Synthetic organogenesis: new paradigms in reconstituting human organ development in vitro

合成器官发生：体外重建人体器官发育的新范例

• 批准号: 10622903
• 负责人: Mijo SIMUNOVIC
• 金额: $ 18.02万
• 依托单位: COLUMBIA UNIV NEW YORK MORNINGSIDE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10622903
• 关键词: 3-Dimensional; Biochemical; Biology; Biomechanics; Blood Vessels; CRISPR screen; Cells; Characteristics; Chemicals; Clustered Regularly Interspaced Short Palindromic Repeats; Coculture Techniques; Colon; Complex; Coupling; Development; Embryo; Embryonic Structures; Endoderm; Endothelial Cells; Ensure; Environment; Feedback; Fetal Development; Gastrocoele; Germ Layers; Goals; Human; Human Development; In Vitro; Light; Maps; Mechanics; Microfluidics; Modeling; Molecular; Morphogenesis; Morphology; Organ; Organogenesis; Organoids; Patients; Pattern; Process; Property; Protocols documentation; Reproducibility; Resolution; Role; SHH gene; Series; Signal Transduction; Source; Structure; System; Techniques; Thymus Gland; Tissue Engineering; Tissues; Transcriptional Regulation; Tube; Work; angiogenesis; bone morphogenetic protein 4; design; fetal; genome editing; human model; human pluripotent stem cell; human stem cells; human tissue; in vitro Model; in vivo; innovative technologies; live cell microscopy; mechanical signal; morphogens; neglect; organ growth; parent grant; pluripotency; programs; rational design; reconstitution; self organization; spatiotemporal; stem cell biology; transcriptomics

PROJECT SUMMARY Organogenesis is a process in which biochemical signals and mechanical cues transform embryonic germ layers into organs during fetal development. Recent advances in stem cell biology created opportunities to differenti- ating human pluripotent stem cells into expandable 3D tissues that contain many of the cellular and functional characteristics of fetal organs, which we call organoids. They hold a tremendous potential to answer longstand- ing questions of human development and one day to serve as a renewable source of patient-specific tissues. However, organoids still only approximatively recapitulate organogenesis as they rely on spontaneous tissue self-organization and the establishment of signaling gradients in unpredictable ways. Common protocols also neglect the contribution of surrounding developing organs, especially from other germ layers, and because they lack the vasculature, they are not scalable. This supplement proposal aims precisely to advance these gaps in the current organoid paradigm; namely, by generating organoids from all three germ layers and by mimicking in vitro angiogenesis to increase their scalability. We focus on the organogenesis of the gut tube, the embryonic structure on which many adjacent organs form, and whose axial patterning strongly depends on the signaling feedback between all three germ layers. We propose to employ microfluidics to generate precise signaling gra- dients so to reproducibly mimic both the signaling and the morphogenesis of gut tube organogenesis in vitro. We will generate carefully assembled components of the primitive gut tube comprising all three germ layers, then expose them to signaling gradients in a highly controlled mechanical environment. Combined with live- cell microscopy, CRISPR editing, and single cell transcriptomics, our multi-organoids will reveal a detailed hi- erarchy of fate choices cells make from pluripotency to regionally segmented gut tube, and they will allow us to make crucial connections between signaling, transcriptional regulation, and tissue morphogenesis. Our work will shed light on the largely unknown regulatory mechanisms by which the three germ layers communicate to generate complex and asymmetric patterns along the body axes.

项目摘要 器官发生是一个生化信号和机械信号改变胚胎胚层的过程 在胎儿发育过程中进入器官。干细胞生物学的最新进展创造了分化的机会， 将人类多能干细胞培养成可扩展的3D组织， 胎儿器官的特征，我们称之为类器官它们拥有巨大的潜力来回答长期以来- 解决人类发展的问题，并有一天成为患者特定组织的可再生来源。 然而，类器官仍然只是近似重演器官发生，因为它们依赖于自发组织 自组织和以不可预测的方式建立信号梯度。通用协议还 忽视周围发育器官的贡献，特别是来自其他胚层的器官，因为它们 缺乏血管系统，它们无法扩展。这一补充提案的目的正是为了在以下方面弥补这些差距： 目前的类器官范例;即通过从所有三个胚层产生类器官，并通过模仿 体外血管生成以增加其可扩展性。我们专注于肠管的器官发生，胚胎 许多相邻器官形成的结构，其轴向图案强烈依赖于信号传导 三个胚层之间的反馈我们建议使用微流体来产生精确的信号gra- 因此，在体外可重复地模拟肠管器官发生的信号传导和形态发生。 我们将产生由三个胚层组成的原始肠管的精心组装的组件， 然后将它们暴露在高度受控的机械环境中的信号梯度中。结合现场- 细胞显微镜，CRISPR编辑和单细胞转录组学，我们的多类器官将揭示一个详细的高， 细胞从多能性到区域分割的肠管的命运选择的多样性，它们将允许我们 在信号传导、转录调控和组织形态发生之间建立起至关重要的联系。我们的工作 将揭示三个种系相互交流的基本上未知的调节机制 沿着身体轴线产生复杂和不对称的图案。