Investigating pulmonary complications due to abnormal collagen/ER stress in Osteogenesis Imperfecta

研究成骨不全症中胶原蛋白/内质网应激异常引起的肺部并发症

• 批准号: 10556308
• 负责人: Jennifer Zieba
• 金额: $ 39万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-15 至 2028-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10556308
• 关键词: Address; Affect; Apoptosis; Basement membrane; Bleomycin; Bone Diseases; Cell Communication; Cell Differentiation process; Cell physiology; Cell secretion; Cells; Chronic; Clinical; Coculture Techniques; Collagen; Collagen Gene; Collagen Type I; Communication; Defect; Deformity; Development; Disease; Environment; Epithelial Cells; Exhibits; Extracellular Matrix; Fibroblasts; Foundations; Fracture; Frequencies; Functional disorder; Future; Gene Expression; Gene Expression Profile; Genes; Histologic; Homeostasis; Impairment; In Vitro; Infection; Knowledge; Lead; Lung; Lung diseases; Mesenchymal; Methodology; Methods; Modeling; Molecular; Molecular Chaperones; Morbidity - disease rate; Morphology; Mus; Mutation; Myofibroblast; Organoids; Osteoblasts; Osteogenesis Imperfecta; Patients; Perinatal mortality demographics; Phenotype; Population; Population Distributions; Predisposition; Proliferating; Proteins; Pulmonary Valve Insufficiency; RNA; Reaction; Recovery; Reporting; Severities; Signal Pathway; Signal Transduction; Structure; Structure of parenchyma of lung; Therapeutic; Tissues; Validation; Work; alveolar epithelium; bone; bone fragility; cell injury; defined contribution; disability; endoplasmic reticulum stress; environmental stressor; experimental study; improved; in vivo; insight; loss of function mutation; lung development; lung injury; mortality; mouse model; mutant; novel; postnatal; postnatal development; pulmonary function; single-cell RNA sequencing; skeletal

Project Summary Osteogenesis imperfecta (OI) is a genetically heterogenous disorder characterized by increased bone fragility leading to fractures and primarily results from defects in the structure and/or the amount of secreted type I collagen. While bone fragility is the primary cause of morbidity in OI, pulmonary compromise is the leading cause of mortality. We and others showed that in OI there is an abnormal bone extracellular matrix (ECM) structure and type I collagen expressing cells manifest ER stress. Pulmonary mesenchymal derived cells also abundantly express type I collagen. We hypothesize that OI negatively impacts the lung through two mechanisms; secretion of an abnormal ECM and chronic ER stress. Using two OI mouse models representing major forms of OI (missense/loss of function mutations in type I collagen), this proposal will address the hypotheses that mutant type I collagen secretion and ER stress produces abnormal pulmonary morphology, affects lung cell differentiation, impairs lung damage recovery, and that altered ECM and ER stress negatively impact signaling pathways. We address these hypotheses via three aims: 1. Determine the effect of type I collagen mutations on lung postnatal homeostasis and cell differentiation/communication. Hypothesis: Mutations in type I collagen genes lead to alterations in OI lung morphology and cell differentiation. Strategy: Using Aga2 and Col1a1+/- mouse models, the lung will be studied at multiple stages of development via histological/immunohistochemical (IHC) for differentiation and ECM composition. Using in vitro epithelial cell/fibroblast co-culture organoid experiments we will define the contribution of abnormal ECM secretion to lung cell differentiation, proliferation, and apoptosis. 2. Determine the effect of ER stress due to type I collagen mutations on lung cell differentiation, tissue homeostasis, and reaction to damage. Hypothesis: ER stress in pulmonary type I collagen expressing cells affect lung cell differentiation and function. Strategy: Using the models from Aim 1, we will determine ER stress levels in pulmonary cells and whether modulating ER stress in vivo with the chaperone 4-PBA can influence cell differentiation and homeostasis. Using organoid experiments, we will define the contribution of chronic ER stress to lung cell differentiation, proliferation, and apoptosis. To study OI lung damage susceptibility, we will perform in vivo treatment of WT, Aga2, and Col1a1+/- mice with bleomycin to observe the effects of cellular damage on OI lung tissue in conjunction with the 4-PBA treatment; 3. Identify changes in lung cell population distribution and gene expression in the context of an abnormal ECM and ER stress. Hypothesis: Lungs with altered ECM and ER stress affect signaling pathways important in cell differentiation and function. Strategy: Using the Aga2 and Col1a1+/- mouse models, single-cell RNA-seq cells/tissues derived from lung will be performed to identify changes in cell development and gene expression correlated with signaling cascades localized to specific lung cell populations. Completion of these aims will reveal causative mechanisms while introducing novel treatment methods of OI pulmonary dysfunction, the major cause of mortality in OI.

项目摘要 成骨不全是一种以骨脆性增加为特征的遗传异质性疾病 导致骨折并主要由结构缺陷和/或分泌的I型胶原的量引起 胶原虽然骨脆性是OI发病的主要原因，但肺损害是主要原因 死亡率。我们和其他人发现，在OI有一个异常的骨细胞外基质（ECM）结构， 表达I型胶原的细胞表现出ER应激。肺间充质来源的细胞也大量存在 表达I型胶原。我们假设OI通过两种机制对肺产生负面影响：分泌 异常细胞外基质和慢性内质网应激的症状使用两种代表OI主要形式的OI小鼠模型 （I型胶原蛋白中的错义/功能丧失突变），该提案将解决突变体 I型胶原分泌和ER应激导致肺形态异常，影响肺细胞 分化，损害肺损伤恢复，改变ECM和ER应激对信号传导产生负面影响， 途径。我们通过三个目标来解决这些假设：1。确定I型胶原突变对 肺出生后体内平衡和细胞分化/通讯。假设：I型胶原蛋白突变 基因导致OI肺形态和细胞分化的改变。策略：使用Aga 2和Col 1a 1 +/- 在小鼠模型中，将通过组织学/免疫组织化学在多个发育阶段研究肺 (IHC)用于分化和ECM组成。使用体外上皮细胞/成纤维细胞共培养类器官 在实验中，我们将确定异常ECM分泌对肺细胞分化，增殖， 和凋亡。2.确定I型胶原突变引起的ER应激对肺细胞分化的影响， 组织稳态和对损伤的反应。假设：肺I型胶原表达中的ER应激 细胞影响肺细胞分化和功能。策略：使用目标1中的模型，我们将确定ER 肺细胞中的应激水平以及用伴侣蛋白4-PBA在体内调节ER应激是否可以 影响细胞分化和体内平衡。使用类器官实验，我们将定义 慢性内质网应激对肺细胞分化、增殖和凋亡的影响。为了研究OI肺损伤的易感性， 我们将用博来霉素对WT、Aga 2和Col 1a 1 +/-小鼠进行体内治疗，以观察细胞凋亡的影响。 结合4-PBA治疗对OI肺组织的损伤; 3.识别肺细胞群的变化 在异常ECM和ER应激的背景下的分布和基因表达。假设：肺 改变的ECM和ER应激影响在细胞分化和功能中重要的信号传导途径。策略： 使用Aga 2和Col 1a 1 +/-小鼠模型，将从肺衍生的单细胞RNA-seq细胞/组织进行纯化。 用于识别与信号级联相关的细胞发育和基因表达的变化 定位于特定的肺细胞群。完成这些目标将揭示因果机制， 介绍OI肺功能障碍的新治疗方法，这是OI死亡的主要原因。