皱纹盘鲍抗氧化基因的克隆及其在营养调控下表达的研究

详细信息    本馆镜像全文|  推荐本文 |  |   获取CNKI官网全文

英文题名：Cloning and Expression Analysis of Antioxidant Genes in Abalone Haliotis Discus Hannai Ino in Response to Nutrition 作者：吴成龙 论文级别：博士 学科专业名称：水生生物 中文关键词：皱纹盘鲍 ; 抗氧化基因 ; 克隆 ; 表达 ; 营养调控 英文关键词：Haliotis discus hannai Ino ; antioxidant gene ; cloning ; expression ; nutation regulation 学位年度：2010 导师：麦康森 学科代码：071004 学位授予单位：中国海洋大学 论文提交日期：2010-04-01

摘要

皱纹盘鲍(Haliotis discus hannai Ino)隶属于软体动物门,腹足纲(Gastropoda),前腮亚纲(Prosobranchia),原始腹足目(Archaeogastropoda),鲍科(Haliotidae),主要分布在我国渤海和黄海,以及日本和朝鲜的附近海域,是世界重要的名贵海水养殖品种。但是近年来由于环境污染、操作不当、投饲频率增加等原因,使得养殖的皱纹盘鲍不断发生各类病害,其产量大幅度下跌,种质出现衰退现象。而环境中各种胁迫因子的增加和鲍鱼免疫力的下降直接导致病原体的高感染率和鲍鱼的高死亡率。但时至今日,皱纹盘鲍的先天性免疫学研究中仍然存在大量的空白,尤其是如何利用皱纹盘鲍的营养免疫机制来提高其免疫力的研究较少。所以深入开展皱纹盘鲍营养免疫机制研究并在此基础上寻找增加皱纹盘鲍抗胁迫能力的有效方法已成为当务之急。
     研究发现,在受到病原体、化学因子、辐射、温度(高温或者低温)变化、溶解氧、营养不良以及其他可能的胁迫条件下,水生生物体会产生大量的活性氧分子。这些活性氧分子能够消灭掉外源入侵者,但同时由于活性氧分子反应的非特异性,它们也会对宿主的细胞、组织和器官造成严重伤害,进而导致皱纹盘鲍生理机能的损伤和免疫系统的破坏。所以,消除皱纹盘鲍体内因过度免疫反应产生的过量氧自由基将能够增强其抵御病原侵染的能力,提高免疫力。为了维持机体健康,避免活性氧对自身细胞造成损伤,阻止由于氧化胁迫引起的疾病和死亡,耗氧生物体在长期的氧胁迫环境以及外界生存环境中形成了一个行之有效的抗氧化体系,以维持体内产生适量ROS和清除多余ROS之间的平衡,进而维持动物机体内氧化和抗氧化之间的动态平衡。其中,包括抗氧化酶系统,如：谷胱甘肽过氧化物酶(GPx)、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)等,以及非酶类抗氧化分子,如硒结合蛋白、铁蛋白、热休克蛋白、维生素C和谷胱甘肽等。
     众所周知,生物体内自由基的产生、清除、利用、损伤及其修复所需物质与能量均直接或间接来源于营养物质及其代谢物,即动物体内营养素及其代谢物是各类自由基产生的物质来源；清除自由基系统的成分均直接或间接来自营养素与膳食中抗氧化剂。因此,营养状况应能维持自由基的产生与清除处于正常动态平衡,内环境处于稳定的还原态,并使各类活性氧的生理作用以及彼此相互作用能正常发挥。此外,大量研究已经证明动物体能够通过营养调控的方式获得更好的氧化还原平衡,如通过获得适量的硒、锌、铁和维生素C等。然而,尚未有关于营养素和皱纹盘鲍抗氧化系统在基因转录水平上进行相互作用研究的报道。此外,克隆和鉴定抗氧化反应相关基因将会帮助我们更好的理解对于营养调控的分子生物学机制。
     在本研究中,我们首先对皱纹盘鲍氧化还原平衡系统中的硒依赖型谷胱甘肽过氧化物酶(Se-GPX),硒结合蛋白(SEBP),热休克蛋白90(HSP90)和铁蛋白(FT)等进行了基因全长克隆,并对其分子结构特征、组织分布特征及其在营养(硒、铁和锌)调控下的表达特征进行了相关分析。此外,我们还应用荧光定量PCR技术,对经过维生素C调控下的皱纹盘鲍抗氧化相关基因(Cu/Zn-SOD,Mn-SOD,CAT,GPX,GST, Tpx, SEBP, HSP26, HSP70和HSP90)和氧化胁迫相关基因(NF-κB)进行了相对定量的表达分析。
     1皱纹盘鲍谷胱甘肽过氧化物酶的基因克隆及其在饲料中不同浓度硒、锌和铁元素处理下的表达分析
     以皱纹盘鲍(Haliotis discus hannai Ino)(平均体长：65.01±0.45 mm)为材料,利用同源克隆技术和RACE技术从皱纹盘鲍肝脏中成功扩增出中硒依赖型谷胱甘肽过氧化物酶(selenium-dependent glutathione peroxidase, HdhGPx)。HdhGPx全长为963bp(Genbank中的注册号为GU254066),包括21bp的5'非编码区、273 bp的3'非编码区和669 bp的开放阅读框。另外,在3'非编码区还存在一个101bp的SECIS作用原件。其中HdhGPx的开放阅读框编码222个氨基酸,预测分子量约为24.76 KDa,理论等电点为7.00；在其N端含有一个23个氨基酸残基编码的信号肽。在HdhGPx的cDNA序列中含有一个典型的终止密码子(235TGA237)用于编译硒半胱氨酸(U72),一个GPx信号基序2(96LGLPCNQF103),一个GPx活化位点基序(179WNFEKF184)。此外,在HdhGPx的氨基酸序列中还存在两个潜在的蛋白质糖基化位点(112NGTE115和132NLTQ135)。在三维空间结构上,HdhGPx显示出与人GPx3具有更高的结构相似性。实时荧光定量PCR检测结果显示,HdhGPx mRNA在肝脏、外套膜、鳃、性腺、肌肉和肾脏中的表达量要显著高于在血细胞中的表达量,而且在肝脏中的表达量为最高。通过分别用不同浓度的微量元素硒(0.15mg/kg,1.32mg/kg和48.7 mg/kg)、锌(6.69mg/kg,33.85 mg/kg和710.63 mg/kg)和铁(29.19 mg/kg,65.7 mg/kg和1267.2 mg/kg)的饲料来饲喂幼鲍(平均体长：13.05±0.25 mm)20周后,分别取其肝脏和血细胞总RNA,利用实时荧光定量PCR技术进行HdhGPx mRNA的表达分析。分析结果显示,在硒处理组,皱纹盘鲍肝胰脏中HdhGPx mRNA的表达量随硒添加量增加而升高。皱纹盘鲍肝胰脏中HdhGPx mRNA的表达量在硒添加量为1.32 mg/kg时表达量最高,但皱纹盘鲍血细胞中HdhGPx mRNA的表达量在硒过量组(48.7 mg/kg)为最高。在锌处理组,皱纹盘鲍肝胰脏中HdhGPx mRNA的表达量升高,但是在锌添加量为33.85mg/kg达到最高。在铁处理组,皱纹盘鲍HdhGPx mRNA的表达量呈现先升高后降低的趋势,即在铁适量组(65.7 mg/kg)为最高,而后在铁过量组(1267.2 mg/kg)肝胰脏中HdhGPx的表达量降低。上述研究结果表明HdhGPx的表达受到饲料中微量元素(硒、锌和铁)添加量的影响,而且可能在提高机体抗氧化能力和防止由微量元素含量过高引起的氧化胁迫中担负重要作用。
     2皱纹盘鲍硒结合蛋白的基因克隆及其在饲料中不同浓度硒、锌和铁元素处理下的表达分析
     利用同源克隆技术和RACE技术从皱纹盘鲍肝脏中成功扩增出中硒结合蛋白(selenium-binding protein, HdhSEBP)。HdhSEBP全长为2071bp(Genbank中的注册号为GU014544),包括55bp的5'非编码区、522 bp的3'非编码区和1494 bp的开放阅读框。其中HdhSEBP的开放阅读框编码497个氨基酸,预测分子量约为55.6 KDa,理论等电点为5.47。BLAST分析结果显示HdhSEBP与其他物种,如哺乳动物、鸟、鱼和无脊椎动物的SEBP氨基酸序列具有高度的相似性。此外,HdhSEBP的氨基酸序列中还存在多个金属结合位点和一个潜在的蛋白质糖基化位点(320NKTV323)。实时荧光定量PCR检测结果显示,HdhSEBP mRNA在外套膜、肾脏、肝脏和血细胞中的表达量要显著高于在鳃、性腺和肌肉中的中的表达量,而且以在外套膜中的表达量为最高。通过分别用不同浓度的微量元素硒(0.15 mg/kg,1.32 mg/kg和48.7 mg/kg)、锌(6.69 mg/kg,33.85 mg/kg和710.63 mg/kg)和铁(29.19 mg/kg,65.7 mg/kg和1267.2mg/kg)的饲料来饲喂幼鲍(最初体长：13.05±0.25 mm)20周后,分别取其肝脏和血细胞总RNA,利用实时荧光定量PCR技术进行HdhSEBP mRNA的表达分析。分析结果显示,在硒处理组,皱纹盘鲍肝胰脏中HdhSEBP mRNA的表达量随硒添加量增加而升高。皱纹盘鲍肝胰脏和血细胞中HdhSEBP mRNA在硒添加量为1.32 mg/kg时表达量均为最高水平。在锌处理组,皱纹盘鲍肝胰脏和血细胞中HdhSEBP mRNA在锌添加量为33.85 mg/kg时表达量均为最高,但是在锌添加量为710.63 mg/kg降低到最低水平。在铁处理组,皱纹盘鲍HdhSEBP mRNA的表达量呈现先升高后降低的趋势,即在铁适量组(65.7 mg/kg)为最高,而后在铁过量组(1267.2 mg/kg)中HdhSEBP的表达量降低。上述研究结果表明HdhSEBP的表达受到饲料中微量元素(硒、锌和铁)添加量的影响,而且可能在提高机体抗氧化能力和防止由微量元素含量过高引起的氧化胁迫中担负重要作用。
     3.皱纹盘鲍热休克蛋白90的基因克隆及其在饲料中不同浓度硒、铁和锌元素处理下的表达分析
     以皱纹盘鲍(Haliotis discus hannai Ino)(平均体长：65.01±0.45 mm)为材料,利用同源克隆技术和RACE技术从皱纹盘鲍肝脏中成功扩增出中热休克蛋白90(heat shock protein 90, HdhHSP90)。HdhHSP90全长为2660 bp (Genbank中的注册号为GU014545),包括96 bp的5’非编码区、377 bp的3’非编码区和2187 bp的开放阅读框。其中HdhHSP90的开放阅读框编码728个氨基酸,预测分子量约为84.134 KDa,理论等电点为4.619。BLAST分析结果显示HdhHSP90与其他物种,如哺乳动物、鸟、鱼和无脊椎动物的HSP90氨基酸序列具有高度的同源性。在HdhHSP90氨基酸序列中含有6个典型的热休克蛋白家族的信号基序(NKEIFLRELISNSSDALDKIR, LGTIAKSGT, IGQFGVGFYSAYLVAE, IKLYVRRVFI, GVVDSEDLPLNISRE, MEEVD)。实时荧光定量PCR检测结果显示,HdhHSP90 mRNA在性腺和血细胞中的表达量较高。通过分别用不同浓度的微量元素硒(0.15 mg/kg,1.32 mg/kg和48.7mg/kg)、锌(6.69 mg/kg,33.85 mg/kg和710.63 mg/kg)和铁(29.19 mg/kg,65.7 mg/kg和1267.2 mg/kg)的饲料来饲喂幼鲍(最初体长：13.05±0.25mm)20周后,分别取其肝脏和血细胞总RNA。荧光定量PCR分析结果显示,在硒处理组,皱纹盘鲍肝胰脏中HdhHSP90 mRNA的表达量随硒添加量增加而升高。皱纹盘鲍肝胰脏和血细胞中HdhHSP90 mRNA在硒添加量为1.32 mg/kg时表达量均为最高水平。在锌处理组,皱纹盘鲍肝胰脏和血细胞中HdhHSP90 mRNA在锌添加量为33.85 mg/kg时表达量均为最高,但是在锌添加量为710.63 mg/kg降低到最低水平。在铁处理组,皱纹盘鲍HdhHSP90 mRNA的表达量呈现先升高后降低的趋势,即在铁适量组(65.7 mg/kg)为最高,而后在铁过量组(1267.2 mg/kg)中HdhHSP90的表达量降低。上述研究结果表明HdhHSP90的表达受到饲料中微量元素(硒、锌和铁)添加量的影响,而且可能在提高机体抗胁迫能力和保护动物机体免受由饲料中微量元素(硒、锌和铁)含量过高引起的氧化胁迫中担负重要作用。
     4皱纹盘鲍铁蛋白的基因克隆及其在饲料中不同浓度铁元素处理下的表达分析
     利用文库构建技术和RACE技术从皱纹盘鲍肝脏中成功扩增出中一个新型的铁蛋白(ferritin, HdhNFT)。HdhNFT全长为915bp(Genbank中的注册号为GU479917),包括103 bp的5'非编码区、296 bp的3'非编码区和516 bp的开放阅读框。其中HdhNFT的开放阅读框编码171个氨基酸,预测分子量约为19.8 KDa,理论等电点为4.79。BLAST分析结果显示HdhNFT与其他物种,如哺乳动物、鸟、鱼和无脊椎动物的FT氨基酸序列具有高度的相似性。此外,HdhNFT的氨基酸序列中还存在一个潜在的蛋白质糖基化位点(27NCSY30)。实时荧光定量PCR检测结果显示,HdhNFT mRNA在肾脏、肝脏、鳃、肌肉和外套膜和血细胞中的表达量要显著高于在性腺和血细胞中的中的表达量,而且以在肾脏中的表达量为最高。通过用不同浓度的微量元素铁(29.19 mg/kg,65.7 mg/kg,1267.2 mg/kg和6264.7 mg/kg)的饲料来饲喂幼鲍(最初体长：13.05±0.25mm)20周后,分别取其肝脏和血细胞总RNA,利用实时荧光定量PCR技术进行HdhSEBP mRNA的表达分析。分析结果显示,皱纹盘鲍HdhNFT mRNA的表达量呈现升高的趋势,即随铁添加量的增加,在铁过量组(6264.7mg/kg)中HdhNFT的表达量最高。上述研究结果表明HdhNFT的表达受到饲料中铁元素添加量的影响,而且可能在提高机体抗氧化能力和防止由铁元素含量过高引起的氧化胁迫中担负重要作用。
     5饲料中添加维生素C对皱纹盘鲍肝胰腺中抗氧化和胁迫相关基因表达的影响
     本研究以皱纹盘鲍(Haliotis discus hannai Ino)(平均体长：84.36±0.24 mm)为研究对象,探讨饲料中添加不同梯度维生素C对皱纹盘鲍肝胰腺组织中抗氧化胁迫相关基因(超氧化物歧化酶,SOD；过氧化氢酶,CAT；谷胱甘肽过氧化物酶,GPX；谷胱甘肽-S-转移酶,GST；硒结合蛋白,SEBP;热休克蛋白26,HSP26；热休克蛋白70,HSP70；热休克蛋白90,HSP90)以及细胞核转录因子Rel/NF-κB等在转录水平上表达的影响。通过用添加了不同浓度维生素C(0.0 mg/kg,70.3 mg/kg,829.8 mg/kg和4967.5 mg/kg)的饲料来饲喂皱纹盘鲍成鲍,在室内流水系统养殖24周后,取其肝脏总RNA,利用实时荧光定量PCR技术进行相关基因的表达水平的分析。分析结果显示,与基础饲料组相比,饲料中添加适量的维生素C (70.3 mg/kg)能够显著提高皱纹盘鲍肝胰腺中抗氧化相关基因(Cu/Zn-SOD,CAT,GST,NFT,HSP26)mRNA的表达量,但是显著降低Mn-SOD,GPX,TPx和SEBP mRNA的表达量。与添加适量的维生素C (70.3 mg/kg)饲料组相比,添加过量的维生素C(829.8 mg/kg和4967.5mg/kg)显著提高皱纹盘鲍肝胰腺中Mn-SOD,GPX,TPx,SEBP,NFT,HSP70,HSP90和NF-κB mRNA的表达量。上述研究结果表明饲料中维生素C能够影响皱纹盘鲍抗氧化胁迫相关基因的表达量,这提示饲料中添加适量的维生素C能够提高机体抗氧化能力,减轻机体内的氧化胁迫程度。但是饲料中添加过量的维生素C会起到氧化剂的作用,从而降低皱纹盘鲍机体的抗胁迫能力。
     总体而之,饲料中添加适量的微量元素(硒、锌和铁)或抗氧化剂(维生素C)能够显著提高皱纹盘鲍抗氧化相关基因的表达量,进而提高机体的抗氧化胁迫的能力。而过量的微量元素(硒、锌和铁)或维生素C会起到氧化剂的效果,会对皱纹盘鲍抗氧化功能造成负面的影响,从而降低抗胁迫能力。
Abalone Haliotis discus hannai Ino are large algivorous marine gastropods, and the most commercially important specie of gastropods in aquaculture for Asia. However, abalone culture has suffered serious problems of mortality from disease outbreaks, environmental contamination and decreased innate immunity of abalone in China and other countries or districts. It is an emergent and necessary work for us to explore the immune system of abalone and to find effective methods to control these diseases or to allieveate stresses. It has been proved that a mass of reactive oxygen species (ROS) could be induced by pathogens, malnutrition, chemicals, radiation, high or cold temperature, ressloved oxygen and other possible stresses. Though ROS can kill foreign invaders, the excessive accumulation of these toxic by-products could cause serious damage to lipids, proteins and nucleic acids. In order to protect cells against the toxicity caused by ROS, animals have evolved protective antioxidant systems, including antioxidant enzymatic systems such as glutathione peroxidase (GPx), superoxide dismutase (SOD) and catalase (CAT), and non-enzymatic systems, such as vitamin C and glutathione, etc.
