Characterization of stem cell-derived human dopaminergic neuron subtypes

加拿大在运动障碍，特别是帕金森病（PD）的研究方面处于领先地位。作为第二大最常见的神经退行性疾病，帕金森病造成了巨大的经济、社会和个人负担。努力发现治疗PD患者的有效方法是当务之急，研究帮助我们更好地了解PD患者大脑中神经元如何丢失对于达到治疗目标至关重要。长期以来，多巴胺能（DA）神经元一直被认为是PD的影响细胞类型；最近的研究结果表明，这种细胞群比我们以前认为的要复杂得多，实际上DA神经元有6种以上的亚型。其中一些亚型比其他亚型更容易患帕金森病。这是一个具有临床意义的重要发现，但我们目前对DA神经元亚型的了解有限。令人难以置信的是，神经科学研究人员现在能够在实验室中从人类患者身上容易产生的干细胞（称为诱导多能干细胞（iPSCs））中产生人类DA神经元。ipsc衍生的DA神经元已成为模拟PD发病和进展的强大工具，并在大规模筛选实验中发现潜在的治疗化合物。然而，我们目前的iPSC-DA神经元生成协议有很大的局限性。这些问题包括分化效率低下（通常，产生的神经元中只有约30%具有DA身份）以及缺乏关于产生哪些DA神经元亚型的知识。在这个建议中，我们将使用高通量化合物筛选和成像方法来提高ipsc衍生的DA神经元的生产效率，并允许生产的亚群可靠的分层。我们的目标是优化现有的协议，这样研究人员就可以直接从iPSCs中产生DA神经元的单个亚组。该提案将利用以下方面的互补专业知识：1)西蒙弗雷泽大学的Julian和Vocadlo实验室，神经科学、细胞命运决定的生物调节和高通量分析方面的专家；2) Erin Knock博士和她在STEMCELL Technologies的神经科学团队，这是一家总部位于温哥华的国际知名公司，在优化iPSC神经元分化策略方面拥有悠久的专业知识。我们的团队在开展这项工作方面具有独特的优势，我们预计这项工作将对PD和运动障碍研究界产生重大影响。

Canada is a leader in research on movement disorders, especially Parkinson's Disease (PD). As the second most common neurodegenerative condition, PD is responsible for substantial economic, societal, and personal burden. Efforts to uncover effective approaches to treat PD patients are a high priority, and research to help us better understand how neurons are lost in the brains of PD patients is essential to reach therapeutic goals. Dopaminergic (DA) neurons have long been known as the cell type affected in PD; recent findings have revealed that this cell population is more complex than we previously thought and there are actually more than six subtypes of DA neurons. Some of these subtypes are more susceptible than others in PD. This is an important discovery with clinical implications, but our current understanding of DA neuron subtypes is limited.Incredibly, neuroscience researchers are now able to generate human DA neurons in the lab from easy to generate stem cells derived from human patients, termed induced pluripotent stem cells (iPSCs). iPSC-derived DA neurons have served as powerful tools to model the onset and progression of PD, and to uncover potential therapeutic compounds in large-scale screening experiments. There are substantial limitations, however, to our current protocols for iPSC-DA neuron generation. These include inefficient differentiation (typically, only ~30% of neurons generated are of DA identity) and a lack of knowledge as to which DA neuron subtypes are produced. In this proposal we will use high throughput compound screening and imaging methods to improve the efficiency of iPSC-derived DA neuron production and permit reliable stratification of the subgroups produced. Our goal is to optimize existing protocols so researchers can directly produce individual subgroups of DA neurons from iPSCs. This proposal will harness the complementary expertise of: 1) the Julian and Vocadlo labs at Simon Fraser University, experts in neuroscience, biological regulation of cell fate determination and high-throughput analysis; and 2) Dr. Erin Knock and her Neuroscience team at STEMCELL Technologies, an internationally renowned Vancouver-based company with long established expertise in optimization of iPSC neuronal differentiation strategies. Our team is uniquely poised to conduct this work that we anticipate will substantially impact the PD and movement disorder research communities.