IPF ECM-Derived Hydrogels Promote the Inflammatory Cytokine Expression of Cultured IPF Fibroblasts and Trigger an Inflammatory and Proliferative Immune Cell Profile

Idiopathic pulmonary fibrosis (IPF) is a chronic disease that causes scarring of lung tissue, leading to a reduction in oxygen transport. At the cellular level, IPF is characterized by overproduction and accumulation of extracellular matrix (ECM) secreted by fibroblasts. Previous work has shown that culturing fibroblasts on stiff 2D surfaces can alter gene expression in both IPF fibroblasts (IPF-F) and normal human lung fibroblasts (NHLF), but fail to recapitulate key signatures of the fibrotic microenvironment in vivo. This compromises their usefulness as model systems in which to interrogate IPF biology and test candidate therapeutics.

To better understand fibroblast biology in IPF, we developed a 3D hydrogel model derived from human fibrotic lung tissue that allows us to study the effects of fibroblast activation and cytokine expression without the confounding influence of mechanosignaling from stiff plastic substrates. Hydrogels were generated from lung tissue via decellularization, lyophilization, pulverization, and digestion of scaffold prior to forming gels. 2D culture served as control. Fibroblast 3D hydrogel cultures were assayed for contractility, imaged via optical microscopy, and analyzed with ImageJ. Conditioned media was collected from fibroblast cultures and used to treat monocytes and macrophages. Gene expression was assayed via quantitative PCR, and protein expression was assayed via Western Blot. The mechanical properties of hydrogels were characterized with rheometry and atomic force microscopy and correlated with contractility of the IPF-F when cultured in our ECM hydrogels, as compared to NHLF. Our results broadly suggest that hydrogels recapitulate the increased inflammatory immune cells as a result of the fibrotic ECM in vitro model and altered mechanical properties in the IPF disease state.

In conclusion, our hydrogels drive expression of inflammatory cytokines that are lost in standard 2D culture. These observations reinforce the need for improved in vitro model systems that retain environmental cues in the IPF lung. Our hydrogel platform will serve as a robust in vitro platform for further investigations of fundamental issues underlying the progression of IPF.