重金属诱导贝类金属硫蛋白研究进展

金属硫蛋白(Metallothionein,MT)是一种低分子量的蛋白质,广泛存在于大多数生物体内,在参与必需金属元素调节和非必需金属元素的解毒等方面具有重要作用。介绍了MT的结构与功能,对贝类MT的诱导机制、解毒机理、降解与金属排除进行概述,探讨了MT诱导合成过程中的一些主要影响因素。由于贝类MT能够通过多途径诱导,而且存在众多的环境因素和自身生理因素的影响,MT的变化规律往往出现较大的波动性。因此,今后的研究重点会是贝类MT作为污染监测指示物的研究、重金属诱导MT数学模型的构建以及贝类MT产品的开发等方面。     

金属硫蛋白基因对转基因鲤及实验动物体内重金属累积的影响

金属硫蛋白(metallothiontine,简称MT),又称重金属结合蛋白,是一种低分子量富含半胱氨酸的金属结合蛋白。锌、铜、镉等多种金属元素均可在体外或体内诱导组织MT大量合成。已有研究表明,MT在微量元素代谢、重金属富集、环境污染的清除等方面起重要作用,因此, 选择恰当的启动子是保证外源基因在转基因鱼中成功表达的最重要因素。在转基因鱼的研究过程中,使用的转基因材料从最初阶段的人及其它哺乳类如牛、羊等的生长激素基因发展到现在的鱼的生长激素基因,启动调控顺序也由小鼠的重金属螯合蛋白(MT-1)基因发展到鱼的重金属结合蛋白基因,如虹鳟金属硫蛋白基因、鲤β-肌动蛋白基因、美洲鲽抗冻蛋白基因以及鲤金属硫蛋白     

重金属离子对凡纳滨对虾肝胰脏MT含量的影响

金属硫蛋白（metallothionein，MT）是一类小分子的蛋白质，能螯合金属离子，调节微量元素（主要为Cu、Zn）贮存、运输和代谢及具有对重金属中毒解毒作用和清除氧自由基的功能。目前，关于重金属胁迫对甲壳动物MT含量的影响国外已有一些研究报道；国内这方面尚未见报道。     

泥蚶金属硫蛋白的鉴定、原核重组表达及其组织细胞分布

采用RT-PCR方法扩增、克隆和鉴定了泥蚶(Tegillarca granosa)的金属硫蛋白(Metallothionein,MT)基因(TgMT)的开放阅读框(ORF)。基于金属硫蛋白氨基酸序列构建的系统进化树表明,泥蚶和魁蚶(Scapharca broughtonii)的亲缘关系最近。利用ORF序列,构建原核重组表达质粒。重组质粒经PCR、酶切鉴定及测序验证后转化大肠杆菌(Escherichia coli)BL21(DE3),经IPTG诱导,表达重组蛋白,重组蛋白的分子量约为28.3 kD。可溶性分析表明重组蛋白主要以可溶性形式存在。用His-tag柱亲和纯化重组蛋白,并利用重组蛋白制备了兔抗血清。Real-time PCR和Western blot实验表明,金属硫蛋白在泥蚶的各个组织中都有表达,并且在消化腺中的表达量最高。采用激光共聚焦显微镜,对泥蚶的消化腺进行免疫组化定位分析,发现泥蚶的MT主要存在于消化腺腺管上皮细胞的胞质中。泥蚶MT的克隆和表达研究为进一步研究该蛋白在环境监测中的作用、探讨细胞解毒的分子生物学机制奠定了基础。     

贝类染色体组工程的研究

染色体组工程的研究,主要是指人工雌核生殖,人工雄核生殖,多倍体的诱导等方面的研究,自从Stanlq等(1981)报道了人工诱导美洲牡蛎(Crassostreavirginica)受精卵获得多倍体以来,海洋经济贝类多倍体育种研究取得了很大进展。除了对各种诱导方法进行了探索和完善外,对多倍体贝类的生物学也进行了较多的研究。研究表明,人工诱导的三倍体贝类具有不育、个体大、生长快、肉质好等优良品性。因此,根据各种经济贝类养殖需要,培育三倍体品种,对促进贝类养殖生产的发展,具有重要的经济价值和应用前景。近年来水产动物染色体组的操作,新的染色体操…     

鱼类金属硫蛋白基因的表达与调控

金属硫蛋白(Metallothionein,MT)是一类能选择性地结合重金属离子的低分子量蛋白质。自Marafante及其同事于1970年证明MT在真骨鱼类中的存在以来,人们就对MT在动物体内和体外的作用和功能进行了广泛的研究。Dunn等(1987)对鱼类MT的研究表明,这类蛋白质长度极为均一,由60~61个氨     

