Dissecting the mechanism of direct neural induction through genomic interrogation

通过基因组询问剖析直接神经诱导的机制

• 批准号: 8047505
• 负责人: Howard Y Chang
• 金额: $ 290.88万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-30 至 2013-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8047505
• 关键词: Action Potentials; Address; Adopted; Adult; Area; Arts; Autologous; Basic Science; Biology; Biomedical Engineering; Biomedical Research; Body Regions; Brain; Brain Diseases; Cell Therapy; Cell Transplantation; Cells; Characteristics; Coculture Techniques; Communities; Complex; Data Set; Disease; Disease model; Electrophysiology (science); Embryo; Ensure; Epigenetic Process; Event; Exhibits; Fibroblasts; Gene Delivery; Gene Expression; Gene Targeting; Generations; Genes; Genetic; Genetic Transcription; Genome; Genomics; Goals; Growth; Heterogeneity; Human; Hypersensitivity skin testing; Immunofluorescence Immunologic; In Vitro; Location; Malignant Neoplasms; Membrane; Methods; Microfluidic Microchips; Modeling; Modification; Molecular; Molecular Profiling; Morphology; Mus; Neurodegenerative Disorders; Neuroglia; Neurons; Oncogenes; Oncogenic; Parkinson Disease; Pathway interactions; Patients; Pattern; Phenotype; Population; Process; Production; Property; Prosencephalon; Publishing; Risk; Safety; Seminal; Skin; Somatic Cell; Synapses; Tail; Techniques; Technology; Testing; Transgenic Organisms; Translating; Translations; Transplantation; Tumor Suppressor Proteins; Tumorigenicity; abstracting; base; brain repair; cancer risk; cell type; clinical application; excitatory neuron; expectation; fetal; genome wide association study; genome-wide; genome-wide analysis; high throughput technology; histone acetyltransferase; histone modification; human subject; induced pluripotent stem cell; inhibitor/antagonist; insight; interdisciplinary approach; mature animal; nervous system disorder; neurogenesis; novel; overexpression; relating to nervous system; research study; stem; stem cell biology; success; tool; trait; transcription factor

DESCRIPTION (provided by applicant): This project covers 3 thematic areas: Applying Genomics and Other High Throughput Technologies, Translating Basic Science Discoveries into New and Better Treatments and Reinvigorating the Biomedical Research Community. Somatic cells are highly stable in adult animals due to robust gene expression patterns, which are stabilized by epigenetic mechanisms. The seminal invention of induced pluripotent stem (iPS) cells, however, provided the surprising conclusion that the differentiated state can be reversed by simple expression of four transcription factors (TFs). This finding proved that even supposedly stable epigenetic modifications of genes are essentially controlled by TFs. We asked whether this concept can be extended to trans-differentiation of one cell type into another, and recently succeeded in converting mouse fibroblasts directly into functional neurons, referred to as induced neuronal (iN) cells, by overexpression of only three lineage-specific TFs. Our findings indicate that TFs suffice to not only reverse a particular pathway of differentiation, but also to redirect the transcriptional regulatory network in a cell into a completely different pathway. This fundamental result answered one of the key open questions in the field, and is the basis of the current proposal. Apart from documenting the dominance of TFs over epigenetic modifications, iN cells could represent an attractive way to derive patient- specific neurons from skin fibroblasts. This may be used to model various neurological diseases or for cell transplantation therapy. This proposal aims to characterize the process of iN cell generation on the molecular level, with the expectation to gain fundamental insights into the biology of the underlying trans-differentiation process. In addition to identifying the molecular events underlying the fibroblast-to-neuron conversion, this study will in particular assess the epigenetic stability of the iN cell state as well as their safety with respect to their potential tumorigenicity, key prerequisites for clinical application of iN cell technologies. Our multidisciplinary approach entails state-of-the art high-throughput sequencing technologies for genome-level interrogation of epigenetic states and transcription, newly developed microfluidic devices enabling genome- wide analyses of small cell populations as well as multiplex gene expression on the single cell level allowing the determination of cellular heterogeneity, electrophysiology, and neurodevelopmental techniques. PUBLIC HEALTH RELEVANCE: This application will develop methods to generate neurons directly from non-neuronal cells, allowing the production of neurons from skin fibroblasts of human patients. Patient-derived neurons could be used for modeling neurological diseases or as cell grafts to treat neurodegenerative diseases like Parkinson's disease.

描述（由申请人提供）：该项目涵盖 3 个主题领域：应用基因组学和其他高通量技术、将基础科学发现转化为新的更好的治疗方法以及重振生物医学研究界。由于强大的基因表达模式，体细胞在成年动物中高度稳定，并通过表观遗传机制稳定。然而，诱导性多能干细胞 (iPS) 的开创性发明提供了令人惊讶的结论：通过四种转录因子 (TF) 的简单表达可以逆转分化状态。这一发现证明，即使是所谓稳定的基因表观遗传修饰本质上也是由转录因子控制的。我们询问这个概念是否可以扩展到一种细胞类型向另一种细胞类型的转分化，并且最近通过仅过表达三种谱系特异性 TF，成功地将小鼠成纤维细胞直接转化为功能性神经元，称为诱导神经元 (iN) 细胞。我们的研究结果表明，转录因子不仅足以逆转特定的分化途径，而且还能将细胞中的转录调控网络重定向到完全不同的途径。这一基本结果回答了该领域的关键开放问题之一，也是当前提案的基础。除了记录转录因子对表观遗传修饰的主导作用外，iN 细胞还可以代表一种从皮肤成纤维细胞中衍生出患者特异性神经元的有吸引力的方法。这可用于模拟各种神经疾病或用于细胞移植治疗。该提案旨在在分子水平上表征 iN 细胞生成过程，期望获得对潜在转分化过程生物学的基本见解。除了确定成纤维细胞向神经元转化的分子事件外，本研究还将特别评估 iN 细胞状态的表观遗传稳定性及其潜在致瘤性的安全性，这是 iN 细胞临床应用的关键先决条件技术。我们的多学科方法需要最先进的高通量测序技术，用于基因组水平的表观遗传状态和转录询问，新开发的微流体装置能够对小细胞群进行全基因组分析，并在单细胞水平上进行多重基因表达允许确定细胞异质性、电生理学和神经发育技术。 公共健康相关性：该申请将开发直接从非神经元细胞产生神经元的方法，从而允许从人类患者的皮肤成纤维细胞产生神经元。源自患者的神经元可用于模拟神经系统疾病或作为细胞移植物来治疗帕金森病等神经退行性疾病。

项目成果

Dissecting direct reprogramming from fibroblast to neuron using single-cell RNA-seq.

使用单细胞 RNA-seq 剖析从成纤维细胞到神经元的直接重编程。

• 发表时间: 2016-06-16
• 影响因子: 64.8
• 作者: Treutlein, Barbara;Lee, Qian Yi;Camp, J Gray;Mall, Moritz;Koh, Winston;Shariati, Seyed Ali Mohammad;Sim, Sopheak;Neff, Norma F;Skotheim, Jan M;Wernig, Marius;Quake, Stephen R
• 通讯作者: Quake, Stephen R

Non-viral Transfection and Direct Reprogramming of Fibroblasts to Neurons and Glia: Importance of Physical and Chemical Microenvironments

成纤维细胞的非病毒转染和直接重编程为神经元和神经胶质细胞：物理和化学微环境的重要性

• 发表时间: 2024-09-13
• 作者: Andrew F Adler