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.

Project Narrative Pulmonary complications are the leading cause of mortality in patients with osteogenesis imperfecta (OI) and the mechanisms behind these complications, specifically reaction to environmental damage, are unknown and not well explored. This proposal will gain insights into the mechanisms causing OI lung insufficiency and completion of the aims will contribute to knowledge regarding the effect of abnormal extracellular matrix secretion and ER stress on lung cell differentiation and damage recovery as well as the effect of in vivo ER stress inhibition treatment. Importantly, our study will identify novel targets for future clinical therapies that could ameliorate OI lung abnormalities, for which there are presently no established treatments.