Cell shape-dependent early responses of fibroblasts to cyclic strain

Abstract

Randomly spread fibroblasts on fibronectin-coated elastomeric membranes respond to cyclic strain by a varying degree of focal adhesion assembly and actin reorganization. We speculated that the individual shape of the cells, which is linked to cytoskeletal structure and pre-stress, might tune these integrin-dependent mechanotransduction events. To this aim, fibronectin circles, squares and rectangles of identical surface area (2000μm(2)) were micro-contact printed onto elastomeric substrates. Fibroblasts plated on these patterns occupied the corresponding shapes. Cyclic 10% equibiaxial strain was applied to patterned cells for 30min, and changes in cytoskeleton and cell-matrix adhesions were quantified after fluorescence staining. After strain, megakaryocytic leukemia-1 protein translocated to the nucleus in most cells, indicating efficient RhoA activation independently of cell shape. However, circular and square cells (with radial symmetry) showed a significantly greater increase in the number of actin stress fibers and vinculin-positive focal adhesions after cyclic strain than rectangular (bipolar) cells of identical size. Conversely, cyclic strain induced larger changes in pY397-FAK positive focal complexes and zyxin relocation from focal adhesions to stress fibers in bipolar compared to symmetric cells. Thus, radially symmetric cells responded to cyclic strain with a larger increase in assembly, whereas bipolar cells reacted with more pronounced reorganization of actin stress fibers and matrix contacts. We conclude that integrin-mediated responses to external mechanical strain are differentially modulated in cells that have the same spreading area but different geometries, and do not only depend on mere cell size.

Using SERS Tags to Image the Three‐Dimensional Structure of Complex Cell Models

“…The use of unlabeled filler rHDF cells allowed us to achieve improved resolution, and hence measurements in the z‐axis, of the different labels (Figures 4B and S12, Supporting Information). We determined the average height of one layer of rHDF cells to be approximately 5 µm (Figure S12, Supporting Information), which is in agreement with published studies 55. However, with an increasing number of rHDF layers, the detected fluorescence signal decreased and a sufficient signal for quantification of fluorescence at the desired resolution could only be observed by increasing the laser power with increasing sample depth (Figure 4C, Supporting Information).…”

“…In addition to the individual cellular morphology, many cell types, including fibroblasts, have been shown to display considerable macro‐scale morphological plasticity, depending on the geometry of their encapsulating structures 55,56. In this context, we studied the effect of culture well geometry on the overall 3D model obtained using rHDF cells.…”

Controlling Cell Geometry Affects the Spatial Distribution of Load Across Vinculin

“…The manipulation of cell geometry, such as confining to circular or square patterns independently of cell spread area, is known to affect many cellular processes, including stem cell differentiation, 31 gene expression, 28 and the response of cells to applied loads. 22,51 Sensing of cell shape is thought to be mostly mediated through mechanically-sensitive signaling within focal adhesions (FAs), the structures that physically link the force generating actin-myosin cytoskeleton to the extracellular matrix. 25 These structures are comprised of transmembrane proteins, called integrins which bind to dense plaques containing more than 180 proteins, the top layer of which is bound to the actinmyosin cytoskeleton.…”

Agarose‐Assisted Micro‐Contact Printing for High‐Quality Biomolecular Micro‐Patterns

“…But most of all, this micropatterning technique has been highlighted because of its ability to pattern biomolecules, thus enabling the design of cellular microenvironments in biological experiments and tissue engineering applications. The geometric constraints confined by micropatterns could influence cellular morphology, differentiation, migration, proliferation, and development . In addition, mimicking biological environment between different types of cells has been also achieved to facilitate the investigation of cell–cell interactions in cultures .…”