Differential expression of lipid metabolism‐related genes and miRNAs in Ctenopharyngodon idella liver in relation to fatty liver induced by high non‐protein energy diets

To explore the mechanism of fatty liver formation induced by high non‐protein energy diets in grass carp (Ctenopharyngodon idella), basal diet and high‐energy diets were fed to juvenile grass carp for 9 weeks. The experimental groups fed on high‐energy diets which included a high‐lipid diet (H‐LIP), a high‐carbohydrate diet (H‐CHO) and a high‐lipid and carbohydrate diet (H‐CL). The control group fed on basal diet. Growth performance, liver fat accumulation, serum biochemical indexes and the expression levels of lipid metabolism‐related genes (SREBP‐1, PPARγ, FAS, ACC1, and LPL) and miRNAs (miR‐33, miR‐122, and miR‐370) were examined at the end of the feeding trial. There were no significant differences in growth rate and feed efficiency among the four groups. However, significant increase in mesenteric and liver fat contents, and lipid droplets in the liver was induced by high‐lipid and high‐carbohydrate diets. There were significant differences in serum biochemical indicators such as AST/ALT, GLB, TG and TP, and liver fatty acid composition between the control and experimental groups. The expression levels of SREBP‐1, PPARγ, FAS, ACC1 and LPL were upregulated, while CPT‐1 was downregulated with the high‐energy treatments. Additionally, the expression levels of miR‐33, miR‐122 and miR‐370 in the liver were higher in the three high‐energy treatments than those in the control (P < 0.05). The results suggest that modifications of lipid metabolism‐related genes and miRNAs may be involved in fatty liver formation induced by high non‐protein energy diets in grass carp.     

Influence of different dietary lipid sources on the growth,tissue fatty acid composition,histological changes and peroxisome proliferator‐activated receptor γ gene expression in large yellow croaker (Pseudosciaena crocea R.)

A 9‐week feeding trial was conducted to investigate the impact of dietary lipid sources on the lipid mechanisms of large yellow croaker by feeding three isonitrogenous and isoenergetic diets containing fish oil (FO), soybean oil (SO) and beef tallow (BT) respectively. The effects of the diets on the growth performance, somatic indices, tissue fatty acid composition, histological changes and peroxisome proliferator‐activated receptor (PPAR)γ expression were evaluated. Experimental diets were all well accepted by fish and no significant (P>0.05) differences were found in the weight gain, growth rate and feed conversion rate. The fatty acid profile of the fish fillet and liver reflected the fatty acid composition of the diets. Specific‐fatty acids were selectively retained, however, in the flesh of the fish; in particular, docosahexaenoic acid and arachidonic acid concentrations were higher than the dietary concentrations. When FO was replaced by SO or BT diets, the reduction in eicosapontaenoic acid in fish tissue was more pronounced, suggesting a preferred utilization of this fatty acid. The consumption of SO displayed intense lipid accumulation in the liver of the fish. The expression of PPARγ increased significantly in the SO group compared with the FO and BT groups (P<0.05).     

Effects of dietary non‐protein energy source levels on growth performance,body composition and lipid metabolism in herbivorous grass carp (Ctenopharyngodon idella Val.)

A study was conducted to determine the effects of dietary non‐protein energy sources on growth, tissue lipid accumulation and lipid metabolism‐related genes expression of grass carp. Triplicate groups of fish were fed for 9 weeks on four isonitrogenous (300 g kg^?1) experimental diets with four levels of non‐protein energy (6.52 kJ g^?1 control diet, 5.32 kJ g^?1 high‐CEL diet, 8.46 kJ g^?1 high‐CHO diet and 8.53 kJ g^?1 high‐LIP diet respectively). Increasing dietary non‐protein energy source levels did not improve the growth, and the high‐CEL diet reduced the growth of grass carp. The high‐CHO diet tended to induce high hepatosomatic index, with high fat and glycogen content of liver. However, the high‐LIP diet caused the high mesenteric fat index, but did not increase liver fat. The mRNA abundance and activities of hepatic lipogenic enzymes were significantly increased in the high‐CHO diet group, whereas the opposite tendencies were observed in the high‐LIP diet group. Peroxisome proliferator‐actived receptor‐α (PPARα) in liver and PPARγ in mesenteric adipose tissue were up‐regulated in the high‐CEL diet group. Lipoprotein lipase (LPL) gene expression was significantly increased both in liver and mesenteric adipose tissue of fish fed the high‐LIP diet, while the LPL gene expression was up‐regulated in liver but down‐regulated in mesenteric adipose tissue of fish fed the high‐CEL diet. These findings suggest that an increase in dietary non‐protein energy sources alters the genes expression of lipid metabolism and increased lipid deposition.     

