PXR介导河口水体中PhACs暴露对阿部鲻鰕虎鱼的化学致敏/致耐性效应及其调控机制

Pharmaceuticals active compunds (PhACs) are a group of potential emerging pollutants. The combined pollution of PhACs may cause the alteration of the fish detoxification system and lead to the changes of other PhACs metabolic dynamics process and PhACs contents in fish. Consequently, the threshold of toxic responses of the fish to PhACs exposure will vary up/down, and ultimatly the susceptibility of the fish to PhACs exposure will be changed. Therefore, toxic impacts of PhACs combined pollution are specially worthy of paying more attenion. The aims in the project mainly focus on the study of the effects of typical PhACs in estuary water (diclofenac and simvastatin) upon the detoxification system of the fish (Mugilogobius abei), including transcriptional factors (PXR, pregnane X receptor), ABC superfamily transporters proteins(e.g. P-gp), P450 enzymes system (e.g. P4501A, P4503A) and Phase II conjugation enzymes (GST, UGT) etc. The responses of PXR to the exposure of PhACs combined pollution and its regulation mechanism upon the detoxification system of the fish will be investigated. The analysis of relationship between the responses of the fish toxic endpoints in individual physiological levels and the changes of its detoxification system in biochemical and molecule levels will be extensively attempted. The successful accomplish of the project will be benefit to unravel the main mechanisms in chemosensitive/chemoresistant effects derived from the combined pollution of PhACs. The works will be helpful to provide scientific basis for the risk assessment of PhACs to estuary ecosystem.

医药活性化合物(PhACs)是一类受到广泛关注的新型污染物。在复合污染状态下，某些PhACs对鱼类解毒系统的干扰破坏可能引起其他类PhACs在鱼体内代谢过程改变，从而使得鱼类对PhACs的毒性反应阈值降低/增高，导致鱼类对该污染物敏感性增高/降低，这种化学致敏/致耐性效应使得PhACs复合污染问题特别值得关注和研究。本项目以河口水体典型PhACs(双氯芬酸、辛伐他汀)和阿部鯔鰕虎鱼为对象，研究PhACs对鱼类解毒代谢系统(包括核受体转录因子PXR，膜转运蛋白P-gp、P450酶系及GST等)的影响及其作用机制；探讨PXR对PhACs暴露的响应变化及其对下游解毒代谢相关靶基因的调控作用；揭示PhACs复合污染对鱼类解毒代谢系统干扰与其化学致敏/致耐性效应之间的关系；阐明PhACs复合污染暴露对鱼类产生化学致敏/致耐性效应的作用机理，为PhACs的生态风险评价提供科学依据。

双氯芬酸和辛伐他汀是广泛使用的典型医用药物，其在水体环境中的低剂量长期暴露可对水生非靶生物产生潜在影响。本项目主要研究双氯芬酸和辛伐他汀暴露对珠江口土著鱼类--阿部鯔鰕虎鱼代表性非靶生物解毒代谢过程的干扰影响及由此变化可能导致的受试生物的化学致敏/致耐性效应改变。研究围绕解毒代谢相关的PXR信号通路、抗氧化相关的Nrf2信号通路及PPAR信号通路，开展了两种药物对阿部鯔鰕虎鱼解毒代谢相关基因在转录水平及酶活性与蛋白水平的响应变化研究及药物暴露剂量－时间－效应关系研究。本研究成功克隆阿布鲻吓唬鱼PXR及其下游主要相关靶基因（CYP1a、CYP3a、GST、UGT、P-gp、Mrp）。同时在研究过程中发现阿布鲻吓唬鱼脂肪代谢有别于普通鱼类，所以对脂肪代谢相关的核转录因子PPAR信号通路及Nrf2抗氧化信号通路进行了探索性研究。完成PPAR信号通路相关基因（RXRα, PGC-1α, SULT, CYP4,）和Nrf2信号通路相关基因（GCLC、SOD、CAT、GPx、Trx）的克隆和序列分析，对构建更全面的阿部鯔鰕虎鱼解毒代谢系统相关基因文库奠定了基础。水环境中环境相关浓度的双氯芬酸和辛伐他汀暴露对阿布鲻鰕鯱鱼的解毒代谢相关PXR信号通路、抗氧化相关Nrf2信号通路和脂代谢相关PPARα信号通路的基因转录表达、酶活及蛋白表达可产生显著影响。不同信号通路相关基因的转录表达与辛伐他汀和双氯芬酸暴露之间存在典型的时间-剂量-效应关系。双氯芬酸暴露对阿布鲻鰕鯱鱼肝脏组织结构及胚胎发育影响相对较弱。但辛伐他汀暴露对阿布鲻鰕鯱鱼肝脏组织结构可造成明显改变，对阿布鲻鰕鯱鱼胚胎发育有显著毒性影响。水体环境中辛伐他汀和双氯芬酸等典型PPCPs类新型污染物对鱼类等非靶生物可造成不同程度的氧化胁迫、胚胎发育毒性，其毒性效应及潜在生态生态风险不容忽视，需进一步深入研究。