Tuesday, June 6, 2017

Spatial distributions of pericellular stiffness in natural extracellular matrices are dependent on cell-mediated proteolysis and contractility

M. Keating, A. Kurup, M. Alvarez-Elizondo, A.J. Levine, E. Botvinick

Bulk tissue stiffness has been correlated with regulation of cellular processes and conversely cells have been shown to remodel their pericellular tissue according to a complex feedback mechanism critical to development, homeostasis, and disease. However, bulk rheological methods mask the dynamics within a heterogeneous fibrous extracellular matrix (ECM) in the region proximal to a cell (pericellular region). Here, we use optical tweezers active microrheology (AMR) to probe the distribution of the complex material response function (α = α′ + α″, in units of µm/nN) within a type I collagen ECM, a biomaterial commonly used in tissue engineering. We discovered cells both elastically and plastically deformed the pericellular material. α′ is wildly heterogeneous, with 1/α′ values spanning three orders of magnitude around a single cell. This was observed in gels having a cell-free 1/α′ of approximately 0.5 nN/µm. We also found that inhibition of cell contractility instantaneously softens the pericellular space and reduces stiffness heterogeneity, suggesting the system was strain hardened and not only plastically remodeled. The remaining regions of high stiffness suggest cellular remodeling of the surrounding matrix. To test this hypothesis, cells were incubated within the type I collagen gel for 24-h in a media containing a broad-spectrum matrix metalloproteinase (MMP) inhibitor. While pericellular material maintained stiffness asymmetry, stiffness magnitudes were reduced. Dual inhibition demonstrates that the combination of MMP activity and contractility is necessary to establish the pericellular stiffness landscape. This heterogeneity in stiffness suggests the distribution of pericellular stiffness, and not bulk stiffness alone, must be considered in the study of cell-ECM interactions and design of complex biomaterial scaffolds.

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