非酯化脂肪酸对原代犊牛肝细胞内质网应激信号通路的影响

通过体外添加一定浓度的非酯化脂肪酸(nonestesterifiedfatty acid,NEFAs)(1.8 mmol/L)处理犊牛原代肝细胞不同时间,运用Western blot和实时荧光定量PCR(qRT-PCR)方法,测定NEFAs对内质网应激(ER stress)标志分子IRE1α磷酸化水平和GRP78、CHOP、XBP-1的mRNA表达水平的影响。结果显示,随着NEFAs作用时间的增加,与0.5h组相比,IRE1α磷酸化水平逐渐升高,NEFAs处理小于5h时,IRE1α磷酸化水平呈时间依赖性增加。并在5h达到最高水平,极显著高于0.5h组(P0.01),此后,磷酸化水平降低。与0.5h组相比,GRP78、CHOP、XBP-1的mRNA的表达水平整体逐渐升高,NEFAs处理9h时极显著高于0.5h时组(P0.01)并达到最高水平。结果表明,一定浓度的NEFAs可诱导奶牛肝细胞发生内质网应激。     

非酯化脂肪酸对体外培养犊牛原代肝细胞中炎性因子表达及其活性的影响

在建立犊牛原代肝细胞体外培养模型的基础上,通过培养液中添加不同浓度的非酯化脂肪酸（Nonesterifiedfatty acids,NEFAs）,运用实时荧光定量PCR和ELISA方法,测定NEFAs对肝细胞中炎性因子TNFα、IL-6和IL-1β的mRNA表达及其活性的影响。结果显示,NEFAs作用肝细胞9h后,与对照组相比,高浓度NEFAs（1．8mmol／L）组肝细胞中TNFα、IL-1β的mRNA表达水平显著增加（P〈0．05）,IL-6的mRNA表达水平在2．4retool／L时极显著增加（P〈0．01）,低浓度NEFAs（0．6、1．2mmol／L）组肝细胞中TNFα、IL-1β和IL-6的mRNA表达水平无显著差异（P〉0．05）;并且TNFα、IL-1G的活性在NEFAs浓度达到1．8、2．4mmol／L显著高于对照组（P〈0．01）,且呈剂量依赖性作用,IL-6的活性在在NEFAs浓度达到1．8mmol／L显著高于对照组（P〈0．01）,在2．4mmol／L也高于对照组,但差异不显著。结果表明,高浓度NEFAs在-定程度上能引起或加剧奶牛肝细胞炎性反应,可能是导致产后奶牛肝功能炎性损伤主要因素。     

非酯化脂肪酸对体外培养犊牛原代肝细胞中炎性因子表达及其活性的影响

在建立犊牛原代肝细胞体外培养模型的基础上,通过培养液中添加不同浓度的非酯化脂肪酸(Nonesterified fatty acids,NEFAs),运用实时荧光定量PCR和ELISA方法,测定NEFAs对肝细胞中炎性因子TNFα、IL-6和IL-1β的mRNA表达及其活性的影响。结果显示,NEFAs作用肝细胞9h后,与对照组相比,高浓度NEFAs(1.8mmol/L)组肝细胞中TNFα、IL-1β的mRNA表达水平显著增加(P<0.05),IL-6的mRNA表达水平在2.4mmol/L时极显著增加(P<0.01),低浓度NEFAs(0.6、1.2mmol/L)组肝细胞中TNFα、IL-1β和IL-6的mRNA表达水平无显著差异(P>0.05);并且TNFα、IL-1β的活性在NEFAs浓度达到1.8、2.4mmol/L显著高于对照组(P<0.01),且呈剂量依赖性作用,IL-6的活性在在NEFAs浓度达到1.8mmol/L显著高于对照组(P<0.01),在2.4mmol/L也高于对照组,但差异不显著。结果表明,高浓度NEFAs在一定程度上能引起或加剧奶牛肝细胞炎性反应,可能是导致产后奶牛肝功能炎性损伤主要因素。     

Orai1对NEFA诱导内质网应激引起犊牛肝细胞脂质沉积的影响

为探究Orai1是否参与非酯化脂肪酸(nonestesterifiedfatty acid,NEFAs)诱导内质网应激引起犊牛肝细胞脂质沉积,本试验首先通过体外添加高浓度NEFAs,运用Western blot和实时荧光定量PCR(qRT-PCR)方法,测定NEFAs对内质网应激标志分子GRP78表达量以及脂肪从头合成ACC1、FAS的表达水平的影响。而后又添加Orai1抑制剂2APB与RNA靶向沉默Orai1。结果显示,抑制或沉默Orai1后NEFAs刺激可明显降低GRP78、Orai1、ACC1、FAS的表达水平以及脂滴含量。结果表明,Orai1参与NEFA诱导的内质网应激引起犊牛肝细胞脂质沉积。     

