Engineered alveolar organoids to understand ECM signaling

工程化肺泡类器官以了解 ECM 信号传导

• 批准号: 10685523
• 负责人: Claudia Loebel
• 金额: $ 24.9万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10685523
• 关键词: Affect; Alveolar; Alveolar Process; Architecture; Area; Basement membrane; Biochemical; Biological Models; Biomedical Research; Biophysics; Cell Separation; Cell Therapy; Cell physiology; Cells; Chemical Engineering; Cicatrix; Complement; Cues; Deposition; Development; Distal; Engineering; Epithelial Cells; Epithelium; Extracellular Matrix; Feedback; Fibrillar Collagen; Fibroblasts; Fibrosis; Foundations; Functional disorder; Future; Goals; Hydrogels; Impairment; In Situ; In Vitro; Inflammation; Injury; Knowledge; Label; Learning; Ligands; Lung; Measures; Mechanics; Mediating; Mesenchymal; Mesenchyme; Metabolic; Methods; Modeling; Monitor; Myofibroblast; Natural regeneration; Organ; Organoids; Outcome; Paracrine Communication; Pathway interactions; Phase; Population; Process; Proliferating; Property; Regenerative capacity; Research; Research Personnel; Signal Pathway; Signal Transduction; Source; Structure; Structure of parenchyma of lung; System; Techniques; Technology; Testing; Tissue Model; Tissues; Training; Transforming Growth Factors; Variant; Work; alveolar epithelium; career; cellular engineering; crosslink; idiopathic pulmonary fibrosis; in vitro Model; in vivo; intercellular communication; matrigel; mouse model; novel strategies; physical separation; profibrotic cytokine; programs; repaired; reparative process; response; spatial relationship; therapeutic evaluation; therapeutic target; tool

PROJECT ABSTRACT Idiopathic pulmonary fibrosis (IPF) of the distal lung is characterized by spatially heterogeneous areas of fibroblasts/myofibroblasts and accumulation of excess extracellular matrix (ECM) that disrupts the alveolar architecture. Alveolar epithelial cells are now thought to directly contribute to the development and progression of fibrosis, but how new ECM deposition impacts alveolar epithelia function and feedbacks to reinforce fibrotic remodeling is unknown. This is due in part to the lack of tools to study ECM dynamics or to directly measure cell fate in response to altered ECM in vivo. Organoid cultures can generate an in vivo-like complement of alveolar tissue; however, current in vitro models depend on the use of Matrigel matrices, which feature variable compositions and is not conducive to controlled manipulations. The overall focus of the proposed work is to connect changes in secreted alveolar ECM cues and epithelial cell function (K99 phase) and epithelia to mesenchyme signaling (R00). During the K99 phase, defined hydrogel matrices will be developed to form alveolar organoids in a defined microenvironment and determine how newly secreted ECM composition and mechanics guide epithelial cell function. Using this platform, the secreted ECM will be externally crosslinked to examine whether ECM stiffening promotes epithelial cells dysfunction. With an understanding of how secreted ECM initiates epithelial cell dysfunction, the second aim will determine how the secreted ECM alters the response of epithelial cells to signals from the mesenchyme during the R00 phase. We will determine if the accumulation of ECM changes the interaction between AT2 cells and mesenchymal cells, and whether this reinforces fibrotic remodeling. Next, microstructured hydrogels will be used to control for spatial relationships and examine the effect of the physical separation of epithelial and mesenchymal cells on epithelial cell function. To understand the bidirectional signaling of cells and their continuously changing surroundings within the alveolar niche, this research will use organoid cultures and engineering approaches to manipulate and deconstruct cell-ECM interactions. The outcomes will comprise identification of new ECM mediated mechanisms involved in alveolarization and reparative processes, and provide new avenues for testing therapeutics modulating paracrine signaling pathways involved in IPF. Importantly, this proposal comprises a rigorous training plan that will build the foundation to advance the applicant’s career in biomedical research. Specifically, the K99 training will consist of learning mouse models, lineage tracing, and primary cell isolation to establish the foundation towards investigating cell-cell signaling using engineered hydrogels during the independent investigator R00 phase.

项目摘要 远端肺的特发性肺纤维化（IPF）的特点是空间异质区域 成纤维细胞/肌成纤维细胞和过量细胞外基质 (ECM) 的积累会破坏肺泡 建筑学。现在认为肺泡上皮细胞直接促进发育和进展 纤维化，但新的 ECM 沉积如何影响肺泡上皮功能和反馈以强化纤维化 改造未知。部分原因是缺乏研究 ECM 动力学或直接测量细胞的工具 体内 ECM 改变的反应。类器官培养可以产生类似体内的肺泡补体 组织;然而，当前的体外模型依赖于基质胶基质的使用，其特征是变量 成分，不利于受控操作。 拟议工作的总体重点是将分泌性肺泡 ECM 线索的变化与 上皮细胞功能（K99 期）和上皮间质信号转导 (R00)。在K99阶段，定义 将开发水凝胶基质以在定义的微环境中形成肺泡类器官并确定 新分泌的 ECM 成分和机制如何指导上皮细胞功能。利用这个平台， 将分泌的 ECM 进行外部交联，以检查 ECM 硬化是否促进上皮细胞生长 功能障碍。了解分泌型 ECM 如何引发上皮细胞功能障碍后，第二个目标是 确定分泌的 ECM 如何改变上皮细胞对间充质信号的反应 R00 阶段。我们将确定 ECM 的积累是否会改变 AT2 细胞与 间充质细胞，以及这是否会增强纤维化重塑。接下来，将使用微结构水凝胶 控制空间关系并检查上皮和上皮的物理分离的效果 间充质细胞对上皮细胞功能的影响。 了解细胞的双向信号传导及其在细胞内不断变化的环境 肺泡生态位，这项研究将使用类器官培养和工程方法来操纵和 解构细胞-ECM 相互作用。结果将包括识别新的 ECM 介导机制 参与肺泡化和修复过程，并为测试治疗提供新途径 调节 IPF 中涉及的旁分泌信号通路。重要的是，该提案包括严格的培训 计划将为推进申请人在生物医学研究领域的职业生涯奠定基础。具体来说，K99 培训将包括学习小鼠模型、谱系追踪和原代细胞分离，以建立 为在独立研究期间使用工程水凝胶研究细胞信号传导奠定了基础 调查员R00阶段。