Microengineered alveolar array lung-on-chip with hydrogel membrane and simulated breathing mechanics for anti-fibrotic testing

We present a precision-engineered lung-on-chip platform that replicates the biomechanical and structural features of the human alveolar microenvironment for respiratory disease modeling and therapeutic evaluation. At the core of the device is a thin, suspended hydrogel membrane composed of biologically relevant collagen and elastin, engineered to mimic the dimensions and mechanical fragility of the native alveolar basement membrane. This membrane supports a geometrically defined array of alveolar units, each capable of undergoing finely controlled, physiologically relevant deflections under cyclic mechanical actuation-emulating the subtle deformations that occur during human breathing. To address the challenges posed by the membrane's mechanical fragility and the requirement for accurately controlled micron-scale deflections, the platform is fabricated using precision injection molding. This manufacturing strategy ensures structural integrity and reproducibility, creating a rigid support structure around the suspended hydrogel membrane. The design is integrated into a SBS microwell plate format, facilitating robust fluidic interfacing, consistent cyclic actuation, and medium-throughput operation. Human alveolar epithelial cells and lung fibroblasts are co-cultured on a membrane and subjected to cyclic biomechanical stress that mimics respiratory movements. We demonstrate that cyclic stretching significantly amplifies fibrotic signaling in the presence of transforming growth factor-beta 1 (TGF-<i>β</i>1), evidenced by increased expression of extracellular matrix (ECM) components such as collagen I, collagen III, and fibronectin. Treatment with the anti-fibrotic drug nintedanib reduced expression of ECM proteins and plasminogen activator inhibitor-1 (PAI-1), validating the system's utility for pharmacological testing. This alveolar array-based lung-on-chip system bridges a critical gap between conventional<i>in vitro</i>models and the physiological complexity of human lung tissue, offering a robust platform for mechanistic studies and preclinical evaluation in pulmonary fibrosis and related disorders.