罗格列酮对非酯化脂肪酸体外诱导的犊牛原代肝细胞线粒体功能紊乱的影响

本试验旨在探讨非酯化脂肪酸(Nonestesterified fatty acid,NEFAs)对体外原代培养犊牛肝细胞线粒体功能以及罗格列酮对NEFAs诱导的线粒体功能紊乱的影响。体外添加NEFAs(1.8mmol/L)处理犊牛原代肝细胞不同时间,运用Western blot和实时荧光定量PCR(qRT-PCR)方法检测线粒体呼吸链复合体IV(COX IV)蛋白水平及mRNA水平的影响。另外选取细胞,罗格列酮(10nmol/L)预处理细胞24h后添加NEFAs(1.8mmol/L)处理9h。运用Western blot和qRT-PCR方法检测PGC-1α、Mfn-2的蛋白表达水平及COX IV mRNA表达水平。结果显示,随着NEFAs作用时间的增加,与0h组相比,NEFAs处理5h以上时COX IV蛋白表达水平极显著降低(P0.01),处理7~12h时COX IV mRNA表达水平呈显著降低(P0.05)。添加罗格列酮后,罗格列酮可极显著增加COX IV mRNA水平,PGC-1α、Mfn-2蛋白表达水平(P0.01)。结果表明,NEFAs可诱导犊牛原代肝细胞发生线粒体功能紊乱,罗格列酮可缓解由NEFAs诱导的犊牛原代肝细胞线粒体功能紊乱。     

靶向沉默CaMKK-AMPK通路对NEFAs介导肝脂损伤的调控机制

为探究Ca2+/钙调素依赖性蛋白激酶(CaMKK)-AMP依赖蛋白激酶(AMPK)对非酯化脂肪酸(NEFAs)介导的AML12细胞肝脂损伤的调控机制,本试验通过添加高浓度的NEFAs刺激AML12细胞,RNA靶向沉默CaMKK,检测不同分组细胞内AMPK及固醇调节原件结合蛋白(SREBP1)的蛋白表达水平以及利用流式细...     

非酯化脂肪酸对奶牛氧化应激的诱导作用及其机制

处于围产期的奶牛由于能量负平衡引起脂质动员,导致大量非酯化脂肪酸(NEFA)释放进入肝脏和血液。当NEFA处于较高浓度时,可改变机体抗氧化系统与氧化系统的平衡状态,并激活核因子κB信号通路及丝裂原活化蛋白激酶信号通路,同时也会抑制核因子E2相关因子2信号通路的活化,诱发机体炎性损伤,最终诱导奶牛产生氧化应激,从而直接影响奶牛养殖业的经济效益。本文主要综述了NEFA对围产期奶牛氧化应激的诱导作用及其机制的研究进展,旨在为奶牛生产中缓解氧化应激、提高免疫机能及促进生产性能的发挥提供理论依据。     

非酯化脂肪酸对原代培养犊牛肝细胞胰岛素信号通路的影响

旨在探讨非酯化脂肪酸(nonestesterified fatty acids,NEFAs)对体外原代培养犊牛肝细胞胰岛素信号通路的影响及SIRT1在其中的调控作用。选择培养48h的原代犊牛肝细胞,添加不同浓度的NEFAs(0、0.6、1.2、1.8mmol/L),继续培养12h,胰岛素处理组在收样前用100nmol/L胰岛素处理30min,每个浓度3个重复。另外选取已培养48h的细胞,用SIRT1抑制剂Nicotinamide(10mmol/L)处理12h,胰岛素处理组在收样前用100 mmol/L胰岛素处理30min。运用免疫印迹Western blot方法,检测NEFAs和Nicotinamide对肝细胞胰岛素信号通路关键蛋白IR和Akt的磷酸化水平及SIRT1蛋白表达水平的影响。结果显示,与对照组相比,1.2、1.8 mmol/L NEFAs组胰岛素刺激的IR磷酸化水平显著降低(P0.01),而Akt的磷酸化程度在NEFAs浓度为0.6 mmol/L时即显著降低(P0.01),同时,SIRT1的蛋白表达水平在NEFAs为1.2、1.8mmol/L显著降低(P0.01);添加SIRT1抑制剂显著降低SIRT1的蛋白表达水平(P0.01),同时胰岛素刺激的Akt磷酸化水平也显著降低(P0.01)。结果表明,高浓度的NEFAs可以损伤体外原代培养肝细胞的胰岛素信号通路,SIRT1在NEFAs损伤的胰岛素信号通路中发挥重要作用。     

