Generation of Cortical Motoneurons from Embryonic Stem Cells

描述（由申请人提供）：大脑皮层中的运动神经元，也称为上运动神经元，通过脑干和脊髓中的下运动神经元控制我们的肌肉运动。上运动神经元的退化导致肌肉进行性痉挛和无力，这是一些衰弱性神经系统疾病的基础，如肌萎缩性侧索硬化症（ALS）、遗传性痉挛性偏瘫和原发性侧索硬化症。这些运动神经元疾病没有有效的治疗方法，主要是由于缺乏对皮质运动神经元如何产生、功能成熟和退化的了解。基于我们从人类胚胎干细胞（ESCs）中成功产生脊髓运动神经元的应用，将探索从自我更新的人类ESCs （H1和H9系，NIH Registry WA01和WA09）中产生上运动神经元的可行性。我们已经建立了一个化学定义的培养系统，将人类ESCs引导到神经上皮细胞，并发现了一个关键的原始神经上皮阶段，用于指定特定区域的神经元亚型。我们将首先通过检测Wnt、Wnt拮抗剂和成纤维细胞生长因子（FGFs）等形态因子的作用，分别、联合或依次诱导背端端脑神经祖细胞分化。这些祖细胞将在神经营养因子和/或靶细胞（下运动神经元）和/或局部环境的存在下进一步分化为运动神经元。体外培养的上运动神经元与下运动神经元的相互作用以及移植到鸡胚胎背端脑后的功能将被评估。因此，该应用程序将从无限的ESCs中产生目前无法获得的上运动神经元，并为研究人类上运动神经元的诞生、功能成熟和病变提供了一种其他难以获得的工具。这些细胞也将为阻止运动神经元变性过程的药物筛选提供靶点，并为上运动神经元相关疾病的潜在未来细胞治疗提供来源。

DESCRIPTION (provided by applicant): motoneurons in the cerebral cortex, also known as upper motoneurons, control our muscle movement through lower motoneurons in the brain stem and spinal cord. Degeneration of upper motoneurons results in progressive spastic and weakness in muscles, which underlies some debilitating neurological disorders, such as amyotrophic lateral sclerosis (ALS), hereditary spastic hemiplegia and primary lateral sclerosis. There is no effective treatment for these motoneuron disorders, for the most part due to the lack of understanding how cortical motoneurons are born, functionally mature and degenerate. This application, based on our success in producing spinal motoneurons from human embryonic stem cells (ESCs), will explore the feasibility of generating upper motoneurons from self-renewing human ESCs (H1 and H9 lines, NIH Registry WA01 and WA09). We have established a chemically defined culture system to direct human ESCs to neuroepithelial cells and discovered a critical primitive neuroepithelial stage for specification of region-specific neuronal subtypes. We will first induce differentiation of dorsal telencephalic neural progenitors by examining the effect of such morphogens as Wnts, Wnt antagonists, and fibroblast growth factors (FGFs), individually, in combination, or sequentially. These progenitors will then be further differentiated to motoneurons in the presence of neurotrophic factors and/or target cells (lower motoneuron) and/or local environment. The function of in vitro generated upper motoneurons will be assessed by their interaction with lower motoneurons in culture and following transplantation to dorsal telencephalon of a chick embryo. This application will thus produce the currently unavailable upper motoneurons from limitless ESCs and offer an otherwise inaccessible tool for studying how human upper motoneurons are born, functionally mature, and become sick. These cells will also provide a target for screening pharmaceuticals that stop the process of motoneuron degeneration and a source for potential future cell therapy for upper motoneuron related diseases.