220 Chapter 9 Hydrogels, which are water-swollen polymeric networks, have been used as an in vitro tool to mimic the 3D organ microenvironment. Synthetic hydrogels, such as those based on dextran [15], and natural hydrogels based on collagen type I [9] have been used for in vitro studies. While synthetic hydrogels provide opportunities to fine-tune the structural arrangement of the fibers and mechanical properties, they lack the biological implications of the altered biochemical microenvironment as found in IPF lungs [16]. Natural hydrogels can provide bioactive cues to cells seeded in them; however, mechanically tuning these hydrogels to mimic a diseased microenvironment is rather limited [17]. Hydrogels made of decellularized organderived ECM can provide an ideal background for addressing these concerns [18]. Decellularized ECMs (dECMs) retain most of the biochemical composition of the native organs and tissues [19], and hydrogels derived from these dECMs have been shown to recapitulate the mechanical properties of their native tissues [18]. Specifically, ECM-derived hydrogels prepared from lung tissue from patients with IPF show the increased stiffness characteristics of native tissue, making these hydrogels an ideal candidate for recapitulating the (fibrotic) microenvironment in vitro [12]. The unexplored interaction between the 3D fibrotic ECM and fibroblasts can therefore be mimicked using such hydrogels to improve our understanding of how the fibrotic response is perpetuated by the feedback from the fibrotic microenvironment itself during IPF. In this study, we hypothesized that an altered microenvironment in fibrotic lungs contributes to perpetuation of fibrosis by inducing profibrotic behavior of lung fibroblasts. To address this, we used an in vitro model using IPF and control lung ECM-derived hydrogels cultured with either IPF or control lung-derived primary fibroblasts in a combined fashion for 7 and 14 days. We investigated the influence of the fibrotic microenvironment on both IPF and control fibroblasts by comparing the ECM remodeling responses of the fibroblasts seeded in IPF and control ECM in 3D. We characterized fibroblast induced changes to the microenvironment with respect to modulation of collagen and glycosaminoglycan (GAG) content, collagen fiber organization and mechanical properties by comparing the fibroblast-originated responses with empty hydrogels.
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