金属硫蛋白对应激猪血液和肌肉中非酯化脂肪酸及肌乳酸含量的影响

采用驱赶法使乳猪产生应激,探讨金属硫蛋白对应激猪血清与肌肉中非酯化脂肪酸及肌乳酸含量的影响.结果表明:在应激状态下,金属硫蛋白可使血清和运动初期肌肉中的非酯化脂肪酸含量升高,肌乳酸的生成降低,因而可增强有氧代谢,改善机体对运动负荷的压力,提高动物的抗应激能力,因此金属硫蛋白对乳猪具有一定的应激保护作用.     

二脒那秦内源性激活血管紧张素转化酶2对细胞脂质沉积的抑制效应及其机制

旨在通过诱导3T3-L1前脂肪细胞转分化为3T3-L1脂肪细胞,探讨二脒那秦（DIZE）内源性激活血管紧张素转化酶2（ACE 2）对细胞脂质沉积的抑制效应及机制。对3T3-L1前脂肪细胞转分化,将分化成功的3T3-L1脂肪细胞分为对照组、DIZE组（12.5、25 μmol·L^-1处理）,siACE2组和siACE2+DIZE组（siACE2干扰后+25 μmol·L^-1 DIZE）,作用48 h,取上清和细胞进行试验：1）测定细胞上清中三酰甘油和葡萄糖含量;2）油红O染色细胞,并进行定量分析;3） Western blot检测细胞内ACE2和脂肪酸合成关键酶或因子FAS、ACC和SREBP-1c及葡萄糖转运蛋白GLUT4与氧化分解关键酶CS的蛋白表达水平。结果显示：1）成功诱导得到了3T3-L1脂肪细胞,前脂肪细胞诱导至第14天,有95%以上细胞内出现脂滴;2）确定了DIZE作用浓度为12.5和25 μmol·L^-1,处理时间为48 h,并用该药物浓度及时间处理3T3-L1脂肪细胞,细胞上清中三酰甘油含量显著降低（P<0.05）,葡萄糖含量显著升高（P<0.05）;25 μmol·L^-1 DIZE处理组细胞脂滴减少,siACE2组无显著变化,siACE2+DIZE组脂滴蓄积介于对照组和DIZE组之间; 25 μmol·L^-1 DIZE组ACE2蛋白水平显著高于对照组（P<0.05）;siACE2组显著下调（P<0.05）,siACE2+DIZE组较siACE2组无明显变化;3）与对照组相比25 μmol·L^-1 DIZE组细胞中FAS、ACC和SREBP-1c蛋白表达下调,siACE2组和siACE2+DIZE组则表达均上调;4）与对照组相比,25 μmol·L^-1 DIZE组GLUT4和CS蛋白表达水平显著上调（P<0.05）,siACE2组和siACE2+DIZE组则均显著下调（P<0.05）。综上所述,DIZE处理通过介导脂肪细胞ACE2的内源性激活改善了脂肪细胞的脂肪沉积。其机理：一方面抑制脂质合成;另一方面促进葡萄糖摄取和氧化代谢,两方面协同减少了脂肪沉积。结果提示,ACE2或DIZE可作为防控脂肪沉积发生和发展的潜在靶点或药物。     

精料中添加酵母培养物对犊牛生长的影响

对荷斯坦牛犊牛饲喂酵母培养物(达农威益康XP),试验分为两组:试验组(A)和对照组(B),每组10头犊牛,试验组在饲喂基础日粮的基础上每天添加酵母培养物20～25g/头,对照组仅饲喂基础日粮。试验为期60d,并测定犊牛的体重、体高、胸围、采食量等参数,以此来反映犊牛的生长发育情况。试验结果表明:添加了酵母培养物的犊牛在增重、体尺方面差异不显著(P>0.05),但平均日增量试验组高于对照组、平均日采食量试验组低于对照组,在生产实践中体现出的效果和经济效益较明显。     

Effect of dietary fish oil on fatty acid deposition and expression of cholesterol homeostasis controlling genes in the liver and plasma lipid profile: comparison of two animal models