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, β-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.     

Oral bovine serum albumin immune‐stimulating complexes improve the immune responses and resistance of large yellow croaker (Pseudosciaena crocea,Richardson, 1846) against Vibrio alginolyticus

This study aimed to investigate effects of bovine serum albumin immune‐stimulating complexes (BSA ISCOMs) on immune‐related genes expression, serum nonspecific immunity and disease resistance of large yellow croaker (Pseudosciaena crocea). Fish were fed diets containing 3.5 ml of BSA ISCOMs per kg feed (experimental group) or 3.5 ml of phosphate‐buffered saline per kg feed (control group) for 1 week. The liver, spleen, head‐kidney tissues were sampled for determining gene expression of myxovirus‐resistant protein (Mx), major histocompatibility complex class II alpha chain (MHC II α), tumour necrosis factor‐alpha (TNF‐α) and interleukin‐10 (IL‐10) 30 and 90 days after feeding. Also, blood samples were collected for determining activities of serum superoxide dismutase (SOD), interferon alpha (IFN‐α), TNF‐α and alkaline phosphatase (ALP). TNF‐α and MHCⅡα gene expression in the liver, spleen, head‐kidney, as well as IFN‐α, TNF‐α and ALP activities in the serum, of experimental fish were significantly higher 30 days after feeding; while only TNF‐α and MHC II gene expression in the head‐kidney remained upregulated 90 days after feeding. The cumulative mortality of the experimental fish was significantly lower than control. This study indicated that BSA ISCOMs improved the immune response and induced protective immunity in large yellow croaker.     

Berberine attenuates oxidative stress and hepatocytes apoptosis via protecting mitochondria in blunt snout bream <Emphasis Type="Italic">Megalobrama amblycephala</Emphasis> fed high-fat diets

High-fat diets may have favorable effects on growth and cost, but high-fat diets often induce excessive fat deposition, resulting in liver damage. This study aimed to identify the hepatoprotective of a Chinese herb (berberine) for blunt snout bream (Megalobrama amblycephala). Fish were fed with a normal diet (LFD, 5 % fat), high-fat diet (HFD, 15 % fat) or berberine-supplemented diets (BSD, 15 % fat with berberine 50 or 100 mg/kg level) for 8 weeks. After the feeding, histology, oxidative status and mitochondrial function of liver were assessed. The results showed that HFD caused fat accumulation, oxidative stress and apoptosis in hepatocytes of fish. Hepatocytes in HFD group appeared to be hypertrophied, with larger liver cells diameter than these of LFD group. Berberine-supplemented diets could attenuate oxidative stress and hepatocytes apoptosis. HFD induced the decreasing mitochondrial complexes activities and bulk density and surface area density. Berberine improved function of mitochondrial respiratory chain via increasing the complex activities. Moreover, the histological results showed that berberine has the potential to repair mitochondrial ultrastructural damage and elevate the density in cells. In conclusion, our study demonstrated that berberine has attenuated liver damage induced by the high fat mainly via the protection for mitochondria.     

禁食对养殖大黄鱼体成分、肌肉脂肪酸组成和血清生化指标的影响

为了探讨禁食对大黄鱼体成分、肌肉脂肪酸组成和血清生化指标的影响,从海上网箱选择规格相近、健康的养殖大黄鱼105尾[平均体质量(249.07±7.28)g]进行禁食实验,分别在实验开始时及禁食后第3、7、14、21、28、35、42和49天采集实验鱼肌肉和血液样品,进行肌肉营养成分和血清生化指标的测定。结果发现,随着禁食时间的延长,大黄鱼肌肉中水分和粗灰分含量呈现升高的趋势,粗脂肪含量呈现下降的趋势,粗蛋白含量在禁食后第21天下降后基本保持稳定;肌肉中饱和脂肪酸(SFA)和单不饱和脂肪酸(MUFA)含量均无显著变化,多不饱和脂肪酸(PUFA)和n-3系列高度不饱和脂肪酸(n-3HUFA)总量呈现下降的趋势。禁食后第3天血清中的谷丙转氨酶(ALT)、谷草转氨酶(AST)、乳酸脱氢酶(LDH)、葡萄糖(GLU)、钾离子(K~+)、钙离子(Ca~(2+))、钠离子(Na~(+)))和氯离子(Cl~-)含量显著上升;甘油三酯(TG)、总胆固醇(CHOL)含量显著下降;血清总蛋白(TP)、高密度脂蛋白胆固醇(HDL-C)、低密度脂蛋白胆固醇(LDL-C)和碱性磷酸酶(ALP)无显著变化。研究表明,随着禁食时间的延长,血清中除了Na~+和Cl~-含量出现波动性上升外,血清中的葡萄糖、蛋白质、脂类、K~+和Ca~(2+)含量以及血清代谢酶活性均出现波动式下降,表明大黄鱼禁食期间主要以脂类作为能量来源,体内营养物质代谢活动强度呈波浪形的起伏变化,随着禁食时间的延长机体代谢活动趋于减弱,但免疫功能和代谢机能未受到明显影响。     

