Explore the vast range of titles available.

The RAC1-WAVE-Arp2/3 signaling pathway generates branched actin networks that power lamellipodium protrusion of migrating cells. Feedback is thought to control protrusion lifetime and migration persistence, but its molecular circuitry remains elusive. Here, we identify PPP2R1A by proteomics as a protein differentially associated with the WAVE complex subunit ABI1 when RAC1 is activated and downstream generation of branched actin is blocked. PPP2R1A is found to associate at the lamellipodial edge with an alternative form of WAVE complex, the WAVE Shell Complex, that contains NHSL1 instead of the Arp2/3 activating subunit WAVE, as in the canonical WAVE Regulatory Complex. PPP2R1A is required for persistence in random and directed migration assays and for RAC1-dependent actin polymerization in cell extracts. PPP2R1A requirement is abolished by NHSL1 depletion. PPP2R1A mutations found in tumors impair WAVE Shell Complex binding and migration regulation, suggesting that the coupling of PPP2R1A to the WAVE Shell Complex is essential to its function. The WAVE regulatory complex activates Arp2/3 at the cell cortex and in membrane protrusions to generate persistent cell migration. Here authors show that PPP2R1A, a scaffold subunit of protein phosphatase 2, associates with an alternative form of the WAVE complex where WAVE, the subunit that activates Arp2/3, is replaced by NHSL1.

Flexible superparamagnetic filaments (‘fleximags’) are very slender elastic filaments, which can be driven by distributed magnetic torques to mimic closely the behaviour of biological flagella. Previously, fleximags have been used as a basis for artificial micro-swimmers capable of transporting small cargos Dreyfus et al. (Nature, vol. 437, 2005, p. 862). Here, we demonstrate how these filaments can be anchored to a wall to make carpets of artificial micro-magnetic cilia with tunable densities. We analyse the dynamics of an artificial cilium under both planar and three-dimensional beating patterns. We show that the dynamics are controlled by a single characteristic length scale varying with the inverse square root of the driving frequency, providing a mechanism to break the fore and aft symmetry and to generate net fluxes and forces. However, we show that an effective geometrical reciprocity in the filament dynamics creates intrinsic limitations upon the ability of the artificial flagellum to pump fluid when driven in two dimensions.

Atherosclerosis is triggered by chronic inflammation of arterial endothelial cells (ECs). Because atherosclerosis develops preferentially in regions where blood flow is disturbed and where ECs have a cuboidal morphology, the interplay between EC shape and mechanotransduction events is of primary interest. In this work we present a simple microfluidic device to study relationships between cell shape and EC response to fluid shear stress. Adhesive micropatterns are used to non-invasively control EC elongation and orientation at both the monolayer and single cell levels. The micropatterned substrate is coupled to a microfluidic chamber that allows precise control of the flow field, high-resolution live-cell imaging during flow experiments, and in situ immunostaining. Using micro particle image velocimetry, we show that cells within the chamber alter the local flow field so that the shear stress on the cell surface is significantly higher than the wall shear stress in regions containing no cells. In response to flow, we observe the formation of lamellipodia in the downstream portion of the EC and cell retraction in the upstream portion. We quantify flow-induced calcium mobilization at the single cell level for cells cultured on unpatterned surfaces or on adhesive lines oriented either parallel or orthogonal to the flow. Finally, we demonstrate flow-induced intracellular calcium waves and show that the direction of propagation of these waves is determined by cell polarization rather than by the flow direction. The combined versatility and simplicity of this microfluidic device renders it very useful for studying relationships between EC shape and mechanosensitivity.

