Mapping the cytoskeletal prestress

Abstract

Cell mechanical properties on a whole cell basis have been widely studied, whereas local intracellular variations have been less well characterized and are poorly understood. To fill this gap, here we provide detailed intracellular maps of regional cytoskeleton (CSK) stiffness, loss tangent, and rate of structural rearrangements, as well as their relationships to the underlying regional F-actin density and the local cytoskeletal prestress. In the human airway smooth muscle cell, we used micropatterning to minimize geometric variation. We measured the local cell stiffness and loss tangent with optical magnetic twisting cytometry and the local rate of CSK remodeling with spontaneous displacements of a CSK-bound bead. We also measured traction distributions with traction microscopy and cell geometry with atomic force microscopy. On the basis of these experimental observations, we used finite element methods to map for the first time the regional distribution of intracellular prestress. Compared with the cell center or edges, cell corners were systematically stiffer and more fluidlike and supported higher traction forces, and at the same time had slower remodeling dynamics. Local remodeling dynamics had a close inverse relationship with local cell stiffness. The principal finding, however, is that systematic regional variations of CSK stiffness correlated only poorly with regional F-actin density but strongly and linearly with the regional prestress. Taken together, these findings in the intact cell comprise the most comprehensive characterization to date of regional variations of cytoskeletal mechanical properties and their determinants.

Local mechanical response in semiflexible polymer networks subjected to an axisymmetric prestress

“…Also marked on this figure are Also shown in the inset to Fig. 3 is the same data plotted on linear axes, demonstrating approximate proportionality between prestress and stiffness, which has also been observed in isolated smooth muscle cells [28]. We note, however, that there is a slight upturn to the model predictions when crossing from linear prestress τ 0 2 c /κπ …”

Mapping intracellular mechanics on micropatterned substrates

“…Our technique allows to measure cell-averaged parameters that can be compared with those obtained with more classical approaches, but also reveals the spatial variations of intracellular mechanics. Previous work has either studied local mechanics without controlling the cell geometry and thus with large cell-tocell variability (22,33,34) or measured regional rigidity variations of the cell surface in controlled geometries using AFM or magnetic twisting cytometry (28,37,38). Central to our method is the use of adhesive micropatterns to standardize the intracellular organization.…”

“…Recently experiments with atomic force microscopy (AFM) (34) and AFM or magnetic twisting cytometry combined with micropatterning (37,38) have been performed but did not discriminate between the mechanical contribution of the prestressed cell cortex and that of the cell interior. In contrast, our approach isolates the contribution of the cell interior.…”

Micropipette Aspiration of Substrate-attached Cells to Estimate Cell Stiffness

“…In contrast to AFM that measures local membrane rigidity, microaspiration provides a global measure of the deformability of a cell. Other methods to estimate cellular biomechanical properties include various bead/particle approaches, such as magnetic twisting cytometry [31][32][33] , particle tracking 34,35 ; (ii) Particle tracking can be used for several different purposes, such as estimating the force that cells exert on the substrate (Traction Force Microscopy) 34 or estimate the stiffness of the internal "deep" layers of the cell by analyzing the motion of the beads or organelles inside the cell 35 . In contrast, optical tweezesrs are used to pull membrane nanotubes (tethers) to estimate cortical tension and membrane-cytoskeleton adhesion 37 .…”