230 Chapter 9 DISCUSSION In this study, we used a 3D in vitro model composed of human lung ECM and human primary lung fibroblasts to assess the influence of the microenvironment on fibroblast responses. We showed that while collagen content and GAG content were unchanged by the fibroblast groups, fiber organization directed by the fibroblasts differed substantially due to influence of the ECM microenvironment. When examining individual collagen fiber characteristics the fibrotic microenvironment did not induce significant changes. However, the global structure of the ECM arrangement, as illustrated by fiber alignment and high density matric proportion was impacted by the nature of the microenvironment. Control fibroblasts did not alter the fiber alignment, while fibrotic fibroblasts increased fiber alignment and this happened to a greater extent in the fibrotic microenvironment. In contrast, control fibroblasts modulated the percentage of high density matrix in a temporal manner in the fibrotic microenvironment, while the IPF fibroblasts reduced the percentage of high density matrix. In addition, control fibroblasts altered the topographical arrangement of the collagen fibers, giving them a greater degree of curvature than that seen with the IPF fibroblasts. The mechanical characteristics of the fibrotic microenvironment were increased by fibroblasts from both control and IPF donors, whereas this change was not seen in the control hydrogels. These findings illustrate that the fibrotic microenvironment imparts a powerful message that drives cellular responses. Overall, our results illustrate that the fibroblast-seeded lung ECM-derived hydrogel model is a powerful in vitro tool for understanding cell interactions with the local microenvironment, and divulging greater knowledge of feedback by the fibrotic ECM and how fibroblasts remodel the microenvironment during this response.
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