     It is well known that all the materials and energy are rooted in nutrients used for the production, elimination, usage, damnification and rehabilitation of ROS.Namely, all components are directly or indirectly resulted from nutrients or antioxidants in the food. Therefore, it is important for optimal nutrition states to well balance the homeostasis between production and elimination of ROS,maintain the reduction states cellular environment, and induce normal function of ROS and each other. In addition, many studies have confirmed that animals could maintain better homeostasis of oxidation/reduction through nutrition regulation with adequate nutrients, including selenium, zinc, iron and vitamin C, etc.However, there has no published literature on the interaction mechanisms between dietary minerals and anti-oxidative responses at gene transcriptional levels in abalone Haliotis discus hannai Ino.In addition, identification and cloning of the genes involved in anti-oxidative response will help us to understand the molecular basis of the innate immune response to the malnutrition stresses.
     In the present study, selenium-dependent glutathione peroxidase(Se-GPX), selenium-binding protein (SeBP), heat shock protein 90 (HSP90) and ferritin (FT) which related to the oxidation/reduction system in abalone Haliotis discus hannai Ino were first cloned and analyzed under the conditions of nutritional regulation respectively.In addition, other relative genes (Cu/Zn-SOD, Mn-SOD, CAT, GST, Tpx, HSP70, HSP26 and NF-κB, etc.)were also detected in abalone fed with graded diets containing vitamin C or a-lipoic acid using real-time PCR assays, respectively.
     1 Molecular cloning, characterization and mRNA expression of selenium-dependent glutathione peroxidase from abalone Haliotis discus hannai Ino in response to dietary selenium, zinc and iron
     A novel selenium-dependent glutathione peroxidase (Se-GPX) was cloned from abalone Haliotis discus hannai Ino (HdhGPx) by homology cloning with degenerate primers and RACE techniques. The full length of HdhGPx cDNA was 963 bp with a 669 bp open reading frame (ORF) encoding 222 amino acids and a 101 bp eukaryotic selenocysteine insertion sequence (SECIS) in 3'untranslated region (UTR).It was showed that HdhGPx has a characteristic codon at 235TGA237 that corresponds to selenocysteine (SeC) as U72. Sequence characterization revealed that HdhGPx contains a characteristic GPx signature motif 2 (96LGLPCNQF103), an active site motif (179WNFEKF184). In Addition, two potential N-glycosylation sites (112NGTE115 and 132NLTQ135) were identified in HdhGPx.3D modeling analysis showed that the overall structure of HdhGPx monomer had more similarity to human GPx3.Relatively higher-level mRNA expression was detected in hepatopancreas, mantle and gonad by real-time PCR assays. The relative expression levels of HdhGPx mRNA in hepatopancreas and haemocytes were detected by real-time PCR in abalone fed with nine different diets containing graded levels of selenium (0.15,1.32 and 48.7 mg/kg), zinc (6.69,33.85 and 710.63 mg/kg) and iron (29.17,65.7 and 1267.2 mg/kg) for 20 weeks, respectively. The results showed that the expression of HdhGPx mRNA increased and reached the maximum at optimal dietary selenium(1.32 mg/kg), zinc (33.85 mg/kg) and iron (65.7 mg/kg), respectively.But HdhGPx mRNA expression levels were down-regulated by excessive dietary selenium (48.7 mg/kg), zinc (710.63 mg/kg) and iron(1267.2 mg/kg) and zinc (33.85 mg/kg) compared with these dietary minerals, respectively. These results indicated that optimal dietary minerals could trigger the mRNA expression of HdhGPx to increase the total antioxidant capacities in abalone.
     2 Molecular cloning, characterization and mRNA expression of selenium-binding protein in abalone (Haliotis discus hannai Ino):response to dietary selenium, iron and zinc
     Selenium-binding protein(SEBP) is believed to play crucial role in controlling the oxidation/reduction in the physiological processes. In this study, the cDNA of selenium-binding protein from abalone Haliotis discus hannai Ino (HdhSEBP) was cloned by homology cloning and rapid amplification of cDNA ends (RACE) technique.The full length of HdhSEBP cDNA was 2071 bp, consisting of a 5'untranslated region (UTR) of 55 bp,a 3'UTR of 522 bp, and an open reading frame (ORF) of 1494 bp. The deduced protein has 497 amino acid residues with a calculated molecular mass of 55.6 kDa and a predicted isoelectric point of 5.47.BLAST analysis reveals that HdhSEBP shares high identities with other known SEBPs from mammal, bird, fish and mollusk, etc.The mRNA expression patterns of HdhSEBP in hepatopancreas and haemocytes were measured by real-time PCR in abalone fed with nine different diets containing graded levels of selenium (0.15,1.32 and 48.7 mg/kg), iron (29.17,65.7 and 1267.2 mg/kg) and zinc (6.69, 33.85 and 710.63 mg/kg) for 20 weeks, respectively. The results showed that the expression of the HdhSEBP mRNA increased and reached the maximum at optimal dietary selenium(1.32 mg/kg), iron (65.7 mg/kg) and zinc (33.5 mg/kg), respectively. Deficient or excessive level of dietary selenium, iron or zinc, respectively, leaded to significant depression of HdhSEBP mRNA. It is concluded that the expression levels of HdhSEBP are probably involved in the regulation of oxidation/reduction homeostasis affected by dietary selenium, iron or zinc.
     3 Molecular Cloning, Characterization and expression analysis of heat shock protein 90 from abalone, Haliotis discus hannai Ino in response to dietary selenium,iron and zinc
     In the present study, the cDNA of Haliotis discus hannai Ino HSP90 (designated HdhHSP90) was cloned by the combination of homology cloning and rapid amplification of cDNA ends (RACE) approaches. The full-length cDNA of HdhHSP90 was of 2660 bp, including an open reading frame (ORF) of 2187bp encoding a polypeptide of 728 amino acids with predicted molecular weight of 84.134kDa and theoretical isoelectric point of 4.619.BLAST analysis revealed that HdhHSP90 shared high similarity with other known HSP90s, and the five conserved amino acid blocks defined as HSP90 protein family signatures were also identified in HdhHSP90, which indicated that HdhHSP90 should be a cytosolic member of the HSP90 family.Relatively higher levels of HdhHSP90 mRNA expression were detected in gonad and haemocytes by real-time PCR assays. The expression levels of HdhHSP90 in haemocytes and hepatopancreas were measured by real-time PCR after abalones were fed with different semipurified diets containing selenium (0.15,1.32 and 48.7 mg/kg), zinc (6.69,33.85 and 710.63 mg/kg) and iron (29.17,65.7 and 1267.2 mg/kg) supplements for 20 weeks, respectively. The results showed that the expression of the HdhHSP90 mRNA increased and reached the maximum at optimal dietary selenium(1.32 mg/kg), iron (65.7 mg/kg) and zinc (33.5 mg/kg), respectively.Deficient or excessive level of dietary selenium, iron or zinc, respectively, leaded to significant depression of HdhHSP90 mRNA.It is concluded that the expression of HdhHSP90 could be affected by dietary minerals and were involved in the anti-oxidation stress functions arose by dietary selenium, iron or zinc.
     4 Transcriptional up-regulation of a novel ferritin homolog in abalone Haliotis discus hannai Ino by dietay iron
     A novel cDNA encoding ferritin (HdhNFT) was cloned from the hepatopancreas of abalone, Haliotis discus hannai Ino. The deduced protein contains 171 amino acid residues with a predicted molecular weight (MW) about 19.8 KDa and theoretical isoelectric point (pI) of 4.792.Amino acid alignment revealed that HdhNFT shared high similarity with other known ferritins. The HdhNFT contained a highly conserved motif for the ferroxidase center, which consists of seven residues of a typical vertebrate heavy-chain ferritin with a typical stem-loop structure. HdhNFT mRNA contains a 27 bp iron-responsive element (IRE) in the 5'-untranslated region. This IRE exhibited 82.14% similarity with abalone H. discus discus and 78.57% similarity with Pacific oyster Crassostrea gigas IREs. By real-time PCR assays, the mRNA transcripts of HdhNFT were found to be higher expressed in kidney, hepatopancreas, gill, mantle and muscle than in haemocytes and gonad. Moreover, mRNA expression levels of HdhNFT in the hepatopancreas and haemocytes were measured by real-time PCR in abalone fed with graded levels of dietary iron (29.2,65.7,1267.2 and 6264.7 mg/kg). Results showed that the expression of the HdhNFT mRNA increased with dietary iron contents. Furthermore, the maximum value of the HdhNFT mRNA was found in the treatment with 6264.7 mg/kg of dietary iron. These data indicated that dietary iron can up-regulate HdhNFT at transcriptional level in abalone Haliotis discus hannai Ino.
     5 Effect of vitamin C supplements on antioxidant defence and stress proteins in hepatopancreas of abalone Haliotis discus hannai Ino
     Studies were conducted to investigate the effects of dietary ascorbic acid on transcriptional expression of antioxidant proteins, stress responsive proteins and nuclear factor Rel/NF-κB in hepatopancreas of abalone Haliotis discus hannai Ino (initial average length:84.36±0.24 mm) using real-time quantitative PCR assays. Four practical diets were formulated to contain 0.0,70.3,829.8 and 4967.5 mg ascorbic acid equivalent kg-1 diet, supplied as L-ascorbyl-2-polyphosphate (LAPP). Each diet was fed to triplicate groups of abalone adult in seawater floating acrylic tank (200 L), and each cage was stocked with 15 abalone adult. Abalone were fed once daily(17:00) to apparent satiation for 24 weeks. The results showed that adequate vitamin C (70.3 mg/kg) could significantly up-regulate expression levels of Cu/Zn-SOD, CAT,GST, NFT and HSP26 in hepatopancreas of abalone fed with dietary vitamin C supplement compared with abalone fed with vitamin C-free diet. But the expression levels of CAT, GST and HSP26 were decreased in abalone fed with excessive dietary vitamin C (70.3 mg/kg) compared with abalone fed adequate dietary vitamin C (70.3 mg/kg).And adequate vitamin C (70.3 mg/kg) could significantly down-regulate expression levels of Mn-SOD, GPX,TPx,SEBP, HSP70, HSP90 in hepatopancreas of abalone compared with abalone fed with vitamin C-free diet. In addition, significant up-regulations of expression levels of Mn-SOD, GPX,TPx, SEBP, NFT, HSP70, HSP90 and Rel/NF-κB were observed in abalone fed with excessive dietary vitamin C (829.8 and 4967.5 mg/kg) compared with abalone fed adequate dietary vitamin C (70.3 mg/kg). It is concluded that adequate dietary vitamin C could affect the expression levels of antioxidant proteins,stress responsive proteins and nuclear factor Rel/NF-KB in hepatopancreas of abalone at transcriptional level. Meanwhile, adequate dietary vitamin C could reduce the oxidative stress partly though the coordinated effect with antioxidant proteins in abalone.
     In conclusion, adequate dietary minerals (selenium, zinc and iron) or antioxidant (vitamin C) could affect the mRNA expression of antioxidant gene, increase the total antioxidant capacities of abalone H. discus hannai, and then eliminate or reduce the oxidative stresses caused by kinds of ROS in abalone H. discus hannai.

引文

Abele D, Puntarulo S. Formation of reactive species and induction of antioxidant defense systems in polar and temperate marine invertebrates and fish. Comp Biochem Physiol A 2004; 138:405-415.
    Abele D. Toxic oxygen:the radical life-giver. Nature,2002; 420:27.
    Aispuro-Hemandez E, Garcia-Orozco KD, Muhlia-Almazan A, Del-Toro-Sanchez L, Robles-Sanchez RM, Hernandez J, et al. Shrimp thioredoxin is a potent antioxidant protein. Comp Biochem Physiol C 2008; 148:94-99.
    Akira S., Uematsu S., Takeuchi O., Pathogen recognition and innate immunity. Cell 2006; 124:783-801.
    Alder MN, Rogozin IB, Iyer LM, Glazko GV, Cooper MD, Pancer Z. Diversity and function of adaptive immune receptors in a jawless vertebrate. Science 2005; 310:1970-1973.
    Apel K, Hirt H. Reactive oxygen species:metabolism, oxidative stress, and signal transduction. Annu Rev Plant Bio 2004;55:373-379.
    Bartosz G.Reactive oxygen species:Destroyers or messengers? Biochem Pharmacol.2009; 77(8):1303-15.
    Berger MM. Can oxidative damage be treated nutritionally? Clin Nutr 2005; 24(2):172-83.
