No Cell Left Behind: Using Embryoid Bodies to Understand Human Biology

不遗余力：利用胚胎体来了解人类生物学

基本信息

批准号：
10427990
负责人：
Yoav Gilad
金额：
$ 28.7万
依托单位：
UNIVERSITY OF CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-07-01 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10427990
关键词：
Acute Adult Affect Autopsy Biological Cardiac Myocytes Cell Culture Techniques Cell Line Cells Collection Complex Controlled Environment DNA Data Development Developmental Process Disease Embryo Environment Ethics Event Funding Opportunities Gene Expression Gene Expression Regulation Genes Genetic Genetic Diseases Genetic Variation Genomics Genotype-Tissue Expression Project Germ Layers Goals Hepatocyte Human Human Biology Human body In Vitro Individual Learning Left Maps Measures Modeling Neurons Nucleic Acid Regulatory Sequences Organoids Phenotype Preparation Problem Solving Process Quantitative Trait Loci Regulation Research Design Resources Sample Size Sampling Source Stem cell pluripotency Suggestion System Testing Time Tissues Untranslated RNA Variant cell transformation cell type design differentiation protocol directed differentiation disorder risk experimental study genetic variant genome wide association study human disease human tissue induced pluripotent stem cell inter-individual variation novel strategies power analysis response single cell technology single-cell RNA sequencing stem cell biology stem cell differentiation stem cells temporal measurement theories trait unethical

项目摘要

Abstract This is a revised R21 proposal submitted in response to funding opportunity announcement PA-18-867, “Novel Approaches for Relating Genetic Variation to Function and Disease”. Most of the genetic variants that are associated with disease lie within non-coding DNA and are thought to affect gene regulation. This has inspired efforts to identify variants that affect gene expression levels (eQTLs) in a wide range of adult tissues. However, most disease-associated SNPs – though they are located in putatively regulatory regions – have not been found to be eQTLs. One reason for this could be that despite large-scale efforts to map eQTLs in diverse sets of tissues (e.g, GTEx), we still have not yet examined gene regulation in the cell types or states most relevant for disease. Many human tissues and cell types, especially those that are present in early development, are inaccessible due to practical or ethical constraints. Thus, the pace of genetic discovery is fundamentally limited by access to relevant human tissues. The discovery that mature human cells can be transformed into stem cells was an important step toward solving this problem. Induced pluripotent stem cells (iPSCs) provide a renewable source of human tissue that can, in theory, develop into any cell type. In practice, however, it can take years to discover how to produce any single tissue from iPSCs using directed differentiation. At the nexus of stem cell biology and emerging single-cell technologies, there is an opportunity to generate and study many, or even most, human cell types simultaneously, all within a single dish. When grown in the proper conditions, stem cells form spontaneously differentiating organoids known as embryoid bodies (EBs). Cells within EBs differentiate asynchronously into cell types originating from all three germ layers, including pluripotent, intermediate, and mature cell types. By applying single-cell RNA-sequencing (scRNA-seq) to cells within EBs, we can jointly identify eQTLs across a multitude of cell types, all within a controlled genetic environment. The use of EBs will also allow us to observe cellular transitions and regulatory events that are not evident in static cell culture.

抽象的这是针对资助机会公告 PA-18-867“Novel 将遗传变异与功能和疾病联系起来的方法”。大多数与疾病相关的遗传变异都存在于非编码 DNA 中，并且被认为这激发了人们努力识别影响基因表达水平（eQTL）的变异。然而，大多数与疾病相关的单核苷酸多态性（SNP）存在于多种成人组织中。推定的监管区域——尚未发现 eQTL，原因之一可能是尽管如此。大规模努力在不同的组织中绘制 eQTL（例如 GTEx），但我们仍然没有检查基因与疾病最相关的细胞类型或状态的调节，尤其是许多人体组织和细胞类型。由于实际或道德限制，早期开发中存在的那些是无法实现的。基因发现的步伐从根本上受到相关人体组织获取的限制。成熟的人体细胞可以转化为干细胞的发现是朝着这一目标迈出的重要一步。诱导多能干细胞（iPSC）提供了一种可再生的人体组织来源。理论上，它可以发育成任何细胞类型，但实际上，可能需要数年时间才能发现如何产生。使用定向分化技术从 iPSC 中提取任何单个组织。在干细胞生物学和新兴单细胞技术的结合中，有机会产生和当在适当的培养皿中生长时，可以同时研究多种甚至大多数人类细胞类型。在这种情况下，干细胞会形成自发分化的类器官，称为胚状体（EB）。 EB 异步分化为源自所有三个胚层的细胞类型，包括通过对细胞应用单细胞 RNA 测序 (scRNA-seq) 来识别多能细胞、中间细胞和成熟细胞类型。在 EB 中，我们可以联合识别多种细胞类型的 eQTL，所有这些都在受控的遗传范围内 EB 的使用还将使我们能够观察非如此的细胞转变和调控事件。在静态细胞培养中很明显。