Spatial and Single Cell Transcriptomics Approach to Understand Neuron-Oligodendrocyte Communication in Human Synaptic Development

了解人类突触发育中神经元-少突胶质细胞通讯的空间和单细胞转录组学方法

• 批准号: 10646970
• 负责人: Hiroko Nobuta
• 金额: $ 20.93万
• 依托单位: RUTGERS BIOMEDICAL AND HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-15 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10646970
• 关键词: Ablation; Action Potentials; Adherent Culture; Antibodies; Architecture; Area; Bar Codes; Biological Assay; Biology; Brain; Cell Culture Techniques; Cell Maturation; Cells; Central Nervous System; Cerebrum; Coculture Techniques; Communication; Communities; Complementary DNA; Data; Data Set; Development; Diphtheria Toxin; Disease; Electrophysiology (science); Formulation; Foundations; Funding; Future; Generations; Genes; Grant; Histologic; Histology; Human; In Vitro; Institutional Review Boards; Intellectual functioning disability; Knowledge; Live Birth; Maps; Measurement; Mediating; Methods; Molecular; Morphology; Multiple Sclerosis; Mus; Myelin; Names; Nervous System Physiology; Neuroglia; Neurologic Dysfunctions; Neurons; Oligodendroglia; Ontology; Pathology; Patients; Pregnancy; Premature Birth; Premature Infant; Protocols documentation; Regulation; Reporting; Resolution; Rodent; Rodent Model; Role; Secure; Signal Transduction; Slice; Slide; Source; Synapses; System; THBS2 gene; Testing; Tissue Donors; Tissues; Universities; Ventricular; WNT5A gene; WNT7A gene; brain tissue; clinical translation; drug candidate; drug testing; effective therapy; experimental study; fetus cell; genetic signature; human tissue; leukodystrophy; neuron development; neuropsychiatric disorder; novel; novel therapeutics; oligodendrocyte precursor; postsynaptic; precursor cell; presynaptic; single-cell RNA sequencing; spatial relationship; synaptic function; transcriptomic profiling; transcriptomics; transmission process; white matter injury

ABSTRACT Oligodendrocytes are the myelinating glia cells of the central nervous system allowing coordinated conduction of action potentials among neurons, thus imperative for proper neurological functions. The vast majority of our knowledge in oligodendrocytes is derived from rodent models. Perhaps for that reason, clinical translation of our knowledge has been limited, leaving many disorders without treatment. In order to understand the fundamental human oligodendrocyte development, pathology, and ultimately to discover drug candidates with higher confidence, therefore, it is necessary to develop protocols in human-based platform. Single-cell RNA sequencing comparing mouse and human oligodendrocyte precursor cells (OPCs) revealed a compelling unique sub- population in human OPCs that does not seem to exist in mouse OPCs. More importantly, gene ontology analysis of this sub-population identified highly expressed signature genes associated with synaptic development, organization, and transmission, suggesting neuron-oligodendrocyte communication during neuronal development. To our knowledge, there are no reports suggested a direct synaptic control by OPCs. Based on these observations, we hypothesize that a sub-population of human OPCs regulates synaptic development. This proposal intends to establish concrete evidence for human-specific OPC sub-population through two pilot experiments to prepare for a future external funding focusing on understanding the molecular mechanisms of human synaptic development controlled by OPCs. First, to define temporal and spatial interaction between human specific OPC sub-population and neurons, we will generate spatial transcriptomic profiling of developing human brain spanning gestational week 10 to 24, when dynamic synaptic development as well as OPC maturation occur. Spatial transcriptomics uses an intact tissue section mounted on a slide that is coated with arrays of barcoded RT primers to create spatially barcoded cDNA, allowing us to obtain transcriptomic data from the entire tissue retaining spatial information. These data will reveal where the OPC sub-population exists in developing brains and what type(s) of neighboring neurons they are interacting with. Second, we will begin to test the function of human specific OPC sub-population using quantitative electrophysiology in a co-culture and slice culture formats that we have optimized. Once achieved, these studies will lay the foundation to study a novel function of human OPC sub-population. Generated transcriptomic datasets will allow us to formulate testable hypotheses for future external grants. In a larger perspective, discovery of a new mechanism in synaptic control may lead to a re-formulation of the pathomechanisms in diseases with suspected involvement of oligodendrocytes, such as intellectual disabilities and neuropsychiatric disorders.

摘要 少突胶质细胞是中枢神经系统的髓鞘胶质细胞，具有协调传导功能。 神经元之间的动作电位的变化，因此对于正常的神经功能是必不可少的。我们的绝大多数人 少突胶质细胞的知识来源于啮齿动物模型。也许正是因为这个原因，我们的临床翻译 知识有限，导致许多疾病得不到治疗。为了更好地理解 人少突胶质细胞的发育、病理，最终发现的候选药物具有较高的 因此，有必要在以人为本的平台上开发协议。单细胞RNA测序 比较小鼠和人的少突胶质前体细胞(OPC)发现了一个引人注目的独特的亚细胞。 人类OPC中似乎不存在于小鼠OPC中的种群。更重要的是，基因本体论分析 在这个亚群中，发现了与突触发育相关的高表达特征基因， 组织和传递，提示神经元-少突胶质细胞在神经元中的通讯 发展。据我们所知，没有报道表明OPC直接控制突触。基于 在这些观察中，我们假设人类OPC的一个亚群调节突触的发育。这 该提案打算通过两个试点为特定于人类的OPC亚群建立具体证据 为未来的外部资金做准备的实验，重点是了解分子机制 由OPC控制的人类突触发育。首先，界定时间和空间的相互作用 人类特定的OPC亚群和神经元，我们将生成发育中的空间转录图谱 人脑跨越妊娠10至24周，此时突触动态发育以及OPC 成熟发生了。空间转录学使用安装在玻片上的完整组织切片，玻片上覆盖有 条形码RT引物的阵列，以创建空间条形码的cDNA，使我们能够从 整个组织都保留了空间信息。这些数据将揭示OPC亚群体存在的位置 发育中的大脑以及他们正在与哪种类型的邻近神经元(S)相互作用。第二，我们将开始 用定量电生理学检测人类特定OPC亚群在共培养中的功能 我们已经优化的切片文化格式。一旦实现，这些研究将为研究 人类OPC亚群的新功能。生成的转录数据集将使我们能够 未来外部赠款的可检验假设。从更大的角度来看，突触中一种新机制的发现 控制可能导致重新表述疑似涉及 少突胶质细胞，如智力障碍和神经精神障碍。