Actomyosin machinery endows cells with contractility at a single cell level. However, within a monolayer, cells can be contractile or extensile based on the direction of pushing or pulling forces exerted by their neighbours or on the substrate. It has been shown that a monolayer of fibroblasts behaves as a contractile system while epithelial or neural progentior monolayers behave as an extensile system. Through a combination of cell culture experiments and in silico modeling, we reveal the mechanism behind this switch in extensile to contractile as the weakening of intercellular contacts. This switch promotes the buildup of tension at the cell-substrate interface through an increase in actin stress fibers and traction forces. This is accompanied by mechanotransductive changes in vinculin and YAP activation. We further show that contractile and extensile differences in cell activity sort cells in mixtures, uncovering a generic mechanism for pattern formation during cell competition, and morphogenesis.

Highlights d Caveolae influence contractile tension in epithelial monolayers d Caveolin-1 depletion enhances epithelial tension via PtdIns(4,5)P 2 signaling d Elevated epithelial tension inhibits oncogenic cell extrusion

The physical cues from the extracellular environment mediates cell signaling spatially and temporally.

Epithelia act as a barrier against environmental stress and abrasion and in vivo they are continuously exposed to environments of various mechanical properties. The impact of this environment on epithelial integrity remains elusive. By culturing epithelial cells on 2D hydrogels, we observe a loss of epithelial monolayer integrity through spontaneous hole formation when grown on soft substrates. Substrate stiffness triggers an unanticipated mechanical switch of epithelial monolayers from tensile on soft to compressive on stiff substrates. Through active nematic modelling, we find that spontaneous half-integer defect formation underpinning large isotropic stress fluctuations initiate hole opening events. Our data show that monolayer rupture due to high tensile stress is promoted by the weakening of cell-cell junctions that could be induced by cell division events or local cellular stretching. Our results show that substrate stiffness provides feedback on monolayer mechanical state and that topological defects can trigger stochastic mechanical failure, with potential application towards a mechanistic understanding of compromised epithelial integrity during immune response and morphogenesis.

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Mixing in a microfluidic system is challenging due to dominant diffusion effects at a microscale (low Reynolds number). In this work, we report the improvement of mixing performance in spiral microchannels of varying cross-sectional geometry and hydraulic diameter. The formation of secondary flow interactions in spiral channels, known as Dean vortices, aid the primary diffusion process. The evolution of these Dean vortices was experimentally visualized along the length of the microchannel by confocal microscopy, and then compared to numerical studies. The cross-sectional geometries of the spiral channels, especially in the case of irregular shapes such as the semi-circular and trapezoidal profiles, were found to be an important factor in tuning the strength of Dean vortices, which in turn dictate the mixing performance, as opposed to diffusion which is more prominent at lower Re. This experiment-based finding has been validated via the evaluation of swirling strength of the working fluid, obtained using a numerical study. The results thus obtained show a mixing performance greater than 90% above a Reynolds number of 20 for most spiral channel designs, making this system suitable for high throughput operation with reduced pressure drop. This work is the first to experimentally and numerically demonstrate, within this operating range (20  <  Re  <  277), the impact on mixing performance in curved microchannels of varying cross-sectional geometries of constant cross-sectional area, and of varying hydraulic diameters for square shaped channels. The capability of these channels to operate at a moderately high Re with enhanced mixing performance and reduced pressure drop would be of great use in large-scale industrial operations, such as complex integrated micro-reactors wherein pressure drop plays a key role.

Actomyosin machinery endows cells with contractility at a single cell level. However, at a tissue scale, cells can show either contractile or extensile behaviour based on the direction of pushing or pulling forces due to neighbour interactions or substrate interactions. Previous studies have shown that a monolayer of fibroblasts behaves as a contractile system while a monolayer of epithelial cells or neural crest cells behaves as an extensile system. How these two contradictory sources of force generation can coexist has remained unexplained. Through a combination of experiments using MDCK (Madin Darby Canine Kidney) cells, and in-silico modeling, we uncover the mechanism behind this switch in behaviour of epithelial cell monolayers from extensile to contractile as the weakening of intercellular contacts. We find that this switch in active behaviour also promotes the buildup of tension at the cell substrate interface through an increase in actin stress fibers and higher traction forces. This in turn triggers a mechanotransductive response in vinculin translocation to focal adhesion sites and YAP (Yes-associated protein) transcription factor activation. Our studies also show that differences in extensility and contractility act to sort cells, thus determining a general mechanism for mechanobiological pattern formation during cell competition, morphogenesis and cancer progression.

In the version of this Article originally published, the captions for Extended Data Figs. 1, 2 and 3 were in the wrong order and did not correspond to their associated figures. The correct captions are listed below and the Article has been corrected accordingly. In addition, the cell line MCF7 was mistakenly written as 'MCF7A' in seven instances in the main text, Methods and Extended Data Fig. 4 caption, and as 'MCF10A' in one instance in the ' Author contributions' section; these errors have now been corrected. Extended Data Fig. 1 | MDCK WT behave as an extensile system. a, Kymograph of a short junction (<10μm) (top) and long junction (>15μm) (bottom) before and after laser ablation. b, Recoil velocity after laser ablation for short (<10μm) (n=9) (N=4), medium (10-15μm) (n=13)(N=4) and long junctions (> 15μm) (n=12) (N=6). n, is the number of junctions ablated and N is the number of independent experiments from which these results were obtained. Error bars represent the standard deviation. ANOVA test was performed leading to *p<0.05, **p<0.01, ***p<0.001 and ****p<0.0001. Scale bars, 20μm. Extended Data Fig. 2 | MDCK WT behaves as an extensile system and MDCK E-cadherin KO behave as a contractile system. a,b, Orientation field (left) and velocity vectors (right) around a single comet-shaped (+1/2) defect (a) and trefoil (-1/2) defect obtained from WT (top) and E-cadherin KO (bottom) monolayers. c,d, Trajectory of several comet (+1/2) (left) and trefoil (-1/2) (right) shaped defects obtained from MDCK WT (c) and MDCK E-cadherin KO (d) monolayers. Scale bars: 40μm. Extended Data Fig. 3 | Fibroblasts behave as a 2D contractile active nematic. a, Average yy-and xy-components of strain rate map around +1/2 defect obtained from experiments (left and middle respectively) and corresponding average flow field (right) (n = 1489 defects from 2 independent experiments) for NIH3T3 cells. Colour code is positive for stretching and negative for shrinkage. b, Average yy (left)-, xy (middle)-and isotropic (right) components of stress around a +1/2 defect obtained from experiments for NIH3T3 (n = 1,428 defects from 2 independent experiments).