Browsing by Author "Zhen-Yu Du"

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    Atorvastatin remodels lipid distribution between liver and adipose tissues through blocking lipoprotein efflux in fish
    (American Physiological Society, 2023-03-01) Rui-Xin Li; Ling-Yun Chen; Samwel Mchele Limbu; Bing Yao; Yi-Fan Qian; Wen-Hao Zhou; Li-Qiao Chen; Fang Qiao; Mei-Ling Zhang; Zhen-Yu Du; Yuan Luo
    The regulation of cholesterol metabolism in fish is still unclear. Statins play important roles in promoting cholesterol metabolism development in mammals. However, studies on the role of statins in cholesterol metabolism in fish are currently limited. The present study evaluated the effects of statins on cholesterol metabolism in fish. Nile tilapia (Oreochromis niloticus) were fed on control diets supplemented with three atorvastatin levels (0, 12, and 24 mg/kg diet, ATV0, ATV12, and ATV24, respectively) for 4 wk. Intriguingly, the results showed that both atorvastatin treatments increased hepatic cholesterol and triglyceride contents mainly through inhibiting bile acid synthesis and efflux, and compensatorily enhancing cholesterol synthesis in fish liver (P < 0.05). Moreover, atorvastatin treatment significantly inhibited hepatic very-low-density lipoprotein (VLDL) assembly and thus decreased serum VLDL content (P < 0.05). However, fish treated with atorvastatin significantly reduced cholesterol and triglycerides contents in adipose tissue (P < 0.05). Further molecular analysis showed that atorvastatin treatment promoted cholesterol synthesis and lipogenesis pathways, but inhibited lipid catabolism and low-density lipoprotein (LDL) uptake in the adipose tissue of fish (P < 0.05). In general, atorvastatin induced the remodeling of lipid distribution between liver and adipose tissues through blocking VLDL efflux from the liver to adipose tissue of fish. Our results provide a novel regulatory pattern of cholesterol metabolism response caused by atorvastatin in fish, which is distinct from mammals: cholesterol inhibition by atorvastatin activates hepatic cholesterol synthesis and inhibits its efflux to maintain cholesterol homeostasis, consequently reduces cholesterol storage in fish adipose tissue.
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    Concentration-dependent effects of 17 β-estradiol and bisphenol A on lipid metabolism, inflammation and antioxidant response in male zebrafish (Danio rerio). Chemosphere, 237: (2019) 124422.
    (Elsevier, 2019-12) Sheng-Xiang Sun; Yun-Ni Zhang; Dong-Liang Lu; Wei-Li Wang; Samwel Mchele Limbu; Dong-Liang Li; Li-Qiao Chen; Mei-Ling Zhang; Zhen-Yu Du
    Environmental estrogenic compounds are important pollutants, which are widely distributed in natural water bodies. They produce various adverse effects on fish, but their concentration-dependent toxicities in fish metabolism and health are not fully understood. This study investigated the effects of 17β-estradiol (E2) and bisphenol A (BPA) at low and high concentrations on lipid deposition, inflammation and antioxidant response in male zebrafish. We measured fish growth parameters, gonad development, lipid contents and the activities of inflammatory and antioxidant enzymes, as well as their mRNA expressions. All E2 and BPA concentrations used increased body weight, damaged gonad structure and induced feminization in male zebrafish. The exposure of zebrafish to E2 and BPA promoted lipid accumulation by increasing total fat, liver triglycerides and free fatty acid contents, and also upregulated lipogenic genes expression, although they decreased total cholesterol content. Notably, zebrafish exposed to low concentrations of E2 (200 ng/L) and BPA (100 μg/L) had higher lipid synthesis and deposition compared to high concentrations (2000 ng/L and 2000 μg/L, respectively). However, the high concentrations of E2 and BPA increased inflammation and antioxidant response. Furthermore, BPA caused greater damage to fish gonad development and more severe lipid peroxidation compared to E2. Overall, the results suggest that the toxic effects of E2 and BPA on zebrafish are concentration-dependent such that, the relative low concentrations used induced lipid deposition, whereas the high ones caused adverse effects on inflammation and antioxidant response.
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    Dietary docosahexaenoic acid reduces fat deposition and alleviates liver damage induced by D-galactosamine and lipopolysaccharides in Nile tilapia (Oreochromis niloticus)
    (Elsevier, 2023-06) Yi-Chan Liu; Samwel Mchele Limbu; Jin-Gang Wang; Mai Wang; Li-Qiao Chen; Fang Qiao; Yuan Luo; Mei-Ling Zhang; Zhen-Yu Du
    Liver health is important to maintain survival and growth of fish. Currently, the role of dietary docosahexaenoic acid (DHA) in improving fish liver health is largely unknown. This study investigated the role of DHA supplementation in fat deposition and liver damage caused by D-galactosamine (D-GalN) and lipopolysaccharides (LPS) in Nile tilapia (Oreochromis niloticus). Four diets were formulated as control diet (Con), Con supplemented with 1 % DHA, 2 % DHA and 4 % DHA diets, respectively. The diets were fed to 25 Nile tilapia (2.0 ± 0.1 g, average initial weight) in triplicates for four weeks. After the four weeks, 20 fish in each treatment were randomly selected and injected with a mixture of 500 mg D-GalN and 10 μL LPS per mL to induce acute liver injury. The results showed that the Nile tilapia fed on DHA diets decreased visceral somatic index, liver lipid content and serum and liver triglyceride concentrations than those fed on the Con diet. Moreover, after D-GalN/LPS injection, the fish fed on DHA diets decreased alanine aminotransferase and aspartate transaminase activities in the serum. The results of liver qPCR and transcriptomics assays together showed that the DHA diets feeding improved liver health by downregulating the expression of the genes related to toll-like receptor 4 (TLR4) signaling pathway, inflammation and apoptosis. This study indicates that DHA supplementation in Nile tilapia alleviates the liver damage caused by D-GalN/LPS through increasing lipid catabolism, decreasing lipogenesis, TLR4 signaling pathway, inflammation, and apoptosis. Our study provides novel knowledge on the role of DHA in improving liver health in cultured aquatic animals for sustainable aquaculture.
