Mammary fibroblasts remodel fibrillar collagen microstructure in a biomimetic nanocomposite hydrogel.

Architecture and microstructure of type I collagen fibers constitute central regulators of tumor invasion with aligned fibers providing a route for migration of stromal and cancer cells. Several different aspects of fibrillar collagen, such as stiffness, density, thickness, and pore size, may regulate migration of cancer cells, but determining effects of any one parameter requires clear decoupling of physical properties of collagen networks. The objective of this work is to develop and apply an in vitro three-dimensional (3D) tumor-extra cellular matrix (ECM) model with tunable physical parameters to define how stromal fibroblasts modulate collagen microstructure to control migration of breast cancer cells. We incorporated two different types of polyhedral oligomeric silsesquioxane (POSS) nano-molecules into a collagen/alginate matrix to induce different mechanisms of gelling. The resultant biomimetic, nanocomposite hydrogels show different collagen fibrillar microstructures while maintaining constant overall matrix stiffness, density, and porosimetry. Spheroids of human mammary fibroblasts embedded in these 3D matrices remodel the collagen network to varying extents based on differences in underlying matrix microstructures. The remodeled collagen matrix shows oriented, thicker fibrillar tracks, facilitating invasion of tumor cells. By decoupling effects of matrix stiffness and architecture, our nanocomposite hydrogels serve as robust platforms to investigate how biophysical properties of tumor environments control key processes regulating tumor progression in breast cancer and other malignancies. STATEMENT OF SIGNIFICANCE: Our manuscript demonstrates a new type of nanocomposite hydrogel with two different gelling mechanisms, produced by incorporating two types of polyhedral oligomeric silsesquioxane (POSS) nano-molecules into a collagen/alginate matrix. The resultant biomimetic hydrogels show different fibrillar collagen microstructures while maintaining constant overall matrix stiffness, density, and porosimetry. These gels allow us to uncouple effects of matrix stiffness versus architecture on migration and invasion of breast cancer cells and stromal fibroblasts. Upon embedding spheroids of human mammary fibroblasts (HMFs) and dissociated 231 breast cancer cells, we showed that HMFs remodeled the collagen network to differing extents dependent on starting matrix microstructures in each hydrogel. The remodeled collagen matrix showed aligned collagen fibers perpendicular to the surface of a spheroid with migrating HMFs following these fibers as occurs in tumors in vivo. To our knowledge, this is the first study showing significant different fibrillar collagen microstructures with constant collagen density and gel stiffness. This study establishes a new type of nanocomposite 3D hydrogels for studies of biophysical and cellular interactions in engineered tumor environments.