Fig. 2
From: Bimodal sensing of guidance cues in mechanically distinct microenvironments
ECM dimensionality and stiffness influence guidance sensing. a MDA-MB-468 breast carcinoma cells sensing and protruding along uniaxial collagen lines that mimic aligned fibers and biaxial cues to mimic fiber networks (left and right halves of each micrograph, respectively; 3D reconstructions) on soft (2.3 kPa) and stiff (50 kPa) substrates under control (DMSO, Supplementary Movies 1,2,3) and blebbistatin-treated conditions. Blebbistatin-treated cells display a dendritic protrusion phenotype in response to uniaxial (Supplementary Movie 4) and biaxial (Supplementary Movie 5) cues on both soft and stiff substrates, suggesting the presence of a bimodal response to guidance cues that depends on cell traction stress magnitudes. b, c Morphometric analysis of cells on soft and stiff uniaxial guidance cues for control conditions (blue) and blebbistatin treatment (red). For both conditions, uniaxial collagen lines induce a robust guidance response with no significant differences in cell protrusion between soft and stiff substrates. All corresponding n values in c are shown in b. d, e Morphometric analysis of cells interacting with biaxial guidance cues in control (blue) and blebbistatin-treated conditions (red), with significantly greater protrusion on stiff versus soft substrates for control cells. Blebbistatin-treated cells display no detectable difference in their biaxial guidance response on soft and stiff substrates. All corresponding n values in e are shown on d. f, g Morpho-mechanical analysis of cells of soft and stiff substrates showing significantly higher values for both R and R/d metrics in response to stiff substrates, representative of higher cell traction forces in response to stiff substrates. In contrast, soft substrates or inhibition of contractile forces result in low R values and mean R/d values near the threshold for elastic cell behavior. All corresponding n values and individual data points in g are shown in f. h Nuclei deformation, where lateral compression results from cell tension alignment, as a reference for contractile force magnitude. Scale bar in a—15 µm. Corresponding n values are shown on the plots. Number of replicates (independent experiments) for all measurements N = 4. Data in c, e, g, h are mean ± s.d.; ns: no significant difference between groups; *p < 0.05, **p < 0.001 (ANOVA)