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    Dietary L-carnitine alleviates the adverse effects caused by reducing protein and increasing fat contents in juvenile largemouth bass (Micropterus salmoides)
    (2022-05-06) Yi-Chan Liu; Samwel Mchele Limbu; Jin-Gang Wang; Jiong Ren; Fang Qiao; Mei-Ling Zhang; Zhen-Yu Du
    Protein ingredients for formulation of fish feeds are expensive and have limited availability. Therefore, reducing dietary protein while increasing dietary fat content is a common practice in rearing carnivorous fish species. However, the ability of dietary L-carnitine to alleviate adverse effects in such diets is currently unknown. This study investigated the role of L-carnitine supplementation in alleviating adverse effects on growth performance, energy metabolism, antioxidant capacity, and inflammation response in juvenile largemouth bass (Micropterus salmoides) fed on a low protein and high fat diet. Three diets were formulated to contain low protein and high fat (LPHF: 420 g kg-1 protein and 150 g kg-1 lipid), LPHF supplemented with L-carnitine (LPHFC: 420 g kg-1 protein and 150 g kg-1 lipid), and a control diet (CON: 480 g kg-1 protein and 130 g kg-1 lipid). The diets were fed to 30 largemouth bass (g) juveniles in triplicates for eight weeks. The results showed that the fish feed on LPHF diet increased hepatosomatic index, visceral somatic index, mesenteric fat index, whole-body crude fat content, serum and liver triglyceride concentrations, and serum non-esterified fatty acid level than those fed on CON diet. Moreover, the fish fed on LPHF diet increased serum alanine aminotransferase activity and liver malondialdehyde content and reduced superoxide dismutase (SOD) activities in the serum and liver. Furthermore, the fish fed on LPHF diet reduced the whole-body crude protein content. Interestingly, feeding the fish on the LPHFC diet decreased fat deposition and liver damage by downregulating the expression of genes related to lipogenesis, inflammation, and increased SOD activity. This study indicates that L-carnitine supplementation in largemouth bass alleviates the adverse effects caused by LPHF diet by decreasing lipogenesis and increasing lipid catabolism. Our study provides novel knowledge on strategies to improve utilization of LPHF diet in cultured aquatic animals.
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    Dietary L-carnitine improves glycogen and protein accumulation in Nile tilapia via increasing lipid-sourced energy supply: an isotope-based metabolic tracking. Aquaculture Reports, 17: 100302.
    (Elsevier, 2020-07) Ling-Yu Li; Dong-Liang Lu; Zhe-Yue Jiang; Samwel Mchele Limbu; Fang Qiao; Liqiao Chen; Meiling Zhang; Zhen-Yu Du
    L-carnitine is a functional aquafeed additive for enhancing lipid catabolism by elevating mitochondrial fatty acid β-oxidation and modulating energy metabolism to provide a “protein sparing effect”. However, results on the effects of dietary l-carnitine on nutrient metabolism in fish are still conflicting. We explored comprehensively the effects of dietary l-carnitine on energy metabolism in Nile tilapia. We fed Nile tilapia for eight weeks with diets supplemented with l-carnitine or not. We conducted metabolic tracking tests by intraperitoneally injecting individual fish with 14C-labeled palmitic acid (PA), glucose (Glu) and an amino acid mixture (AAs). After the feeding trial, insignificant growth-promoting effect of l-carnitine was obtained in treated fish. However, l-carnitine significantly reduced the lipid content in whole body and muscle accompanied by increasing the free carnitine concentration and fatty acid β-oxidation efficiency. Moreover, l-carnitine elevated concentrations of serum glucose, pyruvate and lactate, and increased glycogen and protein deposition in muscle. These results suggest that ingested glucose and protein prefer to be reserved in carnitine-fed fish with sufficient fatty acids oxidation for energy. Nevertheless, after a 14C-labeled single nutrient injection, carnitine-fed fish showed a higher oxidation rate of [1-14C]-PA, d-[1-14C]-Glu and l-[14C (U)]-AAs. Our study indicates that, the effects of l-carnitine on nutrient metabolism are correlated with the abundance of individual macronutrients such that an inadequate lipid supply would cause dietary l-carnitine supplementation to elevate higher breakdown of glucose and protein for energy generation. The present study provides new insights on the regulation mechanism of l-carnitine on nutrient metabolism in fish.