Dietary salmon milt deoxyribonucleic acid prevents hepatosteatosis in mice

With the increased incidence of lifestyle-related diseases, prevalence of nonalcoholic fatty liver disease (NAFLD) is increasing at an unprecedented rate, leading to rising interest in studying the role of functional foods in its prevention. This study demonstrated the efficacy of salmon milt deoxyribonucleic acid (SM DNA) in preventing liver dysfunction in a diet-induced obesity model. Seven-week-old male C57BL/6J mice were fed normal diet (ND), high-fat diet (HFD), and HFD supplemented with 3% SM DNA (HFD?+?SM DNA) for 12 weeks. Increased body weight, fat-to-body weight ratio, and fat accumulation in HFD-fed mice demonstrated successful establishment of an obesity model. The HFD group showed increased alanine aminotransferase (ALT) levels, a marker of liver function, and NAFLD activity score (NAS) based on histological observations showed steatosis and mild inflammation but no fibrosis, indicating NAFLD but no nonalcoholic steatohepatitis. In the HFD?+?SM DNA group, the improved superoxide dismutase activity and NAFLD activity score suggested that SM DNA prevented liver dysfunction while maintaining antioxidant capacity and suppressing intrahepatocyte fat droplet formation. SM DNA attenuated fat accumulation in the liver and improved hepatic function. This study explored the potential of SM DNA as a functional food to prevent NAFLD and other visceral obesity-related diseases.

     

Short‐term starvation in silver pomfret (Pampus argenteus): molecular effects on lipid mobilization and utilization

To investigate the effects of short‐term starvation on lipid metabolism, juvenile silver pomfret (Pampus argenteus) with an initial weight of 18.1 ± 1.53 g were starved for 6 days. Fish were sampled on days 0, 2, 4 and 6 of starvation (S0, S2, S4 and S6, respectively) and then analysed. The results showed that short‐term starvation induced a significant decrease in visceral index, hepatosomatic index. The crude lipid content was also significantly decreased by 42.22% and 82.96% on S2 and S6 respectively. In addition, short‐term starvation significantly decreased the plasma triacylglycerol, non‐esterified fatty acid and low‐density lipoprotein cholesterol. Moreover, short‐term starvation significantly increased the expressions of hormone‐sensitive lipase (HSL) and adipose triglyceride lipase (ATGL) at the mRNA and protein levels in the muscle and the ATGL expression at the protein level in the liver. The expressions of fatty acid translocase (CD36) and plasma membrane fatty acid‐binding protein (FABPpm) at the protein level in the muscle were significantly increased by short‐term starvation, while the expressions of fatty acid transport protein 1 (FATP1), FABPpm and lipoprotein lipase (LPL) at the protein level in the liver were significantly increased by short‐term starvation. Furthermore, the expressions of peroxisome proliferator activated receptor (PPAR) α and PPARγ at the mRNA and protein levels in the muscle were significantly elevated by short‐term starvation. These findings suggested that short‐term starvation increased lipid mobilization and utilization possibly through activation of lipolysis‐related genes (HSL and ATGL), lipid uptake‐related genes (LPL, CD36, FATP1 and FABPpm) and PPARs.     

