Molecular profile of cranial motor neurons

描述(由申请人提供)：我们主要对运动神经元对肌萎缩侧索硬化症(ALS)的特殊易感性感兴趣。肌萎缩侧索硬化症导致运动神经元的选择性变性，尽管导致肌萎缩侧索硬化症的因素也存在于其他对肌萎缩侧索硬化症具有抵抗力的中枢神经系统神经元中。即使在运动神经元中，也有不同程度的ALS易感性。例如，脑干中的脑神经I11、IV和VI核对ALS具有相对的抵抗力，而面神经核运动神经元不可避免地屈从于变性。由于肌萎缩侧索硬化症中只有部分神经元退化，我们推测可能存在运动神经元的特定特征，从而赋予易感性。识别和表征独特的分子可以降低或增强运动神经元的变性易感性，对于理解和最终治疗运动神经元疾病至关重要。沿着这些思路，了解特定运动神经元组的基因和蛋白质表达谱对于了解它们对运动神经元病的影响的敏感性至关重要。因此，我们建议利用基因芯片和实时定量聚合酶链式反应的方法，在ALS的小鼠模型--正常小鼠和铜/锌超氧化物歧化酶(SOD1)突变小鼠中，通过基因表达谱方法寻找在脑运动神经元中唯一表达的基因。这样的研究将使我们能够了解运动神经元特有的分子结构和功能，并将建立一个研究运动神经元疾病基因表达变化的筛选系统。这项研究的具体目标概述如下。 具体目的1.通过基因表达谱鉴定运动神经元特异性基因表达。我们将进行细胞分选和激光捕获显微切割，以分离在颅运动神经核运动神经元表达的mRNAs。通过DNA微阵列和实时荧光定量聚合酶链式反应，将在正常和SOD1突变小鼠中鉴定和比较运动神经元中特异表达的基因。具体目的2.明确运动神经元特异性基因在正常和SOD1突变小鼠脑运动核团中的差异表达。我们将通过原位杂交来确定特定目的1中确定的运动神经元特异性基因在颅运动核中的表达模式。这些基因的差异表达水平将进一步通过实时荧光定量聚合酶链式反应进行定量。

DESCRIPTION (provided by applicant): We are interested primarily in the specific susceptibility of motor neurons to Amyotrophic Lateral Sclerosis (ALS). ALS results in selective degeneration of motor neurons despite that factors contributing to ALS are also present in other CNS neurons that are generally resistant to ALS. Even among motor neurons there are differing degrees of susceptibility to ALS. For example, cranial nerve nuclei Ill, IV, and VI in the brainstem are relatively resistant to ALS, while facial nucleus motor neurons inevitably succumb to degeneration. Since only subsets of neurons degenerate in ALS, we hypothesize that there may be specific features of motor neurons that confer susceptibility. Identification and characterization of unique molecules that either diminish or enhance a motor neuron's vulnerability to degeneration are critical to understanding and eventually treating motor neuron diseases. Along these lines, knowledge of the gene and protein expression profiles of a specific motor neuron group is critical to understanding their susceptibility to the effects of motor neuron disease. Therefore, we propose to identify genes uniquely expressed in cranial motor neurons by gene profiling using microarray and real-time PCR methods in normal and Cu/Zn superoxide dismutase (SOD1) mutant mice, a mouse model of ALS. Such a study will allow us to understand the molecular architecture and function specific to motor neurons and also will establish a screening system for studying gene expression alterations in motor neuron diseases. The Specific Aims of this study are outlined below. Specific Aim 1. To identify motor neuron-specific gene expression by gene profiling. We will undertake cell sorting and laser capture microdissection to isolate mRNAs expressed in motor neurons of cranial motor nuclei. The genes expressed specifically in motor neurons will be identified and compared in normal and SOD1 mutant mice by DNA microarray and real-time PCR. Specific Aim 2. To define differential expression of motor neuron-specific genes in cranial motor nuclei of normal and SOD1 mutant mice. We will determine by in situ hybridization the expression patterns of motor neuron-specific genes in cranial motor nuclei identified in Specific Aim 1. Differential expression levels of these genes will be further quantified by real-time PCR.