Figure 4.
Effect of mechanical resistance on the invasion of MDA-MB-231 cells into Matrigel networks. (A) The leftmost graph shows the average value of permanent deformation for cells invading Matrigel networks of thickness T = 10 (red), 20 (green), and 30 μm (blue). The second graph shows the apparent elastic modulus of the Matrigel encountered by the cells for each gel thickness, Eapp. The nominal elastic modulus of the gel is 400 Pa. Higher values of Eapp of thinner gels reflect an increase in gel stiffness caused by the proximity of the rigid glass coverslip. The last three graphs show the hcrit, ϕ3D,crit, and τzz,crit for cells invading Matrigel networks of varying thickness calculated in the range γ = 0.05–0.2. Values of hcrit, and ϕ3D,crit increase with mechanical resistance while the compressive traction stresses at which the cells switch from elastic to permanent deformation of the matrix,τzz,crit, remain constant. Stars denote statistically significant differences among groups using the Kruskal-Wallis nonparametric analysis of variance (★p < 0.005; ∗p < 0.05). Error bars indicate the 5% confidence interval of the mean (n = 73, 88, and 161 for T = 10, 20, and 30 μm respectively). (B–D) Confocal images of cells invading a 6-μm-thick Matrigel network at ϕ3D ≤ 5° with varying postplating time (30–90 min). At left are three xy images showing F-actin staining (green) and beads (white) within the network, and at right are the corresponding xz sections showing F-actin (green) and MT1-MMP (red) staining. At t = 30 min, plasma membrane blebbing was observed in cells similar to invasion into 30-μm-thick Matrigel (xy plane 1–3). MT1-MMP was detected within the cytoplasm (xz plane). At t = 60 min, the blebs were found to divert along the Matrigel surface, as indicated by arrowheads (xy plane 1–3). MT1-MMP was again detected within the cytoplasm (xz plane). At t = 90 min, cells adopted a spread morphology with the appearance of actin stress fibers at the basal domain of the cell (xy plane 1–3). The MT1-MMP translocated to the cellular periphery (xz plane). (E) The presence of invadopodia is observed through colocalization of cortactin and F-actin at discrete locations. (Insets in the xz-plane images (D and E)) Magnification of the location of invadopodia formation indicated by white arrowheads. Scale bars (both horizontal and vertical), 10 μm. To see this figure in color, go online.