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    Dietary L-carnitine supplementation recovers the increased pH and hardness in fillets caused by high-fat diet in Nile tilapia (Oreochromis niloticus)
    (Elsevier, 2022-07-15) Zhi-Yong Zhang; Samwel Mchele Limbu; Si-Han Zhao; Li-Qiao Chen; Yuan Luo; Mei-Ling Zhang; Fang Qiao; Zhen-Yu Du
    The wide use of high-fat diet (HFD) causes negative effects on flesh quality in farmed fish. l-carnitine, a lipid-lowering additive, enhances mitochondrial fatty acid β-oxidation. However its roles in alleviating the effects of HFD on flesh quality in fish are unknown. We fed Nile tilapia with medium-fat diet (MFD, 6% dietary lipid), high-fat diet (HFD, 12% dietary lipid) and HFCD supplemented with l-carnitine (HFCD + 400 mg/kg l-carnitine) for 10 weeks. The HFD-fed fish had higher fat deposition, pH value, myofiber density and flesh hardness than those fed on MFD. However, feeding the fish with the HFCD improved lipid catabolism, which increased significantly lactic acid content and myofiber diameter in muscle, thus reduced pH and hardness values. HFCD also reduced endoplasmic reticulum stress and myofiber apoptosis caused by HFD in the fish. Our study suggests that dietary l-carnitine supplementation alleviates the negative effects of HFD on flesh quality of farmed fish.
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    Dietary sodium acetate improves high-fat diet utilization through promoting differential nutrients metabolism between liver and muscle in Nile tilapia (Oreochromis niloticus)
    (Elsevier, 2023-02-25) Wen-Hao Zhou; Samwel Mchele Limbu; Rui-Xin Li; Yuan Luo; Jiong Ren; Fang Qiao; Mei-Ling Zhang; Zhen-Yu Du
    igh-fat diet (HFD) often causes many negative effects and impairs fish growth. Short-chain fatty acids (SCFAs) such as acetates modify metabolic disorders and regulate body homeostasis. However, the effects of sodium acetate on alleviating HFD in fish is currently unknown. This study investigated the role of using dietary sodium acetate to alleviate adverse effects caused by HFD in fish. Three replicates (thirty fish each) of 4.8 ± 0.30 g Nile tilapia (Oreochromis niloticus) were fed with control diet (Con), high-fat diet (HFD) or HFD containing sodium acetate diet (HFD + NaAc) for eight weeks. After the feeding trial, Nile tilapia fed with HFD increased significantly tissue lipid deposition, reduced insulin sensitivity and suppressed glucose and lipid metabolism in both liver and muscle, accompanied with signs of metabolic disorders and liver damage. Moreover, HFD feeding inhibited muscle protein synthesis and impaired fish growth performance. However, the fish fed on HFD + NaAc improved significantly oxidative stress, inflammation, apoptosis and injury in liver compared to those fed on HFD. More importantly, dietary sodium acetate supplementation enhanced insulin sensitivity, promoted glucose catabolic utilization and protein synthesis in muscle through activation of insulin and mTOR signaling pathways, respectively, and markedly improved the growth performance. In contrast, dietary sodium acetate promoted hepatic pentose phosphate pathway, hepatic glycogen accumulation, and activated lipid catabolism to alleviate hepatic lipid deposition. Our study illustrates that sodium acetate alleviates differentially the adverse effects induced by feeding Nile tilapia with HFD in muscle and liver. SCFAs such as acetate may be used for improving HFD utilization in fish nutrition.
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    Different effects of two dietary levels of tea polyphenols on the lipid deposition, immunity and antioxidant capacity of juvenile GIFT tilapia (Oreochromis niloticus) fed a high-fat diet. Aquaculture, 542: 736896. https://doi.org/10.1016/j.aquaculture.2021.736896
    (Elsevier, 2021-05-11) Yu-Cheng Qian; Xue Wang; Jiong Ren; Jie Wang; Samwel Mchele Limbu; Rui-Xin Li; Wen-Hao Zhou; Fang Qiao; Mei-Ling Zhang; Zhen-Yu Du
    Long-term feeding of fish with a high-fat diet (HFD) causes excess fat deposition and an impairment of immune function. In the present study, we aimed to determine whether dietary tea polyphenols (TPs) would ameliorate the adverse effects of HFD-feeding in GIFT tilapia. Juvenile GIFT tilapias (5.4 ± 0.9 g) were raised in twelve 200-L tanks (three tanks per diet, 20 fish per tank) and fed a control diet (6% fat, 36% protein), an HFD (12% fat, 36% protein), or an HFD supplemented with 50 mg/kg or 200 mg/kg TP for 8 weeks. The fish were hand-fed 5% of their body weight per day in three feeds, and maintained at 28 ± 1 °C under a 14-h light/10-h dark cycle. The fish in each tank were bulk weighed and counted fortnightly, and the daily feed amount was adjusted accordingly. At the end of the trial, the cumulative survival rate was calculated, and the weight gain and feed conversion ratio were calculated according to the bulk weight of fish in each tank. Tissues were collected from nine fish per diet, their organs were weighed, and biochemical and molecular indices were subsequently measured. HFD-feeding significantly increased lipid deposition, reduced cumulative survival from 96% to 75%, reduced hepatic alkaline phosphate activity (AKP) and serum total antioxidant capacity (T-AOC); and reduced the hepatic expression of immunoglobulin M (IgM), transforming growth factor-beta (TGF-β) and glutathione-S-transferase (GST) genes versus the control diet. The addition of TPs at 50 or 200 mg/kg both ameliorated the HFD-induced increase in lipid droplets in the liver (50 mg/kg TP from 40.83% to 17.27%; 200 mg/kg TP to 25.33%), and increased the cumulative survival rate of the tilapia. The addition of 50 mg/kg TP had a marked effect increasing cumulative survival to 90%, and increasing the activities of serum acid phosphatase (ACP), T-AOC; and IgM, TGF-β, nuclear factor-κB (NF-κB), superoxide dismutase (SOD), and GST gene expression to the highest level of the HFD-fed groups. The 50 mg/kg TP-containing diet also significantly increased the hepatic expression of carnitine palmitoyltransferase 1 alpha (CPT1α) versus the control diet. In contrast, the tilapia fed an HFD supplemented with 200 mg/kg TPs had the lowest expression of adipose triglyceride lipase, hormone-sensitive lipase, CPT1α, fatty acid synthase and acetyl-CoA carboxylase alpha genes of any of the groups, which implies that the lower and higher levels of TP supplementation have differing effects on lipid metabolism. The 200 mg/kg supplement had lower cumulative survival rate (82%), and smaller effects on serum ACP and hepatic AKP activities than the 50 mg/kg dose, and had no significant effect on serum T-AOC or the expression of IgM, TGF-β, GST, or NF-κB genes in the tilapia. These results indicate that the beneficial effects of TPs on the lipid metabolism and health of fish fed an HFD are dose-related. Moreover, they are likely to be largely mediated through lipid catabolism.