Dietary n‐3 highly unsaturated fatty acids affect the biological and serum biochemical parameters,tissue fatty acid profile,antioxidation status and expression of lipid‐metabolism‐related genes in grass carp,Ctenopharyngodon idellus

A 95‐day feeding trial was conducted to determine the effects of n‐3 highly unsaturated fatty acids (n‐3 HUFA) on the growth, antioxidation and lipid metabolism in grass carp (Ctenopharyngodon idellus). Two isonitrogenous and isolipidic diets were formulated with either lard oil (LO) or fish oil (FO) as the main lipid source. The results showed that the intraperitoneal fat (IPF) ratio was significantly lower (P < 0.05) in FO group. The concentration of n‐3 HUFA in muscle, hepatopancreas and IPF was significantly higher in FO group (P < 0.05). The serum low‐density lipoprotein (LDL) content was significantly lower (P < 0.05), and glucose (GLU) content was significantly higher (P < 0.05) in FO group. The serum total superoxide dismutase (T‐SOD) activity was significantly higher (P < 0.05) in FO group, consistent with the serum malondialdehyde (MDA) content. The gene expression of IPF fatty acid synthase (FAS), acetyl‐CoA carboxylase (ACC), sterol regulatory element‐binding protein (SREBP‐1) and peroxisome proliferator‐activated receptor γ (PPARγ) was significantly lower (P < 0.05) and that of peroxisome proliferator‐activated receptor α (PPARα) was significantly higher (P < 0.05) in FO group compared with LO group. Similar trends were found in the hepatopancreas, except for PPARγ. It is suggested that n‐3 HUFA could inhibit lipid accumulation in grass carp by affecting the expression of lipid‐metabolism‐related genes.     

Effects of dietary tea polyphenols on growth,biochemical and antioxidant responses,fatty acid composition and expression of lipid metabolism related genes of large yellow croaker (Larimichthys crocea)

The study was conducted to investigate the effects of dietary tea polyphenols (TP) on growth performance, biochemical and antioxidants responses, fatty acid composition, and lipid metabolism‐related gene expressions of large yellow croaker (Larimichthys crocea). Four diets were formulated with different levels of TP (0.00%, 0.01%, 0.02% and 0.05%). Results showed that growth performance of L. crocea were not different among dietary treatments. Compared with the control group, fish in 0.02% TP group had lower body and hepatic lipid content and lower total cholesterol content. The minimum content of triglycerides and low‐density lipoprotein‐cholesterol were found in 0.05% TP group. Hepatic n‐6 PUFA and n‐3 PUFA were significantly higher in TP supplementation groups. Malondialdehyde content was lower in TP supplementation groups, and superoxide dismutase activity was higher in 0.01% TP group than the control group. The mRNA expressions of carnitine palmitoyltransferase1, acyl‐CoA oxidase and peroxisome proliferators‐activated receptor α were up‐regulated in 0.01% and 0.02% TP groups, while lipoprotein lipase expression was down‐regulated in TP supplementation groups than the control group. Results suggested that 0.01%–0.02% TP supplementation could reduce the deposition of liver lipid of L. crocea caused by high‐lipid diet, which might be due to the increase in lipid oxidation related gene expressions.     

柱形病对黄颡鱼血液生化指标的影响

为探究柱形病对黄颡鱼(Pelteobagrus fulvidraco)生理机能的影响,随机选取患病和健康黄颡鱼各10尾,采集其血液,静置后离心分离血清以测定血液生化指标如皮质醇、乳酸(LD)含量,乳酸脱氢酶(LDH)、碱性磷酸酶(ALP)活性,葡萄糖、白蛋白(ALB)、总蛋白(TP)、尿素氮(BUN)、总胆固醇(TCHO)、高密度脂蛋白胆固醇(HDLC)、低密度脂蛋白胆固醇(LDLC)含量,谷草转氨酶(AST)、谷丙转氨酶(ALT)活性。结果显示:患病黄颡鱼血清皮质醇浓度和LDH、AST活性均极显著高于健康鱼,LD含量和ALT活性显著高于健康鱼。与健康鱼相比,患病黄颡鱼血清葡萄糖含量极显著降低,而HDLC、LDLC、TCHO、ALB、TP、BUN浓度和ALP活性无显著差异。健康鱼肝细胞呈近圆形,形状较规则,细胞界线清晰,细胞核多位于细胞中央,呈圆形或椭圆形,肝细胞以中央静脉为中心形成放射状排列。患病鱼肝细胞出现局部坏死、空泡化,形成坏死灶,伴有大量中性粒细胞、淋巴细胞浸润。结果表明:柱状黄杆菌感染可引起黄颡鱼鱼体显著的应激反应,并引起鱼体肝脏损伤。     

Effects of total fish oil replacement to vegetable oils at two dietary lipid levels on the growth,body composition,haemato‐immunological and serum biochemical parameters in caspian brown trout (Salmo trutta caspius Kessler, 1877)

