【作者】 周继术；

【导师】 王建华； Bente E.Torstensen；

【作者基本信息】 西北农林科技大学 ， 临床兽医学， 2010， 博士

【摘要】 鲤属我国的传统淡水杂食性鱼类,大西洋鲑属海洋肉食性鱼类,该类鱼均能为人类提供优质蛋白源及人类必需的高不饱和脂肪酸。鱼油中的高不饱和脂肪酸(highly unsaturated fatty acid, HUFA)能预防人类心脑血管疾病等,已成为人类保健食品的开发对象。脂质也是动物生长繁殖的必需营养元素,在水产饲料中添加鱼油是传统的油脂添加类型,可为鱼类的生命活动提供能量和必需脂肪酸,但鱼油及其HUFA对鱼类尤其是淡水鲤的营养及健康养殖(如机体抗氧化系统等)的作用效果及机制还未充分研究。同时,随着全球鱼类资源的衰竭,鱼油供应逐渐短缺,鱼油价格的持续上涨,严重影响到了水产饲料行业的可持续发展,尤其对饲料中油脂添加水平可达30%的肉食性大西洋鲑的饲料行业产生了较大影响,寻找鱼油替代品已成为研究热点,其中较为廉价的植物油则成为替代鱼油的首选目标。然而,植物油以及鱼油的脂肪酸组成有较大差异,如鱼油富含HUFA而植物油则缺乏等特点,这将使植物油替代对水产动物脂质的营养作用和代谢过程产生影响,但相关研究尚需进一步探讨。为此,本论文开展了一系列试验研究,旨在为鱼油及HUFA对淡水鱼类脂质营养代谢的影响及健康养殖提供参考资料,同时为肉食性海水鱼类饲料中植物油替代鱼油的可能性提供理论数据,为水产饲料行业的可持续发展提供有益参考。1.鱼油对鲤生长及脂质代谢的影响为探讨鱼油对鲤生长、脂质代谢的影响,进行了鱼油饲养及HUFA灌喂两个试验。试验1,将尾均重(43.0±2.8) g鲤随机分为2组,每组两个重复,每重复19尾,分别饲以鱼油(鱼油组)和豆油(对照组)为脂肪源的两种半纯化饲料。试验2,以剂量750 mg HUFA/kg鱼体重对鲤(23.5±0.5)g进行一次灌喂并于灌喂后0,1,2,4,6,12,24 h取肝胰脏样品。每个处理3个重复。结果表明:试验1中:①鱼油组饲料转化效率、绝对生长率等与对照组差异不显著;②鱼油组腹腔脂肪指数显著高于对照组(P< 0.05),但肥满度、空壳比率等生物学性状与对照组无显著差异;③鱼油组肝胰脏苹果酸脱氢酶(MDH)、脂蛋白酯酶(LPL)活性均显著低于对照组(P< 0.05),肝胰脏葡萄糖6磷酸脱氢酶(G6PDH)活性与对照组差异不显著。试验2中:HUFA组肝胰脏MDH活性于2 h、4 h显著低于对照组(P< 0.05),而在8 h、16 h、32 h两组间差异不显著。结果显示:鲤能较好利用鱼油和豆油用于生长;鱼油可抑制鲤的肝胰脏脂质合成及外源脂质进入肝胰脏,影响脂质在组织间的分配。这些作用与其富含的HUFA有关。2.二十二碳六烯酸对鲤机体抗氧化能力的影响为了研究二十二碳六烯酸(Docosahexaenoic acid;DHA)对鲤机体抗氧化能力的影响,进行了DHA灌喂(短期)、DHA饲养(长期)和DHA与肝细胞离体培养三个试验。DHA灌喂试验方法参照试验1。在DHA饲养试验中,将尾均重(14.81±0.13)g鲤平均分为四个组,每组三个重复,每个重复30尾鱼。各组分别饲以6 %及12 %油脂水平下添加0及1.5% DHA的四组饲料。饲养试验在室内循环流水水族箱(200L/箱)中进行,饲养期为74天。试验结束后对鲤肝胰脏及血清进行采样并进行相关抗氧化指标的测定。在DHA细胞培养试验中,对1kg体重的上市规格鲤肝细胞进行分离和培养并用0.152mol/L DHA制剂进行处理,以无DHA处理为对照,于0,0.5,1,2,4,8 h后分别收取肝细胞样,测定其相关抗氧化指标。抗氧化指标分别为总抗氧化能力(T-AOC)、总超氧化物歧化酶(T-SOD)、谷胱甘肽S转移酶(GST)、过氧化氢酶(CAT)活性、丙二醛(MDA)含量等。结果表明:①DHA组鲤肝胰脏T-AOC水平在灌喂后的各时间点上均高于对照组,且在32 h时达到显著水平(P<0.05)。②两个油脂水平下,DHA组鲤肝胰脏T-AOC水平、CAT、SOD活性及MDA含量DHA组与对照组间差异均不显著(P>0.05);血清T-AOC水平、SOD活性DHA组与对照组间差异也不显著(P>0.05);12%油脂水平下DHA组GST活性显著低于对照组(P<0.05),但6 %油脂水平下GST活性在两组间差异不显著(P>0.05)。③鲤肝细胞用DHA分别处理0.5 h和1 h后,MDA含量与对照差异不显著(P>0.05),而肝细胞在DHA分别处理2 h、4 h、8 h后,MDA含量显著高于对照组(P<0.05),且在4 h、8 h时达极显著水平(P<0.01);DHA组T-SOD活性在处理后4 h时显著高于对照组(P<0.05),而在其它时间点上DHA组与对照组间差异均不显著(P>0.05)。研究表明:DHA直接或短期作用下,鲤肝胰脏或肝细胞处于氧化应激状态,其抗氧化能力诱导性升高;而DHA长期饲喂后,在正常油脂水平下机体表现出对DHA造成的氧化应激的适应,高油脂水平下DHA的添加则可能通过加剧机体氧化应激状态,造成了抗氧化能力的下降。本研究所采用的短期灌喂、长期饲养以及肝细胞离体培养相结合的研究手段可用于外源性物质对鱼类生理活性的评定。3.大西洋鲑肝细胞对长链脂肪酸的吸收及细胞内代谢的研究3.1大西洋鲑肝细胞对油酸吸收随时间的变化特点目的:选择大西洋鲑肝细胞对油酸的最佳吸收时间。方法:油酸(60μM)与[1-14C]油酸按60:1比例混合在L-15不完全培养液(无胎牛血清的L-15完全培养液)中,与培养12 h的贴壁大西洋鲑肝细胞与共培养(0.2μCi/培养瓶),分别于0.5、1、2、4、6、12和24 h后吸取培养液以终止细胞的吸收反应,并收取细胞样。用闪烁计数器测定细胞及培养液放射性,同时用氯仿:甲醇(2:1)提取细胞及培养液内脂质,用高效薄层色谱(HPTLC)分离各类脂质(磷脂、一酰、二酰甘油、三酰甘油及自由脂肪酸),碘蒸汽内显色并与标准品对照以识别并标记出各类脂质。刮取各类脂质于闪烁瓶中,用闪烁计数器测定其放射性。通过油酸与[1-14C]油酸的比例,换算出油酸的吸收量。结果:0~2 h内细胞对油酸的吸收线性增加,4~6 h增长缓慢,12 h以后逐渐达到吸收的饱和状态。结果表明: 2 h属于大西洋鲑肝细胞对油酸吸收的最佳有效培养时间。3.2不同油酸培养浓度下大西洋鲑肝细胞对油酸吸收的变化特点目的:选择大西洋鲑肝细胞对油酸吸收的最佳油酸培养浓度。