Increased Stiffness Inhibits Invadopodia Formation and Cell Migration in 3D

Abstract

Cancer cells typically invade through basement membranes (BMs) at key points during metastasis, including primary tumor invasion, intravasation, and extravasation. Cells extend invadopodia protrusions to create channels in the nanoporous BM through which they can invade, either via proteolytic degradation or mechanical force. Increased matrix stiffness can promote cancer progression, and two-dimensional (2D) culture studies indicate that increased stiffness promotes invadopodia degradation activity. However, invadopodia can function mechanically, independent of their degradative activity, and cells do not form fully matured invadopodia or migrate in the direction of the invadopodia in 2D environments. Here, we elucidated the impact of matrix stiffness on the mechanical mode of invadopodia activity of cancer cells cultured in three-dimensional BM-like matrices. Invadopodia formation and cell migration assays were performed for invasive breast cancer cells cultured in mechanically plastic, nanoporous, and minimally degradable interpenetrating networks of reconstituted BM matrix and alginate, which presented a range of elastic moduli from 0.4 to 9.3 kPa. Across this entire range of stiffness, we find that cells form mature invadopodia that often precede migration in the direction of the protrusion. However, at higher stiffness, cells form shorter and more transient invadopodia and are less likely to extend invadopodia overall, contrasting with results from 2D studies. Subsequently, cell migration is diminished in stiff environments. Thus, although previous studies indicate that increased stiffness may promote malignant phenotypes and the degradative activity of invadopodia, our findings show that increased stiffness physically restricts invadopodia extension and cell migration in three-dimensional, BM-like environments.

Recursive feedback between matrix dissipation and chemo-mechanical signaling drives oscillatory growth of cancer cell invadopodia

“…The heatmap indicates that the invadopodia extension length decreases as elastic modulus increases (purple arrow in Figure 4H) due to the larger resistance encountered by invadopodia in stiffer ECMs. Interestingly, by utilizing the same type of IPN hydrogels of varying elastic stiffnesses (low stiffness [LS], medium stiffness [MS], high stiffness [HS], and very high stiffness [VHS]; see STAR Methods) (Chang et al, 2020), we found that cells have shorter invadopodia lengths in stiffer ECMs (Figures 4F and 4G). By varying the model parameter associated with stiffness (i.e., elastic modulus E 0 ), we can quantitatively fit the model to the experimental data.…”

“…Alginate molecular weight (MW) and cross-linker concentration were varied in order to form a set of IPNs with initial elastic moduli of around 1.8 kPa but varying mechanical plasticity (Wisdom et al, 2018). In addition, calcium crosslinking of the alginate matrix was varied to form a set of IPNs with different elastic moduli, i.e., low stiffness 0.4 kPa (LS), medium stiffness 1.5 kPa (MS), high stiffness 4.4 kPa (HS), and very high stiffness 9.3 kPa (VHS) (Chang et al, 2020).…”

“…Poorly segmented cells and partially imaged cells at frame edges were excluded. Further details can be referred to one previously published work (Chang et al, 2020). Objected-oriented bounding box lengths for each cell was exported from Imaris and analyzed with custom MATLAB code, and three box lengths a; b and c (note aRbRc) for three principal axes can be obtained (Figure S5A).…”

Controlled Fabrication of Bioactive Microtubes for Screening Anti-Tongue Squamous Cell Migration Drugs

“…It is time to improve traditional cell culture to better simulate the complex environment in vivo (Pampaloni et al, 2007;Duval et al, 2017). The 3D cell culture models have emerged for studying cancer cell invasion so far, which better imitates the internal environment (Chung et al, 2009;Engel et al, 2015;Wagner et al, 2019;Chang et al, 2020). Due to the variability of morphology and the excellent biocompatibility, hydrogels are widely used in many kinds of research about cell migration in 3D environments (Sugimoto et al, 2018).…”

Cell Migration Regulated by Spatially Controlled Stiffness inside Composition‐Tunable Three‐Dimensional Dextran Hydrogels

“…[31,30,32] Chang et al showed that increased stiffness physically restricted invadopodia extension and cell migration in 3D, which presented inconsistent tendency compared to the results in 2D. [33] Vasudevan et al proposed biomimetic ECMs with independently tunable stiffness to demonstrate the distinct cell migratory patterns due to differences in matrix stiffness. [29] Hadden et al fabricated polyacrylamide hydrogels with graded stiffness to study how human adipose-derived stem cells performed in such environments, and the results showed that stem cells tended to move toward stiffer regions.…”