weights and protein contents of hepatopancreas and intestine, folds heights, activities of alkaline phosphatase (AKP), γ- glutamyl transpeptidase and Na(+)/K(+)-ATPase in all intestinal segments, glutamate-oxaloacetate transaminase (GOT) and glutamate-pyruvate transaminase (GPT) activities in hepatopancreas, and 4E-BP2 gene expression in muscle, hepatopancreas and intestinal segments were significantly enhanced by Thr (p<0.05). However, the plasma ammonia concentration and TOR gene expression decreased (p<0.05). In vitro, Thr supplement significantly increased cell numbers, protein content, the activities of GOT, GPT, AKP and Na(+)/K(+)-ATPase, and protein synthesis rate of enterocytes, and decreased LDH activity and ammonia content in cell medium (p<0.05). In conclusion, Thr improved growth, digestive and absorptive capacity, enterocyte proliferation and differentiation, and protein synthesis and regulated TOR and 4E-BP2 gene expression in juvenile Jian carp. The dietary Thr requirement of juvenile Jian carp was 16.25 g/kg diet (51.3 g/kg protein) based on quadratic regression analysis of PWG.