    Beutler B. Innate immunity:an overview. Mol Immunol 2004; 40:845-59.
    Bilbao E, de Cerio OD, Cajaraville MP, Cancio I. Cloning and expression pattern of peroxisomal enzymes in the mussel Mytilus galloprovincialis and in the thicklip grey mullet Chelon labrosus:Generation of new tools to study peroxisome proliferation. Marine Environmental Research 2006; 62(S1):S109-S112.
    Bogdan C, ROllinghoff M, Diefenbach A. Reactive oxygen and reactive nitrogen intermedia-tes in innate and specific immunity. Curr Opin Immunol 2000; 12:64-76.
    Boutet I, Tanguy A, Moraga D. Response of the Pacific oyster Crassostrea gigas to hydrocar-bon contamination under experimental conditions. Gene 2004; 329:147-57.
    Brouwer M, Brouwer TH, Grater W. Brown-Peterson. Replacement of a cytosolic copper/zinc superoxide dismutase by a novel cytosolic manganese superoxide dismutase in crustaceans that use copper (haemocyanin) for oxygen transport. Biochem J.2003; 374:219-228.
    Brouwer M, Hoexum Brouwer T, Grater W., Syring R. Crustacea that are dependent on copper for 02 transport have a Cu-specific metallothionein and have replaced Cu/Zn-SOD with MnSOD. Comp Biochem Phys A.2000; 126(1): 18-18(1).
    Bugge DM, Hegaret H, Wikfors GH, Allam B. Oxidative burst in hard clam (Mercenaria mercenaria) haemocytes. Fish Shellfish Immunol 2007;23(1):188-196.
    Calder PC, Kew S. The immune system:a target for functional foods? Br J Nutr 2002; 88 (Suppl 2):S165-77.
    Campa-Cordova AI, Hernandez-Saaverdra NY, De Philippis R. Ascencio F. Generation of superoxide anion and SOD activity in haemocytes and muscle of American white shrimp (Litopenaeus vannameii) as a response to β-glucan and sulphated polysaccharide. Fish Shellfish Immunol.2002; 12:353-366.
    Cetinkaya O, Silig Y, Cetinkaya S, Demirezer LO. The effects of Rumex patientia extract on rat liver and erythrocyte antioxidant enzyme system. Pharmazie 2002; 57(7):487-8.
    Chang C, Su MS, Chen HY, Liao IC. Dietary beta-1,3-glucan effectively improves immunity and survival of Penaeus monodon challenged with white spot syndrome virus. Fish Shellfish Immunol.20Q3;15(4):297-310.
    Chen Y, Zhang YX, Fan TJ, Meng L, Ren GC, Chen SL. Molecular identification and ex-pression analysis of the natural killer cell enhancing factor (NKEF) gene from turbot (Scophthalmus maximus). Aquaculture 2006; 261:1186-1193.
    Cheng W, Tung YH, Chiou TT, Chen JC. Cloning and characterisation of mitochondrial manganese superoxide dismutase (mtMnSOD) from the giantfreshwater prawn Macrobrachium rosenbergii. Fish Shellfish Immunol. 2006a;21(4):453-466.
    Cheng W, Tung YH, Liu CH, Chen JC. Molecular cloning and characterisation of copper/zinc superoxide dismutase (Cu,Zn-SOD) from the giant freshwater prawn Macrobrachium rosenbergii.Fish Shellfish Immunol.2006b; 21(1): 102-112.
    Cimen MY.Free radical metabolism in human erythrocytes. Clin Chim Acta 2008; 390:1-11.
    Cossu C, Doyotte A, Jacquin MC, Babut M, Exinger A, Vasseur P. Glutathione reductase, selenium-dependent glutathione peroxidase, glutathione levels, and lipid peroxidation in freshwater bivalves, Unio tumidus, as biomarkers of aquatic contamination in field studies. Ecotoxicol Environ Saf 1997;38(2):122-31.
    Crapo JD, Oury T, Rabouille C, Slot JW, Chang LY. Copper,zinc superoxide dismutase is primarily a cytosolic protein in human cells. Proc Natl Acad Sci U S A.1992;89(21):10405-10409.
    Cressey D. Future fish. Nature,2009,485:398-400.
    David E, Tanguy A, Moraga D. Peroxiredoxin 6 gene:A new physiological and genetic indicator of multiple environmental stress response in Pacific oyster Crassostrea gigas. Aquatic Toxicology 2007; 84(3):389-398.
    De Zoysa M, Pushpamali WA, Oh C, Whang I, Kim SJ, Lee J. Transcriptional up-regulation of disk abalone selenium dependent glutathione peroxidase by H2O2 oxidative stress and Vibrio alginolyticus bacterial infection. Fish Shellfish Immunol 2008;25:446-457.
    Dondero F, Piacentini L, Marsano F, Rebelo M, Vergani L, Venier P, Viarengo A. Gene transcription profiling in pollutant exposed mussels (Mytilus spp.) using a new low-density oligonucleotide microarray. Gene 2006; 376: 24-36.
    Dong WR, Xiang LX, Shao JZ. Cloning and characterisation of two natural killer enhancing factor genes (NKEF-A and NKEF-B) in pufferfish, Tetraodon nigroviridis. Fish Shellfish Immunol 2007; 22(1-2):1-15.
    Doyen P, Bigot A, Vasseur P, Rodius F. Molecular cloning and expression study of pi-class glutathione S-transferase (pi-GST) and selenium-dependent glutathione peroxidase (Se-GPx) transcripts in the freshwater bivalve Dreissena polymorpha. Comp Biochem Physiol C 2008; 147(1):69-77.
    Doyen P, Vasseur P, Rodius F. Identification, sequencing and expression of selenium-dependent glutathione peroxidase transcript in the freshwater bivalve Unio tumidus exposed to Aroclor 1254. Comp Biochem Physiol C,2006; 144: 122-129.
    Doyotte A, Cossu C, Jacquin MC, Babut M, Vasseur P. Antioxidant enzymes, glutathione and lipid peroxidation as relevant biomarkers of experimental or field exposure in the gills and the digestive gland of the freshwater bivalve Unio tumidus. Aquatic Toxicology 1997; 39(2):93-110.
    Ekanayake PM, Kang HS, Zyosa MD, Jee YH,Lee YH, Lee JH. Molecular cloning and characterization of Mn-superoxide dismutase from disk abalone (Haliotis discus discus). Comp Biochem Physiol B 2006; 145: 318-324.
    Ekanayake PM, Zoysa MD, Kang HS, Wan Q, Jee YH, Lee YH, Kim SJ, Lee JH. Cloning, characterization and tissue expression of disk abalone (Haliotis discus discus) catalase. Fish Shellfish Immunol.2008; 24(3):267-78.
    Ezekowitz RA, Hoffmann JA. The blossoming of innate immunity. Curr Opin Immunol 1998; 10:9-11.
    Fang YZ, Yang S, Wu GY.Free Radicals, antioxidants, and nutrition. Nutrition,2002; 18:872-879.
    FAO. The State of World Fisheries and Aquaculture 2008. FAO,2009
    Fearon DT, Locksley RM.The instructive role of innate immunity in the acquired immune response. Science 1996; 272: 50-53.
    Finkel T, Holbrook NJ. Oxidants, oxidative stress and the biology of ageing. Nature.2000; 408(6809):239-47.
    Finkel T. Oxygen radicals and signaling. Curr Opin Cell Biol.1998; 10(2):248-253.
    Fridovich I. Superoxide dismutases. An adaptation to a paramagnetic gas. J Biol Chem 1989; 264:7761-4.
    Geret F, Serafim A, Barreira L, Bebianno MJ. Response of antioxidant systems to copper in the gills of the clam Ruditapes decussatus. Mar Environ Res 2002;54(3-5):413-7.
    Giorgio M, Migliaccio E, Orsini F, Paolucci D, Moroni M, Contursi C, Pelliccia G,Luzi L, Minucci S, Marcaccio M, Pinton P,Rizzuto R, Bernardi P, Paolucci F, Pelicci PG Electron Transfer between Cytochrome c and p66Shc Generates Reactive Oxygen Species that Trigger Mitochondrial Apoptosis. Cell 2005; 122(2):221-233.
    Gomez-Anduro GA, Barillas-Mury CV, Peregrino-Uriarte AB, Gupta L, Gollas-Galvan T, Hernandez-Lopez J, Yepiz-Plascencia G The cytosolic manganese superoxide dismutase from the shrimp Litopenaeus vannameii: Molecular cloning and expression. Dev Comp Immunol 2006; 30(10):893-900.
    Goodall CP, Bender RC, Broderick EJ, Bayne CJ. Constitutive differences in Cu/Zn superoxide dismutase mRNA levels and activity in hemocytes of Biomphalaria glabrata (Mollusca) that are either susceptible or resistant to Schistosoma mansoni (Trematoda). Mol Biochem Parasitol.2004; 137(2):321-328.
    Gotia S, Popovici I, Hermeziu B. Antioxidant enzymes levels in children with juvenile rheumatoid arthritis. Rev Med Chir Soc Med Nat Iasi 2001;105(3):499-503.
    Ha EM, Oh CT, Bae YS, Lee WJ.2005a, A direct role for dual oxidase in Drosophila gut immunity. Science,310: 847-50.
    Ha EM, Oh CT, Ryu JH, Bae YS, Kang SW, Jang IH, Brey PT, Lee WJ. An antioxidant system required for host protection against gut infection in Drosophila. Dev Cell 2005b; 8:125-132.
    Hoffmann JA, Kafatos FC, Janeway CA, Ezekowitz RAB. Phylogenetic perspectives in innate immunity. Science 1999; 284:1313-1318.
    Hoyles L, Vulevic J. Diet, immunity and functional foods. Adv Exp Med Biol.2008; 635:79-92.
    Hwang Y, Jeong M, Park JS, Kim MH, Lee DB, Shin BA, Mukaida N, Ellis LM, Kim HR, Ahn BW. Interleukin-1 bold italic beta stimulates IL-8 expression through MAP kinase and ROS signaling in human gastric carcinoma cells. Oncogene 2004; 23:6603-6611.
    Jezek P, Hlavata L. Mitochondria in homeostasis of reactive oxygen species in cell, tissues, and organism. In J Biochem Cell Biol 2005;37(12):2478-2503.
    Kamata H, Manabe T, Oka S, Kamata K, Hirata H. Hydrogen peroxide activates I kappaB kinases through phosphorylation of serine residues in the activation loops. FEBS Lett 2002; 519:231-7.
    Kang SW, Rhee SG, Chang TS, Jeong W, Choi MH.2-Cys peroxiredoxin function in intracellular signal transduction, therapeutic implications.Trends Mol Med 2005; 11(12):571-578.
    Khachik F, Carvalho L, Bernstein PS, Muir GJ, Zhao DY, Katz NB.Chemistry, distribution, and metabolism of tomato carotenoids and their impact on human health. Exp Biol Med 2002; 227(10):845-51.
    Kim IH, Kim K, Rhee SG. Induction of an antioxidant protein of Saccharomyces cerevisiae by O2, Fe3+, or 2-mercaptoethanol. Proc Natl Acad Sci USA.1989; 86:6018-6022.
    Kim KY, Lee SY, Cho YS, Bang IC, Kim KH,Kim DS, Nam YK. Molecular characteri-zation and mRNA expression during metal exposure and thermal stress of copper/zinc-and manganese superoxide dismutases in disk abalone, Haliotis discus discus. Fish Shellfish Immunol 2007;23:1043-1059.
    Klaunig JE, Kamendulis LM. The role of oxidative stress in carcinogenesis. Annu Rev Pharmacol Toxicol 2004; 44: 239-67.
    Labreuche Y, Lambert C, Soudant P, Boulo V, Huvet A, Nicolas JL.2006. Cellular and molecular hemocyte responses of the Pacific oyster, Crassostrea gigas, following bacterial infection with Vibrio aestuarianus strain 01/32. Microbes Infect.8(12-13):2715-24.
    Lee KS, Kim SR, Park NS, Kim I, Kang PD, Sohn BH, Choi KH, Kang SW, Je YH,Lee SM, Sohn HD, Jin BR. Characterization of a silkworm thioredoxin peroxidase that is induced by external temperature stimulus and viral infection. Insect Biochem. Mol Biol 2005;35(1):73-84.
    Li C, Ni D, Song L, Zhao J, Zhang H, Li L. Molecular cloning and characterization of a catalase from Zhikong scallop Chlamys farreri. Fish Shellfish Immunol 2008;24(1):26-34.
    Limon-Pacheco J, Gonsebatt ME. The role of antioxidants and antioxidant-related enzymes in protective responses to environmentally induced oxidative stress. Mutation Research.2009; 674:137-147.
    Liu CH, Tseng MC, Cheng W. Identification and cloning of the antioxidant enzyme, glutathione peroxidase, of white shrimp, Litopenaeus vannamei, and its expression following Vibrio alginolyticus infection. Fish Shellfish Immunology 2007; 23(1),34-45.
    Maningas MBB, Koyama T, Kondo H, Hirono I, Aoki T. A peroxiredoxin from kuruma shrimp, Marsupenaeus japonicus, inhibited by peptidoglycan. Dev Com Immunol 2007; 2008; 32(3):198-203.
    McEligot AJ, Yang S, Meyskens FL Jr. Redox regulation by intrinsic species and extrinsic nutrients in normal and cancer cells. Annu Rev Nutr 2005;25:261-295.
    Merle PL, Sabourault C, Richier S, et al. Catalase characterization andimplication in bleaching of a symbiotic sea anemone. Free Radic Biol Med 2007; 42:236-246.
    Mourente G,Diaz-Salvago E, Bell J G, et al. Increased activities of hepatic antioxidant defence enzymes in juvenile gilthead sea bream (Sparus aurata L.)fed dietary oxidized oil:attenuation by dietary vitamin E. Aquaculture 2002; 214:343—361.
    Munoz M, Cedeno R, Rodriguez J, Van der Knaap WPW, Mialhe E, Bachere E. Measurement of reactive oxygen intermediate production in haemocytes of the penaeid shrimp, Penaeus vannamei. Aquaculture 2000; 191:89-107.
    Murphy R, DeCoursey TE. Charge compensation during the phagocyte respiratory burst. Biochim Biophys Acta 2006; 1757(8):996-1011.
    Muruganandan S, Gupta S, Kataria M, Lal J, Gupta PK. Mangiferin protects the streptozo-tocin-induced oxidative damage to cardiac and renal tissues in rats. Toxicology 2002;176(3):165-73.
    Nakamura H, Nakamura K, Yodoi J. Redox regulation of cellular activation. Annu Rev Immunol 1997; 15:351-69.
    Ni DJ, Song LS, Gao Q, Wu LT, Yu YD. The cDNA cloning and mRNA expression of superoxide dismutase (SOD) gene in scallop Chlamys farreri. Fish Shellfish Immunol 2007; 23:1032-1042.
    Nordberg J, Arner ES. Reactive oxygen species, antioxidants, and the mammalian thioredoxin system. Free Radic Biol Med.2001;31(11):1287-312.
    Okado-Matsumoto A, Matsumoto A, Fujii J, Taniguchi N. Peroxiredoxin IVis a secretable protein with heparin-binding properties under reduced conditions. J Biochem (Tokyo) 2000; 127(3):493-501.
