A developmental engineering toolbox for large-scale tissue engineering

用于大规模组织工程的发育工程工具箱

• 批准号: 10456084
• 负责人: Alex Hughes
• 金额: $ 39.81万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10456084
• 关键词: 3-Dimensional; Address; Adhesives; Affect; Breast; CRISPR screen; Cells; Chronic Kidney Failure; Complex; Congenital Abnormality; DNA; Development; Developmental Process; Disease; Duct (organ) structure; Engineering; Extracellular Matrix; Gel; Homeostasis; In Vitro; Kidney; Kidney Diseases; Location; Lung; Methods; Modeling; Morphogenesis; Organ; Organoids; Pattern; Pharmacotherapy; Phenotype; Philosophy; Pluripotent Stem Cells; Positioning Attribute; Prostate; Renal function; Research Personnel; Resolution; Seeds; Shapes; Standardization; Structure; System; Technology; Tissue Engineering; Tissue Model; Tissues; Urinary tract; Variant; base; body system; cell type; collecting tubule structure; genetic risk factor; human disease; human tissue; tissue support frame; whole genome

PROJECT SUMMARY Many diseases in complex, hierarchically organized tissues such as the breast, lung, and prostate have been difficult to address, because they are a product of complex multicellular dynamics. For example, congenital diseases of the kidney are staggeringly common. Around a third of all birth defects are associated with problems in kidney and urinary tract development, but researchers have few options for capturing the full functional complexity of this organ system outside of the body. This is because current kidney models are either 2D, single cell-type approximations, or are organoid models with more cellular diversity, but with little of the long-range spatial structure that is crucial for kidney function. The Hughes lab aims to solve two critical engineering barriers to the development of better in vitro human tissue models. First, we aim to standardize and vastly increase the throughput of organoid- based phenotypic screens related to human disease. Second, we aim to bring an entirely new philosophy to tissue engineering, in which tissue scaffolds are not built in final form, but rather as immature “seeds” that are guided through developmental transitions in structure that mimic those of their target tissue. These transitions morph flat tissue scaffolds into final tissue forms that achieve defined shapes, cell distributions, and ECM compaction and alignment patterns in 3D that establish a new way of building hierarchical tissues like the kidney. To the first aim, we propose to re-engineer our cell DNA “velcro” cell and organoid patterning technology. This technology allows us to precisely pattern multiple cell types with single-cell resolution at the interface with organotypic gel layers, yet its throughput is currently limited. We will apply a photopatterning approach in which cell-adhesive ssDNA strands can be patterned in millions of locations simultaneously, a key requisite for whole-genome organoid screens. Secondly, we propose high-throughput pluripotent stem cell patterning and culture technologies that reduce inter-organoid variation, to enable whole genome CRISPR- based screening for genetic risk factors of disease, using kidney organoids as a prototypical system. To the second aim, we build upon our recent description of dynamic tissue scaffolds to position organoids in 3D using autonomously folding gels that couple their niches through tracts of dynamically remodeled ECM. Using these centimeter-scale, 3D organoid patterning capabilities, we envision an analogy between the branching pattern of the kidney collecting duct network and the edge networks of “flasher” origami patterns. By controlling the morphogenesis of these patterns, we seek to engineer the progressive formation of a contiguous collecting duct network between locally self-organizing tissue niches. Rather than directly building tissues in a final, yet immature form, we believe that building hierarchical tissues by guided morphogenesis presents a transformative opportunity for modeling tissue homeostasis and disease.

项目摘要 复杂的，分层组织的组织中的许多疾病，例如乳房，肺和前列腺 difficalt增加了，因为它们是复杂的多细胞动力学的产物 肾脏的疾病非常普遍。 肾脏和尿道发育中的问题，但是研究人员几乎没有选择可以捕获。 身体外部的器官系统的功能复杂性是因为当前的肾脏模型是 2D，单细胞类型的近似值，或者是具有更多细胞多样性的器官模型，但很少 对于肾功能至关重要的远程空间结构。 休斯实验室旨在解决两个关键的工程障碍，以开发更好 体外人体组织模型。 基于与人类疾病有关的表型筛选。 组织工程，其中组织支架不是最终形式的，而是不成熟的“种子” 指导着靶向组织的结构中的艰难发育过渡 将平坦的组织支架变成最终组织形式，以实现定义的形状，细胞分布和ECM 3D中的压实和对齐方式建立了一种新的构建等级组织的方式 肾。 为了第一个目的，我们建议重新设计我们的细胞DNA“ velcro” Cellcro“ Cellcro”细胞和类器官图案 技术。 目前，我们将使用器官凝胶层的接口，我们将应用一个照相。 在数百万个位置，可以将细胞粘附性s​​sDNA链图案化的方法 全基因组器官筛选的必要条件。 减少甲基机间变异的图案和文化技术，以使整个基因组CRISPR- 基于疾病的筛查因素，使用肾脏器官作为原型系统 第二个目的，我们以最新的描述动态组织支架来定位器官 3D使用自主折叠凝胶，其小甲基菌群进行动态重塑的ECM。 使用这些厘米尺度的3D器官图案化能力，我们想象在您之间的类比 肾脏收集尘埃网络和“闪光灯”折纸的边缘网络。 控制这些模式的形态发生，我们试图设计 在局部自组织的组织中，连续收集的管道网络而不是直接 以最终但不成熟的形式建造组织，我们认为通过指导构建分层组织 形态发生提出了对组织稳态和疾病进行建模的变革性统一性。