Cells react highly sensitive to mechanical and structural influences of their environment by mechanosensing. Micropatterning allows the precise regulation of forces by changing the shape of the cell. In order to record the adaptation to the pattern shape and the changes in viscoelastic properties in detail and over a longer period of time, the VELOMIR sensor is ideally suited. The live cell measurements were recorded at 3000 frames per second over 10 minutes. Here we show the difference in passive microrheology measurements of mouse embryonic fibroblasts with incorporated 1 μm polystyrene beads cultured on three differently patterned shapes in a range of sizes.
Cells actively probe the mechanical properties of their environment and adapt their physiological state accordingly. For this fundamental task, they exert forces on their environment using their actomyosin machinery. This machinery consists of protein fibers consisting of dynamically assembling and dissembling actin units, so called actin stress fibers, on which motor proteins act to generate forces. This machinery is also used to react to external forces applied to cells. Transduction of forces from the actomyosin complexes to the extracellular space occurs via adaptive multi-protein complexes, so called focal adhesions. Focal adhesions re-structure depending on biochemical and mechanical signals. The distance of the extracellular matrix to the actomyosin fibers is determined by the structure of these complexes. Monitoring this distance reveals cellular adaptions to force. Here, we use metal induced energy transfer (MIET) to monitor the distance of the extracellular matrix to focal adhesions for understanding force transduction through focal adhesions. We manipulate the cells either with drugs influencing cellular force generation or using an atomic force microscope. We find that high forces lead to a multistep-restructuring of focal adhesions, rendering force transduction more efficient.
Cells adapt their actin cytoskeletons architecture to structural cues of the environment in all three dimensions. Nevertheless, how manipulating cell shape influences the actin cytoskeletons z-dimension is unstudied, but crucial for an understanding of the mutual influence of cell shape, cell tension and actin architecture. To study the effect of shape on the z-dimension of the actin cytoskeleton we combine metal-induced energy transfer as a super-resolution technique with micropatterning. This allows us not only to precisely manipulate the shape of the cell but also to regulate forces by changing the shape while studying specific actin structures with super-resolution.
Focal adhesions function are cellular anchoring points to the extracellular matrix and enable cells to sense and exert forces on their environment . They are complex structures consisting of a multitude of different proteins. Despite the important role of the focal adhesion complex in cellular adhesion, its structure and mechanoresponse remain difficult to resolve. Knowing the exact position of the proteins in the focal adhesion complex under strain is necessary to understand their working principle. For a detailed analysis of the focal adhesion architecture coupled with force response, we require a method to measure small distances with super resolution precision while manipulating force acting on the cell. To meet this challenge, we couple life-cell atomic force spectroscopy with Metal Induced Energy Transfer (MIET) to resolve forces with pN and with nanometer accuracy. Here, we show an initial analysis of how forces are transduced from the extracellular space to the actin cytoskeleton.
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