The orientation of extracellular matrix (ECM) fibers, resulting in anisotropic microarchitecture, is ubiquitous in mammalian tissues. This specialized ECM architecture is essential for various biological processes, including tissue organization, wound healing, inflammation, and tumor metastases. The ECM topography regulates cell shape and cell migration by modulating the cell-ECM interaction. In particular, anisotropic ECM governs the directional migration of cells in both normal and pathological conditions. Cells respond to the anisotropy by adjusting their protrusions, aligning the actin filaments, and restricting the focal adhesions along the ECM fibers. In addition, cells regulate their mechanical properties to migrate through confined architecture of the ECM. However, the impact of anisotropic ECM on the cell membrane viscoelasticity in a 3D matrix is less reported.
In this study, we design a 3D collagen matrix that recapitulates the anisotropic ECM and examine the relationship between the anisotropy of the matrix and the anisotropy of the cell membrane. Using unidirectional freezing, we fabricated an array of microchannels (~50 µm channel width) filled with collagen. MCF-7 cells embedded in these collagen matrices display asymmetric morphology and directional migration. Optical tweezer microrheology demonstrates the anisotropic viscoelasticity of the collagen due to confinement. Notably, the collagen matrix elastic modulus is greater along the confinement direction. Next, by attaching tracer particles to the cell membrane, we quantified the cell membrane viscoelasticity under anisotropic confinement. We found that the viscous and elastic modulus of the cell membrane exhibits significant anisotropy. Moreover, the cell membrane is softer under anisotropic confinement despite the local environment being five times stiffer. Finally, to understand the role of the actin cytoskeleton in regulating the membrane viscoelasticity, we performed the OT measurements after actin depolymerization. Our results show that membrane viscoelasticity is unaffected by actin depolymerization for the cells under confinement. Together these findings suggest that ECM anisotropy correlates with cell migration and cell membrane anisotropy in an actin-independent manner.