    Peterson KJ, Butterfield NJ. Origin of the Eumetazoa:testing ecological predictions of molecular clocks against the Proterozoic fossil record. Proc Natl Acad Sci USA 2005;102(27):9547-52.
    Peterson KJ, Lyons JB, Nowak KS, Takacs CM, Wargo MJ, McPeek MA.Estimating metazoan divergence times with a molecular clock. Proc. Natl.Acad. Sci. U.S.A.2004;101(17):6536-6541.
    Pipe RK, Porte C, Livingstone DR. Antioxidant enzymes associated with the blood cells and haemolymph of the mussel Mytilus edulis. Fish Shellfish Immunology 1993; 3:221-233.
    Rhee SG,Chae HZ, Kim K. Peroxiredoxins:a historical overview and speculative preview of novel mechanisms and emerging concepts in cell signaling. Free Radic Biol Med 2005; 38(12):1543-1552.
    Rhee SG,Kang SW, Jeong W,Chang TS, Yang KS, Woo HA. Intracellular messenger function of hydrogen peroxide and its regulation by peroxiredoxins. Curr Opin Cell Biol 2005; 17(2):183-189.
    Rhee SG Cell signaling, H2O2,a necessary evil for cell signaling. Science 2006; 312(5782):1882-1893.
    Roch P.Defense mechanisms and disease prevention in farmed invertebrates. Aquaculture 1999; 172(1-2):125-145.
    Seifried HE, Anderson DE, Fisher El, Milner JA. A review of the interaction among dietary antioxidants and reactive oxygen species. J Nutr Biochem.2007; 18(9):567-79.
    Seifried HE. Oxidative stress and antioxidants:a link to disease and prevention. J Nutr Biochem.2007; 18(3):168-171.
    Shinohara R, Mano T, Nagasaka A, Hayashi R, Uchimura K, Nakano I, Watanabe F, Tsugawa T, Makino M, Kakizawa H. Lipid peroxidation levels in rat cardiac muscle are affected by age and thyroid status. J Endocrinol 2000; 164: 97-102.
    Sies H. Oxidative stress. In:Encyclopedia of stress. (Fink G.ed),2000; Vol.3, pp.102-105. Academic press, San Diego, CA.
    Tavares-Sanchez OL, Gomez-Anduro GA, Felipe-Ortega X, Islas-Osuna MA, Sotelo MRR, Barillas-Mury C, Yepiz-Plascencia G Catalase from the white shrimp Penaeus (Litopenaeus) vannamei:molecular cloning and protein detection. Comp Biochem Physiol B Biochem Mol Biol.2004; 138(4):331-337.
    Utke K, Bergmann S, Lorenzen N, Kollner B, Ototake M, Fischer U. Cell-mediated cytotoxicity in rainbow trout, Oncorhynchus mykiss, infected with viral haemorrhagic septicaemia virus. Fish Shellfish Immunol.2007; 22(3): 182-196.
    Valko M, Leibfritz D, Moncol J, Cronin MT, Mazur M, Telser J.Free radicals and antioxidants in normal physiological functions and human disease. Int J Biochem Cell Biol 20Q7; 39(1):44-84.
    Valko M, Rhodes CJ, Moncol J, Izakovic M, Mazur M. Free radicals, metals and antioxidants in oxidative stress-induced cancer. Chem Biol Interact 2006;160:1-40.
    Weisiger RA, Fridovich I. Mitochondrial Superoxide Dismutase. Site of synthesis and intramitochondrial localization. J Biol Chem 1973;248(13):4793-4796.
    Winston GW, DiGiulio RT. Prooxidant and antioxidant mechanisms in aquaticorganisms. Aquat Toxicol 1991;19: 137-161.
    Yildirim O, Buyukbingol Z. Effects of supplementation with a combination of cobalt and ascorbic acid on antioxidant enzymes and lipid peroxidation levels in streptozocin-diabetic rat liver. Biol Trace Elem Res 2002; 90(1-3):143-54.
    Zelck UE, Janje B, Schneider O. Superoxide dismutase expression and H2O2 production by hemocytes of the trematode intermediate host Lymnaea stagnalis (Gastropoda). Dev Comp Immunol 2005; 29(4):305-314.
    Zhang Q, Li F, Wang B, Zhang I, Liu Y, Zhou Q, Xiang J. The mitochondrial manganese superoxide dismutase gene in Chinese shrimp Fenneropenaeus chinensis:Molecular cloning, characterization and expression. Dev Com Immunol 2007;31:429-440.
    Zhang Q, Li F, Zhang J, Wang B, Gao H, Huang B, Jiang H, Xiang J. Molecular cloning, expression of a peroxiredoxin gene in Chinese shrimp Fenneropenaeus chinensis and the antioxidant activity of its recombinant protein. Mol Immunol 2007;44:3501-3509.
    Zhang Q, Li F, Zhang X, Dong B, Zhang J, Xie Y, Xiang J.cDNA cloning, characterization and expression analysis of the antioxidant enzyme gene, catalase, of Chinese shrimp Fenneropenaeus chinensis. Fish Shellfish Immunol.2008; 24(5):584-91.
    曹道俊,杨毅军.氧自由基与临床.中国医药科技出版社,2000.
    付京花,张文兵,麦康森,冯秀妮,徐玮,刘付志国维生素D对皱纹盘鲍生长和体组织抗氧化反应的影响.高技术通讯.2006；16(12)：1306-1311.
    郭碧花.活性氧诱导细胞凋亡作用机理的研究进展.川北医学院学报,2002；17(4)：166-169.
    林晓明,李勇.高级营养学.北京大学医学出版社.2004；pp,174-200.
    刘晓华,曹俊明,吴建开,周萌,赵红霞,蓝汉冰,谢从新.饲料中添加谷胱甘肽对凡纳滨对虾肝胰腺抗氧化指标和脂质过氧化物含量的影响.水产学报.2007；31(2),235-240.
    麦康森,艾庆辉,徐玮,刘付志国.水产养殖中的环境胁迫及其预防——营养学途径.中国海洋大学学报.2004；34(5)：767-774.
    牟海津,江晓路,刘树清.免疫多糖对栉孔扇贝酸性磷酸酶、碱性磷酸酶和超氧化物歧化酶活性的影响.青岛海洋大学学报.1999；29(3)：462-468.
    万敏,麦康森,马洪明,徐玮,刘付志国.硒和维生素E对皱纹盘鲍血清抗氧化酶活力的影响.水生生物学报.2003；28(5)：496-503.
    徐芝勇.细胞内活性氧与肿瘤.生物学通报.2004；39(9),24-25.
    于善谦,王洪海,朱乃硕,叶荣.《免疫学导论》.高等教育出版社.施普林格出版社.1999.
    Almar M, Otero L., Santos C., Gallego JG.,1998. Liver glutathione content and glutathione-dependent enzymes of two species of freshwater fish as bioindicators of chemical pollution. J. Environ. Sci. Health.33,769-783.
    Anantharamaiah GM.,Mishra VK.,Garber DW., Datta G., Handattu SP., Palgunachari MN., Chaddha M., Navab M., Reddy S.T., Segrest JP., Fogelman AM.,2007.Structural requirements for antioxidative and anti-inflammatory properties of apolipoprotein A-I mimetic peptides. J. Lipid. Res.48(9),1915-1923.
    Arthur JR.,2000. The glutathione peroxidases. Cell. Mol. Life. Sci.57,1825-1835.
    Berry MJ., Banu L., Chen Y., Mandel SJ., Kieffer JD., Harney JW., Larsen PR.,1991.Recognition of UGA as a selenocysteine codon in type I deiodinase requires sequences in the 3'untranslated region. Nature.353,273-276.
    Bogdan C., Rollinghoff M., Diefenbach A.,2000. Reactive oxygen and reactive nitrogen intermediates in innate and specific immunity. Curr. Opin. Immunol.12,64-76.
    Cabreiro F., Picot C.R., Perichon M., Friguet B., Petropoulos I.,2009. Overexpression of Methionine Sulfoxide Reductases A and B2 Protects MOLT-4 Cells Against Zinc-Induced Oxidative Stress. Antioxid. Redox. Sign.11(2), 215-226.
    Calabrese V., Lodi R.,Tonon C., D'Agata V., Sapienza M., Scapagnini G.,Mangiameli A., Pennisi G.,Stella AM., Butterfield DA.,2005. Oxidative stress, mitochondrial dysfunction and cellular stress response in Friedreich's ataxia. J. Neurol. Sci.233(1-2),145-162.
    Chen H., Mai KS., Zhang WB., Liufu ZG., Xu W., Tan BP.,2005. Effects of dietary pyridoxine on immune responses in abalone,Haliotis discus hannai Ino. Fish. Shellfish. Immunol.19:241-252.
    Cheng W., Juang FM., Chen JC.,2004. The immune response of Taiwan abalone Haliotis diversicolor supertexta and its susceptibility to Vibrio parahaemolyticus at different salinity levels. Fish. Shellfish. Immunol.16,295-306.
    Chu FF., Doroshow JH., Esworthy RS.,1993. Expression, characterization, and tissue distribution of a new cellular selenium-dependent glutathione peroxidase, GSHPx-GI. J. Biol. Chem.268,2571-2576.
    Chung SS., Kim M., Youn BS., Lee NS., Park JW., Lee IK., Lee YS., Kim JB., Cho YM., Lee HK., Park KS.,2009. Glutathione peroxidase 3 mediates the antioxidant effect of peroxisome proliferator-activated receptor gamma in human skeletal muscle cells. Mol. Cell. Biol.29(1),20-30.
    Cossu C., Doyotte A., Jacquin MC.,Babut M., Exinger A., Vasseur P.,1997. Glutathione reductase, selenium-dependent glutathione peroxidase, glutathione levels, and lipid peroxidation in freshwater bivalves, Unio tumidus, as biomarkers of aquatic contamination in field studies. Ecotoxicol. Environ. Saf.38,122-131.
    Dandapat J., Chainy GBN., Rao KJ.,2000. Dietary vitamin-E modulates antioxidant defence system in giant freshwater prawn, Macrobrachium rosenbergii. Comp. Biochem. Physiol. C 127,101-115.
    De Zoysa M., Pushpamali WA., Oh C., Whang I., Kim SJ.,Lee J.,2008. Transcriptional up-regulation of disk abalone selenium dependent glutathione peroxidase by H2O2 oxidative stress and Vibrio alginolyticus bacterial infection. Fish. Shellfish. Immunol.25,446-457.
    Depege N., Drevet J., Boyer N.,1998. Molecular cloning and characterization of tomato cDNAs encoding glutathione peroxidase-like proteins. Eur. J. Biochem.253,445-451.
    Doyen P., Bigot A., Vasseur P., Rodius F.,2008. Molecular cloning and expression study of pi-class glutathione S-transferase (pi-GST) and selenium-dependent glutathione peroxidase(Se-GPx) transcripts in the freshwater bivalve Dreissena polymorpha. Comp. Biochem. Physiol. C 147,69-77.
    Doyen P., Vasseur P., Rodius F.,2006. Identification, sequencing and expression of selenium-dependent glutathione peroxidase transcript in the freshwater bivalve Unio tumidus exposed to Aroclor 1254. Comp. Biochem. Physiol. C 144,122-129.
    Fang YZ., Yang S.,Wu GY.,2002. Free Radicals, antioxidants, and nutrition. Nutrition.18,872-879.
    Fu JH.,Zhang WB., Mai KS., Feng XN.,Xu W., Liufu ZG., Ma HM.,Ai QH., Effects of vitamin E on antioxidant enzyme activities and fatty acid compositions in juvenile abalone Haliotis discus hannai Ino. J. Shellfish. Res. 26(3),809-814.
    Fu JH, Zhang WB, Mai KS, Feng XN, Xu W, Liufu ZG, Tan BP. Effects of dietary vitamin A on antioxidant responses of abalone Haliotis discus hannai Ino. Acta. Oceanol. Sin.2006;25,141-150.
    Genestra M. Oxyl radicals, redox-sensitive signalling cascades and antioxidants. Cell. Signal.2007; 19:1807-1819.
    Ghyselinck NB, Dufaure JP. A mouse cDNA sequence for epididymal androgen-regulated proteins related to glutathione peroxidase. Nucleic Acids Res 1990; 18:7144.
    Grundener-Culemann E, Martin GW, Harney JW, Berry M. Two distinct SECIS structures capable of directing selenocysteine incorporation in eukaryotes. RNA 1998; 5,625-635.
    Hatfield DL., Gladyshev VN.,2002. How selenium has altered our understanding of the genetic code. Mol. Cell. Biol. 22,3565-3576.
    Hooper C., Day R., Slocombe R., Handlinger J., Benkendorff K.,2007. Stress and immune responses in abalone: limitations in current knowledge and investigative methods based on other models. Fish. Shellfish. Immunol.22, 363-739.
    Jo PG.,Choi YK., Choi CY.,2008. Cloning and mRNA expression of antioxidant enzymes in the Pacific oyster, Crassostrea gigas in response to cadmium exposure. Comp. Biochem. Physiol. C 147,460-469.
    Karasu C.,2000. Time course of changes in endothelium-dependent and-independent relaxation of chronically diabetic aorta:role of reactive oxygen species. Eur. J. Pharmacol.392,163-173.
    Kenet G.,Freedman J., Shenkman B., Regina E., Brok-Simoni F., Holzman F., Vavva F., Brand N., Michelson A., Trolliet M., Loscalzo J., Inbal A.,1999. Plasma glutathione peroxidase deficiency and platelet insensitivity to nitric oxide in children with familial stroke. Arterioscler. Thromb. Vasc. Biol.19,2017-2023.
    Kindermann B., Doring F., Fuchs D., Pfaffl MW., Daniel H.,2005. Effects of increased cellular zinc levels on gene and protein expression in HT-29 cells. BioMetals.18,243-253.
    Koyama H., Omura K., Ejima A., Kasanuma Y., Watanabe C.,Satoh H.1999. Separation of selenium-containing proteins in human and mouse plasma using tandem high-performance liquid chromatography columns coupled with inductively coupled plasma-mass spectrometry. Anal Biochem.267,84-91.
    Kryukov GV., Castellano S.,Novoselov SV., Lobanov AV., Zehtab O., Guigo R., Gladyshev VN.,2003. Characterization of mammalian selenoproteomes. Science.300,1439-1443.
    Latreche L.,Jean-Jean O.,Driscoll DM.,Chavatte L.,2009. Novel structural determinants in human SECIS elements modulate the translational recoding of UGA as selenocysteine. Nucleic. Acids. Res.37(17),5868-5880.
    Liu CH., Tseng MC., Cheng W.,2007. Identification and cloning of the antioxidant enzyme, glutathione peroxidase, of white shrimp, Litopenaeus vannamei, and its expression following Vibrio alginolyticus infection. Fish. Shellfish. Immunol.23,34-45.
    Livak KJ., Schmittgen TD.,2001.Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods.25,402-408.
    Mai KS., Ai QH., Xu W., Liufu ZG.,2004. Stress in aquaculture and its prevention with emphasis on nutritional methods. J. Ocean. Univ. China.34(5),767-74. (in Chinese with English abstract)
    Mai KS., Mercer JP., Donlon J.,1995. Comparative studies on the nutrition of two species of abalone, Haliotis tuberculata L. and Haliotis discus hannai Ino. Ⅲ. Response of abalone to various levels of dietary lipids. Aquaculture 134; 65-80.
