Morphological adaptations in breast cancer cells as a function of prolonged passaging on compliant substrates

Abstract

Standard tissue culture practices involve propagating cells on tissue culture polystyrene (TCP) dishes, which are flat, 2-dimensional (2D) and orders of magnitude stiffer than most tissues in the body. Such simplified conditions lead to phenotypical cell changes and altered cell behaviors. Hence, much research has been focused on developing novel biomaterials and culture conditions that more closely emulate in vivo cell microenvironments. In particular, biomaterial stiffness has emerged as a key property that greatly affects cell behaviors such as adhesion, morphology, proliferation and motility among others. Here we ask whether cells that have been conditioned to TCP, would still show significant dependence on substrate stiffness if they are first pre-adapted to a more physiologically relevant environment. We used two commonly utilized breast cancer cell lines, namely MDA-MB-231 and MCF-7, and examined the effect of prolonged cell culturing on polyacrylamide substrates of varying compliance. We followed changes in cell adhesion, proliferation, shape factor, spreading area and spreading rate. After pre-adaptation, we noted diminished differences in cell behaviors when comparing between soft (1 kPa) and stiff (103 kPa) gels as well as rigid TCP control. Prolonged culturing of cells on complaint substrates further influenced responses of pre-adapted cells when transferred back to TCP. Our results have implications for the study of stiffness-dependent cell behaviors and indicate that cell pre-adaptation to the substrate needs consideration.

Matrix stiffness-sensitive long noncoding RNA NEAT1 seeded paraspeckles in cancer cells

“…Analysis of the aspect ratio (major axis/minor axis) revealed no changes in this parameter when cells were grown on different stiffness, with the exception being a slight increase in the aspect ratio for the two metastatic cell lines 143B and MDA-MB-231, when cultured on stiff 40 kPa hydrogels compared with soft 3 kPa hydrogels ( Figure 3 D). Consistent with our findings, previous work reported that breast cancer cells grown on stiff substrates may appear more elongated in shape, potentially assisting epithelial-to-mesenchymal transition and thus promoting metastasis ( Syed et al. , 2017 ).…”

Adipose stem cells exhibit mechanical memory and reduce fibrotic contracture in a rat elbow injury model

“…40 Additionally, investigations evaluating mechanical memory in breast cancer cell lines (eg, MDA-MD-231, MCF-10A) showed that soft priming decreased proliferation rate and migration speed compared to cells cultured only on stiff substrates. 54,55 Thus, some cellular functions exhibit mechanical memory. In the current study, the effect of mechanical memory on ASC activity was evaluated by the quantification of individual cell ECM production and cytokine secretion.…”

Measuring microenvironment-tuned nuclear stiffness of cancer cells with atomic force microscopy

“…Cells cultured on these ECM-coated PA gels are used for AFM based stiffness measurement. The underlying ECM-coated PA gel mimics the bulk stiffness of the microenvironment and can be tuned accordingly using different concentrations of acrylamide and bis-acrylamide ( Syed et al., 2017 ; Tse and Engler, 2010 ). For our experiments, cells were grown on collagen-coated ~4.5 kPa PA gels and nuclear stiffness was measured by probing live cells in the culture media using an AFM cantilever.…”

“… Timing: 2 h Acrylamide/bis-acrylamide stock solution preparation: Prepare a stock solution of acrylamide and bis-acrylamide in ultra-pure (UP) water. PA gels of varying stiffness can be prepared by mixing different concentrations of acrylamide and bis-acrylamide as per previous publications ( Syed et al., 2017 ; Tse and Engler, 2010 ). In our studies, we have mixed 1.25 mL of 40% acrylamide solution and 0.75 mL of 2% (w/v) bis-acrylamide in 8 mL of UP water that produces a stiffness ~4.5 kPa ( Tse and Engler, 2010 ).…”