Mouse lung organoid responses to reduced, increased, and cyclic stretch

This study developed two mechanotransduction models using postnatal day 5 mouse lung organoids (mLOs) generated from EpCAM-positive epithelial cells co-cultured with lung fibroblasts to emulate lung developmental and disease-relevant mechanical environments. First, static perturbations were used to model increased and decreased strain: forskolin-induced swelling increased organoid cross-sectional area (~+59% at 20 h) and increased proliferation (higher PCNA-positive cells), while physical disruption decreased organoid size (~-68% at 20 h) and downregulated lung developmental programs. Second, a biaxial cyclic stretch model was created by embedding individual organoids in Matrigel within a custom silicone insert and applying 20 h of cyclic stretch. Cyclic stretch (5% stretch; 10.25% change in area; 0.1 Hz) drove distinct transcriptomic responses versus static stretch and no stretch, including upregulation of extracellular matrix organization and integrin-mediated signaling and a shift toward mesenchymal lineage-associated gene sets (smooth muscle/fibroblast) with downregulation of epithelial-associated gene sets. Functionally, cyclic stretch increased the proportion of aSMA-positive cells and aSMA/PDGFRa-double-positive cells, supporting a role for cyclic mechanical inputs in promoting mesenchymal maturation programs relevant to postnatal lung development and mechanically altered disease states such as congenital diaphragmatic hernia and fetal tracheal occlusion contexts.