Effects of endocrine disruptors on developing brain, with special reference to gene expression.

为了研究子宫内双酚A（BPA）暴露对发育中的大脑的影响，我们采用分子生物学和免疫组织化学方法研究了胚胎期暴露于BPA的胎鼠大脑。从妊娠初至胚胎期12.5、14.5、16.5、18.5天，对妊娠期ICR小鼠分别给予双酚a。胎儿端脑被处理为包含超过30,000个小鼠基因片段的微阵列进行全局基因分析。给予上述处理的怀孕小鼠5-溴-2'-脱氧尿嘧啶（BrdU, 12.5mg.kg），剂量分别为E12.5、14.5、16.5和18.5。分别在注射BrdU后1小时或2-3天后进行剖宫产。将脑固定并包埋于蜡中，然后进行BrdU、蛋白二硫异构酶（PDI）和其他细胞分化标志物的免疫组织化学染色。cDNA微阵列分析显示，大约2%的基因上调（bpa处理小鼠的2倍差异被认为是显著的），其中包括发育调节基因，如Numb和Lnx家族。在E12.5 ~ 16.5范围内，皮质板、底板、背侧新皮质中间带和海马CA均可见PDI免疫反应性。PDI免疫反应性在皮质板呈放射状分布。脑远端壁brdu阳性细胞密度从E12.5增加到14.5，表明BPA加速了干细胞的增殖和神经元的过早迁移。在bpa处理的小鼠中，tuj1阳性的突出纤维显示出更突出的束状化。我们的观察表明，在胚胎早期到中期暴露于BPA会影响小鼠的大脑发育。

To investigate the effects of in utero exposure to bisphenol A(BPA) on developing brain, we have studied fetal mouse brains that were exposed to BPA during embryonic period with molecular biological as well as immunohistochemical methods. BPA was given so to pregnant ICR mice, from the beginning of gestation to embryonic day 12.5, 14.5, 16.5, 18.5, respectively. Fetal telencephalons were processed for global gene analysis with microarrays containing more than 30,000 mouse gene fragments. Pregnant mice that were treated as mentioned above were given 5-bromo-2'-deoxyuridine (BrdU, 12.5mg.kg) ip, at E12.5, 14.5, 16.5, and 18.5. Fetuses were obtained by caesarean section, 1h after BrdU injection or 2-3days after. The brains were fixed and embedded in wax, followed by immunohistochemical staining for BrdU as well as protein disulfide isomerase(PDI) and other cell-differentiation markers. cDNA microarray analyses revealed up-regulation (2-fold difference in BPA-treated mice was regarded as being significant) of approximately 2% of genes, which included developmentally regulated genes such as Numb, and Lnx family. PDI immunoreactivity was discernible in the cortical plate, subplate, intermediate zone of dorsal neocortex and CA of hippocampus, from E12.5 to 16.5. PDI immunoreactivity was prominent in the cortical plate as radial distribution. BrdU-positive cell density in telencephalic wall was increased in all zones of developing cortex, from E12.5 to 14.5, which suggests accelerated stem cell proliferation and premature neuronal migration by BPA. TUJ1-positive projecting fibers showed more prominent fasciculation in BPA-treated mice. Our observation indicates that exposure to BPA at early to mid-embryonic period affects brain development in mice.