方法:油酸(0, 37.5, 75, 150, 300, 600μM)与[1-14C]油酸按30:1比例分别混合在L-15不完全培养液中,与培养12 h的贴壁大西洋鲑肝细胞共培养(各浓度下放射性分别为0, 0.13, 0.24, 0.49, 0.98, 1.98μCi/培养瓶), 2 h后按3.1所述方法取样并进行样品放射性测定和计算油酸吸收量。结果:大西洋鲑肝细胞对油酸呈饱和吸收状态,饱和吸收常数(Km)为47μM,最大吸收速度(Vmax)为2.3 nmol h-1 million cells-1。表明: 37.5μM油酸浓度可用于大西洋鲑肝细胞对油酸的吸收试验。3.3大西洋鲑肝细胞吸收油酸的抑制试验目的:了解大西洋鲑肝细胞对油酸的吸收过程。方法:膜蛋白抑制剂细胞前处理培养液(前处理液)的配制:将选取的膜蛋白抑制剂,环氧糊精,根皮素,二异氰酸二磺酸(DIDS),硫基琥珀酰亚胺马来酰亚胺油酸酯(SSO)和硫基琥珀酰亚胺马来酰亚胺丁酸酯(SSMB)分别溶于微量二甲基亚砜(DMSO),再将抑制剂溶液溶入L-15不完全培养液中,制成10 mM环氧糊精, 250μM根皮素, 250μM DIDS, 200μM SSO和200μM SSMB前处理培养液(DMSO<0.05 % )。在前处理培养液中分别加入按30:1比例混合的非放射性油酸(37.5μM)与[1-14C]油酸组成的混合油酸,制成吸收-抑制培养液。培养12 h的贴壁大西洋鲑肝细胞与细胞前处理液共培养0.5 h后,将该培养液弃去,换为吸收-抑制培养液(0.3μCi/培养瓶), 2 h后按3.1所述方法收取细胞及培养基样品并进行样品放射性测定和计算油酸吸收量(nmol h-1 million cells-1)。结果:大西洋鲑肝细胞对油酸吸收均受到膜蛋白抑制剂的抑制,其中根皮素的抑制作用最强(28 %),DIDS和SSMB次之(14 %),SSO与环氧糊精最低(8 %~10 %)。结果表明:大西洋鲑肝细胞对油酸的跨膜吸收是膜蛋白介导的主动吸收与被动扩散共同作用的过程。3.4大西洋鲑肝细胞对五种长链脂肪酸的吸收及细胞内代谢特点的研究目的:研究大西洋鲑肝细胞对五种长链脂肪酸的跨膜吸收机制及细胞内代谢特点。方法:同3-3。五种长链脂肪酸分别为18:1n-9,18:2n-6, 18:3n-3, 20:5n-3和22:6n-3。膜蛋白抑制剂细胞前处理培养液制作方法同3-3。膜蛋白抑制剂为根皮素, DIDS和SSO。每种脂肪酸分别含有该三种膜蛋白抑制剂的吸收-抑制培养液。培养12 h的贴壁大西洋鲑肝细胞与细胞前处理液共培养0.5 h后,将培养液弃去,换为吸收-抑制培养液(0.24~0.36μCi/培养瓶), 2 h后按3.1所述方法终止吸收反应,取得细胞及培养液样品,同时进行样品放射性测定,用放射性(dpm)的相对值(吸收率;相对于添加到培养液中的总放射性)计算油酸吸收及胞内代谢。结果:大西洋鲑肝细胞对20:5n-3吸收率最高,为15 %,22:6n-3居中,为13 %,18:1n-9、18:2n-6和18:3n-3吸收率最低,为10 %~ 11 %。根皮素对20:5n-3吸收的抑制作用最强(67 %),对18:1n-9吸收的抑制作用最弱(13 %),对18:3n-3、22:6n-3和18:2n-6吸收的抑制作用居中,分别为46 %、35 %和25 %。在DIDS作用下五种长链脂肪酸的吸收均被抑制,但仅对18:2n-6、18:3n-3和22:6n-3的吸收抑制显著,吸收抑制率分别为16 %、30 %和16%。SSO对五种长链脂肪酸的吸收均无显著抑制。18:3n-3在细胞内非酯化脂肪酸的分布最高,为68 %,其次是18:1n-9、18:2n-6和20:5n-3 (61%~67 %),22:6n-3最低(50 %)。18:3n-3, 18:1n-9和18:2n-6在细胞内磷脂的分布最低(20 %~23 %),20:5n-3居中(27 %),22:6n-3最高(38 %)。长链脂肪酸在胞内中性脂质的分布无显著差别。18:3n-3氧化量最高,为吸收量的7 %;20:5n-3、22:6n-3和18:2n-6略低,为吸收量的3 %左右;18:1n-9的氧化量最低,低于吸收量的1 %。结果表明:大西洋鲑肝细胞对长链脂肪酸的跨膜吸收是膜蛋白介导的主动吸收与被动扩散共同作用的过程。不同链长及饱和性的脂肪酸在肝细胞的吸收及胞内代谢有较大差别。4.植物油替代鱼油对大西洋鲑肝细胞脂肪酸吸收及胞内代谢的影响目的:探讨大西洋鲑肝细胞脂肪酸组成被饲料油源脂肪酸组成改变后,对油酸吸收和细胞内油酸代谢的影响。方法:大西洋鲑幼鲑分别饲喂以鱼油和植物油为油源的两种饲料,5个月后,大西洋鲑肝细胞脂肪酸组成分别被油源改变,分离肝细胞,与油酸共培养2 h后,测定细胞对油酸的吸收量及油酸在细胞内的代谢情况。具体测定方法同3-3。所选膜蛋白抑制剂及配伍为250μM根皮素, 250μM DIDS, 200μM SSO,400μM SSO,SSO +根皮素,200μM SSMB。培养12 h的贴壁大西洋鲑肝细胞与细胞前处理液共培养0.5 h后,将培养液换为吸收-抑制培养液(0.3μCi/培养瓶),2 h后中止吸收,采取细胞及培养液样品,然后测定样品放射性并计算油酸吸收量(nmol h-1 million cells-1),同时采用RT-PCR方法测定与脂肪酸代谢相关的转运蛋白质或酶的基因表达量。结果:鱼油与植物油组肝细胞对油酸的吸收均无显著差别,但抑制剂对植物油组肝细胞油酸吸收有显著抑制作用而对鱼油组油酸吸收无显著影响。经吸收进入细胞内的油酸,于非酯化脂肪酸(自由脂肪酸,FFA)和磷脂(PL)的分布在鱼油和植物油组间无显著差别。但植物油表现出减少了油酸在中性脂质的分布,促进了油酸β氧化。抑制剂对油酸在细胞内的代谢无显著影响。植物油显著增加了FAT/CD36,脂肪酸结合蛋白(FABP3),Δ6与Δ5去饱和酶的基因表达,但对FATP, FABP10和肉碱棕榈酸转移酶I (CPT I)基因表达无显著影响。植物油显著增加了应激与抗氧化防御基因的表达,如谷胱甘肽过氧化物酶4,热应激蛋白70,金属硫蛋白和泛素,而抑制剂对这几种蛋白的基因表达无显著影响。结果表明:肝细胞膜脂肪酸组成在一定程度上影响了细胞对脂肪酸的吸收和胞内代谢,但肝细胞对脂肪酸的跨膜吸收表现为由膜蛋白介导的主动吸收与被动扩散的共同作用,同时该作用不受细胞膜脂肪酸组成的影响。更多还原

