US Environmental Protection Agency, Research Triangle Park, NC 27711, USA.
Toxicol Appl Pharmacol. 2010 May 15;245(1):1-8. doi: 10.1016/j.taap.2010.02.008. Epub 2010 Feb 19.
Polybrominated diphenyl ethers (PBDEs) are used as additive flame retardants and have been detected in human blood, adipose tissue, and breast milk. Both in vitro and in vivo studies have shown that the effects of PBDEs are similar to the known human developmental neurotoxicants such as polychlorinated biphenyls (PCBs) on a molar basis. Previously, we reported that PBDE mixtures and congeners, perturbed calcium homeostasis which is critical for the development and function of the nervous system. In the present study, we tested whether environmentally relevant PBDE/PCB mixtures and congeners affected mitogen-activated protein kinase (MAPK) pathways, which are down-stream events of calcium signaling in cerebellar granule neuronal cultures. In this study, phosphorylated extracellular signal-regulated kinase (pERK)1/2, a widely studied MAPK cascade and known to be involved in learning and memory, levels were quantitated using western blot technique with phospho-specific antibodies. Glutamate (a positive control) increased pERK1/2 in a time- and concentration-dependent manner reaching maximum activation at 5-30min of exposure and at doses > or =10microM. Both Aroclor 1254 (a commercial penta PCB mixture) and DE-71 (a commercial penta PBDE mixture) elevated phospho-ERK1/2, producing maximum stimulation at 30min and at concentrations > or =3microg/ml; Aroclor 1254 was more efficacious than DE-71. DE-79 (an octabrominated diphenyl ether mixture) also elevated phospho-ERK1/2, but to a lesser extent than that of DE-71. PBDE congeners 47, 77, 99, and 153 also increased phospo-ERK1/2 in a concentration-dependent manner. The data indicated that PBDE congeners are more potent than the commercial mixtures. PCB 47 also increased phospho-ERK1/2 like its structural analog PBDE 47, but to a lesser extent, suggesting that these chemicals affect similar pathways. Cytotoxicity, measured as %LDH release, data showed that higher concentrations (>30microM) and longer exposures (>30min) are required to see cell death. These results show that PBDE mixtures and congeners activate MAPK pathway at concentrations where no significant cytotoxicity was observed, suggesting that perturbed intracellular signaling including MAPK pathway might be involved in the initiation of adverse effects, including learning and memory, related to these persistent chemicals.
多溴联苯醚（PBDEs）被用作添加型阻燃剂，已在人体血液、脂肪组织和母乳中被检测到。体外和体内研究均表明，PBDEs 的影响与已知的人类发育神经毒物（如多氯联苯（PCBs））在摩尔基础上相似。此前，我们报道 PBDE 混合物和同系物扰乱了钙稳态，而钙稳态对神经系统的发育和功能至关重要。在本研究中，我们测试了环境相关的 PBDE/PCB 混合物和同系物是否会影响丝裂原活化蛋白激酶（MAPK）途径，该途径是小脑颗粒神经元培养物中钙信号的下游事件。在这项研究中，使用磷酸化特异性抗体通过 Western blot 技术定量了磷酸化细胞外信号调节激酶（pERK）1/2 的水平，磷酸化细胞外信号调节激酶（pERK）1/2 是一种广泛研究的 MAPK 级联反应，已知与学习和记忆有关。谷氨酸（阳性对照）以时间和浓度依赖的方式增加了 pERK1/2，在暴露 5-30 分钟和剂量≥10μM 时达到最大激活。Aroclor 1254（一种商用五氯联苯混合物）和 DE-71（一种商用五溴联苯醚混合物）均升高了磷酸化 ERK1/2，在 30 分钟和浓度≥3μg/ml 时产生最大刺激；Aroclor 1254 比 DE-71 更有效。DE-79（一种八溴二苯醚混合物）也升高了磷酸化 ERK1/2，但程度低于 DE-71。PBDE 同系物 47、77、99 和 153 也以浓度依赖的方式增加了磷酸化 ERK1/2。数据表明 PBDE 同系物比商业混合物更有效。PCB 47 像其结构类似物 PBDE 47 一样增加了磷酸化 ERK1/2，但程度较小，表明这些化学物质影响相似的途径。细胞毒性，以 LDH 释放的百分比表示，数据表明，在观察到细胞死亡之前，需要更高的浓度（>30μM）和更长的暴露时间（>30 分钟）。这些结果表明，在没有明显细胞毒性的浓度下，PBDE 混合物和同系物激活了 MAPK 途径，这表明细胞内信号的紊乱，包括 MAPK 途径，可能与这些持久性化学物质引起的学习和记忆等不良效应的发生有关。
