Establishment of the evaluation system for endocrine-disrupting chemimals using stickleback

刺鱼内分泌干扰物评价体系的建立

• 批准号: 18510060
• 负责人: NAGAE Masaki
• 金额: $ 2.65万
• 依托单位: Nagasaki University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2006
• 资助国家: 日本
• 起止时间: 2006 至 2007
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18510060/
• 关键词: Biomarker; Stickleback; Endocrine-disrupting chemical

Male stickleback produce a glue protein named "spiggin" that is used as a cementing substance for the building of its nest. The synthesis of spiggin is strongly controlled by androgen. Therefore, spiggin is a useful biomarker to evaluate androgenic activity of environmental chemicals, in addition to vitellogenin, precursor of yolk protein produced from the liver by the estrogenic stimulation. In this study, we established the highly sensitive quantification system for both spiggin and vitellogenin to evaluate sex-hormonal effect of chemicals.Highly sensitive quantification system for spiggin mRNA was established using real-time RT-PCR technique. This system enables precise and high sensitive quantification. Vitellogenin, biomarker for estrogenic stimulation, was also purified by several chromatographic techniques. Specific antibody against vitellogenin was also raised. Furthermore, vitellogenin quantification system was established using the method of chemiluminescent-linked immunosorb … More ent assay. This system also enables high sensitive quantification same with spiggin assay system. In addition to these preparations for quantification systems for the biomarkers, the detail condition for in vivo exposure test using stickleback was determined. Using MT as a control androgen, we performed exposure test. As a result, only 3 days exposure at a MT concentration 0.1 and 1 μg/L was efficient for spiggin mRNA expression in kidney. Next, MT exposure was performed under three different water temperature (5, 10 and 15℃) to know the effect of water temperature on spiggin mRNA expression. Exposure at 15℃ induced maximal MT effect on spiggin mRNA expression, the others, particular in exposure at 5℃, markedly diminished MT effect. This result suggests that at least water temperature from 10℃ and up is necessary to perform high sensitive detection of androgenic activity using spiggin as a biomarker.The present established systems will be powerful tools for the evaluation of the endocrine disrupting potency of chemicals. Less

雄性刺鱼会产生一种名为“spiggin”的胶蛋白，用作筑巢的胶结物质。 spiggin 的合成受到雄激素的强烈控制。因此，除了卵黄蛋白原（由雌激素刺激肝脏产生的卵黄蛋白前体）之外，spiggin 也是评估环境化学物质雄激素活性的有用生物标志物。在本研究中，我们建立了spiggin和卵黄原蛋白的高灵敏定量系统来评估化学物质的性激素效应。利用实时RT-PCR技术建立了spiggin mRNA的高灵敏定量系统。该系统可实现精确且高灵敏度的定量。卵黄蛋白原（雌激素刺激的生物标志物）也通过多种色谱技术进行纯化。还产生了针对卵黄原蛋白的特异性抗体。此外，采用化学发光联免疫吸附法建立了卵黄蛋白原定量系统。该系统还能够实现与 spiggin 测定系统相同的高灵敏度定量。除了生物标志物定量系统的这些准备之外，还确定了使用刺鱼的体内暴露测试的详细条件。使用MT作为对照雄激素，我们进行了暴露试验。结果，仅 3 天的 MT 浓度 0.1 和 1 μg/L 暴露对肾脏中 spiggin mRNA 的表达有效。接下来，在三种不同的水温（5℃、10℃和15℃）下进行MT暴露，以了解水温对spiggin mRNA表达的影响。 15℃暴露对spiggin mRNA表达的MT效应最大，其他的，特别是5℃暴露，MT效应显着减弱。这一结果表明，使用spiggin作为生物标记物进行雄激素活性的高灵敏度检测需要至少10℃及以上的水温。目前建立的系统将成为评估化学物质内分泌干扰效力的有力工具。较少的

项目成果

Highly sensitive detection of spiggin mRNA for the evaluation of androgenic potency of chemicals

高灵敏度检测 spiggin mRNA，用于评估化学品的雄激素效力

• 发表时间: 2007
• 作者: Masanobu;Sakai;et. al.
• 通讯作者: et. al.

Detection and assessment of androgenic potency of endocrine-disrupting chemicals using three-spined stickleback,Gasterosteus aculeatus

三刺棘鱼内分泌干扰物雄激素效力的检测和评估

• 发表时间: 2007
• 期刊: Environmental Sciences 14
• 作者: Tomita;T;Yamaguchi Y;yamauchi k;Amasaki Y;Kakinuma S;Yamauchi K;Amasaki S;Kakinuma S;Kakinuma S;柿沼 志津子;柿沼志津子;柿沼志津子;柿沼志津子;柿沼志津子;山内一己;山内一己;甘崎佳子;尚奕;小玉陽太郎;島田義也;島田義也;Kakinuma S.;Kakinuma S.;Kakinuma S;Kakinuma S;Yamauchi K;Yamauchi K;Amasaki S;Shang Y;Kodama Y;Shimada Y;Shimada Y;Masaki Nagae
• 通讯作者: Masaki Nagae

Detection and assessment of androgenic potency of endocrine-disrupting chemicas using three-spined stickleback, Gasterosteus aculeatus

使用三刺棘鱼检测和评估内分泌干扰化学物质的雄激素效力

• 发表时间: 2007
• 期刊: Environmental Sciences 14
• 作者: Ioanna;Katsiadaki;et. al.;Masaki Nagae
• 通讯作者: Masaki Nagae

イトヨゲノムデータベースに基づくアンドロゲン特異的タンパク(スピギン)mRNAの高感度検出系構築

基于刺鱼基因组数据库的雄激素特异性蛋白（spigin）mRNA高灵敏检测系统的构建

• 发表时间: 2007
• 作者: Ioanna;Katsiadaki;et. al.;Masaki Nagae;Ioanna Katsiadaki;坂井 正頌
• 通讯作者: 坂井 正頌

Establishment of highly sensitive quantification sysytem of spiggin mRNA based on stickleback genome database information

基于刺鱼基因组数据库信息的spiggin mRNA高灵敏定量系统的建立

• 发表时间: 2006
• 作者: Masanobu;Sakai;et. al.
• 通讯作者: et. al.