【Abstract】 Common carp (Cyprinus carpio L.) is omninorous fish and is traditionally edible fresh water fish in China. Atlantic salmon (Salmo salar L.) is carnivorous fish living in sea water. They are all high quality protein and essential fatty acid resources for human being. With the finding that fish oil, being rich in highly unsaturated fatty acid (HUFA), is useful in preventing human being from cardiovascular and cerebrovascular disease, which induced the production of health food of fish oil. Oil is also necessary for growth and reproduction of fish and fish oil traditionally was added in their feeds in providing essential fatty acid and energy. However the effect of fish oil and HUFA on the nutrition and healthy aquculture (e.g. Anti-oxidation system), of fish, especially fresh water fish, has not been sufficiently elucidated. In the same time, the fact that fish stocks in the world are limited induces the limited supplement of fish oil and stimulates the price of fish oil, which further influence the sustainable development of the aqua-feeds, especially of the feeds of Atlantic salmon which requires 30 % of adding of fish oil. Vegetable oil is cheaper and more richful than fish oil then the substitution of fish oil with vegetable oil has been emphasized in aquaculture feeds. However the fatty acid composition of fish oil and vegetable oil is different, for example fish oil is rich in HUFA but vegetable oil is scarce of it, which probably influences the nutrion and metabolism of oil in fish and limited dataes were given to shed light on it. The present experiments were carried out to understand the nutrition and metabolism of fish oil and HUFA, especiall in common carp and also to clarify the possibility of replacing dietary fish oil with vegetable oil in Atlantic salmon feeds.1. Influence of fish oil on growth and lipid metabolism in Common carp (Cyprinus carpio L.)In order to examine the influence of fish oil and the HUFA (Highly unsaturated fatty acid) on growth and lipid metabolism in common carp (Cyprinus carpio) a dietary trial and a HUFA orally administration trial were conducted in the present study. In the front trial, semi-purified diets supplemented with fish oil (FO) and soybean oil (VO) were used and the indexes of growth and the activity of lipid metabolic enzymes in hepatopancreas were assayed. In second trial, hepatopancreas were sampled after fish being orally administered with HUFA and the activity of malate dehydrogenase (MDH) was assayed. Dietary oil sources had no impact on feed conversion ratio, absolute growth rate and relative growth rate. Common carp in FO group had significantly higher intraperitoneal fat body (IPF) ratio than that in VO group. Dietary oil sources had no significant