Improving Brain Organoid Models by Mediating Metabolic Dysregulation

通过调节代谢失调改善脑类器官模型

• 批准号: 10509424
• 负责人: Madeline Andrews
• 金额: $ 24.9万
• 依托单位: ARIZONA STATE UNIVERSITY-TEMPE CAMPUS
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-18 至 2024-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10509424
• 关键词: Address; Adopted; Alzheimer' s Disease; Animal Model; Benchmarking; Biological Assay; Biological Models; Blood Vessels; Brain; Cell Differentiation process; Cell Line; Cell Transplantation; Cells; Cellular Stress; Cerebral cortex; Characteristics; Chemicals; Complex; Culture Media; Data; Data Set; Defect; Derivation procedure; Development; Disease; Ensure; Environment; Excision; Gene Expression; Genes; Glucose; Glycolysis; Goals; Growth; Human; Immune; Impairment; In Vitro; Instruction; Knowledge; LIF gene; Laboratories; Link; Mediating; Metabolic; Metabolic stress; Metabolism; Methods; Modeling; Molecular; Molecular Profiling; Morphology; Mus; Neurodegenerative Disorders; Neuroglia; Neurons; Neurophysiology - biologic function; Neurosciences; Normal Cell; Organoids; Oxygen; Parkinson Disease; Pathway interactions; Phenotype; Physiological; Play; Pluripotent Stem Cells; Population; Process; Proteins; Protocols documentation; Proxy; Publishing; Regulation; Reporting; Role; Signal Pathway; Signaling Molecule; Specificity; Stem Cell Development; Stress; Transplantation; Vulnerable Populations; Work; base; cell cortex; cell type; cognitive capacity; endoplasmic reticulum stress; gene network; genetic signature; gliogenesis; human model; improved; in vivo; nervous system disorder; neurodevelopment; neurogenesis; organoid transplantation; prevent; programs; relating to nervous system; response; single-cell RNA sequencing; small molecule; stem cells; tool; translational study; validation studies

Project Summary There is currently an unmet need for accurate model systems of the human brain to study its cellular and molecular features. The cerebral cortex regulates our cognitive capacity, yet the cellular diversity, circuit formation, and function that establish this potential, largely remains a mystery. The cortex is expanded in humans compared to other species; it contains more cellular diversity and abundance, making model organisms limited for translational studies. Brain organoids provide access to human cells for experimental manipulation and recent studies have suggested that organoid models may act as a proxy for the human brain when studying developmental trajectories, circuitry, and a variety of complex neurological diseases. However, using single cell RNA sequencing we performed validation studies of cortical organoid models compared to primary human cortex cells and we discovered several significant deficiencies in organoid cells. First, organoids do not make the same refined cell types with clear gene expression programs observed during normal cortex development. They also lack molecular maturation networks observed endogenously. As organoid cells do not make the specific, refined cell types we observe in the human brain, these models are severely limited when aiming to model circuit activity and disease. Circuits are comprised of highly specialized neuronal subtypes that target one another to wire defined connections and ultimately produce appropriate physiological activity. We require organoid models that make specific cell types that can mature enough to form the building blocks of canonical circuits to enable the study of neural connectivity moving forward. In addition to cell type deficiencies, all organoids assayed, regardless of derivation method, express high levels of metabolic stress. Using transplantation studies, we identified that cellular stress is linked to the in vitro environment, it negatively impacts cell type identity, and it can be reversed by removal from culture conditions. My proposed studies will directly address the current technical limitations of organoid models by intervening in the metabolic dysregulation with the goal of improving cell type specification and maturation. First, I will target glycolytic and endoplasmic reticulum stress pathways, using small molecules to inhibit these processes and by modulating glucose and oxygen concentrations. I will also utilize knowledge of relevant signaling pathways in human cortex development, such as the LIF signaling pathway, which is an important regulator of a human-enriched population of cortical stem cells. Second, I will work to identify the relevant non-neural cells required for normal metabolic regulation endogenously, which are absent from organoid models. To work toward this aim I will use transplantation of human organoid-derived cells into the mouse cortex environment and transplant relevant vascular and immune cells into cortical organoids in vitro. Together, these studies will improve current technical limitations in metabolism and cell type impairment to ensure that organoids more accurately reflect the cellular diversity of the human cortex and provide a tractable model system for the study of human neuroscience.

项目摘要 目前对人脑的精确模型系统的需求尚未得到满足，以研究其细胞和 分子特征。大脑皮层调节我们的认知能力，但细胞多样性，电路 建立这种潜力的形成和功能在很大程度上仍然是一个谜。人类的大脑皮层扩张 与其他物种相比，它含有更多的细胞多样性和丰富性，使模式生物受到限制 进行翻译研究。脑有机化合物为实验操作和最近的人类细胞提供了通道 研究表明，有机体模型在研究人类大脑时可能会起到替代作用 发育轨迹、环路和各种复杂的神经疾病。然而，使用单个电池 RNA测序：我们对皮质器官模型进行了验证研究，并与原始人类皮质进行了比较 细胞，我们发现类器官细胞中有几个明显的缺陷。首先，有机化合物不会产生同样的效果。 在正常皮质发育过程中观察到的具有清晰基因表达程序的精致细胞类型。他们也 内源性观察到缺乏分子成熟网络。由于类器官细胞不会使特定的、精致的 我们在人脑中观察到的细胞类型，这些模型在模拟电路活动时受到严重限制 和疾病。电路由高度专门化的神经元亚型组成，它们相互瞄准并连线。 明确的连接并最终产生适当的生理活动。我们需要有机类模型 使特定的细胞类型足够成熟，以形成规范电路的构建块，从而使 神经连通性的研究进展。除了细胞类型缺陷外，所有的有机物都被检测出来了， 无论采用何种派生方法，都表现出高水平的代谢应激。通过移植研究，我们 确认细胞应激与体外环境有关，它会对细胞类型认同产生负面影响，而且它 可以通过从培养条件中移除来逆转。我提议的研究将直接解决当前 通过干预代谢失调来改善有机物模型的技术局限性 细胞类型规范和成熟。首先，我将针对糖酵解和内质网应激途径， 使用小分子来抑制这些过程，并通过调节葡萄糖和氧气的浓度。这就做 也利用人类皮质发育中的相关信号通路的知识，如LIF信号 途径，这是一个重要的调节人类丰富的皮质干细胞群体。第二，我会 努力确定正常代谢调节所需的相关非神经细胞，这些细胞是 没有出现在器官模型中。为了达到这个目标，我将使用人类器官来源的细胞移植 将相关的血管和免疫细胞移植到皮质类器官中 体外培养。总之，这些研究将改善目前在新陈代谢和细胞类型损伤方面的技术限制，以 确保有机化合物更准确地反映人类大脑皮层的细胞多样性，并提供易于处理的 人类神经科学研究的模型系统。