    Mai KS., Tan BP.,2000. Iron methionine (FeMet) and iron sulfate (FeSO4) as sources of dietary iron for juvenile abalone, Haliotis discus hannai Ino. J Shellfish Res 19(3),861-868.
    Mix H., Lobanov AV., Gladyshev VN.2007. SECIS elements in the coding regions of selenoprotein transcripts are functional in higher eukaryotes. Nucleic. Acids. Res.35,414-423.
    Mohankumar K., Ramasamy P.,2006. White spot syndrome virus infection decreases the activity of antioxidant enzymes in Fenneropenaeus indicus. Virus. Research.115,69-75.
    Murphy R., DeCoursey T.E.,2006. Charge compensation during the phagocyte respiratory burst. Biochim. Biophys. Acta.1757(8),996-1011.
    Owuor ED., Kong ANT.,2002. Antioxidants and oxidants regulated signal transduction pathways. Biochem. Pharmacol. 64,765-770.
    Papanikolaou G., Pantopoulos K.,2005. Iron metabolism and toxicity. Toxicol. Appl. Pharmacol.202,199-211.
    Ramos-Vasconcelos GR., Hermes-Lima M.,2003. Hypometabolism, antioxidant defenses and free radical metabolism in the pulmonate land snail Helix aspersa. J. Exp. Biol.206,675-685.
    Ren B., Huang W., Akesson B., Ladenstein R.,1997. The crystal structure of seleno-glutathione peroxidase from human plasma at 2.9 A resolution. J. Mol. Biol.268(5),869-885.
    Ren Q., Sun RR., Zhao XF., Wang JX.,2009. A selenium-dependent glutathione peroxidase (Se-GPx) and two glutathione S-transferases (GSTs) from Chinese shrimp (Fenneropenaeus chinensis). Comp. Biochem. Physiol. C. 149,613-623.
    Roch P.,1999. Defense mechanisms and disease prevention in farmed invertebrates. Aquaculture.172(1-2),125-145.
    Rotruck JT, Pope AL, Ganther HE, Swanson AB, Hafeman DG, Hoekstra WG. Selenium:biochemical role as a component of glutathione peroxidase. Science 1973;179:588-590.
    Schwarz KB.,1996. Oxidative stress during viral infection:a review. Free. Radic. Biol. Med.21(5),641-649.
    Shen Q., Chu FF.,Newburger PE.,1993. Sequences in the 3'-untranslated region of the human cellular glutathione peroxidase gene are necessary and sufficient for selenocysteine incorporation at the UGA codon. J. Biol. Chem.268, 11463-11469.
    Song ZR., Ji RX., Yan SF., Chen CS., Zhoug YP., Jiang YH.,Ni ZM.,2000. A spherovirus resulted in mass mortality of Haliotis diversicolor aquatilis. J. Fish. China.24 (5),463-466. (in Chinese with English abstract)
    Swiergosz-Kowalewska R., Bednarska A., Kafel A.,2006. Glutathione levels and enzyme activity in the tissue of bank vole Clethrionomys glareolus chronically exposed to a mixture of metal contaminants. Chemosphere.65,963-974.
    Takahashi K., Cohen HJ.1986. Selenium-dependent glutathione peroxidase protein and activity:immunological investigations on cellular and plasma enzymes. Blood 68:640-645.
    Tamura K., Dudley J.,Nei M., Kumar S.,2007. MEGA4:Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0. Mol. Biol. Evol.24(8),1596-1599.
    Tan BP., Mai KS.,2001.Zinc methionine and zinc sulfate as sources of dietary zinc for juvenile abalone, Haliotis discus hannai Ino. Aquaculture.; 192,67-84.
    Tan JM.,Lancaster M.,Hyatt A., van Driel R., Wong F., Warner S.,2008. Purification of a herpes-like virus from abalone (Haliotis spp.) with ganglioneuritis and detection by transmission electron microscopy.J. Virol. Meth.149, 338-341.
    Thorpe GW., Gong CS.,Alic VJ., Higgins VJ., Dawes IW.,2004. Cells have distinct mechanisms to maintain protection against different reactive oxygen species:oxidative-stress-response genes. Proc. Natl. Acad. Sci. USA.101, 6564-6569.
    Ursini F., Maiorino M., Briglius-Flohe R., Aumann KD., Roveri A., Schomburg D., Flohe L.1995. Diversity of glutathione peroxidases. Method. Enzymol.252,38-53.
    Valavanidis A., Vlahogianni T., Dassenakis M., Scoullos M.,2006. Molecular biomarkers of oxidative stress in aquatic organisms in relation to toxic environmental pollutants. Ecotoxicol. Environ. Saf.64,178-189.
    Wan M., Mai KS., Ma HM.,Xu W., Liufu ZG.,2004. Effect of dietary delenium and Vitamin E on antioxidant enzyme activities in Abalone, Haliotis discus hannai Ino. Acta. Hydrobio. Sinica.28 (5),496-503.(in Chinese with English abstract)
    Woo S., Yum S., Park HS., Lee TK., Ryu JC.,2009. Effects of heavy metals on antioxidants and stress-responsive gene expression in Javanese medaka (Oryzias javanicus). Comp. Biochem. Physiol. C 149,289-299.
    Wool GD.,Reardon CA., Getz GS.,2008. Apolipoprotein A-I mimetic peptide helix number and helix linker influence potentially anti-atherogenic properties. J.Lipid. Res.49,1268-1283.
    Wu XS., Huang KH.,Wei CW., Chen F., Pan CL.,2010. Regulation of cellular glutathione peroxidase by different forms and concentrations of selenium in primary cultured bovine hepatocytes. J. Nutri. Biochem.21,153-161.
    Yeh SP., Liu KF., Chiu ST., Jian SJ., Cheng W., Liu CH.,2009. Identification and cloning of a selenium dependent glutathione peroxidase from giant freshwater prawn, Macrobrachium rosenbergii. Fish. Shellfish. Immunol.27, 181-191.
    Yu BP.,1994. Cellular defenses against damage from reactive oxygen species. Physiol. Rev.74,139-162.
    Zhang WB., Mai KS., Xu W., Ma HM.,2003. Metabolic responses to dietary cholecalciferol and phosphorus in abalone Haliotis discus hannai ino. J.Exp. Zool.299A,110-117.
    Zhou LZH., Johnson AP, Rando TA.,2001.NF-κB and AP-1 mediate transcriptional responses to oxidative stress in skeletal muscle cells. Free. Radic. Biol. Med.31(11),1405-1416.
    Bakheet T, Frevel M, Williams BR, Greer W, Khabar KS. ARED:human AU-rich element-containing mRNA database reveals an unexpectedly diverse functional repertoire of encoded proteins. Nucleic Acids Res 2001;29:246-54.
    Bansal MP, Cook RG, Danielson KG, Medina D. A 14-Kilodalton Selenium-binding protein in mouse liver is fatty acid-binding protein. J Biol Chem 1989; 264:13780-4.
    Barbouti A, Doulias PT, Zhu BZ, Balz Frei B, Galaris D. Intracellular iron, but not copper, plays a critical role in hydrogen peroxide-induced DNA Damage. Free Radic Biol Med 2001;31(4):490-8.
    Calabrese, V.,Lodi, R., Tonon, C.,D'Agata, V., Sapienza, M., Scapagnini, G., Mangiameli, A., Pennisi, G., Stella, A.M., Butterfield, D.A.,2005. Oxidative stress, mitochondrial dysfunction and cellular stress response in Friedreich's ataxia. J. Neurol. Sci.233(1-2),145-162.
    Chen G, Wang H, Miller CT, Thomas DG, Gharib TG, Misek DE, Giordano TJ, Orringer MB, Hanash SM, Beer DG. Reduced selenium-binding protein 1 expression is associated with poor outcome in lung adenocarcinomas. J Pathol 2004;202:321-9.
    Chen H, Mai KS, Zhang WB, Liufu ZG, Xu W, Tan BP.2005. Effects of dietary pyridoxine on immune responses in abalone,Haliotis discus hannai Ino. Fish Shellfish Immunol 2005;19:241-52.
    Chen Q, Adams CC, Usack L, Yang J, Monde RA, Stern DB.An AU-rich element in the 3'untranslated region of the spinach chloroplast petD gene participates in sequence-specific RNA-protein complex formation. Mol Cell Biol 1995; 15:2010-8.
    Cheng W, Juang FM, Chen JC. The immune response of Taiwan abalone Haliotis diversicolor supertexta and its susceptibility to Vibrio parahaemolyticus at different salinity levels. Fish Shellfish Immunol 2004; 16:295-306.
    Chivers PT, Prehoda KE, Raines RT. The CXXC motif:a rheostat in the active site. Biochemistry 1997; 36:4061-6.
    Clark LC, Combs GF, Tumbull BW, Slate EH, Chalker DK, Chow J, Davis LS, Glover RA, Graham GF, Gross EG, Krongrad A, Lesher JL, Park K, Sanders BB Jr, Smith CL, Taylor JR. Effects of selenium supplementation for cancer prevention in patients with carcinoma of the skin. J Am Med Assoc 1996; 276:1957-63.
    Clegg MS, Hanna LA, Niles BJ, Momma TY, Keen CL. Zinc deficiency-induced cell death. IUBMB Life 2005; 57: 661-9.
    Evans P, Halliwell B. Micronutrients:oxidant/antioxidant status. Brit J Nutr 2001;85 (suppl):67S-74S.
    Fang YZ, Yang S, Wu GY. Free Radicals, antioxidants, and nutrition. Nutrition 2002; 18:872-9.
    Felton SP, Landolt ML, Grace R. Effects of selenium dietary enhancement on hatchery-reared coho salmon, Oncorhynchus kisutch (Walbaum), when compared with wild coho:hepatic enzymes and seawater adaptation evaluated. Aquac Res 1996; 27:135-42.
    Fomenko DE, Gladyshev VN. Identity and functions of CxxC-derived motifs. Biochemistry 2003;42:11214-25.
    Fu JH, Zhang WB, Mai KS, Feng XN, Xu W, Liufu Z, Ma HM, Ai QH. Effects of vitamin E on antioxidant enzyme activities and fatty acid compositions in juvenile abalone Haliotis discus hannai Ino. J Shellfish Res 2007; 26(3): 809-14.
    Fu JH, Zhang WB, Mai KS, Feng XN, Xu W, Liufu Z, Tan BP. Effects of dietary vitamin A on antioxidant responses of abalone Haliotis discus hannai Ino. Acta Oceanol Sin 2006a; 25:141-50.
    Hamilton SJ. Review of residue-based selenium toxicity thresholds for freshwater fish. Ecotoxicol Environ Saf 2003;56: 201-10.
    Hidalgo MC, Exposito A, Palma JM, de la Higuera M. Oxidative stress generated by dietary Zn-deficiency studies in rainbow trout(Oncorhynchus mykiss). Int J Biochem Cell Biol 2002; 34(2):183-93.
    lshida T, Tasaki K, Fukuda A, Ishii Y, Oguri K. Induction of a cytosolic 54 kDa protein in rat liver that is highly homologous to selenium-binding protein. Environ Toxicol Pharmacol 1998; 6:249-55.
    Ishii Y, Hatsumura M, Ishida T, Ariyoshi N, Oguri K. A coplanar PCB induces a selenium-binding protein as a major cytosolic protein rat liver. Chemosphere 1996a; 32:509-15.
    Ishii Y, Hatsumura M, Ishida T, Ariyoshi N, Oguri K. Significant induction of a 54-KD protein in rat liver with homologous alignment to mouse selenium-binding protein by a coplanar polychlorinated biphenyl 3,4,5,3', 4'-pentachlorobiphenyl and 3-methylcholanthrene. Toxicol Lett 1996b; 87:1-9.
    Jamba L, Nehru B, Bansal MP. Redox modulation of selenium-binding proteins by cadmium exposures in mice. Mol Cell Biochem 1997; 177(1/2):169-75.
    Janero DR. Malondialdehyde and thiobarbituric acid-reactivity as diagnostic indices of lipid peroxidation and peroxidative tissue injury. Free Radic Biol Med 1990; 9:515-40.
    Jeong J Y, Wang Y, Sytkowski AJ. Human selenium-binding protein-1 (hSP56) interacts with VDU1 in a selenium-dependent manner. Biochem Biophys Res Commun 2009; 379:583-8.
    Jo PG, Choi YK, Choi CY.Cloning and mRNA expression of antioxidant enzymes in the Pacific oyster, Crassostrea gigas in response to cadmium exposure. Comp Biochem Physiol C 2008; 147:460-9.
    Kim H, Kang HJ, You KT, Kim SH, Lee KY, Kim TI, Kim C, Song SY, Kim HJ, Lee C. Suppression of human selenium-binding protein 1 is a late event in colorectal carcinogenesis and is associated with poor survival. Proteomics 2006;6:3466-76.
    Lemire J, Mailloux R, Appanna VD. Zinc toxicity alters mitochondrial metabolism and leads to decreased ATP production in hepatocytes. J Appl Toxicol.2008; 28:175-82.
    Li T, Yang W, Li M, Byun DS, Tong C, Nasser S, Zhuang M, Arango D, Mariadason JM, Augenlicht LH. Expression of selenium-binding protein 1 characterizes intestinal cell maturation and predicts survival for patients with colorectal cancer. Mol Nutr Food Res 2008; 52:1289-99.
    Liu SY, Stadtman TC. Heparin-binding properties of seleniumcontaining thioredoxin reductase from HeLa cells and human lung adenocarcinoma cells. Proc Natl Acad Sci U S A 1997; 94:6138-41.
    Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods 2001;25:402-8.
    Mai KS, Mercer JP, Donlon J. Comparative studies on the nutrition of two species of abalone, Haliotis tuberculata L. and Haliotis discus hannai Ino. Ⅲ. Response of abalone to various levels of dietary lipids. Aquaculture 1995; 134: 65-80.
    Mai KS, Tan BP. Iron methionine (FeMet) and iron sulfate (FeSO4) as sources of dietary iron for juvenile abalone, Haliotis discus hannai Ino. J Shellfish Res 2000;19(3):861-8
    Ong ST, Ho JZS, Ho B, Ding JL. Iron-withholding strategy in innate immunity. Immunobiology 2006; 211:295-314.
    Ozment CP, Turi JL. Iron overload following red blood cell transfusion and its impact on disease severity. Biochim Biophy Acta 2009; 1790:694-701.
    Papanikolaou G, Pantopoulos K. Iron metabolism and toxicity. Toxicol. Appl. Pharmacol.2005; 202:199-211.
    Papp LV, Lu J, Holmgren A, Khanna KK. From Selenium to Selenoproteins:Synthesis, Identity, and Their Role in Human Health. Antioxid Redox Sign.2007; 9(7):775-806.
    Pass HI, Liu Z, Wali A, Bueno R, Land S, Lott D, Siddiq F, Lonardo F, Carbone M, Draghici S. Gene expression profiles predict survival and progression of pleural mesothelioma. Clin Cancer Res 2004; 10:849-59.
    Porat A, Sagiv Y, Elazar Z. A 56-kDa Selenium-binding protein participates in intra-golgi protein transport. J Biol Chem 2000; 275:14457-65.
    Rao L, Puschner B, Prolla TA. Gene expression profiling of low selenium status in the mouse intestine, transcriptional activation of genes linked to DNA damage, cell cycle control and oxidative stress. J Nutr 2001;131:3175-81.