effect on condition factor, gutted body ratio, carcass ratio, muscle ratio, hepatosomatic index and relative intestine length. Common carp in FO group than that in VO group had lower activity of MDH, lipoprotein lipase (LPL) and glucose-6-phosphate dehydrogenase (G-6-P-DH). This was further certified by the lower activity of MDH in HUFA group at 2 h and 4 h in HUFA administration study. This suggested that common carp can utilize fish oil and vegetable oil for growth equally well although the lipid metabolism was affected by dietary oil sources.2. Effect of DHA on anti-oxidation capacity in Common carp (Cyprinus carpio.L)To study the effect of DHA (22:6n-3, Docosahexaenoic acid) on anti-oxidation capacity in Common carp, DHA orally administration study, DHA long term (74 d) dietary study and incubation of hepatocytes with DHA study were carried on. The total anti-oxidation capacity (T-AOC), total superoxide dismutase (T-SOD), glutathione S-transferase (GST), catalase (CAT), maleic dialdehyde (MDA) of hepatopancreas or hepatocytes were determined respectively. T-AOC in both DHA and control group increased before 4 h of post-administration then declined till 32 h. T-AOC in DHA group was higher than that in control group especially at 32 h time point of post-administration indicating that common carp endured oxidative stress and the anti-oxidation capacity of hepatopancreas was induced to increase. MDA, CAT, SOD and T-AOC of the hepatopancreas and SOD as well as T-AOC of the serum were not significantly different between DHA and control group except that GST of serum in DHA group was significantly lower than that in control group in 12 % lipid level but not in 6 % lipid level. The dietary trial suggests that common carp is acclimatized to the oxidative stress caused by the DHA in long term feeding of DHA. MDA in both DHA and control group increased in the beginning of incubation with hepatocytes then decreased swiftly. MDA decreased not so fast in DHA group as that in control group, therefore MDA in DHA group was significantly higher when the cells were incubated with DHA for 2, 4 and 8 h but not for 0.5 or 1 h. T-SOD was also decreased with longer incubation time in both of the two groups and there was no significant difference between them except at 4 h incubation where T-SOD was significantly higher in DHA group compared with control. The in vitro trial suggests that hepatocytes incubated with DHA experience higher oxidative stress and the anti-oxidation capacity of hepatocytes is therefore induced to increase.3. Uptake and intracellular metabolism of long chain fatty acids in Atlantic salmon (Salmo salar L.) hepatocytes To elucidate if the trans-membrane uptake of fatty acid was