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    A global analysis on the systemic effects of antibiotics in cultured fish and their potential human health risk: A review. Reviews in Aquaculture, 13(2): 1015–1059
    (Wiley, 2021-03) Samwel Mchele Limbu; Liqiao Chen; Meiling Zhang; Zhen-Yu Du
    The rearing of fish in intensive systems reduces their immunity leading to eruption of diseases, consequently prompting the use of antibiotics. Antibiotics are currently used in cultured fish globally to treat/prevent diseases. On the one hand, antibiotics used in aquaculture help to improve fish health by killing or inhibiting pathogenic bacteria proliferation. However, on the other hand, they cause multiple side effects. To date, information on systemic effects of antibiotics in cultured fish is scattered and their potential human health risk remains elusive. This review critically analyses and discusses the available information in the literature on the positive and negative effects of antibiotics in cultured fish and their potential human health risk via fish consumption. The review found 44 different antibiotics dominated by oxytetracycline were studied in various fish species worldwide. Antibiotics used in global aquaculture either decrease or increase fish growth and survival rates. They induce oxidative stress, which affect antioxidant and detoxification responses and cause host–microbiota dysbiosis. These changes compromise the fish immunity system, which ultimately lead to cellular damage. The toxicities generated inhibit the fish aerobic glycolysis, suppress lipogenesis and fatty acid β‐oxidation while increase energy demand by initiating gluconeogenesis and anaerobic glycolysis. Consumption of fish treated with antibiotics causes human health risk. Future studies are required on agents to restore dysfunctions induced by antibiotics in cultured fish, while attempts to limit their utilisation in aquaculture production are underway.
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    Gnotobiotic models: Powerful tools for deeply understanding intestinal microbiota-host interactions in aquaculture. Aquaculture, 517: 734800. https://doi.org/10.1016/j.aquaculture.2019.734800.
    (Elsevier, 2020-02-25) Meiling Zhang; Chengjie Shan; Fang Tan; Samwel Mchele Limbu; Liqiao Chen; Zhen-Yu Du
    Intestinal microbiota plays profound roles in host nutrition, physiology, and evolution. The development of DNA sequencing technologies has increased dramatically research on fish intestinal microbiota. However, most studies conducted so far have focused on the microbial structure and diversity. Studies targeting the exact function of commensal microbes in aquatic animals are still scarce, which limits the application of microbiota related knowledge in aquaculture. Gnotobiotic models (animals cultured in axenic conditions or with defined microbial lineages) are excellent tools for identifying the molecular interactions between intestinal microbiota and host, which drive studies of microbiota from correlation to causality in mammals. In recent decades, gnotobiotic fish models have been established and applied in aquaculture research. This review summarizes the colonization conditions in gnotobiotic zebrafish model and its application in understanding intestinal microbiota-host interactions in aquaculture. Furthermore, methods and research progress on other gnotobiotic models including freshwater and marine fishes, molluscs and crustaceans are also discussed. Application of gnotobiotic models in aquaculture has deepened our understanding of the relationship between the host and intestinal microbiota, which will facilitate the modulation of intestinal microbiota for production of healthy animals and sustainable development of aquaculture.
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    High carbohydrate diet partially protects Nile tilapia from oxytetracycline-induced side effects. Environmental Pollution, 256: 113508.
