Figure 4

Biophysical properties of E and R cells. (a-b) Reactive oxygen species of HCT-8 E and R cells detected by staining with 5-(and-6)-carboxy-2′, 7′-dichlorodihydrofluorescein diacetate. Left: Fluorescent pictures. Right: DIC pictures. Scale bar: 20 μm. (c) Comparison of ROS production in E and R cells. (d) Comparison of basement membrane invasiveness of E and R cells. R is HCT-8 R cells harvested from 20 kPa gels on after 17 days culture. E is HCT-8 cells cultured on PS for 10 days. (e) Phase-contrast photomicrographs of HCT-8 R (top) and E cells (bottom) cultured on PS prior to harvesting and invasion assay. Scale bar: 100 μm. (f) Assembled 4x4 picture set of invaded HCT-8 E cells after 48 hr incubation. Assembled 4x4 picture set of invaded HCT-8 R cells after 48 hr incubation (purple foci indicates invaded cells). (g-h) Comparison of the anchorage independent growth and viability of HCT-8 E (g) and R cells after culture on non-functionalized gels for 8 days. The percentages of viable E and R cells were determined by Trypan-blue staining (i). Red arrows in (g) indicate the dead cells. (j) Atomic force microscopy measurement of stiffness of HCT-8 cells before and after E-R transition. The square-dot and circle-dot curves represent the force vs. indentation displacement of HCT-8 E and R cells, respectively. The data were fitted using an improved Hertz model to extract the Elastic modulus of cells. (j) Comparison of the cell elasticity of E and R cells. (k) Comparison of actin organization in E and R cells on PS. Rhodamine phalloidin (520/650, red) was used specifically stain F-actin filaments. The spatial fluorescent intensity distributions of actin expression across E and R cells body are indicated by dashed lines a and b). Scale bar: 10 μm.