Biomimetic Topological\nMicropattern Arrays Regulate\nthe Heterogeneity of Cellular Fates in Lung Fibroblasts between Fibrosis\nand Invasion

Idiopathic pulmonary fibrosis (IPF) is characterized\nby persistent\ntissue injury, dysregulated wound healing, and extracellular matrix\n(ECM) deposition by myofibroblasts (MFs) through the fibroblast-to-myofibroblast\ntransition (FMT). Implicit in the FMT process are changes in the ECM\nand cellular topology, but their relationship with the lung fibroblast\nphenotype has not been explored. We engineered topological mimetics\nof alignment cues (anisotropy/isotropy) using lung decellularized\nECM micropattern arrays and investigated the effects of cellular topology\non cellular fates in MRC-5 lung fibroblasts. We found that isotropic\nMRC-5 cells presented changes of the cytoskeleton, increased cell–cell\nadhesions and a multicellular architecture with increased overlap,\nchanges in actin–myosin development, and enhanced focal adhesion\nand cell junction with random alignment. Besides, anisotropic fibroblasts\nwere activated into a regular phenotype with an ECM remodeling profile.\nIn contrast, isotropic fibroblasts developed a highly invasive phenotype\nexpressing molecules, including CD274/programmed death-ligand 1 (PD-L1),\ncellular communication network factor 2 (CCN2)/connective tissue growth\nfactor (CTGF), hyaluronan synthase 2 (HAS2), and semaphorin 7A (SEMA7A),\nbut with downregulated matrix genes. Moreover, isotropic fibroblasts\nalso showed higher expressions of Ki-67 and cyclin D1 (CCND1), resistance\nto apoptosis/senescence, and decreased autophagy. The topology regulated\nthe cellular heterogeneity and resulted in positive feedback between\nchanges in the cellular phenotype and the ECM structure, which may\naggravate fibrosis and lead to a priming of malignant microenvironment\nduring carcinogenesis. Using the versatile platform of micropattern\narray, we can not only visualize the interaction mechanism between\ncells and the ECM but also select potential clinical targets for diagnosis\nand therapeutics.