    (Elsevier, 2020-01) Samwel Mchele Limbu; Han Zhang; Yuan Luo; Meiling Zhang; Zhen-Yu Du
    Antibiotics used in global aquaculture production cause various side effects, which impair fish health. However, the use of dietary composition such as carbohydrate, which is one of the dominant components in fish diets to attenuate the side effects induced by antibiotics, remains unclear. We determined the ability of high carbohydrate diet to protect Nile tilapia (Oreochromis niloticus) from oxytetracycline-induced side effects. Triplicate groups of thirty O. niloticus (9.50 ± 0.08 g) were fed on medium carbohydrate (MC; 335 g/kg) and high carbohydrate (HC; 455 g/kg) diets without and with 2.00 g/kg diet of oxytetracycline (80 mg/kg body weight/day) hereafter, MCO and HCO for 35 days. Thereafter, we assessed growth performance, hepatic nutrients composition and metabolism, microbiota abundance, immunity, oxidative and cellular stress, hepatotoxicity, lipid peroxidation and apoptosis. To understand the possible mechanism of carbohydrate protection on oxytetracycline, we assessed the binding effects and efficiencies of mixtures of medium and high starch with oxytetracycline as well as the MCO and HCO diets. The O. niloticus fed on the MCO and HCO diets had lower growth rate, nutrients utilization and survival rate than those fed on the MC and HC diets, respectively. Dietary HCO increased hepatosomatic index and hepatic protein content of O. niloticus than MCO diet. The O. niloticus fed on the HCO diet had lower mRNA expression of genes related to protein, glycogen and lipid metabolism compared to those fed on the MCO diet. Feeding O. niloticus on the HCO diet increased innate immunity and reduced pathogenic bacteria, pro-inflammation, hepatotoxicity, cellular stress and apoptosis than the MCO diet. The high starch with oxytetracycline and HCO diet had higher-oxytetracycline binding effects and efficiencies than the medium starch with oxytetracyline and MCO diet, respectively. Our study demonstrates that, high carbohydrate partially protects O. niloticus from oxytetracycline-induced side effects by binding the antibiotic. Incorporating high carbohydrate in diet formulation for omnivorous fish species alleviates some of the side effects caused by antibiotics.
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    Inhibition of lipophagy suppresses lipid metabolism in zebrafish liver cells, Frontiers in Physiology, 10; Article 1077; 1-9.
    (Frontiers Media SA, 2019-08-21) Jing Wang; Si-Lan Han; Dong-Liang Lu; Ling-Yu Li; Samwel Mchele Limbu; Dongliang Li; Meiling Zhang; Zhen-Yu Du
    Lipophagy degrades lipid droplets (LDs) through the lysosomal degradative pathway, thus plays important roles in regulating lipid metabolism in mammals. However, information on the existence and functions of lipophagy in fish lipid metabolism is still limited. In the present study, we confirmed the existence of lipophagy by observing the structures of LDs sequestered in autophagic vacuoles in the zebrafish liver cell line (ZFL) via electronic microscopy. Moreover, starved cells increased the mRNA expression of the microtubule-associated protein 1A/1B light chain 3 beta (LC3), which is a marker protein for autophagy and protein conversion from LC3-I to LC3-II. Inhibiting autophagy with chloroquine increased significantly the LDs content and decreased fatty acid β-oxidation and esterification activities in the ZFL cells cultured in the fed state. Furthermore, inhibiting autophagy function downregulated the mRNA expression of the genes and their proteins related to lipid metabolism. Altogether, the present study verified the existence of lipophagy and its essential regulatory roles in lipid metabolism in fish cells.
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    Inhibition of pyruvate dehydrogenase kinase increases carbohydrate utilization in Nile tilapia by regulating PDK2/4-PDHE1α axis and insulin sensitivity
    (Elsevier, 2022-06-24) Yuan Luo; Wen-Hao Zhou; Rui-Xin Li; Samwel Mchele Limbu; Fang Qiao; Li-Qiao Chen; Mei-Ling Zhang; Zhen-Yu Du
    Pyruvate dehydrogenase kinases (PDKs)-pyruvate dehydrogenase E1α subunit (PDHE1α) axis plays an important role in regulating glucose metabolism in mammals. However, the regulatory function of PDKs-PDHE1α axis in the glucose metabolism of fish is not well known. This study determined whether PDKs inhibition could enhance PDHE1α activity, and improve glucose catabolism in fish. Nile tilapia fingerlings (1.90 ± 0.11 g) were randomly divided into 4 treatments in triplicate (30 fish each) and fed with control diet without dichloroacetate (DCA0) (38% protein, 7% lipid and 45% corn starch) and the control diet supplemented with DCA, which inhibits PDKs through binding the allosteric sites, at 3.75 (DCA3.75), 7.50 (DCA7.50) and 11.25 g/kg (DCA11.25), for 6 wk. The results showed that DCA3.75, DCA7.50 and DCA11.25 significantly increased weight gain, carcass ratio and protein efficiency ratio (P < 0.05) and reduced feed efficiency (P < 0.05) of Nile tilapia. To investigate the effects of DCA on growth performance of Nile tilapia, we selected the lowest doseDCA3.75 for subsequent analysis. Nile tilapia fed on DCA3.75 significantly reduced the mesenteric fat index, serum and liver triglyceride concentration and total lipid content in whole fish, and down-regulated the expressions of genes related to lipogenesis (P < 0.05) compared to the control. The DCA3.75 treatment significantly improved glucose oxidative catabolism and glycogen synthesis in the liver, but significantly reduced the conversion of glucose to lipid (P < 0.05). Furthermore, the DCA3.75 treatment significantly decreased the PDK2/4 gene and protein expressions (P < 0.05), accordingly stimulated PDHE1α activity by decreasing the phosphorylated PDHE1α protein level. In addition, DCA3.75 treatment significantly increased the phosphorylated levels of key proteins involved in insulin signaling pathway and glycogen synthase kinase 3β (P < 0.05). Taken together, the present study demonstrates that PDK2/4 inhibition by using DCA promotes glucose utilization in Nile tilapia by activating PDHE1α, and improving insulin sensitivity. Our study helps to understand the regulatory mechanism of glucose metabolism for improving dietary carbohydrate utilization in farmed fish.
