PROJECT SUMMARY Mucociliary clearance (MCC) is a critical mechanical defense mechanism of the human respiratory system. Poor MCC is a fundamental feature of many inherited and acquired respiratory diseases including primary ciliary dyskinesia (PCD), asthma, chronic bronchitis, and cystic fibrosis (CF). Due to the complex organization of the lung, it is largely unknown how defects of the ciliary machinery change functional MCC and how regional variability of airway epithelial structure, including cell type proportions and ciliary beat parameters, affect local MCC pathophysiology. These knowledge gaps dramatically impair our ability to predict the degree of pulmonary dysfunction imparted by specific ciliary defects, and to understand the airway region-specific onset observed in many lung diseases. While impaired MCC is a pre-determined functional consequence of PCD and other chronic lung diseases, to date, there is no established in vitro model that is able to accurately predict the relationship between genotype, cilia motility, MCC, and respiratory phenotype and, therefore, many genotype-phenotype relationships remain unexplained. Our transdisciplinary research program is designed to address an unmet need to understand how a) region-specific airway organization and b) ciliopathy- causing genotypes, impact MCC. To achieve this we will complete specific aims designed to: 1) use ex vivo lung tissues, that retain their in vivo epithelial organization, as models of ciliated airway epithelia to comprehensively evaluate biomechanical structure-function relationships in large and small airways (Aim 1); 2) use established in vitro models of the human tracheo-bronchial (large) airways to determine a minimal set of structural and functional parameters that are conserved between in vitro and ex vivo ciliated tissues (Aim 2); 3) apply state-of-the-art physics-based computational approaches to develop an in silico model that will be able to predict structure-function relationships of ciliated tissues (Aim 3) and 4) use the minimal set of parameters that define functional MCC in both in vitro and ex vivo models as input into the in silico model to predict regional- specific changes in MCC due to ciliary defects in both large and small airways (All Aims). Our specific objectives build to our long-term goal of combining in vitro and in silico models as a preclinical, precision medicine tool for evaluating small molecule or gene-editing therapeutics toward more targeted and efficient treatment regimens of pulmonary diseases characterized by poor mucociliary clearance.