海洋贝类四倍体的研究进展与展望

近几年来，海洋贝类三倍体的研究进展迅速，用四倍体与二倍体杂交的方法理论上能产生１００％的三倍体，从而方便、高效、稳定地生产三倍体，克服直接诱导的不足，开辟了三倍体诱导研究中的一个崭新的方向。四倍体也因此逐步成为海洋贝类染色体操作技术中的热点。一、贝类四倍体的研究现状贝类四倍体研究历史较短，自从Stephens和Downing（１９８８）报道了对长牡蛎四倍体的研究，至今仅有十几年，与人工诱导贝类三倍体相比，人工诱导四倍体的技术难度更大。因为诱导贝类三倍体是通过阻止第一或第二极体的释放来增加一个染色体组，而诱导贝…     

微波消解/ICP-MS法同时测定贝类产品中16种金属元素的含量

本文建立了微波消解-电感耦合等离子体质谱(ICP-MS)同时测定贝类产品中Cr、Mn、Fe、Co、Ni、Cu、Zn、As、Se、Mo、Ag、Cd、Ba、Hg、Tl和Pb等16种金属元素含量的方法.采用微波消解法对样品进行消解,通过在线加入内标校正基体效应,利用ICP-MS测定贝类产品中16种金属元素的含量.测定元素校正曲线的相关系数均在0.9992以上,检出限为6.64× 10-4 ～0.0851 μg· L-1.对牡蛎样品平行测定6次,各元素相对标准偏差(RSD)均小于5％.利用该方法分析了标准参考物质扇贝(GBW10024)中的16种金属元素,其测定值与参考值吻合.该方法具有快速、简单、精密度和准确性良好等优点,适用于贝类产品多种金属元素的同时测定.     

海洋生物体多环芳烃污染残留及其健康风险评价研究

多环芳烃（PAHs）主要来源于煤、石油等矿物燃料及其他有机物的不完全燃烧和裂解过程，是一类广泛存在于环境中具有潜在致畸、致癌和致突变效应的持久性有机污染物。PAHs引起的污染问题越来越受到人们的关注，已经被世界许多国家列为有机污染物的研究重点。海洋环境中的PAHs可以通过生物积累和食物链传递作用进入鱼类、贝类等海洋生物体中，不仅影响海洋生物的生存繁衍，破坏海洋生态环境，还会通过水产品影响人类健康。文章重点对近年来国内外鱼类、贝类等海洋生物体PAHs残留分析方法、残留水平的影响因素、来源解析以及人体暴露健康风险评价等方面进行了综述，同时指出了目前研究还存在的一些问题，为今后的研究提供了疗向。     

双壳贝类呼吸代谢的研究进展

呼吸代谢是生物能量学研究的重要内容之一。依据近年来国内外的研究成果,综述了关于贝类呼吸代谢的研究方法,讨论了影响贝类耗氧率的生物因素和非生物因素,展望了贝类呼吸代谢的研究和应用前景。     

Cadmium,lead and metallothionein contents in cultivated mussels (Mytilus galloprovincialis) from the Gulf of Naples (Southern Italy)

The concentration of cadmium and lead ions was determined in digestive gland and mantle of mussels (Mytilus galloprovincialis) from five aquaculture stations located in the Gulf of Naples (Southern Italy, Tyrrhenian Sea). Metallothionein (MT) levels in the same tissues were also evaluated. This gulf represents a greatly urbanized area, characterized by an important commercial port and past industrial activities, terminated in the last decade of the 20th century. The results were compared with those obtained from mussels cultivated in a more pristine neighbouring area. Data demonstrate that the amount of both metals found in mussels harvested at the highly anthropogenic sites in the Gulf of Naples was comparable with that found in mussels from the more naturalistic site. The content of both cadmium and lead was not related with that of MT. Indeed, the amount of both metals in the digestive gland was greater than in the mantle, whereas the MT preferentially accumulated in the mantle. Possibly, the MT content in mantle was associated to the physiological function of the tissue, rather than heavy metals exposure.     

Metallothionein and metal levels in liver,gills and kidney of <Emphasis Type="Italic">Sparus aurata</Emphasis> exposed to sublethal doses of cadmium and copper

The levels of metallothionein (MT), a biomarker of metal exposure, and of Cd and Cu, known as MT inducers, were investigated in Sparus aurata intraperitoneally injected with 500 μg/kg of Cu and Cd for 2 days. MT and metal concentrations (Cd and Cu) were determined in liver, gills and kidney. MT levels were significantly increased in all investigated tissues, with the highest value in liver of Cu as Cd-treated fishes (3.56-fold and 3.3- fold, respectively). Metal concentrations were statistically different between all tissues. Highest metal concentrations were in the liver. The higher metal concentrations and MT induction levels support the main role of MT in metal homeostasis and detoxification.     