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    Lipolysis and lipophagy play individual and interactive roles in regulating triacylglycerol and cholesterol homeostasis and mitochondrial form in zebrafish
    (Elsevier, 2021-06-08) Si-Lan Han; Yu-Cheng Qian; Samwel Mchele Limbu; Jing Wang; Li-Qiao Chen; Mei-Ling Zhang; Zhen-Yu Du
    Neutral lipases-mediated lipolysis and acid lipases-moderated lipophagy are two main processes for degradation of lipid droplets (LDs). However, the individual and interactive roles of these metabolic pathways are not well known across vertebrates. This study explored the roles of lipolysis and lipophagy from the aspect of neutral and acid lipases in zebrafish. We established zebrafish strains deficient in either adipose triglyceride lipase (atgl−/−; AKO fish) or lysosomal acid lipase (lal−/−; LKO fish) respectively, and then inhibited lipolysis in the LKO fish and lipophagy in the AKO fish by feeding diets supplemented with the corresponding inhibitors Atglistatin and 3-Methyladenine, respectively. Both the AKO and LKO fish showed reduced growth, swimming activity, and oxygen consumption. The AKO fish did not show phenotypes in adipose tissue, but mainly accumulated triacylglycerol (TAG) in liver, also, they had large LDs in the hepatocytes, and did not stimulate lipophagy as a compensation response but maintained basal lipophagy. The LKO fish reduced total lipid accumulation in the body but had high cholesterol content in liver; also, they accumulated small LDs in the hepatocytes, and showed increased lipolysis, especially Atgl expression, as a compensatory mechanism. Simultaneous inhibition of lipolysis and lipophagy in zebrafish resulted in severe liver damage, with the potential to trigger mitophagy. Overall, our study illustrates that lipolysis and lipophagy perform individual and interactive roles in maintaining homeostasis of TAG and cholesterol metabolism. Furthermore, the interactive roles of lipolysis and lipophagy may be essential in regulating the functions and form of mitochondria.
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    Mitochondrial fatty acid β-oxidation inhibition promotes carbohydrate catabolism and protein deposition through energy homeostasis remodelling. The Journal of Nutrition, nxaa187, https://doi.org/10.1093/jn/nxaa187.
    (Oxford University Press, 2019-09) Ling-Yu Li; Jia-Min Li; Li-Jun Ning; Dong-Liang Lu; Yuan Luo; Qiang Ma; Samwel Mchele Limbu; Dong-Liang Li; Li-Qiao Chen; Irfan J. Lodhi; Pascal Degrace; Mei-Ling Zhang; Zhen-Yu Du
    Background Fish cannot use carbohydrate efficiently and instead utilize protein for energy supply, thus limiting dietary protein storage. Protein deposition is dependent on protein turnover balance, which correlates tightly with cellular energy homeostasis. Mitochondrial fatty acid β-oxidation (FAO) plays a crucial role in energy metabolism. However, the effect of remodeled energy homeostasis caused by inhibited mitochondrial FAO on protein deposition in fish has not been intensively studied. Objectives This study aimed to identify the regulatory role of mitochondrial FAO in energy homeostasis maintenance and protein deposition by studying lipid, glucose, and protein metabolism in fish. Methods Carnitine-depleted male Nile tilapia (initial weight: 4.29 ± 0.12 g; 3 mo old) were established by feeding them with mildronate diets (1000 mg/kg/d) for 6 wk. Zebrafish deficient in the carnitine palmitoyltransferase 1b gene (cpt1b) were produced by using CRISPR/Cas9 gene-editing technology, and their males (154 ± 3.52 mg; 3 mo old) were used for experiments. Normal Nile tilapia and wildtype zebrafish were used as controls. We assessed nutrient metabolism and energy homeostasis–related biochemical and molecular parameters, and performed 14C-labeled nutrient tracking and transcriptomic analyses. Results The mitochondrial FAO decreased by 33.1–88.9% (liver) and 55.6–68.8% (muscle) in carnitine-depleted Nile tilapia and cpt1b-deficient zebrafish compared with their controls (P < 0.05). Notably, glucose oxidation and muscle protein deposition increased by 20.5–24.4% and 6.40–8.54%, respectively, in the 2 fish models compared with their corresponding controls (P < 0.05). Accordingly, the adenosine 5′-monophosphate–activated protein kinase/protein kinase B–mechanistic target of rapamycin (AMPK/AKT-mTOR) signaling was significantly activated in the 2 fish models with inhibited mitochondrial FAO (P < 0.05). Conclusions These data show that inhibited mitochondrial FAO in fish induces energy homeostasis remodeling and enhances glucose utilization and protein deposition. Therefore, fish with inhibited mitochondrial FAO could have high potential to utilize carbohydrate. Our results demonstrate a potentially new approach for increasing protein deposition through energy homeostasis regulation in cultured animals.
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    More simple more worse: Simple carbohydrate diets cause alterations in glucose and lipid metabolism in Nile tilapia (Oreochromis niloticus).