    She Y M, Narindrasorasak S, Yang SY, Spitale N, Roberts EA, Sarkar B. Identification of Metal-binding Proteins in Human Hepatoma Lines by Immobilized Metal Affinity Chromatography and Mass Spectrometry. Mol Cell Proteomics 2003;2:1306-18.
    Song LS, Zou HB, Chang YQ, Xu W, Wu LT. The cDNA cloning and mRNA expression of a potential selenium-binding protein gene in the scallop Chlamys farreri. Dev Comp Immunol 2006; 30:265-73.
    Song ZR, Ji RX, Yan SF, Chen CS, Zhoug YP, Jiang YH, Ni ZM. A spherovirus resulted in mass mortality of Haliotis diversicolor aquatilis. J Fish China 2000; 24 (5):463-6. (in Chinese with English abstract)
    Tamura K, Dudley J, Nei M, Kumar S. MEGA4:Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0. Mol Biol Evol 2007; 24(8):1596-9.
    Tan BP, Mai KS. Zinc methionine and zinc sulfate as sources of dietary zinc for juvenile abalone, Haliotis discus hannai Ino. Aquaculture 2001;192:67-84.
    Tan JM, Lancaster M, Hyatt A, van Driel R, Wong F, Warner S. Purification of a herpes-like virus from abalone (Haliotis spp.) with ganglioneuritis and detection by transmission electron microscopy. J Virol Meth 2008; 149: 338-41.
    Teo M, Tan L, Lim L, Manser E. The tyrosine kinase ACK1 associates with clathrin-coated vesicles through a binding motif shared by arrestin and other adaptors. J Biol Chem 2001;276:18392-8.
    Urvoas A, Amekraz B, Moulin C, Clainche L, Stocklin R, Moutiez M. Analysis of the metal-binding selectivity of the metallochaperone CopZ from Enterococcus hirae by electrospray ionization mass spectrometry. Rapid Commun Mass Spectrom 2003;17:1889-96.
    Walker KW, Lyles MM, Gilbert HF. Catalysis of oxidative protein folding by mutants of protein disulfide isomerase with a single active-site cysteine. Biochemistry 1996; 35:1972-80.
    Wan M, Mai KS,Ma HM,Xu W, Liufu ZG. Effect of dietary delenium and Vitamin E on antioxidant enzyme activities in Abalone, Haliotis discus hannai Ino. Acta Hydrobio Sinica 2004; 28 (5):496-503.(in Chinese with English abstract)
    Wang WF, Mai KS, Zhang WB, Ai QH, Yap CF, Li HT, Liufu ZG.Effects of dietary copper on survival, growth and immune response of juvenile abalone, Haliotis discus hannai Ino. Aquaculture 2009; 297:122-7.
    Wernimont AK, Huffman DL, Lamb AL, O'Halloran TV, Rosenzweig AC. Structural basis for copper transfer by the metallochaperone for the Menkes/Wilson disease proteins. Nat Struct Biol 2000; 7:766-71.
    Xue JP, Xu YP, Jin LJ, Liu G,Sun YX, Li SY, Zhang JC. Effects of traditional Chinese medicine on immune responses in abalone,Haliotis discus hannai Ino. Fish Shellfish Immunol 2008;24:752-8.
    Yang M, Sytkowski AJ.Differential expression and androgen regulation of the human selenium-binding protein gene hSP56 in prostate cancer cells. Cancer Res 1998; 58(14):3150-3.
    Zhang WB, Mai KS, Xu W, Ma HM. Metabolic responses to dietary cholecalciferol and phosphorus in abalone Haliotis discus hannai ino. J Exp Zool 2003;299A:110-117.
    刘宝娟.饲料中的铁、锌和铜对皱纹盘鲍(Haliotis discus hannai Ino)生长、代谢反应和抗氧化作用的影响.2009.中国海洋大学博士学位毕业论文.
    王薇芳.皱纹盘鲍(Haliotis discus hannai Ino)矿物元素(K、Mg、Se、Cu)及糖类营养生理的研究.2008.中国海洋大学博士学位毕业论文.
    Ali A, Bharadwaj S, O'Carroll R, Ovsenek N. HSP90 interacts with and regulates the activity of heat shock factor 1 in Xenopus oocytes. Mol Cell Biol 1998; 18:4949-60.
    Ali A, Krone PH, Pearson DS, Heikkila JJ. Evaluation of stress inducible hsp90 gene expression as a potential molecular biomarker in Xenopus laevis. Cell Stress Chaperon 1996; 1:62-69.
    Beck R, Verrax J, Gonze T, Zappone M, Pedrosa RC, Taper H,Feron O, Calderon PB. Hsp90 cleavage by an oxidative stress leads to its client proteins degradation and cancer cell death. Biochem Pharmacol 2009; 77:375-83.
    Bernelli-Zazzera A, Cairo G, Schiaffonati L, Tacchini L. Stress proteins and reperfusion stress in the liver. Ann N Y Acad Sci,1992;663:120-124.
    Byrd CA, Bommann W, Erdjument-Bromage H, Tempst P, Pavletich N, Rosen N, Nathan CF, and Ding AH. Heat shock protein 90 mediates macrophage activation by Taxol and bacterial lipopolysaccharide. Proc Natl Acad Sci USA. 1999;96:5645-5650.
    Calabrese V, Lodi R, Tonon C, D'Agata V, Sapienza M, Scapagnini G, Mangiameli A, Pennisi G, Stella AM, Butterfield DA. Oxidative stress, mitochondrial dysfunction and cellular stress response in Friedreich's ataxia. J Neurol Sci. 2005;233(1-2):145-62.
    Cara JB, Aluru N, Moyano FJ, Vijayan MM.Food-deprivation induces HSP70 and HSP90 protein expression in larval gilthead sea bream and rainbow trout. Comp Biochem Physiol B 2005; 142:426-431.
    Chen H, Mai KS, Zhang WB, Liufu ZG, Xu W, Tan BP.2005. Effects of dietary pyridoxine on immune responses in abalone, Haliotis discus hannai Ino. Fish & Shellfish Immunology.19:241-252.
    Cheng W, Juang FM, Chen JC. The immune response of Taiwan abalone Haliotis diversicolor supertexta and its susceptibility to Vibrio parahaemolyticus at different salinity levels. Fish Shellfish Immunol 2004; 16:295-306.
    Cheung J, Smith DF. Molecular chaperone interactions with steroid receptors:an update. Mol. Endocrinol 2000; 14: 939-946.
    Chiosis G, Vilenchik M, Kim J,Solit D. Hsp90:the vulnerable chaperone. Drug Discov Today 2004; 9:881-8.
    Choi YK, Jo PG, Choi CY. Cadmium affects the expression of heat shock protein 90 and metallothionein mRNA in the Pacific oyster, Crassostrea gigas. Comp Biochem Physiol C 2008; 147:286-92.
    Clegg MS, Hanna LA, Niles BJ, Momma TY, Keen CL. Zinc deficiency-induced cell death. IUBMB Life 2005; 57: 661-9.
    Craig EA.1985. The heat shock response. CRC Crit Rev Biochem.18:239-80.
    Csermely P, Schnaider T, Soti C, Prohaszka Z, Nardai G. The 90-kDa molecular chaperone family:structure, function, and clinical applications. A comprehensive review. Pharmacol Ther 1998; 79:129-68.
    Dean RT, Hunt JV, Grant AJ, Yamamoto Y, Niki E. Free radical damage to proteins:the influence of the relative localization of radical generation, antioxidants, and target proteins. Free Radic Biol Med 1991;11:161-8.
    Dean, R. T., Hunt, J. V., Grant, A. J., Yamamoto, Y., Niki, E. Free radical damage to proteins:the influence of the relative localization of radical generation, antioxidants, and target proteins. Free Radic Biol Med 1991;11(2): 161-8.
    Donati YR, Kantengwa S, Polla BS.Phagocytosis and heat shock response in human monocytes-macrophages. Pathobiology 1991;59:156-61.
    Dunderski J. Cellular stress response:Defence against metal toxicity. Jugoslov med biohem 2004; 23(1):1-9.
    Encomio VG, Chu FL. Characterization of heat shock protein expression and induced thermotolerance in P. marinus parasitized eastern oysters:lab and field studies. J Shellfish Res 2004; 23:289-90.
    Evans P, Halliwell B. Micronutrients:oxidant/antioxidant status. Brit J Nutr 2001;85(suppl):67S-74S.
    Fukuda A, Osawa T, Oda H, Tanaka T, Toyokuni S, Uchida K. Oxidative Stress Response in Iron-Induced Acute Nephrotoxicity:Enhanced Expression of Heat Shock Protein 90. Biochem Biophys Res Commun 1996; 219(1): 76-81.
    Gao Q, Song L, Ni D, Wu L, Zhang H, Chang Y. cDNA cloning and mRNA expression of heat shock protein 90 gene in the haemocytes of Zhikong scallop Chlamys farreri. Comp Biochem Physiol B 2007; 147:704-715.
    Gao Q, Zhao J M, Song L S, Qiu LM, Yu Y D, Zhang H, Ni D J. Molecular cloning, characterization and expression of heat shock protein 90 gene in the haemocytes of bay scallop Argopecten irradians. Fish Shellfish Immunol 2008; 24:379-385.
    Geist J, Werner I, Eder KJ, Leutenegger CM. Comparisons of tissue-specific transcription of stress response genes with whole animal endpoints of adverse effect in striped bass (Morone saxatilis) following treatment with copper and esfenvalerate. Aquatic Toxicology 2007; 85:28-39.
    Gupta RS.1995. Phylogenetic analysis of the 90 kD heat shock family of protein sequence and an examination of the relationship among animals, plants and fungi species. Mol Biol Evol 1995;12:1063-73.
    Gupta RS. Phylogenetic analysis of the 90 kD heat shock family of protein sequences and an examination of the relationship among animals, plants, and fungi species. Mol Biol Evol 1995;12,1063-1073.
    Gustavsson N., Kokke B.P., Anzelius B., Boelens W.C. Sundby C. Substitution of conserved methionines by leucines in chloroplast small heat shock protein results in loss of redox response but retained chaperone-like activity. Protein Sci.2001;10:1785-1793.
    Hamilton SJ. Review of residue-based selenium toxicity thresholds for freshwater fish. Ecotoxicol Environ Saf 2003; 56: 201-10.
    Hooper C, Day R, Slocombe R, Handlinger J, Benkendorff K. Stress and immune responses in abalone:limitations in current knowledge and investigative methods based on other models. Fish Shellfish Immunol.2007; 22:363-79.
    Huang HC, LeeWC, Lin JH, Huang HW, Jian SC, Mao SJT, Yang PC, Huang TY,Liu YC. Molecular cloning and characterization of porcine cDNA encoding a 90-kDa heat shock protein and its expression following hyperthermia. Gene 1999; 226:307-15.
    Jiang SG, Qiu LH, Zhou FL, Huang JH, Guo YH, Yang K. Molecular cloning and expression analysis of a heat shock protein (Hsp90) gene from black tiger shrimp (Penaeus monodon). Mol Biol Rep 2009; 36:127-34.
    Kalmar B, Greensmith L. Induction of heat shock proteins for protection against oxidative stress. Adv Drug Delivery Rev 2009; 61:310-318.
    Lees-Miller, S. P., Anderson, C. W.1989. The human double-stranded DNA-activated protein kinase phosphorylates the 90-kDa heat-shock protein, hsp90 alpha at two NH2-terminal threonine residues. J Biol Chem 1989; 264(29): 17275-80
    Li FH, Luan W, Zhang CS, Zhang JQ, Wang B, Xie YS, Li SH, Xiang JH. Cloning of cytoplasmic heat shock protein 90 (FcHSP90) from Fenneropenaeus chinensis and its expression response to heat shock and hypoxia. Cell Stress Chaperon 2009a; 14:161-72.
    Li YY, Zhang T, Schwartz SJ, Sun DX.New developments in Hsp90 inhibitors as anti-cancer therapeutics Mechanisms, clinical perspective and more potential. Drug Resistance Updates 2009b; 12:17-27.
    Limon-Pacheco J., Gonsebatt ME.2009. The role of antioxidants and antioxidant-related enzymes in protective responses to environmentally induced oxidative stress. Mutation Research.674:137-147.
    Livak KJ, Schmittgen TD.2001.Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods.25:402-408.
    Mai KS, Ai QH., Xu W, Liufu ZG. Stress in aquaculture and its prevention with emphasis on nutritional methods. J Ocean Univ China.2004; 34(5):767-774. (in Chinese with English abstract)
    Mai KS, Mercer JP, Donlon J.1995. Comparative studies on the nutrition of two species of abalone, Haliotis tuberculata L. and Haliotis discus hannai Ino. Ⅲ. Response of abalone to various levels of dietary lipids. Aquaculture.134: 65-80.
    Malago JJ, Koninkx JF, van Dijk JE. The heat shock response and cytoprotection of the intestinal epithelium. Cell Stress Chaperones.2002; 7(2):191-9.
    Martello LB, Friedman CS, Tjeerdema RS.2000. Combined effects of pentachlorophenol and salinity stress on phagocytic and chemotactic function in two species of abalone. Aquatic toxicology.49:213-225.
    McLaughlin SH, Smith HW, Jackson SE. Stimulation of the weak ATPase activity of human hsp90 by a client protein. J Mol Biol.2002;315(4):787-98.
    Meyer P, Prodromou C, Hu B, Vaughan C, Roe SM, Panaretou B, Piper PW, Pearl LH. Structural and functional analysis of the middle segment of hsp90:implications for ATP hydrolysis and client protein and cochaperone interactions. Mol Cell.2003;11(3):647-58.
    Morimoto R I, Tissieres A, Cesrgopoulos C.1993.Cell in stress:transcriptional activation of heat shock genes. Science 1993;259:1409-10.
    Morimoto RI. Cell stress:transcriptional activation of heat shock genes. Science 1993; 259:1409-1410.
    Moseley P. Stress proteins and the immune response. Immunopharmacology 2000; 48:299-302.
    Nardai G., Sass B.,Eber J., Orosz G. Csermely P. Reactive cysteines of the 90 kDa heat shock protein, Hsp90. Arch Biochem Biophys.2000; 384:59-67.
    Neckers L, Ivy SP. Heat shock protein 90. Curr Opin Oncol 2003; 15:419-24.
    Ozment CP, Turi JL. Iron overload following red blood cell transfusion and its impact on disease severity. Biochim Biophy Acta 2009; 1790:694-701.
    Palmisano AN,Winton JR, Dickhoff WW. Tissue-specific Induction of Hsp90 mRNA and plasma cortisol response in chinook salmon following heat shock, seawater challenge, and handling challenge. Mar Biotechnol 2000; 2: 329-338.
    Panaretou B, Siligardi G, Meyer P, Maloney A, Sullivan JK, Singh S, Millson SH, Clarke PA, Naaby-Hansen S, Stein R, Cramer R, Mollapour M, Workman P, Piper PW, Pearl LH, Prodromou C. Activation of the ATPase activity of hsp90 by the stress-regulated cochaperone ahal.Mol Cell 2002; 10(6):1307-18.
    Papanikolaou G, Pantopoulos K. Iron metabolism and toxicity. Toxicol Appl Pharmacol 2005; 202:199-211.