protein-mediated, the uptake of oleic acid (OA; 18:1n-9) was investigated in vitro in Atlantic salmon (Salmo salar L.) hepatocytes. Firstly, to find the optimal time of OA incubation hepatocytes cultured overnight was incubated with 61μM OA for 0.5, 1, 2, 4, 6, 12 and 24 h respectively (0.2μCi/flask). Secondly, to establish the OA incubation concentration for optimal OA uptake the hepatocytes were incubated with 37.5, 75, 150, 300 and 600μM OA for 2 h with a fixed molar ratio between unlabelled and radiolabelled OA of 30:1. OA uptake revealed saturation kinetics with Km values of 47μM and Vmax values of 2.3 nmol h-1 million cells-1. To identify whether trans-membrane OA uptake in hepatocytes was mainly passive or protein mediated, hepatocytes were pre-incubated with membrane protein inhibitors, cyclodextrin, phloretin, diisothiocyanodisulfonic acid (DIDS), sulfo-N-succinimidyl 4-maleimido-oleic acid ester (SSO) and sulfo-N-succinimidyl 4-maleimido-butyric acid ester (SSMB). The OA uptake was significantly reduced by inhibitors with phloretin giving the highest inhibition (28%) and SSO the lowest (8%). While the other inhibitors, cyclodextrin, DIDS and SSMB, showed the intermediate inhibition between 9.8 and 14.5 %. Our results suggest that trans-membrane OA uptake in Atlantic salmon hepatocytes is due to both saturable and inhibitable protein mediated uptake, as well as passive uptake processes. To investigate the uptake and intracellular metabolism of 18:1n-9, 18:2n-6, 18:3n-3, 20:5n-3 and 22:6n-3, Atlantic salmon (Salmo salar L.) hepatocytes incubated overnight were firstly pretreated with known membrane protein inhibitors, phloretin, diisothiocyanodisulfonic acid (DIDS), sulfo-N-succinimidyl 4-maleimido-oleic acid ester (SSO), for 0.5h, then 37.5μM of these fatty acids were added respectively for another 2 hours incubation (0.24- 0.36μCi/flask). Hepatocytes (controls) incubated with fatty acids but without inhibitors were run concurrently. Radioactivity recovered in cells, cellular lipid classes andβ-oxidation products were measured. 20:5n-3 was highest taken up (15 %), 22:6n-3 being the intermediate (13 %), then 18:1n-9, 18:2n-6 and 18:3n-3 was lowest taken up (10%~11 %). Phloretin had the highest inhibition on uptake of 20:5n-3 (67 %) and lowest inhibition on 18:1n-9 (13 %), having the intermediate inhibition on 18:3n-3, 22:6n-3 and 18:2n-6 (46 %, 35 %, and 25 %, respectively). Uptake of 18:2n-6 and 18:3n-3 but not of the other fatty acids was significantly decreased in the presence of DIDS. SSO showed no inhibition on these fatty acids uptake. 