Abstract
When stretched, cells cultured on 2D substrates share a universal softening and fluidization response that arises from poorly understood remodeling of well-conserved cytoskeletal elements. It is known, however, that the structure and distribution of the cytoskeleton is profoundly influenced by the dimensionality of a cell’s environment. Therefore, in this study we aimed to determine whether cells cultured in a 3D matrix share this softening behavior and to link it to cytoskeletal remodeling. To achieve this, we developed a high-throughput approach to measure the dynamic mechanical properties of cells and allow for sub-cellular imaging within physiologically relevant 3D microtissues. We found that fibroblast, smooth muscle and skeletal muscle microtissues strain softened but did not fluidize, and upon loading cessation, they regained their initial mechanical properties. Furthermore, microtissue prestress decreased with the strain amplitude to maintain a constant mean tension. This adaptation under an auxotonic condition resulted in lengthening. A filamentous actin cytoskeleton was required, and responses were mirrored by changes to actin remodeling rates and visual evidence of stretch-induced actin depolymerization. Our new approach for assessing cell mechanics has linked behaviors seen in 2D cultures to a 3D matrix, and connected remodeling of the cytoskeleton to homeostatic mechanical regulation of tissues.
Highlights
When stretched, cells cultured on 2D substrates share a universal softening and fluidization response that arises from poorly understood remodeling of well-conserved cytoskeletal elements
Within the Microtissue Vacuum-Actuated Stretcher (MVAS)-Force, 3T3 fibroblast cells self-assembled around the cantilevers into dense, highly organized, three-dimensional constructs that morphologically resembled tissue
We presented a new high-throughput approach for assessing both dynamic cell mechanics and for visualization of remodeling at the sub-cellular level in response to stretch within physiologically relevant 3D microtissue cultures
Summary
Cells cultured on 2D substrates share a universal softening and fluidization response that arises from poorly understood remodeling of well-conserved cytoskeletal elements. It is known, that the structure and distribution of the cytoskeleton is profoundly influenced by the dimensionality of a cell’s environment. Heartbeat and movement, cells in our body experience cyclic mechanical stretch, which in turn, creates continually unsteady forces at focal adhesions, across the cell membrane, along cytoskeletal filaments and through the nucleus[1,2] In a cell, these forces direct functional and phenotypic behaviors by generating conformational changes, and thereby, alter ligand-receptor affinities[1,2]. Cells grown on stiff substrates tend to have their actin cytoskeleton arranged into dense stress fibers, and are stiffer, more solid-like and under greater pre-stress when compared to cells on softer substrates[20,21,22]
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