围产期低剂量双酚A暴露对小鼠子代脑细胞的影响以及与儿童学习记忆改变的关系

Bisphenol A (BPA) and its analogues, monomers of the production of polycarbonate plastic and epoxy resins, are environmental xenoestrogenic endocrine disruptors. When enter human body, BPA will interferer the development of brain and alter the behavior of children. Embryo and infants are sensitive to it due to the body development. But there does not exist evidence to show that BPA exposure is related with the altered behavior of children. In this study, we plan to simulate the conditions of children’s long-term exposure to BPA by using young mice and pregnant female mice and study the relationships between them. After oral exposure to different levels of BPA during perinatal period, the DNA damage effect of BPA exposure on the brain cell for young mice and the offspring of mice will be assessed by neuron counting, comet assay, flow cytometry ; Y-maze experiments are conducted to test the altered abilities of learning and memory after BPA exposure; Moreover, long term potentiation experiments and the measurements of neurotransmitter concentrations in brain by liquid chromatography coupled with tandem mass spectrometer will be used to explore the mechanism. By comparing the concentrations of BPA and its analogues in urine between healthy children and unhealthy children with mental diseases, the relationships between urinary BPA levels and altered behavior, the associations of BPA exposure and the altered learning and memory ability of children would be surveyed and the dose-effects model is exploreed to be established between them. We hope to find the clinical evidences that low dose exposure to BPA chronically during perinatal period would damage/influence children’s brain development. The study is a combination of environmental analytical chemistry, toxicology and neurophysiology to explore the effects of low-dose and long-term exposure to BPA on the brain cell and children' learning and memory ability during perinatal period.

双酚A（BPA）及其衍生物是一种内分泌干扰物，妨碍脑部发育，改变儿童行为方式。胚胎及婴幼儿对BPA摄入格外敏感，但尚无证据表明环境BPA暴露与儿童学习记忆改变存在直接关联。本研究拟采用动物实验和人群生物监测相结合的方法，研究低浓度BPA暴露与儿童脑细胞损伤和学习记忆行为改变间的关系。围生期母鼠BPA暴露后，通过神经元计数、单细胞凝胶电泳/流式细胞术评价脑细胞DNA损伤及生长周期、海马区神经元树突棘形态变化说明围产期BPA暴露对小鼠后代脑细胞的神经毒性及剂-效关系；Y-迷宫实验检验子代小鼠BPA暴露后学习记忆行为的改变，并用长时程增强实验和神经递质因子阐述其影响机制；比较学习障碍疾患和健康儿童尿中BPA类化合物浓度，探究BPA类化合物暴露和临床儿童学习记忆因子间的剂-效关系，从临床角度寻找低浓度长期BPA暴露与儿童学习记忆改变之间的关系。

双酚A（BPA）是一种塑料添加剂，具有内分泌干扰效应，有研究初步显示其具有神经毒性。但低尚无证据表明环境剂量BPA暴露与儿童学习记忆能力下降或行为学改变存在直接关联。因此本研究采用动物实验和病例-对照人群生物监测相结合的方法，研究低浓度BPA暴露与儿童脑细胞损伤和学习记忆行为改变间的关系及其机制。在此过程中，解决以下两个关键科学问题：（1）低浓度围产期 BPA 暴露是否损伤神经细胞，或改变神经递质从而导致子代小鼠学习记忆行为改变；（2）尿中 BPA 浓度与儿童学习记忆障碍之间是否具有相关性？经过四年的执行期顺利达成所有研究计划，主要结论有以下3条：1）围产期、低剂量BPA暴露对后代小鼠的学习记忆能力的干扰和神经发育的影响呈现性别及代系差异性：BPA暴露损害F1代雄鼠的学习记忆，造成F1代雄鼠脑细胞DNA损伤，显著降低F2代雄鼠海马CA1和DG区神经元数目；2）低剂量BPA暴露损伤子代小鼠空间学习记忆能力的机制与海马神经元树突生长发育和神经递质稳态的扰乱效应有关。3）患有注意力缺陷多动症（ADHD）儿童尿中BPA浓度显著高于健康对照组儿童；随着BPA浓度的增加，男孩和女孩患ADHD的风险比值均逐渐增加，且男孩高于女孩；尿中BPA S浓度与DNA氧化损伤标志物（8-OHdG）浓度显著正相关，且存在剂量-效应关系。此部分与动物实验部分形成了交叉验证。我们执行本研究的过程中发现了BPA暴露导致的神经毒性和学习记忆能力下降具有性别差异性，因此我们拓展了研究的广度和深度，主要采用不同细胞（细胞系和原代海马细胞）为模型，研究了BPA暴露导致的神经毒性的性别差异性机制，目前仍处于继续深入的过程中，初步的结论有：1）无论是原代细胞还是细胞系，BPA暴露均呈现出显著的神经毒性效应，包括增加活性氧水平，导致炎症因子升高，促进细胞凋亡，抑制细胞增殖等现象；2）BPA与雌激素受体结合后，性别差异性诱导人神经母细胞瘤细胞氧化应激、线粒体凋亡和细胞焦亡；3）抗氧化剂EGCG对其造成的神经细胞毒性有保护作用。