    (Elsevier, 2022-03-15) Wen-Hao Zhou; Chen-Chen Wu; Samwel Mchele Limbu; Rui-Xin Li; Li-Qiao Chen; Fang Qiao; Yuan Luo; Mei-Ling Zhang; Tao Han; Zhen-Yu Du
    Dietary carbohydrates are widely used in aquafeeds as cheap energy source and improve quality of feeds. However, comprehensive assessment of the effects of dietary carbohydrate complexity on fish metabolism and liver health is currently lacking. This study investigated the effect of replacing 50% of starch by using sucrose, fructose and glucose. Four isonitrogenic (400 g/kg protein) and isolipidic (60 g/kg fat) feeds containing starch (polysaccharide), sucrose (disaccharide) and fructose and glucose (monosaccharides) were fed to three replicates of 4.5 ± 0.30 g Nile tilapia (Oreochromis niloticus) for eight weeks. Afterwards, growth performance, feed utilization efficiency, glucose and lipid metabolism and liver health were evaluated. The results showed that the Nile tilapia fed on monosaccharide diets had significantly lower growth and feed utilization efficiency than those fed on disaccharide and polysaccharide diets. The Nile tilapia fed on monosaccharide diets reduced significantly lipid deposition, hepatosomatic index (HSI), serum alanine transaminase (ALT) and aspartate aminotransferase (AST) and expression of lipid metabolism genes than those fed on polysaccharide diet. The Nile tilapia fed on disaccharide diet increased significantly the serum insulin, muscle glycogen, and muscle glycogen synthase (gs) gene expression compared with those fed on polysaccharide diet. Interestingly, the Nile tilapia fed on monosaccharide diets reduced significantly the glycogen content while they increased insulin, glucose and the expression of liver gluconeogenesis genes such as glucose-6-phosphatase (g6pase) and phosphoenolpyruvate carboxykinase (pepck) than those fed on polysaccharide diet. Contrary, the Nile tilapia fed on disaccharide diet downregulated liver glucose catabolism and lowered serum triglyceride (TG) levels than those fed on polysaccharide but enhanced muscle peroxisome proliferator activated receptor alpha (pparα) and carnitine palmitoyltransferase 1a (cpt1a) gene expression, liver gluconeogenesis and pentose phosphate pathways. These results indicate that Nile tilapia utilizes better dietary polysaccharides and disaccharides than monosaccharides. Feeding Nile tilapia on monosaccharide diets caused insulin resistance and glucose metabolism disorders manifested by hyperinsulinism and hyperglycemia, respectively. The carbohydrate complexity affects the nutritional metabolism and liver health of fish.
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    Reduced fatty acid β-oxidation improves glucose catabolism and liver health in Nile tilapia (Oreochromis niloticus) juveniles fed a high-starch diet. Aquaculture, 535 (2021) 736392. https://doi.org/10.1016/j.aquaculture.2021.736392
    (Elsevier, 2021-03-30) Ling-Yu Li; Yue Wang; Samwel Mchele Limbu; Jia-Min Li; Fang Qiao; Li-Qiao Chen; Mei-Ling Zhang; Zhen-Yu Du
    Fish are poor users of dietary carbohydrates and often display prolonged hyperglycemia and fat deposition after feeding high digestible carbohydrate diets. Recently, fatty acid β-oxidation (FAO) inhibition has been reported to increase glucose oxidation in fish. Therefore, this study tested the assumption that the inhibition of FAO with mildronate (MD, a carnitine synthesis inhibitor) might also increase glucose utilization and alleviate adverse effects induced by high starch diet (HSD) in Nile tilapia, Oreochromis niloticus. Nile tilapia juveniles (6.13 ± 0.11 g) were cultured in nine 200-L tanks (30 fish per tank) and divided into three groups (three tanks per group). The fish were fed twice a day (9:00 and 18:30) at 4% body weight by using a normal starch diet (NSD, 30% corn starch), a HSD (45% corn starch), or a HSD supplemented with MD (25 g/kg of diet, HSD + MD) for eight weeks. These three feeds contained approximately 35.8% protein and 6.4% lipid. The fish each tank were weighed every two weeks, and the feeding amount was adjusted accordingly. After the feeding trial, the fish fed on HSD showed higher hepatosomatic index (HSI), visceral somatic index (VSI), serum triglyceride concentration and whole-body and tissue (liver and muscle) lipid contents than those fed on NSD. The fish fed on HSD also had higher relative area of vacuolation in the liver, hepatic malondialdehyde (MDA) content, and aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activities in the serum than those fed on NSD. Moreover, the fish fed on HSD increased serum glucose and insulin concentrations, and hepatic lactate, pyruvate and glycogen contents, but reduced whole-body protein content and dietary protein utilization than those fed on NSD, indicating that HSD induced fat deposition, liver damage, glucose intolerance and lowered protein-sparing effect. However, the fish fed on HSD + MD decreased hepatic carnitine content and FAO activity, attenuated the indexes related to fat deposition and liver damage, improved blood glucose clearance and whole-body protein deposition than those fed on HSD, suggesting that the adverse effects caused by HSD were reversed after FAO inhibition. Furthermore, the fish fed on HSD down-regulated the expression of genes associated with glucose uptake, glycolysis, FAO process, and lipolysis compared to those fed on HSD + MD and NSD, yet up-regulated lipogenic and proteolytic genes. These data suggested that inhibition of FAO improved glucose utilization and alleviated the HSD-induced adverse effects in Nile tilapia. This work demonstrates that, modifying mitochondrial FAO activity regulates the ability of fish to adapt to HSD intake through remodeling energy homeostasis. Our study provides new insights into improving carbohydrate utilization in aquatic animals.