The objective of the present study was to compare hepatic fatty acid deposition, plasma lipid level and expression of cholesterol homeostasis controlling genes in the liver of rats (Wistar Albino; n = 32) and pigs (Large White × Landrace; n = 32) randomly assigned into two groups of 16 animals each and fed 10 weeks the diet with either 2.5% of fish oil (F; source of eicosapentaenoic and docosahexaenoic acid, EPA+DHA) or 2.5% of palm oil (P; high content of saturated fatty acids; control). F‐rats deposited in the liver three times less EPA, but 1.3 times more DHA than F‐pigs (p < 0.05). Dietary fish oil relative to palm oil increased PPARα and SREBP‐2 gene expression much strongly (p < 0.01) in the pig liver in comparison with the rat liver, but expression of Insig‐1 and Hmgcr genes in the liver of the F‐pigs relative to the expression of these genes in the liver of the P‐pigs was substantially lower (p < 0.01 and p < 0.05 respectively) as compared to rats. When plasma lipid concentration in the F‐animals was expressed as a ratio of the plasma concentration in the P‐counterparts, dietary fish oil decreased HDL cholesterol less (p < 0.01), but LDL cholesterol and triacylglycerols more (p < 0.05 and p < 0.001 respectively) in rats than in pigs: more favourable effect of fish oil on rat plasma lipids in comparison with pigs can therefore be concluded. Concentration of total cholesterol and both its fractions in the rat plasma was negatively correlated (p < 0.01) with hepatic DHA, but also with unsaturated myristic and palmitic acid respectively. It has been concluded that regarding the similarity of the plasma lipid levels to humans, porcine model can be considered superior; however, using this model, dietary fish oil at the tested amount (2.5%) was not able to improve plasma lipid markers in comparison with saturated palm oil.     

Effect of microbial phytase on energy availability,and lipid and protein deposition in growing swine

Two experiments were conducted to determine the effect of phytase on energy availability in pigs. In Exp. 1, barrows (initial and final BW of 26 and 52 kg) were allotted to four treatments in a 2 x 2 factorial arrangement. Corn-soybean meal (C-SBM) diets were fed at two energy levels (2.9 and 3.2 x maintenance [M]) with and without the addition of 500 phytase units/kg of diet. The diets contained 115% of the requirement for Ca, available P (aP), and total lysine, and Ca and aP were decreased by 0.10% in diets with added phytase. Pigs were penned individually and fed daily at 0600 and 1700, and water was available constantly. Eight pigs were killed and ground to determine initial body composition. At the end of Exp. 1, all 48 pigs were killed for determination of carcass traits and protein and fat content by total-body electrical conductivity (TOBEC) analysis. Six pigs per treatment were ground for chemical composition. In Exp. 2, 64 barrows and gilts (initial and final BW of 23 and 47 kg) were allotted to two treatments (C-SBM with 10% defatted rice bran or that diet with reduced Ca and aP and 500 phytase units/kg of diet), with five replicate pens of barrows and three replicate pens of gilts (four pigs per pen). In Exp. 1, ADG was increased (P < 0.01) in pigs fed at 3.2 x M. Based on chemical analyses, fat deposition, kilograms of fat, retained energy (RE) in the carcass and in the carcass + viscera, fat deposition in the organs, and kilograms of protein in the carcass were increased (P < 0.10) in pigs fed the diets at 3.2 vs. 2.9 x M. Based on TOBEC analysis, fat deposition, percentage of fat increase, and RE were increased (P < 0.09) in pigs fed at 3.2 x M. Plasma urea N concentrations were increased in pigs fed at 3.2 x M with no added phytase but were not affected when phytase was added to the diet (phytase x energy, P < 0.06). Fasting plasma glucose measured on d 28, ultrasound longissimus muscle area (LMA), and 10th-rib fat depth were increased (P < 0.08) in pigs fed phytase, but many other response variables were numerically affected by phytase addition. In Exp. 2, phytase had no effect (P > 0.10) on ADG, ADFI, gain:feed, LMA, or 10th-rib fat depth. These results suggest that phytase had small, mostly nonsignificant effects on energy availability in diets for growing pigs; however, given that phytase increased most of the response variables measured, further research on its possible effects on energy availability seems warranted.     

基于DISCERN算法筛选影响肉鸡脂肪沉积的关键基因

选取法国高低脂2组7周龄肉鸡腹部脂肪的RNA測序转录组数据中的17 934个基因,采用差分稀疏调节网络(differential sparsE regulatory network,DISCERN)算法对相应数据进行分析,以期筛选出影响肉鸡腹部脂肪沉积的关键基因,并识别肉鸡不同基因表达数据之间网络差异关系,推断基因调控...     