    Papp LV, Lu J, Holmgren A, Khanna KK. From Selenium to Selenoproteins:Synthesis, Identity,, and Their Role in Human Health. Antioxid Redox Sign.2007; 9(7):775-806.
    Pearl LH, Prodromou C. Structure and mechanism of the Hsp90 molecular chaperone machinery. Annu Rev Biochem. 2006;75:271-94.
    Picard D. Heat-shock protein 90, a chaperone for folding and regulation. Cell Mol Life Sci 2002; 59:1640-1648.
    Pritchard K A, Ackerman A W, Gross E R, Stepp D W, Shi Y, Fontana J T, Baker J E, Sessa W C.2001. Heat Shock Protein 90 Mediates the Balance of Nitric Oxide and Superoxide Anion from Endothelial Nitric-oxide Synthase. J Biol Chem.276:17621-17624.
    Pritchard KA Jr, Ackerman AW, Gross ER, Stepp DW, Shi Y, Fontana JT, Baker JE, Sessa WC. Heat Shock Protein 90 Mediates the Balance of Nitric Oxide and Superoxide Anion from Endothelial Nitric-oxide Synthase. J Biol Chem. 2001;276(21):17621-17624.
    Quraishi H, Brown IR. Expression of heat shock protein 90 (hsp90) in neural and nonneural tissues of the control and hyperthermic rabbit.Exp Cell Res 1995; 219:358-63.
    Ramaglia V, Harapa GM, White N, Buck LT. Bacterial infection and tissue-specific Hsp72,-73 and-90 expression in western painted turtles. Comp Biochem Physiol C 2004; 138:139-48.
    Ramaglia, V., Harapa, G. M.,White, N.,Buck, L. T. Bacterial infection and tissue-specific Hsp72,-73 and-90 expression in western painted turtles. Comp Biochem Physiol C 2004; 138(2):139-48.
    Rayman M P.2000. The importance of selenium to human health. Lancet.356,233-241.
    Redaelli CA, Tian YH, Schaffner T, Ledermann M, Baer HU, Dufour JF. Extended preservation of rat liver graft by induction of heme oxygenase-1.Hepatology.2002; 35(5):1082-1092.
    Ritchie, K. P., Keller, B. M., Syed, K. M.,Lepock, J.R. Hyperthermia (heat shock)-induced protein denaturation in liver, muscle and lens tissue as determined by differential scanning calorimetry. Int J Hyperthermia 1994; 10(5):605-18.
    Robert, J.,2003. Evolution of heat shock protein and immunity. Dev Comp Immunol.27:449-464.
    Roch P. Defense mechanisms and disease prevention in farmed invertebrates. Aquaculture 1999; 172(1-2):125-145.
    Sappal R, Burka J, Dawson S, Kamunde C. Bioaccumulation and subcellular partitioning of zinc in rainbow trout (Oncorhynchus mykiss):Cross-talk between waterborne and dietary uptake. Aquatic Toxicology 2009; 91:281-90.
    Scheufler C, Brinker A, Bourenkov G, Pegoraro S, Moroder L, Bartunik H, et al. Structure of TPR domain-peptide complexes:critical elements in the assembly of the Hsp70-Hsp90 multichaperone machine. Cell 2000; 101:199-210.
    Shenkin A. The key role of micronutrients. Clin Nutr 2006; 25:1-13.
    Song Y, Cardounel AJ, Zweier JL, Xia Y. Inhibition of superoxide generation from neuronal nitric oxide synthase by heat shock protein 90:implications in NOS regulation. Biochemistry.2002;41(34):10616-22.
    Soti C., Racz A., Csermely P. A nucleotide-dependent molecular switch controls ATP binding at the C-terminal domain of Hsp90:N-terminal nucleotide binding unmasks a C-terminal binding pocket. J Biol Chem.2002; 277: 7066-7075.
    Tamura K, Dudley J, Nei M, Kumar S. MEGA4:Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0. Mol Biol Evol 2007;24(8):1596-9.
    Theodoraki, MA., Mintzas, AC. cDNA cloning, heat shock regulation and developmental expression of the hsp83 gene in the Mediterranean fruit fly Ceratitis capitata. Insect Mol Biol 2006; 15(6):839-852.
    Wan M, Mai KS, Ma HM, Xu W, Liufu ZG.Effect of dietary delenium and Vitamin E on antioxidant enzyme activities in Abalone, Haliotis discus hannai Ino. Acta Hydrobio Sinica 2004; 28 (5):496-503. (in Chinese with English abstract)
    Wu LT, Chu KH.Characterization of heat shock protein 90 in the shrimp Metapenaeus ensis:evidence for its role in the regulation of vitellogenin synthesis. Mol Reprod Dev 2008; 75:952-959.
    Yang Y, Liu B, Dai J, Srivastava P K, Zammit D J, Lefrancois L and Li Z H.2007. Heat Shock Protein gp96 Is a Master Chaperone for Toll-like Receptors and Is Important in the Innate Function of Macrophages. Immunity.26(2): 215-226.
    Yavelsky V., Vais O.,Piura B., Wolfson M., Rabinovich A., Fraifeld V. The role of Hsp90 in cell response to hyperthermia. J Thermal Biol 2004; 29:509-514.
    Yu BP. Cellular defenses against damage from reactive oxygen species. Physiol Rev 1994; 74:139-62.
    Yu ZJ, Yang XD, Wang K. Metal ions induced heat shock protein response by elevating superoxide anion level in HeLa cells transformed by HSE-SEAP reporter gene. Toxicology 2006; 223:1-8.
    Zhang WB, Mai KS, Xu W, Ma HM. Metabolic responses to dietary cholecalciferol and phosphorus in abalone Haliotis discus hannai ino. J Exp Zool 2003; 299A:110-117.
    Zhang XY, Zhang MZ, Zheng CJ, Liu J, Hu HJ. Identification of two hsp90 genes from the marine crab, Portunus trituberculatus and their specific expression profiles under different environmental conditions. Comp Biochem Physiol C 2009; 150:465-73.
    Zou J, Guo Y, Guettouche T, Smith DF, Voellmy R. Repression of heat shock transcription factor HSF1 by HSP90 (HSP90 complex) that forms a stress-sensitive complex with HSF1.Cell 1998; 94:471-480.
    任宝波,王玉艳,王纯净,李士泽,杨焕民.HSP70家族的分类及基因结构与功能.动物医学进展2005；26(1)：98-101.
    吴顺华,成军,郑玉建.热休克蛋白家族与肝癌的关系.世界华人消化杂志.2005；13(14)：1725-1730.
    Aziz N, Munro HN. Iron regulates ferritin mRNA translation through a segment of its 5'untranslation region. Proc Natl Acad Sci USA 1987;84:8478-8482.
    Beck G, Ellis TW, Habicht GS, Schluter SF, Marchalonis JJ. Evolution of the acute phase response:iron release by echinoderm (Asterias forbsei) coelomocytes, and cloning of an echinoderm ferritin molecule. Dev Comp Immunol 2002;26:11-26.
    Beinert H, Kennedy MC, Stout CD. Aconitase as iron-sulfur protein, enzyme, and iron-regulatory protein. Chem Rev 1996;96:2335-2373.
    Bevilacqua MA, Faniello MC, Russo T, Cimino F, Costanzo F. Transcriptional regulation of the human H-ferritin encoding gene (FERH) in G418-treated cells:role of the B-box-binding factor. Gene 1994; 141:287-291.
    Coluccidamato LG, Ursini MV, Colletta G, Cirafici A, Defranciscis V. Thyrotropin stimulates transacription from the ferritin heavy chain promoter. Biochem Biophys Res Commun.1989;165:506-511.
    De Zoysa M, Lee J. Two ferritin subunits from disk abalone(Haliotis discus discus) Cloning, characterization and expression analysis. Fish Shellfish Immunol 2007; 23(3):624-635.
    Durand JP, Goudard F, Barbot C, Pieri J, Fowler SW, Cotret O. Ferritin and hemocyanin:210Po molecular traps in marine fish oyster lobster. Mar Ecol Prog Ser 2002; 233:199-205.
    Durand JP, Goudard F, Pieri J, Escoubas JM, Schreiber N, Cadoret JP. Crassostrea gigas ferritin:cDNA sequence analysis for two heavy chain type subunits and protein purification. Gene 2004; 338:187-195.
    Emerit J, Beaumont C, Trivin F. Iron metabolism, free radicals, and oxidative injury. Biomed Pharmacother 2001;55: 333-339.
    Epsztejn S, Glickstein H, Picard V, Slotki IN, Breuer W, Beaumont C, Cabantchik ZI. H-ferritin subunit overexpression in erythroid cells reduces the oxidative stress response and induces multidrug resistance properties. Blood 1999; 94: 3593-3603.
    Geiser DL, Mayo JJ, Winzerling JJ. The unique regulation of Aedes aegypti larval cell ferritin by iron. Insect Biochem Mol Biol 2007; 37:418-429.
    Georgieva T, Dunkov BC, Dimov S, Ralchev K, Law JH. Drosophila melanogaster ferritin:cDNA encoding a light chain homologue, temporal and tissue specific expression of both subunit types. Insect Biochem Mol Biol 2002; 32: 295-302.
    Ghiselli A, Serafini M, Natella F, Scaccini C. Total antioxidant capacity as a tool to assess redox status:critical review and experimental data. Free Radic Biol Med 2000; 29:1106=1114.
    Ghosh S, Hevi S, Chuck SL. Regulated secretion of glycosylated human ferritin from hepatocytes. Blood 2004; 103: 2369-2376.
    Harrison PM, Arosio P.1996. The ferritins:molecular properties, iron storage function and cellular regulation. Biochem Biophys Acta 1275:161-203.
    Hentze MW, Muckenthaler MU, Andrews NC. Balancing acts:molecular control of mammalian iron metabolism. Cell 2004;117:285-297.
    Le NT, Richardson DR. The role of iron in cell cycle progression and the proliferation of neoplastic cells. Biochim Biophys Acta 2002; 1603:31-46.
    Li M, Saren G, Zhang S.Identification and expression of a ferritin homolog in amphioxus Branchiostoma belcheri Evidence for its dual role in immune response and iron metabolism. Comp Biochem Physiol B 2008; 150(3): 263-270.
    Liu J,Zhang WB,Mai KS,Fu JH,Fen XN,Ma HM,Xu W,Liufu ZG. Construction of mantle cDNA subtractive library of Haliotis discus hannai lno fed with vitamin E-deficiency diet. J Fishery Sciences China 2007; 14(3): 383-389. (in Chinese with English abstract)
    Liu X, Theil EC. Ferritins:dynamic management of biological iron and oxygen chemistry. Acc Chem Res 2005; 38: 167-175.
    Livak KJ, Schmittgen TD.2001.Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods 25:402-408.
    Mai KS, Tan BP. Iron methionine (FeMet) and iron sulfate (FeSO4) as sources of dietary iron for juvenile abalone, Haliotis discus hannai Ino. J Shellfish Research 2000;19(3):861-868.
    Ong DS, Wang L, Zhu Y, Ho B, Ding JL. The response of ferritin to LPS and acute phase of Pseudomonas infection. J Endotoxin Res.2005;11:267-280.
    Ong ST, Ho JZS, Ho B, Ding JL. Iron-withholding strategy in innate immunity. Immunobiology 2006; 211:295-314.
    Orino K, Eguchi K, Nakayama T, Yamamoto S, Watanabe K. Sequencing of cDNA clones that encode bovine ferritin H and L chains. Comp Biochem Physiol B 1997; 118(3):667-673.
    Orino K, Lehman L, Tsuji Y, Ayaki H, Torti SV, Torti FM. Ferritin and the response to oxidative stress. Biochem J 2001; 357:241-247.
    Pham DQD, Winzerling JJ, Dodson MS, Law JH, Transcriptional control is relevant in the modulation of mosquito ferritin synthesis by iron. Eur J Biochem.1999; 266:236-240.
    Rocha ER, Smith CJ. Transcriptional regulation of the Bacteroides fragilis ferritin gene (ftnA) by redox stress. Microbiology 2004; 150 (7):2125-34.
    Rouault TA, Tong WH.Iron-sulphur cluster biogenesis and mitochondrial iron homeostasis. Nat Rev Mol Cell Biol 2005; 6:345-351.
    Scaccabarozzi A, Arosio P, Weiss G, et al. Relationship between TNF-a and iron metabolism in differentiating human monocytic THP-1 cells. Br J Haematol 2000;110:978-84.
    Tamura K, Dudley J, Nei M, Kumar S. MEGA4:Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0. Mol Biol Evol 2007;24(8):1596-1599.
    Theil EC. Ferritin:structure, gene regulation and cellular function in animals, plants and microorganisms. Annu Rev Biochem 1987; 56:289-315.
    Torti FM, Torti SV. Regulation of ferritin genes and proteins. Blood 2002; 99:3505-3516.
    Torti SV, Kwak EL, Miller SC, Miller LL, Ringold GM, Myambo KB, Young AP, Torti FM. The molecular cloning and characterization of murine ferritin heavy chain, a tumor necrosis factor inducible gene. J Biol Chem 1988; 263: 12638-44.
    Tran TN, Eubanks SK, Schaffer KJ, Zhou CY, Linder MC. Secretion of ferritin by rat hepatoma cells and its regulation by inflammatory cytokines and iron. Blood 1997; 90:4979-4986.
    Wang KJ, Ren HL, Xu DD, Cai L, Yang M. Identification of the up-regulated expression genes in hemocytes of variously colored abalone (Haliotis diversicolor Reeve,1846) challenged with bacteria. Dev Comp Immunol 2008; 32:1326-47.
    Zhang D, Dimopoulos G, Wolf A, Minana B, Kafatos FC, Winzerling JJ. Cloning and molecular characterization of two mosquito iron regulatory proteins. Insect Biochem Mol Biol 2002; 32:579-589.
    Zhang J, Li F, Wang Z, Zhang X, Zhou Q, Xiang J. Cloning, expression and identification of ferritin from Chinese shrimp, Fenneropenaeus chinensi. J Biotechnol 2006; 125:173-184.
    Zhang WB, Mai KS, Xu W, Ma HM. Metabolic responses to dietary cholecalciferol and phosphorus in abalone Haliotis discus hannai ino. J Exp Zool 2003b; 299A:110-117.
    Zhang Y, Meng Q, Jiang T, Wang H, Xie L, Zhang R. A novel ferritin subunit involved in shell formation from the pearl oyster (Pinctada fucata). Comp Biochem Physiol B 2003a; 135:43-54.
    Ames BN, Shigenaga M,Hagen TM.Oxidants, antioxidants, and the degenerative diseases of aging. Proc Natl Acad Sci (USA)1993;90:7915-7922.
    Anbarasu K, Chandran MR. Effects of ascorbic acid on the immune response of the catfish, Mystus gulio (Hamilton), to different bacterins of Ameromonas hydrophila. Fish Shellfish Immunol 2001; 11:347-355.
    Arrigo AP, Virot S, Chaufour S, Firdaus W, Kretz-Remy C,Diaz-Latoud C.HSP26 consolidates intracellular redox homeostasis by upholding glutathione in its reduced form and by decreasing iron intracellular levels. Antioxid Redox Signal 2005;7(3-4):414-22.
    Arrigo AP. Small stress proteins:Chaperones that act as regulators of intracellular redox state and programmed cell death. Biol Chem 1998; 379:19-26.
