Development of forebrain organoid platform for modelling human cortical neurogenesis

 描述(申请人提供)：在过去的十年中，干细胞领域的一项重大突破是将人类体细胞重新编程为诱导多能干细胞或IPSCs的技术的发展。除了能够从培养皿中的人IPSCs获得特定的细胞类型作为2D培养外，该领域的快速进步还使从类似于整个发育中的器官(包括肠道、肾脏、视网膜和大脑皮层)的IPSCs产生3D培养或有机类物质成为可能。人类器官类化合物提供了一个独特的机会，可以在类似于人体器官发生的培养系统中模拟器官发育。此外，器官培养提供了对影响多种细胞类型的疾病进行建模和研究非细胞自主效应的机会。我实验室的工作重点是使用小鼠模型和IPSC模型进行神经发育；我们的长期目标是了解正常大脑发育和神经发育疾病的潜在机制，并帮助开发合理的治疗策略。尽管脑有机化合物在模拟脑发生和脑疾病方面有着巨大的希望，但目前可用的技术存在几个主要限制，包括高成本、低重复性和高变异性，这些限制了我们进行定量分析和广泛应用 技术我们最近开发了一种新的方法，通过将用于产生脑有机体的关键组件微型化，从而大大减少了材料、细胞培养介质、空间和成本。在这个探索性项目中，我们建议进一步标准化前脑特定有机物质的生产，并优化细胞培养条件，以实现定向和持续的生长。作为原则的证明，我们将使用这个系统来测试15q11.2微缺失导致癫痫易感性的皮质神经发生异常的假设。15q11.2微缺失是癫痫的一个重要遗传危险因素。我们相信，我们的方法将改变器官发生模型，并有助于识别与疾病相关的生物学过程，这些过程在2D单层培养中很难概括。

 DESCRIPTION (provided by applicant): A major breakthrough in the stem cell field over the last decade has been the development of technology to reprogram human somatic cells into induced pluripotent stem cells or iPSCs. In addition to the ability to derive specific cell types from human iPSCs in dish as 2D cultures, rapid progress in the field has made it possible to generate 3D cultures, or organoids, from iPSCs resembling whole developing organs, including intestinal, kidney, retinal, and cerebral cortex. Human organoids provide a unique opportunity to model organ development in a culture system that is similar to human organogenesis in vivo. Furthermore, organoid cultures provide the opportunity to model diseases that affect multiple cell types and to investigate non-cell- autonomous effects. The work in my laboratory focuses on neural development using both mouse models and iPSC models; and our long term goal is to understand mechanisms underlying normal brain development, neurodevelopmental diseases and to aid in the development of rational therapeutic strategies. Despite the tremendous promise of cerebral organoids to model brain genesis and brain diseases, there are several major limitations of the currently available technology, including high cost, low reproducibility and high variability, that limit our ability for quantitative analyses and broad application of the technology. We have recently developed a new approach by miniaturizing the critical components used to generate cerebral organoids, which allows for a dramatic reduction in materials, cell culture media, space and costs. In this exploratory project, we propose to further standardize forebrain specific organoid production and optimize cell culture conditions for directed and sustained growth. As a proof-of-principle, we will use this system to test the hypothesis that 15q11.2 microdeletion, a prominent genetic risk factor for epilepsy, leads to aberrant cortical neurogenesis for seizure susceptibility. We believe that our approach will transform organogenesis modeling and facilitate the identification of disease-relevant biological processes that are difficult to recapitulate in 2D monolayer cultures.