Substitution of milk protein with isolated soy protein in calf milk replacers

The influence of replacement of milk protein by isolated soy protein on digestion and pancreatic enzyme secretion was determined in nine Holstein male calves. Calves (average weight 47 kg) were fitted with permanent re-entrant pancreatic and a T-type cannula in the distal ileum at 6 to 10 d of age. Following a 2-wk recuperation period, the calves were fed three milk replacers in a triplicated 3 x 3 latin square. Experimental diets consisted of a control, in which 100% of the CP originated from spray-dried skim milk powder (SM), and the test diets, in which 50% (SM/ISP) or 100% (ISP) of the skim milk protein was replaced by isolated soy protein. Each experimental period lasted 2 wk. Replacement of SM protein by ISP decreased (P less than .05) the digestibilities of protein and most amino acids. Ileal digestibilities of total indispensable amino acids for SM, SM/ISP and ISP diets were 82.1, 75.8 and 61.8%, respectively, and total tract digestibilities of total indispensable amino acids were 90.0, 82.6 and 74.0%, respectively. Including ISP did not affect (P greater than .05) the volume of secretion of pancreatic juice, protein or chymotrypsin; however, the secretion of trypsin decreased (P less than .05). Reduction in trypsin secretion may be responsible, in part, for the lower amino acid digestibilities in milk replacers containing isolated soy protein.     

Studies on the cytopathology of bovine adenoviruses in calf testis and calf kidney cell cultures

The sequential development of intranuclear inclusions in calf kidney and calf testis cells infected with nine bovine adenovirus (BAV) serotypes is described. Haematoxylin and Eosin (H & E), immunofluorescent and electron microscope (EM) studies indicated two distinct subgroups of viruses. Serotypes 1, 2, 3, 7 and 9 comprised subgroup-1, while types 4, 5, 6 and 8 comprised subgroup-2. Differences were noted in the early stages of infection. With subgroup-1 viruses, irregular patches of eosinophilic material were first to appear, followed by small refractile inclusions and basophilic inclusions. In EM studies, the eosinophilic material was thought to correspond to the irregular type II inclusions, and the refractile bodies to type I inclusions. Eventually a basophilic inclusion, consisting of aggregated virus-associated inclusions and virus particles, was formed in the centre of the nucleus.With subgroup-2, the refractile inclusions were more prominent, larger, and were the first to appear. These were thought to correspond to type I inclusions, which were larger and denser than with subgroup-1. Circular basophilic bodies developing later were similar in size and distribution to type II inclusions, which with subgroup-2 viruses were seen in prominent circular or lobulated aggregates.Several other types of inclusion including tubular structures and paracrystals, which have thus far not been reported in BAV infected cells, are described.     

Serum trimethylamine-N-oxide and gut microbiome alterations are associated with cholesterol deposition in the liver of laying hens fed with rapeseed meal

Sinapine derived from cruciferous plants could be converted into trimethylamine by intestinal microbiota. Its metabolite, trimethylamine N-oxide (TMAO), is closely linked to increased risk of cardiovascular disease and fat deposition in mammals. Hens fed with rapeseed meal (RSM) suffered from fatty liver hemorrhage syndrome (FLHS). This study was conducted to investigate whether RSM-induced fatty liver is due to TMAO via altering microbiota composition and diversity. At 33 weeks of age, 600 laying hens were randomly divided into 5 treatment groups, namely control and 14% RSM treatment groups (DY5, with 16.2% erucic acid [EA] and 74.66% glucosinolate [Gl] contents; MB1, with 3.50% EA and 43.23% Gl contents; DY6, with 6.7% EA and 22.67% Gl contents; XH3, with 44.60% EA and 132.83% Gl contents) for 8 weeks. Results revealed that 3 hens died due to liver hemorrhage after ingesting 14% RSM diet. The 14% RSM decreased serum low-density lipoprotein cholesterol (LDL-C) content (P < 0.01) while tended to increase serum TMAO content compared to the control group (P = 0.08). The 14% RSM diet increased red oil O optical density (P < 0.01), and increased total cholesterol (TC) and LDL-C content in the liver (P < 0.01, and P < 0.01, respectively). The 14% RSM decreased liver total bile acid (TBA) content compared to the control (P < 0.01). The DY6 had a higher TBA content in the liver than the XH3 (P < 0.01). The 14% RSM decreased mRNA abundance of liver X receptors alpha (LXR-α, P = 0.01), and increased mRNA abundance of sterol response element binding protein 2 (SREBP-2, P = 0.04). Results revealed that the in-feed RSM could alter richness and diversity of cecal microbiota compared to the control (P < 0.05). Liver TC content and serum TMAO showed a negative relationship with Proteobacteria and Actinobacteria (P = 0.04). In conclusion, 14% RSM increased liver TC and induced high liver score of FLHS, which was possibly associated with the altered cecal microbiota composition, increased serum TMAO levels and LXR-α and SREBP-2 expressions.