Physical network regimes of 3D fibrillar collagen networks trigger invasive phenotypes of breast cancer cells.

The mechanical characteristics of the extracellular environment are known to significantly influence cancer cell behavior in vivo and in vitro. The structural complexity and viscoelastic dynamics of the extracellular matrix (ECM) pose significant challenges in understanding its impact on cancer cells. Herein, we report distinct regulatory signatures in the invasion of different breast cancer cell lines into three-dimensional (3D) fibrillar collagen networks, caused by systematic modifications of the physical network properties. By reconstituting collagen networks of thin fibrils, we demonstrate that such networks can display network strand flexibility akin to that of synthetic polymer networks, known to exhibit entropic rubber elasticity. This finding contrasts with the predominant description of the mechanics of fibrillar collagen networks by an enthalpic bending elasticity of rod-like fibrils. Mean-squared displacement analysis of free-standing fibrils confirmed a flexible fiber regime in networks of thin fibrils. Furthermore, collagen fibrils in both networks were softened by the adsorption of highly negatively charged sulfonated polymers and colloidal probe force measurements of network elastic modulus again proofed the occurrence of the two different physical network regimes. Our cell assays revealed that the cellular behavior (morphology, clustering, invasiveness, matrix metalloproteinase (MMP) activity) of the 'weakly invasive' MCF-7 and 'highly invasive' MDA-MB-231 breast cancer cell lines is distinctively affected by the physical (enthalpic/entropic) network regime, and cannot be explained by changes of the network elastic modulus, alone. These results highlight an essential pathway, albeit frequently overlooked, how the physical characteristics of fibrillar ECMs affect cellular behavior. Considering the coexistence of diverse physical network regimes of the ECM in vivo, our findings underscore their critical role of ECM's physical network regimes in tumor progression and other cell functions, and moreover emphasize the significance of 3D in vitro collagen network models for quantifying cell responses in both healthy and pathological states.