Williams Madeline, Prem Smrithi, Zhou Xiaofeng, Matteson Paul, Yeung Percy Luk, Lu Chi-Wei, Pang Zhiping, Brzustowicz Linda, Millonig James H, Dicicco-Bloom Emanuel
Department of Neuroscience and Cell Biology, Rutgers Robert Wood Johnson Medical School.
Center for Advanced Biotechnology and Medicine, Department of Neuroscience and Cell Biology, Rutgers Robert Wood Johnson Medical School.
J Vis Exp. 2018 Mar 2(133):56628. doi: 10.3791/56628.
Human brain development proceeds through a series of precisely orchestrated processes, with earlier stages distinguished by proliferation, migration, and neurite outgrowth; and later stages characterized by axon/dendrite outgrowth and synapse formation. In neurodevelopmental disorders, often one or more of these processes are disrupted, leading to abnormalities in brain formation and function. With the advent of human induced pluripotent stem cell (hiPSC) technology, researchers now have an abundant supply of human cells that can be differentiated into virtually any cell type, including neurons. These cells can be used to study both normal brain development and disease pathogenesis. A number of protocols using hiPSCs to model neuropsychiatric disease use terminally differentiated neurons or use 3D culture systems termed organoids. While these methods have proven invaluable in studying human disease pathogenesis, there are some drawbacks. Differentiation of hiPSCs into neurons and generation of organoids are lengthy and costly processes that can impact the number of experiments and variables that can be assessed. In addition, while post-mitotic neurons and organoids allow the study of disease-related processes, including dendrite outgrowth and synaptogenesis, they preclude the study of earlier processes like proliferation and migration. In neurodevelopmental disorders, such as autism, abundant genetic and post-mortem evidence indicates defects in early developmental processes. Neural precursor cells (NPCs), a highly proliferative cell population, may be a suitable model in which to ask questions about ontogenetic processes and disease initiation. We now extend methodologies learned from studying development in mouse and rat cortical cultures to human NPCs. The use of NPCs allows us to investigate disease-related phenotypes and define how different variables (e.g., growth factors, drugs) impact developmental processes including proliferation, migration, and differentiation in only a few days. Ultimately, this toolset can be used in a reproducible and high-throughput manner to identify disease-specific mechanisms and phenotypes in neurodevelopmental disorders.
人类大脑发育通过一系列精确编排的过程进行，早期阶段的特征是细胞增殖、迁移和神经突生长；后期阶段则以轴突/树突生长和突触形成为特征。在神经发育障碍中，这些过程中的一个或多个通常会受到干扰，导致大脑形成和功能异常。随着人类诱导多能干细胞（hiPSC）技术的出现，研究人员现在拥有大量的人类细胞，这些细胞几乎可以分化为任何细胞类型，包括神经元。这些细胞可用于研究正常大脑发育和疾病发病机制。许多使用hiPSC来模拟神经精神疾病的方案使用终末分化的神经元或使用称为类器官的3D培养系统。虽然这些方法在研究人类疾病发病机制方面已被证明具有不可估量的价值，但也存在一些缺点。将hiPSC分化为神经元和生成类器官是漫长且成本高昂的过程，这可能会影响可评估的实验数量和变量。此外，虽然有丝分裂后的神经元和类器官允许研究与疾病相关的过程，包括树突生长和突触形成，但它们排除了对增殖和迁移等早期过程的研究。在神经发育障碍，如自闭症中，大量的遗传和死后证据表明早期发育过程存在缺陷。神经前体细胞（NPC）是一种高度增殖的细胞群体，可能是一个合适的模型，用于研究个体发育过程和疾病起始问题。我们现在将从研究小鼠和大鼠皮质培养物中的发育中学到的方法扩展到人类NPC。使用NPC使我们能够在短短几天内研究与疾病相关的表型，并确定不同变量（如生长因子、药物）如何影响包括增殖、迁移和分化在内的发育过程。最终，这个工具集可以以可重复和高通量的方式用于识别神经发育障碍中疾病特异性的机制和表型。
