六溴环十二烷通过星形胶质细胞介导神经发育毒性的机制研究

Hexabromocyclododecanes (HBCDs), as a kind of added flame retardants, are widely used in polystyrene foam, upholstery, textiles and electronic products. They are easily released into the environment, and due to its non-biodegradable and biologically cumulative characters they have been detected in food and human breast milk. Researches suggest that they may cause a range of diseases like DDT and PCBs, but the health risk assessment about HBCDs is still in its beginning. Research has shown that HBCDs have neurodevelopmental toxicity, which may be associated with its interference effects on peripheral thyroid hormone. Our previous studies first discovered HBCDs are toxic to astrocytes. Together with previous studies, we assume that HBCDs maybe directly act on the astrocytes by thyroid hormone-dependent mechanisms, affecting their development and function, causing developmental disorders of the nervous system. Based on this assume, our project intends to explore the neural development toxic effects of HBCDs and regulation mechanism from organ, cell and molecular levels, using animal and cell experiments by neural behavior test, organ and cell morphology, gene and protein expression analysis, and RNA interference technology. This project will help to find new targets for prevention and cure HBCDs neural development toxicity, and also lay a theoretical foundation for the neural development toxicity research of thyroid hormone interferent.

六溴环十二烷(HBCDs)为添加型阻燃剂，广泛应用于聚苯乙烯泡沫材料、室内装潢纺织品和电子产品中，易释放进入环境，因其难降解而具有生物累积性，在食品和母乳中被检出。研究表明其可能与DDT和多氯联苯一样，引起一系列疾病，目前对HBCDs的健康风险评价尚处于起步阶段。已有研究表明HBCDs具有神经发育毒性，初步认为与其外周甲状腺激素干扰作用有关。我们前期工作中首次发现HBCDs具有星形胶质细胞毒作用，结合已有的文献分析，我们推测HBCDs通过甲状腺激素依赖机制直接作用于星形胶质细胞，影响其发育和功能状态，从而导致神经系统发育障碍。基于此设想，本课题拟采用细胞和动物实验相结合，通过神经行为学、组织及细胞形态学、基因和蛋白表达分析等，从整体-细胞-分子三个层面探讨HBCDs神经发育毒性作用与机制，为HBCDs神经发育毒性的防治提供新靶标，也为研究甲状腺激素干扰物神经发育毒性作用的机制奠定基础。

六溴环十二烷(HBCDs)是一种添加型阻燃剂，主要用于聚苯乙烯颗粒和建筑材料中以降低其可燃性。HBCDs具有亲脂性和生物累积性，已经成为环境介质和生物群中无处不在的污染物，在人血液和母乳中被检出。现有少量研究表明HBCDs具有内分泌干扰毒性和神经毒性。本年度我们重点研究了HBCDs对星形胶质细胞的毒性作用及作用机制。首先利用原代培养的星形胶质细胞考察不同剂量HBCDs暴露对细胞增殖与活力的影响，结果表明HBCDs剂量依赖性地降低细胞活力。进一步实验结果证实HBCDs引起细胞内活性氧生成增多，抗氧化酶SOD和GSH-Px活力下降、脂质过氧化产物MDA水平上升，氧化损伤导致细胞发生凋亡和坏亡；其次利用酶联免疫方法检测凋亡相关蛋白Bax、Bcl-2和caspase-3表达，蛋白印迹方法检测P38信号通路激活情况，结果表明HBCDs暴露引起促凋亡蛋白Bax表达量增高，抑凋亡蛋白Bcl-2表达降低，Caspase-3和P38信号通路被激活；此外ELISA法检测HBCDs暴露对星形胶质细胞分泌的细胞因子的影响， 结果显示HBCDs引起星形胶质细胞分泌的层粘连蛋白 (laminin) 和纤连蛋白(fibronectin)表达降低，白介素-6 (IL-6) 和肿瘤坏死因子 (TNF-α) 表达增高；最后利用HBCDs暴露的星形胶质细胞条件介质加入到皮层神经元培养液中，利用神经元特异蛋白MAP-2免疫细胞染色发现其对神经元具有损伤作用。本研究证明HBCDs引起星形胶质细胞损伤，发生凋亡现象，损伤的星形胶质细胞抗氧化能力下降，分泌的炎症因子增多，进而对神经元造成损害，影响神经发育。本研究对于深入理解HBCDs暴露对星形胶质细胞毒性的作用机制，以及后续研究HBCDs通过星形胶质细胞发挥神经发育毒性机制具有重要意义。