    Avanzo JL, de Mendonca CX Jr, Pugine SM, de Cerqueira Cesar M. Effect of vitamin E and seleniumon resistance to oxidative stress in chicken superficial pectoralis muscle. Comp Biochem Physiol C Toxicol Pharmacol.2001;129(2):163-173.
    Bai J, Cederbaum AI. Overexpression of catalase in the mitochondrial or cytosolic compartment increases sensitivity of HepG2 cells to tumor necrosis factor-alpha-induced apoptosis. J Biol Chem 2000;275(25):19241-192499.
    Bendich A.,Machlin LJ, Scandurra O, Burton GW, Wayner DDM. The antioxidant role of vitamin C. Adv. Free Rad Biol Med 1986;2:4109-4444.
    Berger TM,Polidor MC, Dabbagh A, Evans PJ, Halliwell B, Morrow JD, Roberts LJ, Jr Frei B. Antioxidant activity of vitamin C in iron-overloaded human plasma. J.Biol. Chem.1997.272: 15656-15660.
    Boop PC. Importance of antioxidant vitamins in immunity and health in animals. Animal Feed Science Technology 1996;59:103-114.
    Brerro-Saby C, Delliaux S, Steinberg JG,Boussuges A, Gole Y, Jammes Y. Combination of 2 oxidant stressors suppresses the oxidative stress and enhances the heat shock protein 27 response in healthy humans. Metabolism Clinical and Experimental 2010, article in press.
    Carr A, Frei B.Does vitamin C act as a pro-oxidant under physiological conditions? FASEB J 1999; 13(9):1007-24.
    Chen H, Mai KS, Zhang WB,Liufu ZG, Xu W, Tan BP.Effects of dietary pyridoxine on immune responses in abalone, Haliotis discus hannai Ino. Fish Shellfish Immunol 2005; 19:241-252.
    Choi YK, Jo PG, Choi CY. Cadmium affects the expression of heat shock protein 90 and metallothionein mRNA in the Pacific oyster, Crassostrea gigas.Comp Biochem Physiol C 2008; 147:286-92.
    Chung SS,Kim M, Youn BS, Lee NS,Park JW, Lee IK, Lee YS, Kim JB, Cho YM,Lee HK,Park KS. Glutathione peroxidase 3 mediates the antioxidant effect of peroxisome proliferator-activated receptor gamma in human skeletal muscle cells. Mol.Cell.Biol.2009; 29(1),20-30.
    Durve VS,Lovell RT.Vitamin C and disease resistance in channel catfish(Ictalurus punctatus). Can. J. Fish. Aquat. Sci.1982.39,948-951.
    Franzellitti S., Fabbri E., Differential HSP70 gene expression in the Mediterraneanmussel exposed to various stressors. Biochem. Biophys. Res. Commun.2005;336,1157-1163.
    Fu J, Zhang W, Mai K, Feng X, Xu W, Liufu Z, Ma H, Ai Q. Effects of vitamin E on antioxidant enzyme activities and fatty acid compositions in juvenile abalone Haliotis discus hannai Ino. J Shellfish Res 2007;26(3):809-14.
    Fu J, Zhang W, Mai K, Feng X, Xu W, Liufu Z, Tan B.Effects of dietary vitamin A on antioxidant responses of abalone Haliotis discus hannai Ino. Acta Oceanol Sin 2006a; 25:141-50.
    Gao Q, Song L, Ni D, Wu L, Zhang H, Chang Y.cDNA cloning and mRNA expression of heat shock protein 90 gene in the haemocytes of Zhikong scallop Chlamys farreri. Comp Biochem Physiol B 2007;147:704-715.
    Gershoff SN. Vitamin C (ascorbic acid):new roles, new requirements. Nutrition Review 1993;51: 313-326.
    Goralska M, Harned J, Grimes AM, Fleisher LN, McGahan MC. Mechanisms by which ascorbic acid increases ferritin levels in cultured lens epithelial cells.Exp Eye Res.1997; 64(3):413-21.
    Halver JE. Vitamin requirement study techniques. J Appl Ichthyol 1995.11:215-224.
    Hardie LJ, Marsden MJ, Fletcher TC, Secombes CJ.In vitro addition of vitamin C affects rainbow trout lymphocyte responses. Fish Shellfish Immunol 1993;3:207-219.
    Hooper C, Day R, Slocombe R, Handlinger J, Benkendorff K.2007.Stress and immune responses in abalone:limitations in current knowledge and investigative methods based on other models. Fish Shellfish Immunol 2007;22:363-739.
    Jo PG, Choi YK, Choi CY. Cloning and mRNA expression of antioxidant enzymes in the Pacific oyster, Crassostrea gigas in response to cadmium exposure. Comp Biochem Physiol. C 2008;147: 460-469.
    Kenet G, Freedman J, Shenkman B, Regina E, Brok-Simoni F, Holzman F, Vavva F, Brand N, Michelson A, Trolliet M, Loscalzo J, Inbal A. Plasma glutathione peroxidase deficiency and platelet insensitivity to nitric oxide in children with familial stroke. Arterioscler Thromb Vasc Biol 1999;19:2017-2023.
    Li N,Karin M.Is NF-B the sensor of oxidative stress? FASEB J.1999;13(10):1137-43.
    Lim C, Lovell RT. Pathology of the vitamin C deficiency syndrome in channel catfish(Ictalurus punctatus).J Nutr 1978; 108:1137-1146.
    Lipschitz DA, Bothwell TH, Seftel HC, Wapnick AA,Charlton RW. The role of ascorbic acid in the metabolism of storage iron.Br J Haematol.1971;20(2):155-63.
    Livingstone DR, Lips F,Martinez PG, Pipe RK.1992. Antioxidant enzymes in the digestive gland of the common mussel Mytilus edulis. Mar Biol 1992; 112:265-276.
    Lopez N,Cuzon G,Gaxiola G, Taboada G, Valenzuela M,Pascual C, Sanchez A,Rosas C. Physiological, nutritional, and immunological role of dietary} 1-3 glucan and ascorbic acid 2-monophosphate in Litopenaeus vannamei juveniles. Aguaculture 2003;224:223-243.
    Mahmoud KZ, Edens FW, Eisen EJ, Havenstein GB. Ascorbic acid decreases heat shock protein 70 and plasma corticosterone response in broilers (Gallus gallus domesticus) subjected to cyclic heat stress. Comp Biochem Physiol B 2004.137(1):35-42.
    Mahmoud KZ, Edens FW, Eisen EJ, Havenstein GB.Effect of ascorbic acid and acute heat exposure on heat shock protein 70 expression by young white Leghorn chickens. Comp Biochem Physiol C 2003;136(4):329-35.
    Mai KS, Ai QH., Xu W, Liufu ZG. Stress in aquaculture and its prevention with emphasis on nutritional methods. J Ocean Univ China.2004; 34(5):767-774.(in Chinese with English abstract)
    Mai KS, Comparative studies on the nutrition of two species of abalone, Haliotis tuberculata L.and Haliotis discus hannai Ino. Ⅶ.Effects of dietary vitamin C on survival, growth and tissue concentration of ascorbic acid. Aquaculture 1998;161:383-392.
    Mai KS, Mercer JP, Donlon J. Comparative studies on the nutrition of two species of abalone, Haliotis tuberculata L. and Haliotis discus hannai Ino.Ⅲ. Response of abalone to various levels of dietary lipids. Aquaculture 1995;134:65-80.
    Mehlen P, Hickey E, Weber LA, Arrigo AP. Large unphosphorylated aggregates as the active form of HSP26 which controls intracellular reactive oxygen species and glutathione levels and generates a protection against TNF alpha in NIH-3T3-ras cells. Biochem Biophys Res Commun 1997; 241: 187-192.
    Mehlen P, Kretz-Remy C, Briolay J, Fostan P, Mirault ME, Arrigo AP. Intracellular reactive oxygen species as apparent modulators of heat-shock protein 27 (HSP26) structural organization and phosphorylation in basal and tumour necrosis factor alpha-treated T47D human carcinoma cells. Biochem J.1995;312 (2):367-375.
    Mehlen P, Kretz-Remy C, Preville X, Arrigo AP. Human HSP26, Drosophila HSP26 and human alpha B-crystallin expression-mediated increase in glutathione is essential for the protective activity of these proteins against TNF alpha-induced cell death. EMBO J 1996; 15:2695-2706.
    Montero D, Marrero M, Izquierdo MS, Robaina L, Vergara JM, Tort L. Effect of vitamin E and C dietary supplementation on some immune parameters of gilthead seabream (Sparus aurata) juveniles subjected to crowding stress. Aquaculture 1999.171:269-278.
    Mourente G, Diaz-Salvago E, Bell J G, et al.Increased activities of hepatic antioxidant defence enzymes in juvenile gilthead sea bream(Sparus aurata L.) fed dietary oxidized oil:attenuation by dietary vitamin E. Aquaculture 2002; 214:343-361.
    Navarre O, Halver J. Disease resistance and humoral antibody production in rainbow trout fed high levels of vitamin C. Aquaculture 1989.79:207-221.
    Orturno J, Cuesta A, Esteban A, Meseguer J. Effect of oral administration of high vitamin C and E dosages on the gilthead seabream (Sparus aurata L.)innate immune system.Vet Immunol Immunopathol 2001;79:167-180.
    Orturno J., Esteban MA.,Meseguer J.Effect of high dietary intake of vitamin C on non-specific immune response of gilthead seabream(Spares aurata L.) Fish Shellfish Immunology.1999.9: 429-443.
    Park JW, Moon C, Yun S, Kim SY, Bae YC, Chun MH, Moon JI.Differential expression of heat shock protein mRNAs under in vivo glutathione depletion in the mouse retina. Neurosci Lett.2007; 413(3):260-4.
    Podmore ID, Griffiths HR, Herbert KE, Mistry N, Mistry P, Lunec J.Vitamin C exhibits pro-oxidant properties. Nature.1998;392(6676):559.
    Preville X, Salvemini F, Giraud S, Chaufour S,Paul C, Stepien G, Ursini MV, Arrigo AP. Mammalian small stress proteins protect against oxidative stress through their ability to increase glucose-6-phosphate dehydrogenase activity and by maintaining optimal cellular detoxifying machinery, Exp. Cell Res.1999; 247:61-78.
    Reddy KV, Kumar TC, Prasad M, et al. Pulmonary lipid peroxidation and antioxidant defenses during exhaustive physical exercise:the role of vitamin E and selenium.Nutrition 1998;14 (5):448-451.
    Ren H, Musch MW, Kojima K, Boone D, Ma A, Chang EB.Short-chain fatty acids induce intestinal epithelial heat shock protein 25 expression in rats and IEC 18 cells. Gastroenterology 2001;121: 631-639.
    Ropeleski MJ, Tang J, Walsh-Reitz MM, Musch MW, Chang EB, Interleukin-11-induced heat shock protein 25 confers intestinal epithelial-specific cytoprotection from oxidant stress. Gastroenterology.2003;124:1358-1368.
    Sakai M. Current research status of fish immunostimulants. Aquaculture 1999.172:63-92.
    Selsby JT, Dodd SL. Heat treatment reduces oxidative stress and protect muscle mass during immobilization. Am J Physiol Regul Integr Comp Physiol 2005;289:R134-9.
    Smith VJ, Brown J H, Hauton C.Immunostimulation in crustaceans:does it really protect against infection. Fish Shellfish Immunol,2003,15(1):71-90.
    van den Berg R, Haenen GR, van den Berg H, Bast A. Transcription factor NF-kappaB as a potential biomarker for oxidative stress. Br J Nutr 2001;86(1):S121-127.
    Verlhac V, Obach R, Gabaudan J, Schuep W, Hole R. Immunomodulation by dietary vitamin C and glucan in rainbow trout (Oncorhynchus mykiss). Fish Shellfish Immunol 1998; 8:409-424.
    waahbφ R, Glette J, Raa-Nilsen E, Sandnes K. Dietary vitamin C, immunity and disease resistance in Atlantic salmon Salmo solar. Fish Physiol Biochem 1993;2:61-73
    Wan M, Mai KS, Ma HM,Xu W, Liufu ZG.Effect of dietary delenium and Vitamin E on antioxidant enzyme activities in Abalone, Haliotis discus hannai Ino. Acta Hydrobio Sinica 2004; 28 (5), 496-503.(in Chinese with English abstract)
    Wang B, Li X, Gou C. Infection of spherical viruses from Haliotis discus hannai Ino. Virol Sin 1997; 12:360-3
    Wang W, Mai K,Zhang W, Ai Q, Yao C, Li H,Liufu Z. Effects of dietary copper on survival,growth and immune response of juvenile abalone, Haliotis discus hannai Ino. Aquaculture 2009; 297: 122-127.
    Whitam M, Fortes MB.Heat shock protein 72:release and biological significance during exercise. Frontiers in Biosci 2008;13:1328-39.
    Wilson RP, Poe WE.,Impaired collagen formation in the scorbutic channel catfish. J. Nutr.1973;103: 1359-1364.
    Wu CL, Mai KS, Zhang WB,Ai QH,Xu W, Wang XJ, Ma HM, Liufu ZG. Molecular cloning, characterization and mRNA expression of selenium-dependent glutathione peroxidase from abalone, Haliotis discus hannai Ino in response to dietary selenium, zinc and iron. Comparative Biochemistry and Physiology Part C:Toxicology and Pharmacology.2010.152:121-132.
    Wu CL, Zhang WB,Mai KS, Liang XF, Xu W, Wang J, Ma HM. Molecular cloning, characterization and mRNA expression of selenium-binding protein in abalone, Haliotis discus hannai Ino: response to dietary selenium, iron and zinc. Fish and shellfish immunology.2010.29:117-125.
    Wyttenbach A, Sauvageot O, Carmichael J, et al. Heat shock protein 27 prevents cellular polyglutamine toxicity and suppresses the increase of reactive oxygen species caused by huntingtin. Hum Mol Genet 2002; 11:1137-51.
    Xue JP, Xu YP, Jin LJ, Liu G, Sun YX, Li SY, Zhang JC. Effects of traditional Chinese medicine on immune responses in abalone, Haliotis discus hannai Ino. Fish Shellfish Immunol 2008; 24: 752-8.
    Ye L, Yu KK, Wang RC. The study on the pathogenic bacteria of the fester disease of cultured juvenile discus abalone. Journal of Fishery Sciences of China.1997; 4:43-8. (in Chinese with English abstract)
    艾春香,陈立侨,高露娇,温小波,江洪波.VC对河蟹血清和组织中超氧化物歧化酶及磷酸酶活性的影响.台湾海峡.2002；21(4)：431-438.
    艾春香,陈立侨,刘晓玲,高露姣,温小波.维生素C对中华绒螯蟹非特异性免疫的影响.水产学报2008；32(2)：249-256.
    李爱杰.维生素c对中国对虾的营养和免疫作用及其最适剂型.粮食与饲料工业.1998.2：33-36.
    刘恒,李光友.免疫多糖对养殖南美白对虾作用的研究.海洋与湖沼1998；29(2)：113-118.
    王嘉.皱纹盘鲍(Haliotis discus hannai Ino) L-古洛糖酸-1,4-内酯氧化酶(GLO)的研究.2009.中国海洋大学博士毕业论文.
    王雷,李光友,毛远兴,等.口服免疫型药物对养殖中国对虾病害防治作用的研究.海洋与湖沼,1994；25(5)：481-486.