Analysis of Glutamatergic Neurons Derived from Patient-Specific iPS Cells

描述(申请人提供)：一些研究表明，精神分裂症(SZ)是由谷氨酸能传递不足引起的。对SZ谷氨酸能通路的分析已经使用了各种不同的实验方法，其中之一是使用死后脑组织或外周血白细胞进行的阵列杂交进行基因表达谱分析。这些研究为SZ的发病机制提供了许多令人振奋的线索。然而，当白细胞被用作大脑中基因表达的替代物时，存在明显的局限性，许多混杂因素使得从尸检样本中解释数据变得困难。此外，这些生物材料来源不能用于电生理分析。因此，研究培养的患者特异性谷氨酸能神经元的分子和电生理事件的能力将极大地增强我们对潜在疾病机制的理解。诱导多能干细胞(IPS)的发现，以及促使它们分化为神经胶质细胞、少突胶质细胞和各种神经元亚型的前景，为解决与解释死后组织数据相关的问题提供了机会。我们将从对照组和SZ患者的毛根角质形成细胞中获得iPS细胞。患者队列将包括具有22q11缺失的SZ患者，22q11缺失是一个遗传同源亚组。诱导的多能干细胞将通过在小鼠ES细胞中建立的程序被诱导分化为谷氨酸能锥体神经元，并将使用大规模并行深度测序(RNA-SEQ)进行表达谱分析，这比基于阵列杂交的方法更敏感。将比较患者和对照组之间的表达情况，以确定SZ特有的差异。由于正在使用的分化方案产生能够形成突触连接的谷氨酸锥体神经元，因此也将进行电生理分析。拟议中的使用分化的、患者特有的iPS细胞的实验将导致开发一种新的方法来研究精神分裂症的潜在分子基础。 公共卫生相关性：这项研究描述了我们对培养精神分裂症患者神经元的兴趣。这些神经元将由所谓的诱导多能干细胞产生，这些干细胞来自皮肤或发根。诱导多能细胞具有分化为多种不同细胞类型的能力，包括神经元。研究精神分裂症患者的神经元将有助于我们了解这种神秘的疾病，并可能导致新的治疗方法。

DESCRIPTION (provided by applicant): A number of studies suggest that schizophrenia (SZ) is caused by a deficiency in glutamatergic transmission. The analysis of glutamatergic pathways in SZ has been addressed using a variety of different experimental approaches, one of which is gene expression profiling using array hybridization carried out with postmortem brain tissue or peripheral blood leukocytes. These studies have provided many exciting clues to SZ pathogenesis. However, there are glaring limitations when leukocytes are used as a surrogate for gene expression in the brain, and many confounding factors make it difficult to interpret data from autopsy specimens. In addition, these sources of biological material cannot be used for electrophysiological analysis. Consequently, the ability to study molecular and electrophysiological events in cultivated patient-specific glutamatergic neurons would greatly enhance our understanding of underlying disease mechanisms. The discovery of induced pluripotent stem (iPS) cells, and the prospect of driving them to differentiate into glia, oligodendrocytes, and various neuronal subtypes, provides the opportunity to address the problems associated with interpreting data from postmortem tissue. We will generate iPS cells from hair root keratinocytes obtained from control subjects and patients with SZ. The patient cohort will include individuals with SZ who have 22q11 deletions, a genetically homogenous subgroup. Induced pluripotent stem cells will be driven to differentiate into glutamatergic pyramidal neurons using a protocol established in mouse ES cells, and expression profiling will be carried out using massively parallel deep sequencing (RNA-seq), which is more sensitive than array hybridization-based methods. Expression profiles between patients and controls will be compared to identify SZ-specific differences. Since the differentiation protocol being used generates glutamatertic pyramidal neurons capable of forming synaptic connections, electrophysiological analyses will be carried out as well. The proposed experiments using differentiating, patient-specific iPS cells will result in the development of a new method for studying the underlying molecular basis of schizophrenia. PUBLIC HEALTH RELEVANCE: This study describes our interest in growing neurons from patients with schizophrenia. The neurons will be generated from so-called induced pluripotent stem cells, which are derived from skin or hair roots. Induced pluripotent cells have the capacity to develop into many different cell types, including neurons. Studying neurons from patients with schizophrenia will contribute to our understanding of this enigmatic illness and possibly lead to new therapies.