多氯联苯（PCBs）对文蛤甲状腺激素干扰机制研究

Thyroid hormones (THs) have an important regulating effect on biological growth and reproduction. Thyroid hormone disrupting effects may seriously affect the biological survival, growth and reproduction of marine organisms. Polychlorinated biphenyls (PCBs) have been listed as one of the twelve persistent organic pollutants (POPs) under the Stockholm, which were also some of the most widely studied thyroid-disrupting chemicals (TDCs). Recently, more attention has been paid to the disturbance of thyroid system by PCBs in fish, which focused on the synthesis, transport and metabolism of THs. However, rarely been reported the disrupting effects on THs pathway of mollusks, especially bivalves. This study used commercialized PCBs mixtures, Aroclor 1254, to figure out the thyroid disruption effects of PCBs under environmental related concentration on M. meretrix, constructing the dynamic defense transcriptome responsive to the analysis of the toxic effect of Aroclor 1254; to probe its role in the synthesis of THs, and the expression levels of genes in synthesis, transport, metabolism and excretion processes. Constructing two recombinant yeast THs systems of TRs and TTR from M. meretrix aims to find out the disruption interaction sites of PCBs, to resolve the mechanism of Thyroid hormone disrupting effects PCBs-induced in clam M. meretrix. This study provides a preliminary basis and new idea for studying screening methods of ecotoxicology assessment of TDCs.

多氯联苯（PCBs）是12种优先控制持久性有机污染物（POPs）之一，也是典型的甲状腺干扰物（TDCs），可能会严重影响生物的生存、生长和繁衍。目前研究较多的是PCBs对鱼类的甲状腺干扰效应，主要集中于对THs的合成、转运和代谢的影响。但PCBs对软体动物特别是双壳贝类的甲状腺激素干扰效应还鲜见报道，其干扰途径和干扰作用靶点都有待研究。本项目结合我国近海PCBs污染现状，选择Aroclor 1254为代表，通过构建Aroclor 1254胁迫下文蛤（M. meretrix）消化盲囊组织动态差异基因表达谱，整体分析Aroclor 1254对文蛤的毒性效应；构建文蛤甲状腺激素受体（TRs）和甲状腺转运蛋白（TTR）的酵母重组系统，查找PCBs的干扰机制；探究Aroclor 1254对文蛤甲状腺激素干扰效应，为筛选出TDCs的海洋生态毒理评价提供新的思路。

多氯联苯（PCBs）作为一种典型的甲状腺干扰物，在近海海域及海洋生物体内的污染状况及毒理学研究备受关注。构建文蛤PacBio全长转录组文库，查找到与甲状腺激素合成、转运、代谢功能相关的基因序列，如钠碘转运蛋白NIS、甲状腺过氧化物酶TPO、甲腺原氨酸脱碘酶Deio、甲状腺激素转运体Mct8等。构建在 Aroclor 1254胁迫下的文蛤成体消化盲囊（10、100、1,000 ng/L）和3月龄稚贝软体部（20、200、2,000 ng/L）差异基因表达谱。在暴露期内，文蛤对低浓度（10 ng/L和20 ng/L）的Aroclor 1254响应更显著，呈上调趋势，且文蛤稚贝对其响应更显著。与文蛤甲状腺激素信号通路相关的促甲状腺激素受体TSHR、TPO、Mct10、Deio、CYP450s、GST、SULT、SOD和ABC转运蛋白等基因的表达被不同程度的抑制或诱导，Aroclor 1254胁迫还会引起文蛤的解毒代谢反应、氧化应激反应、免疫系统反应等。构建文蛤消化盲囊组织初级cDNA文库，文库库容> 约 2×106cfu，重组率大于 100%。获得甲状腺激素受体TRβ基因部分同源序列和甲状腺转运蛋白TTR基因的同源序列的部分。通过激素水平、生理生化和qRT-PCR分析，作为THs结构类似物，Aroclor 1254暴露会引起文蛤体内一定程度的Aroclor 1254的蓄积，并对文蛤体内甲状腺激素水平造成紊乱，总T3/T4水平在暴露早期上升，后期显著降低；暴露期内，Deio活力被抑制，TPO、SULT和UGT的活力被诱导，加速了T4向T3的转化；Deio、Mct8、RXRA的转录水平有升高。Aroclor 1254暴露干扰了文蛤甲状腺激素信号通路，在激素水平、酶活力水平和转录水平均具有干扰效应，推测其干扰机制与甲状腺激素的合成、转运、代谢等过程均相关，但PCBs对文蛤甲状腺激素干扰机制还需要进一步的研究。