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    The reduction of lipid-sourced energy production caused by ATGL inhibition cannot be compensated by activation of HSL, autophagy, and utilization of other nutrients in fish. Fish Physiology and Biochemistry, 47: 173–188
    (Springer, 2021-02) Si-Lan Han; Yan Liu; Samwel Mchele Limbu; Li-Qiao Chen; Mei-Ling Zhang; Zhen-Yu Du
    The adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL)–mediated lipolysis play important roles in lipid catabolism. ATGL is considered the central rate-limiting enzyme in the mobilization of fatty acids in mammals. Currently, severe fat accumulation has been commonly detected in farmed fish globally. However, the ATGL-mediated lipolysis and the potential synergy among ATGL, HSL, and autophagy, which is another way for lipid breakdown, have not been intensively understood in fish. In the present study, we added Atglistatin as an ATGL-specific inhibitor into the zebrafish diet and fed to the fish for 5 weeks. The results showed that the Atglistatin-treated fish exhibited severe fat deposition, reduced oxygen consumption, and fatty acid β-oxidation, accompanied with increased oxidative stress and inflammation. Furthermore, the Atglistatin-treated fish elevated total and phosphorylation protein expressions of HSL. However, the free fatty acids and lipase activities in organs were still systemically reduced in the Atglistatin-treated fish, and the autophagy marker LC3 was also decreased in the liver. On the other hand, glycogenolysis was stimulated but blood glucose was higher in the Atglistatin-treated fish. The transcriptomic analysis also provided the hint that the protein turnover efficiency in Atglistatin-treated fish was likely to be accelerated, but the protein content in whole fish was not affected. Taken together, ATGL plays crucial roles in energy homeostasis such that its inhibition causes loss of lipid-sourced energy production, which cannot be compensated by activation of HSL, autophagy, and utilization of other nutrients.
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    The responses of germ-free zebrafish (Danio rerio) to varying bacterial concentrations, colonization time points and exposure duration. Frontiers in Microbiology 10; Article 2156; 1 - 13.
    (Frontiers Media SA, 2019-09-18) Fang Tan; Samwel Mchele Limbu; Ye Qian; Fang Qiao; Zhen-Yu Du; Meiling Zhang
    Colonizing germ-free (GF) zebrafish with specific bacterial species provides the possibility of understanding the influence on host biological processes including gene expression, development, immunity, and behavioral responses. It also enlightens our understanding on the host-microbe interactions within the physiological context of a living host. However, the responses of GF zebrafish to various colonization conditions such as bacterial concentrations, colonization time points, and exposure duration remain unclear. To address this issue, we explored the responses of GF zebrafish by using two bacterial species at varying concentrations, colonization time points and exposure duration. Therefore, we mono-associated GF zebrafish with Escherichia coli DH5α or Bacillus subtilis WB800N at concentrations ranging from 102 to 107 CFU/ml either at 3 day post fertilization (dpf) or 5 dpf for 24 or 48 h. We evaluated the responses of GF zebrafish by analyzing the survival rate, colonization efficiency, nutrients metabolism, intestinal cell proliferation, innate immunity, stress, and behavior responses by comparing it to conventionally raised zebrafish (CONR) and GF zebrafish. The results indicated that the final bacteria concentrations ranging from 102 to 104 CFU/ml did not cause any mortality when GF mono-associated larvae were exposed to either E. coli DH5α or B. subtilis WB800N at 3 or 5 dpf, while concentrations ranging from 106 to 107 CFU/ml increased the mortality, particularly for 5 dpf owing to the decrease in dissolved oxygen level. The E. coli DH5α mainly induced the expression of genes related to nutrients metabolism, cell proliferation and immunity, while B. subtilis WB800N mainly upregulated the expression of genes related to immunity and stress responses. Moreover, our data revealed that GF zebrafish showed higher levels of physical activity than CONR and the microbial colonization reduced the hyperactivity of GF zebrafish, suggesting colonization of bacteria affected behavior characteristics. This study provides useful information on bacterial colonization of GF zebrafish and the interaction between the host and microbiota.
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    Vitellogenin 1 is essential for fish reproduction by transporting DHA-contained phosphatidylcholine from liver to ovary
    (Elsevier, 2023-04) Sheng-Xiang Sun; Yi-Chan Liu; Samwel Mchele Limbu; Dong-Liang Li; Li-Qiao Chen; Mei-Ling Zhang; Zhan Yin; Zhen-Yu Du
    Vitellogenins (Vtgs) are essential for female reproduction in oviparous animals, yet the exact roles and mechanisms remain unknown. In the present study, we knocked out vtg1, which is the most abundant Vtg in zebrafish, Danio rerio via the CRISPR/Cas 9 technology. We aimed to identify the roles of Vtg1 and related mechanisms in reproduction and development. We found that, the Vtg1-deficient female zebrafish reduced gonadosomatic index, egg production, yolk granules and mature follicles in ovary compared to the wide type (WT). Moreover, the Vtg1-deficient zebrafish diminished hatching rates, cumulative survival rate, swimming capacity and food intake, but increased malformation rate, and delayed swim bladder development during embryo and early-larval phases. The Vtg1-deficiency in female broodstock inhibited docosahexaenoic acid-enriched phosphatidylcholine (DHA-PC) transportation from liver to ovary, which lowered DHA-PC content in ovary and offspring during larval stage. However, the Vtg1-deficient zebrafish increased gradually the total DHA-PC content via exogeneous food intake, and the differences in swimming capacity and food intake returned to normal as they matured. Furthermore, supplementing Vtg1-deficient zebrafish with dietary PC and DHA partly ameliorated the impaired female reproductive capacity and larval development during early phases. This study indicates that, DHA and PC carried by Vtg1 are crucial for female fecundity, and affect embryo and larval development through maternal-nutrition effects. This is the first study elucidating the nutrient and physiological functions of Vtg1 and the underlying biochemical mechanisms in fish reproduction and development.