This research aimed to evaluate the effects of two dietary fat levels [low fat (LF) (10%), high fat (HF) (20%)] and sources [fish oil (FO), vegetable oil (VO)] on the growth and some physiological parameters of Caspian brown trout fingerlings for 60 days. Tuna oil or blends of canola and soybean oils (85:15) were added to diets to design four feeds namely LFFO, HFFO, LFVO and HFVO according to the fat levels and sources. The fish fed the LFFO diet had lower weight gain than the other fish (P<0.05). The total n‐6 fatty acids increased in fish fed diets with the blends of VO, while the total n‐3 fatty acids decreased in these fish (P<0.05). Serum lysozyme activity was higher in fish fed the HFVO diet than the other fish (P<0.05). Serum glucose, total cholesterol, triglyceride and very low‐density lipoprotein were lower in fish fed LFFO than the other fish (P<0.05). The present study demonstrates that in terms of fish growth, VOs can be used as an alternate source of dietary fat, whereas fish health and nutritional value are improved with the LFFO diet. According to these results, a partial substitution of FO by VO in high‐level fat diets is suggested for long‐term feeding of Caspian brown trout.     

斑马鱼营养性脂肪肝模型构建

近年来我国集约化水产养殖业发展势头强劲,而水产动物营养性疾病之鱼类脂肪肝的发病也逐渐迅猛起来,逐渐成为制约水产养殖业发展的因素之一,实验拟通过高脂饲料诱导建立斑马鱼营养性脂肪肝模型,为鱼类营养性脂肪肝相关研究工作的开展奠定基础。该实验分别以基础饲料和16%高脂饲料投喂1月龄斑马鱼,在投喂后第14、30、90天3个时间点取样,采用肝脏组织进行H.E染色、透射电镜、masson染色组织切片观察斑马鱼肝脏脂肪变性及纤维化程度;通过荧光定量PCR技术(q RT-PCR)检测斑马鱼肝脏脂合成及脂解相关基因:二酰甘油酰基转移酶基因(DGAT)、过氧化物酶体增殖物激活受体基因(PPAR)、胆固醇调节元件1基因(SREBP1)、脂肪酸合成酶基因(FAS)、诱导细胞死亡脱氧核糖核酸裂解因子样效应因子基因(CIDE)、激素敏感脂肪酶基因(HSL)、甘油三酯水解酶基因(ATGL)的表达水平;采用甲醇–氯仿法提取斑马鱼肝脏脂肪并检测肝脏中甘油三酯的含量。结果显示,在高脂饲料诱导第30天,光镜下可见发生弥漫性肝细胞脂肪变性,脂合成基因显著上调,甘油三脂的含量显著增加;诱导第90天,经透射电镜及masson染色组织切片观察结果证实,斑马鱼肝脏组织由于纤维化病变而使得肝组织功能受损,肝脏的脂合成基因及甘油三脂含量显著降低。研究表明,实验设计的高脂饲料对肝脏的损伤作用显著,30 d的投喂即可诱导发生脂肪肝,90 d投喂斑马鱼肝脏出现肝纤维化,此过程中肝脏脂合成基因表达量及甘油三酯含量先升高后降低,可知斑马鱼肝脏的损伤是由高脂饲料诱导的脂肪大量堆积而引起的,该营养性模型的成功构建在一定程度上弥补了以往构建斑马鱼高脂模型的缺陷,为研究鱼类营养性脂肪肝的发生机制提供了支撑。     

Influence of dietary biotin levels on growth and non‐specific immune response in large yellow croaker,Larimichthys crocea R

A 9‐week feeding experiment was conducted to determine the effect of dietary biotin levels on growth performance and non‐specific immune response of large yellow croaker. Fish (6.16 ± 0.09 g) were fed twice daily to apparent satiation with diets containing 0.00 (as the basal diet), 0.01, 0.05, 0.25, 1.24 and 6.22 mg biotin kg^?1 diet. Results showed that fish fed the basal diet had the lowest survival rate, and fish fed 0.05 mg kg^?1 dietary biotin achieved significantly higher final weight and weight gain. Dietary biotin levels had no significant influence on carcass crude lipid, moisture and ash content, but significantly influenced the carcass crude protein. Liver biotin concentration significantly increased with the supplementation of biotin, but no tissue saturation was found within the supplementation scope of biotin. Broken‐line regression analysis of weight gain showed that juvenile large yellow croaker requires a minimum dietary biotin of 0.039 mg kg^?1 for maximal growth. The analyses of serum parameters showed that the moderate‐ (0.05 mg kg^?1) and high‐dose (6.22 mg kg^?1) dietary biotin significantly improved both lysozyme and alternative complement pathway activities, indicating dietary biotin within a certain range could improve the non‐specific immune response of large yellow croaker.