Cancer-associated fibroblasts are major architects of the tumour stroma, where their aligned, elongated morphology forms a capsule that mechanically restrains tumour expansion. However, it is unclear how this supracellular organization emerges and persists. Here we show that fibroblasts generate a fibronectin matrix that progressively acquires the same nematic order as the cell layer, and that this matrix in turn feeds back to immobilize both cells and topological defects. Using long-term live imaging, traction force microscopy, matrix microfabrication and hydrodynamic modelling, we find that this reciprocal coupling induces an ageing process in which cellular flows and defect motion slow dramatically and ultimately freeze. Despite this arrest, the monolayer remains active, with defects concentrating contractile forces that may represent mechanical weak points. Disrupting fibronectin production fluidizes the capsule, reactivates defect dynamics and compromises its barrier-like function. These findings reveal a self-organizing mechanism by which fibroblasts and their matrix co-evolve to create a mechanically stable, yet active, stromal architecture with direct implications for tumour dissemination.

Epithelial tissues maintain organ integrity while continuously remodeling during morphogenesis, repair, and disease. At high cell densities, these tissues often appear mechanically arrested in a disordered, solid-like state, raising the question of how they retain the ability to reorganize. Here, we show that, unlike thermal glasses, dense epithelial tissues do not exhibit caging behavior but instead behave as a complex fluid. Cells display subdiffusive creep together with Fickian yet non-Gaussian dynamics and compressed exponential relaxation, hallmarks of stress-driven fluidity. This fluidity arises from the tissue’s structural and mechanical organization rather than from cell division or extrusion, which only transiently enhance local dynamics. Fast-moving cells organize into collective, anisotropic clusters whose spatial heterogeneity correlates with local structural entropy and soft vibrational modes. Together, these findings reveal a hidden fluidity in densely packed epithelia that supports mechanical stability while preserving the capacity for remodeling during development, wound healing, and early tumor invasion.

Intracellular transport of macromolecules is crucial for the proper functioning of most cellular processes. Although intracellular crowding is known to strongly alter macromolecule mobility, how cytoplasmic structures physically modulate diffusion remains largely unexplored. Here, we investigated the mechanisms by which cytoplasmic crowding controls diffusivity using live-cell experiments and porous media modeling approaches. Confocal microscopy combined with fluorescence recovery after photobleaching and fluorescence correlation spectroscopy measurements revealed an anticorrelation between free green fluorescent protein diffusivity and the heterogeneous cytoplasmic structure abundance in live mammalian cells. This motivated the development of a multiscale model, where the cytoplasm is treated as a hierarchical porous medium with nanometric and micrometric obstacles. Numerically solving the model allowed us to predict the effective cytoplasmic diffusion coefficient for various obstacle volume fractions and to identify tortuous and porous hydrodynamic hindrances as key diffusion reduction mechanisms. Comparison with our experimental results highlighted the importance of hydrodynamic interactions between diffusing molecules and nanometric obstacles. Importantly, we found that the effective cytoplasmic diffusivity was not dependent on specific intracellular regions but rather on the local intracellular obstacle volume fraction. Finally, the model was extended to predict the diffusivity of larger macromolecules, showing excellent agreement with literature data for several macromolecules and cell lines. This study provides insights into the physical mechanisms impeding intracellular diffusion, demonstrating the potential of porous media modeling approaches to predict transport mechanisms in dynamic or heterogeneous intracellular structures, as in cell motility, blebbing, and apoptosis.

Cells within biological tissue are constantly subjected to dynamic mechanical forces. Measuring the internal stress of tissues has proven crucial for our understanding of the role of mechanical forces in fundamental biological processes like morphogenesis, collective migration, cell division, or cell elimination and death. Previously, we have introduced Bayesian inversion stress microscopy (BISM), which relies on measuring cell-generated traction forces in vitro and has proven particularly useful to measure absolute stresses in confined cell monolayers. We further demonstrate the applicability and robustness of BISM across various experimental settings with different boundary conditions, ranging from confined tissues of arbitrary shape to monolayers composed of different cell types. Importantly, BISM does not require assumptions on cell rheology. Therefore, it can be applied to complex heterogeneous tissues consisting of different cell types, as long as they can be grown on a flat substrate. Finally, we compare BISM to other common stress measurement techniques using a coherent experimental setup, followed by a discussion on its limitations and further perspectives.