Abstract
Cell rigidity sensing—a basic cellular process allowing cells to adapt to mechanical cues—involves cell capabilities exerting force on the extracellular environment. In vivo, cells are exposed to multi-scaled heterogeneities in the mechanical properties of the surroundings. Here, we investigate whether cells are able to sense micron-scaled stiffness textures by measuring the forces they transmit to the extracellular matrix. To this end, we propose an efficient photochemistry of polyacrylamide hydrogels to design micron-scale stiffness patterns with kPa/µm gradients. Additionally, we propose an original protocol for the surface coating of adhesion proteins, which allows tuning the surface density from fully coupled to fully independent of the stiffness pattern. This evidences that cells pull on their surroundings by adjusting the level of stress to the micron-scaled stiffness. This conclusion was achieved through improvements in the traction force microscopy technique, e.g., adapting to substrates with a non-uniform stiffness and achieving a submicron resolution thanks to the implementation of a pyramidal optical flow algorithm. These developments provide tools for enhancing the current understanding of the contribution of stiffness alterations in many pathologies, including cancer.
Highlights
In order to address cell stiffness sensing at subcellular scales, we designed culture supports with micron-scaled stiffness patterns based on the formulation of an innovative fast curing, water-based, photosensitive solution of acrylamide/bis acrylamide precursors (Figure 1)
As the reticulation process occurred in a water-based solution, the diffusion of the monomers and the photoinitiator contributed to the final reticulation [44]
In order to resolve these micron-scaled modulations of the rigidity, we developed an algorithm that could detect the fluorescent markers embedded in the hydrogel with a submicron resolution
Summary
Many technologies have arisen that build soft environments for cell cultures. Their aim is either to mimic the in-vivo mechanical environment [4] or to enhance a specific cellular response, such as cell differentiation using cell mechanosensitivity [5]. To this end, soft hydrogels or elastomers have been developed with specific mechanical properties to support 2D or 3D cell culture. Polyacrylamide may be replaced by Nanomaterials 2022, 12, 648.
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