Synthetic hydrogels to study formation and maintenance of intestinal crypts

用于研究肠隐窝的形成和维持的合成水凝胶

• 批准号: 9981736
• 负责人: KRISTI S. ANSETH
• 金额: $ 40.17万
• 依托单位: UNIVERSITY OF COLORADO
• 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 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9981736
• 关键词: 3-Dimensional; Ablation; Actomyosin; Address; Adult; Architecture; Basement membrane; Biological; Biological Assay; Cell Differentiation process; Cell Lineage; Cell Proliferation; Cell Shape; Cell physiology; Cells; Complex; Coupled; Cues; Development; Developmental Process; Disease; Dose; Doxorubicin; Drug Delivery Systems; Drug Screening; Encapsulated; Enterocytes; Epithelial; Epithelial Cells; Epithelium; Event; Excision; Genetic; Growth; Hydrogels; In Vitro; Injury; Intestinal Neoplasms; Intestines; Investigation; Kinetics; LGR5 gene; Laser Scanning Confocal Microscopy; Lead; Light; Link; Maintenance; Mechanics; Mediating; Methods; Modeling; Modification; Morphogenesis; Mus; Natural regeneration; Nuclear; Organ; Organoids; Outcome; Paneth Cells; Peptide Hydrolases; Pharmacology; Phenotype; Physiological; Play; Process; Proliferating; Property; Reaction; Relaxation; Research; Role; Shapes; Signal Transduction; Stress; Structure; System; Testing; Time; Tissue Transplantation; Variant; Verteporfin; Villus; Withdrawal; blebbistatin; cell type; clinical translation; crypt cell; drug candidate; drug efficacy; inhibitor/antagonist; innovation; intestinal crypt; intestinal epithelium; matrigel; mechanical properties; mechanotransduction; notch protein; progenitor; reconstitution; response; response to injury; sarcoma; scaffold; self organization; spatiotemporal; stem; stem cell population; stem cells; stemness; tool; viscoelasticity

ABSTRACT Intestinal organoid models hold great promise as a tool to study intestinal development and disease, screen drug candidates, or even produce transplantable tissue in vitro. Current culture methods for growth of intestinal organoids rely almost exclusively on Matrigel, but Matrigel’s loosely-defined and variable composition makes clinical translation nearly impossible and obstructs fundamental investigations into the role of key matrix factors on organoid formation. While intestinal stem cells (ISCs) grown in Matrigel have a tremendous capacity for self- organization into functionally sophisticated intestinal organoid structures, the self-organization principles are also responsible for introducing variability and stochastic organoids that also differ from the native organ in multiple aspects. In the proposed research, we aim to develop tunable hydrogel matrices for ISC expansion, colony formation, and differentiation to form crypts. Unique to our materials is the ability to regulate the ISC microenvironment spatiotemporally using photochemical reactions, and we propose to use photoadaptable hydrogels to test hypotheses related to ISC mechanosensing and its effects on organoid growth (Aim 1); the role of local matrix stiffness on organoid shape, cell proliferation, and crypt formation (Aim 2); and the plasticity of crypt cells during their response to a stress or injury (Aim 3). We hypothesize that exogenous control of matrix properties can be used to support efficient ISC organoid growth, and subsequently mimic cell-mediated crypt formation and remodeling. The proposed material systems will allow us to not only study and direct the formation of the crypt-villus architectures that are physiologically relevant, but also test maintenance of these structures in response to dynamic changes in matrix properties corresponding to developmental processes, as well as crypt regeneration after injury. Specifically, we propose to: 1. Investigate the role of matrix mechanical properties and signaling on intestinal stem cells (ISCs) and their growth into spherical organoids. 2. Understand how spatial changes in hydrogel mechanics permit ISCs to undergo progenitor commitment and subsequent differentiation into functional cell types. and 3. Investigate the role of uniform and spatially variant cell-matrix interactions on the de-differentiation of lineage specific epithelial cells and crypt regeneration after injury.

抽象的 肠道类器官模型作为研究肠发展和疾病的工具，筛查药物有很大的希望 候选者，甚至在体外产生了可移植的组织。肠道生长的当前培养方法 类器官几乎完全依赖于Matrigel，但Matrigel的零星和可变成分使得 临床翻译几乎是不可能的，并且阻碍了对关键基质因子的作用的基本研究 在器官形成上。肠道中生长的肠道干细胞（ISC）具有巨大的自我能力 组织为功能复杂的肠道器官结构，自组织原理也是 负责引入可变性和随机器官，这些器官在多个中也与天然器官不同 方面。在拟议的研究中，我们旨在为ISC扩展，菌落开发可调水凝胶材料 形成和形成隐窝的区分。我们材料独有的是调节ISC的能力 微环境在空间上使用光化学反应，我们建议使用光化 水凝胶测试与ISC机制有关的假设及其对器官生长的影响（AIM 1）；角色 在器官形状，细胞增殖和加密形成上的局部基质刚度（AIM 2）；和可塑性 隐窝对压力或损伤的反应期间（AIM 3）。我们假设对基质的外源控制 特性可用于支持有效的ISC器官生长，然后模仿细胞介导的加密货币 形成和重塑。拟议的材料系统将使我们不仅可以研究和指导形成 在物理上相关的地下村架构，但也测试了这些结构的维护 对与发育过程相对应的矩阵属性的动态变化的响应以及加密 受伤后再生。具体而言，我们提出：1。研究基质机械性能的作用 并在肠道干细胞（ISC）及其生长到球形器官上发出信号。 2。了解如何 水凝胶力学的空间变化允许ISC进行祖细胞承诺，然后 分化为功能细胞类型。和3。研究均匀和空间变体细胞矩阵的作用 损伤后谱系特异性上皮细胞和加密再生的脱不同的相互作用。