Effect of Ethanol on Cell Proliferation

了解乙醇对神经干细胞的影响对于解开缺陷的病因学结至关重要 胎儿酒精谱系障碍（FASD）FASD是一个引人注目的问题，因为它是一个主要的 发育性精神功能障碍的原因，影响到所有活产婴儿的2%（事实上，它是精神障碍的主要原因。 在美国的发育迟缓），它提供了对其他（通常是共病）精神健康障碍病因的见解 (e.g.,注意力缺陷多动障碍和自闭症）。大脑的组成和大小 通过神经干细胞的增殖以及衍生物的数量和谱系。不正确的数字或 神经元亚群的平衡可能是精神功能障碍的基础。因此，我们将测试乙醇 影响发育中的中枢神经系统中产生的细胞的循环行为和命运。 大脑皮层由兴奋性投射神经元和抑制性局部投射神经元组成， 回路神经元（LCN）。这些来源于产前两个不同的增殖区域。PN来自背侧 端脑包括两个区：脑室区（VZ）及其衍生物，脑室下区 （SZ）。大多数皮质LCN产生于端脑腹侧，内侧神经节隆起（MGE）。 VZ/SZ和MGE细胞的命运在两步过程中定义。(1)决定细胞是否保留 和（2）表型（例如，神经元的类型）被定义。在上一期间 支持，我们表明，转化生长因子（TGF）β 1是神经干细胞增殖的关键调节因子。 本项目将探讨乙醇对细胞增殖动力学的影响，以及对 细胞命运具体目标1和2将使用体内模型确定乙醇对循环活性的影响 (cell周期动力学和退出）和皮质中神经干细胞对TGF β受体的表达/活化的影响 增殖区在它们的发育过程中，神经干细胞表达同源框基因产物Pax 6和/或Pax 8。 Tbr 2.这些蛋白质定义了从VZ中的（Pax 6+）干细胞到（Tbr 2+）中间祖细胞的转变 细胞期这种过渡对于表层皮层的发育至关重要（例如，胼胝体突起的起源） 这是产前暴露于乙醇的目标。具体目标3将使用两种类型的培养物（器官型切片 保留体内样组织和从VZ/SZ或MGE收获的神经干细胞系），以探索 乙醇对TGF β 1调节的细胞增殖和命运决定的影响。此外，我们还将鉴定出 被乙醇和/或TGF β 1上调和下调并沉默（甲基化）。在具体目标4中，神经干 将细胞移植（同位或异位）以确定乙醇和/或TGF β 1对 遗传和环境的贡献，以确定循环行为和定义细胞的命运。 与此同时，新的目标将使用三个互补的模型，以获得关键的洞察机制，定义 细胞增殖和命运以及乙醇对这些关键因素的影响。

Understanding the effects of ethanol on neural stem cells is critical to unraveling the etiological knots of deficits associated with fetal alcohol spectrum disorder (FASD). FASD is a compelling problem because it is a major cause of developmental mental dysfunction affecting ~2% of all live births (indeed, it is the chief cause of mental retardation in the USA), and it provides insights into the etiology of other (often co-morbid) mental health disorders (e.g., attention deficit hyperactivity disorder and autism). The composition and size of the brain are established by the proliferation of neural stem cells and by the numbers and lineage of the derivatives. Improper numbers or balance of neuronal subpopulations can underlie mental dysfunction. Thus, we will test the hypothesis that ethanol affects the cycling behavior and fates of cells generated in the developing central nervous system. Cerebral cortex is composed of two types of neurons: excitatory projection neurons (PNs) and inhibitory local circuit neurons (LCNs). These derive prenatally from two distinct proliferative regions. PNs come from the dorsal telencephalon which comprises two zones: the ventricular zone (VZ) and its derivative, the subventricular zone (SZ). Most cortical LCNs are generated in the ventral telencephalon, in the medial ganglionic eminence (MGE). The fates of VZ/SZ and MGE cells are defined in a two-step process. (1) It is decided whether the cells remain in the cycling population and (2) the phenotype (e.g., the type of neuron) is defined. During the previous period of support, we showed that transforming growth factor (TGF) beta1 is a key regulator of neural stem cell proliferation. The present project will explore the effects of ethanol on dynamics of cell proliferation and on the definition of cell fate. Specific Aims 1 and 2 will use an in vivo model to determine the effects of ethanol on the cycling activity (cell cycle kinetics and exit) and on the expression/activation of TGFbeta receptors by neural stem cells in the cortical proliferative zones. During their development, neural stem cells express a homeobox gene product(s), Pax6 and/or Tbr2. These proteins define the transition from (Pax6+) stem cells in the VZ to an (Tbr2+) intermediate progenitor cell stage. This transition is critical for the development of superficial cortex (e.g., the origin of callosal projections) which is a target of prenatal exposure to ethanol. Specific Aim 3 will use two types of cultures (organotypic slices that retain in vivo-like organization and lines of neural stem cells harvested from the VZ/SZ or MGE) to explore the effects of ethanol on TGFbeta1-regulated cell proliferation and fate decisions. In addition, we will identify genes that are up- and down-regulated and silenced (methylated) by ethanol and/or TGFbeta1. In Specific Aim 4, neural stem cells will be transplanted (homotopically or heterotopically) to determine the effects of ethanol and/or TGFbeta1 on genetic and environmental contributions to determining cycling behavior and to defining cell fate. In concert, the novel Aims will use three complementary models to gain critical insight into mechanisms defining cell proliferation and fate and the effects of ethanol on these critical factors.