18:3n-3 was highestβ-oxidised (7 % of uptake), 20:5n-3, 22:6n-3 and 18:2n-6 being the intermediate (about 3 %), and 18:1n-9 was leastβ-oxidised (<1 %). The intracellular free fatty acid (FFA) distribution was highest with 18:3n-3 (68 % of cellular lipids), followed with 18:1n-9, 18:2n-6 and 20:5n-3 (61-67 %), and it was lowest with 22:6n-3 (50 %). The order of fatty acids incorporation into phospholipids (PL) was reverse to that of FFA, indicating that 22:6n-3 was most incorporated into PL (38 %), followed by 20:5n-3 (27 %), then lowest by 18:2n-6, 18:1n-9 and 18:3n-3 (20 %~23 %). No difference was found in the incorporation of these FAs into neutral lipids.4. Replacing dietary fish oil with vegetable oil affect oleic acid uptake and metabolism in hepatocytes of Atlantic salmon (Salmo salar L.)The aim was to investigate how uptake and metabolism of 18:1n-9 (oleic acid; OA) in Atlantic salmon (Salmo salar L.) hepatocyte is affected by dietary oil sources. Atlantic salmon post smolt was fed diets containing either 100% fish oil (FO) or vegetable oil (VO) for 5 months to produce hepatocytes with typical FO and VO fatty acid composition. Then OA uptake and metabolism in isolated hepatocytes were studied by incubating with 37.5μM OA for 2 h (0.3μCi/flask) with and without (control) membrane protein inhibitors; phloretin, sulfo-N-succinimidyl 4-maleimido-oleic acid ester, diisothiocyanodisulfonic acid, and sulfo-N-succinimidyl 4-maleimido-butyric acid ester. OA uptake was not different in control cells between FO and VO fed fish. The effect of adding membrane protein inhibitors, however, was significantly different with OA uptake being inhibited in VO but not in FO hepatocytes. OA incorporation into cellular neutral lipids was significantly lower and the recovery in cellularβ-oxidation products was significantly higher in VO compared to FO hepatocytes. Intracellular free OA and OA incorporation into phospholipids was not different between the two groups. Inhibitors did not affect OA cellular distribution. The mRNA expression of FAT/CD36, fatty acid binding protein 3 and 10 (FABP3 and FABP10),Δ6 desaturase andΔ5 desaturase were significantly higher in VO than in FO hepatocytes, whereas the expression of FATP, FABP10 and carnitine palmitoyl transferase I (CPT I) was not different between FO and VO hepatocytes. The expression of genes related to stress responses and antioxidant defence; glutathione peroxidase 4, heat shock protein 70, metallothionein and ubiquitin were significantly higher in VO than in FO hepatocytes, however the incubation with inhibitors did not affect the expression these genes. Overall, the results indicate that hepatocyte fatty acid composition is involved in the regulation of fatty acid uptake and intracellular fatty acid metabolism whereas inhibition of OA uptake did not influence intracellular metabolism.更多还原