Effect of cadmium binding form on rabbit aminolevulinate dehydratase in high molecular weight fraction of rainbow trout liver

Enzyme inhibition owing to cadmium (Cd) accumulation in the soluble high molecular weight fraction (HMF) of the liver from rainbow trout was investigated by measuring the inhibition of aminolevulinate dehydratase (ALAD). Factors affecting ALAD, which reacts specifically to metal, were examined before measuring ALAD activity. Although enzyme inhibition by Cd in the HMF was confirmed in the excretion 2 (E2) group fed Chinese parsley (CP), it was weaker than that in the excretion 1 (E1, control) group, probably due to the chelation of Cd by some substances contained in CP. Based on the absorption spectrum, the proportion of Cd bound to cysteine thiol radicals in the HMF of the E2 group was greater than that of E1. The HMF might detoxify harmful metals to some extent because it contains relatively higher levels of cysteine residues than metallothionein (MT) and shows comparatively weak enzyme inhibition. The results of the present study demonstrated that, in addition to MT, HMF is involved in the detoxification of harmful heavy metals in fish.     

Cadmium detoxification effect of Chinese parsley Coriandrum sativum in liver and kidney of rainbow trout Oncorhynchus mykiss

Although fish feed supplemented with Chinese parsley (CP), Coriandrum sativum, does not affect the health of rainbow trout Oncorhynchus mykiss, it has the remarkable effect of reducing the uptake of Cd (12–17% compared with controls without CP). Here, we measured the change in the amount of metallothionein (MT) in the liver and kidney using gel permeation chromatography-high-performance liquid chromatography (TSK GEL SW 3000 column). The increase in MT content over time in the Cd-fed groups corresponded to the change in Cd accumulation. Gel permeation was performed with soluble fractions from the fish liver and kidney, and changes in the Cd, Cu, and Zn concentrations over time for each fraction, and the relationship between the form of Cd that accumulated in tissues and its toxicity were measured. A nontoxic MT fraction of 7,000 Da and a high-molecular-weight fraction of approximately 60,000 Da were isolated from the soluble fractions. The induction of nontoxic MT–Cd was higher in the CP-supplemented groups than in the control groups. The synergetic actions of several compounds in CP may contribute to bind MT, thereby preventing accumulation of harmful heavy metals and essential metals in fish.     

A Novel Metallothionein Gene from Mussel,Hyriopsis cumingii: Identification and Expression under Lanthanum Exposure

Metallothionein (MT) is a superfamily of cysteine‐rich, metal‐binding, low‐molecular‐weight proteins that participate in the storage, transportation, and metabolism of the trace metal elements. In this study, a novel MT gene from Hyriopsis cumingii was cloned (designated HcMT1); it shared 59.2% identity with previously published MT sequence of H. cumingii and 56.1–97.3% identity with other MT genes from mollusks. The expression level of HcMT1 in various tissues was determined by real‐time polymerase chain reaction. Relatively higher expression levels were observed in digestive gonad, mantle, and gill, and the expression level in hemocytes was significantly upregulated by lanthanum (La³⁺) exposure. The nucleotide sequence of HcMT1 amplified from genomic DNA is identical to the cDNA template group under La³⁺ exposure, which indicated that HcMT1 is an intronless gene. These results indicate that HcMT1 may play an important role in La³⁺ metabolism.     

Lack of relationship between virulence of Aeromonas salmonicida and the putative virulence factors: A-layer, extracellular proteases and extracellular haemolysins

Abstract The role of A-layer (A), protease (P) and haemolysin (H) as virulence factors of Aeromonas salmonicida, the aetiological agent of fish furunculosis, was a investigated using three strains of the bacterium. Strain MT004 lacked the A-Iayer (A⁻) and produced extracellular caseinase and gelatinase (P⁺) and haemolysin (T-lysin; H⁺). Strain MT028 was A⁻, P⁻ and H⁻, and strain MT048 was A⁺, P⁺ and H⁻. The pathology and LD50 produced in rainbow trout by cells or extracellular products (ECP) of each strain were determined. The ECP was produced by two different methods where the protease and haemolytic activities differed in relative levels, or when the protease of MT004 ECP was inhibited by the serine protease inhibitor PMSF. The results indicate that the presence of A-layer is not essential, at least for a moderate degree of virulence; that in vitro production of extracellular proteases is not a requisite of virulent strains; that presence of protease and haemolysin in ECP can be correlated with the development of certain lesions and a rapid time to death but cannot be correlated with the lethal toxicity of the ECP. The authors conclude that an as-yet unrecognized component of ECP is responsible for killing fish.