Blood vessels expand and contract actively, while continuously experiencing dynamic external stresses from the blood flow. The mechanical response of the vessel wall is that of a composite material: its mechanical properties depend on a diverse set of cellular mechanical components, which change dynamically as cells respond to external stress. Mapping the relationship between these underlying cellular processes and emergent tissue mechanics is an on-going challenge, in particular in endothelial cells. Here we use a microstretcher mimicking the native environment of blood vessels to assess both the mechanics and cellular dynamics of an endothelial tube in response to a physiological increase in luminal pressure. The characterization of the instantaneous monolayer elasticity reveals a strain-stiffening, actin-dependent and substrate-responsive behavior. In response to a maintained pressure increase, the tissue displays a fluid-like expansion, accompanied by the reorientation of cell shape and of actin fibers. This actin-driven reorientation depends on focal adhesions and adherens junctions, two key mechanosensors. We introduce a mechanical model coupling actin fiber nematodynamics with active and elastic tension generation by actin fibers in the endothelium, which recapitulates the response to pressure of endothelial tubes.

In the microvasculature, blood flow-derived forces are key regulators of vascular structure and function. Consequently, the development of hydrogel-based microvessel-on-chip systems that strive to mimic the in vivo cellular organization and mechanical environment has received great attention in recent years. However, despite intensive efforts, current microvessel- on-chip systems suffer from several limitations, most notably failure to produce physiologically relevant wall strain levels. In this study, a novel microvessel-on-chip based on the templating technique and using luminal flow actuation to generate physiologically relevant levels of wall shear stress and circumferential stretch is presented. Normal forces induced by the luminal pressure compress the surrounding soft collagen hydrogel, dilate the channel, and create large circumferential strain. The fluid pressure gradient in the system drives flow forward and generates realistic pulsatile wall shear stresses. Rigorous characterization of the system reveals the crucial role played by the poroelastic behavior of the hydrogel in determining the magnitudes of the wall shear stress and strain. The experimental measurements are combined with an analytical model of flow in both the lumen and the porous hydrogel to provide an exceptionally versatile user manual for an application-based choice of parameters in microvessels-on-chip. This unique strategy of flow actuation adds a dimension to the capabilities of microvessel-on-chip systems and provides a more general framework for improving hydrogel-based in vitro engineered platforms.

Pericytes are mural cells of the microvasculature, characterised by their elongated distinct shape. Pericytes span along the axis of the vessels they adhere to, therefore they experience extreme lateral and longitudinal confinement. Pericyte shape is key for their function during vascular regulation and their spatial distribution is established by cell migration during the embryonic stage and maintained through controlled motility in the adult. However, how pericyte morphology is associated with migration and function remains unknown. We use micropatterns to mimic pericyte adhesion to vessels, and to reproduce in vitro the shapes adopted by pericytes in vivo. We show that lateral confinement controls cell shape and produces in vivo-like phenotype. Modelling the pericyte as an incompressible linear elastic material predicts strain and shape of pericytes as a function of lateral confinement. Pericyte kinetics on both laterally confining lanes, and longitudinally constraining motifs is described by dry friction theory. Pericytes are capable of crossing gaps of different sizes. The percentage of crossings is correctly predicted by the likelihood of a fluctuating system to overcome an energy barrier. Our joint experimental and theoretical approach demonstrates the effect of in vivo-like geometrical confinement on pericyte morphology and migration which is accurately described by dry friction theory.Competing Interest StatementThe authors have declared no competing interest.

The RAC1-WAVE-Arp2/3 signaling pathway generates branched actin networks that power lamellipodium protrusion of migrating cells. Feedback is thought to control protrusion lifetime and migration persistence, but its molecular circuitry remains elusive. Using proteomics, we identified PPP2R1A among proteins differentially associated with the WAVE complex subunit ABI1 when RAC1 was activated and downstream generation of branched actin was blocked. PPP2R1A was found to associate at the lamellipodial edge with a novel form of WAVE complex, the WAVE Shell Complex (WSC), that contains NHSL1 instead of the Arp2/3 activating subunit WAVE as in the canonical WAVE Regulatory Complex (WRC). PPP2R1A was required for persistence in random and directed migration assays and for RAC1-dependent actin polymerization in cell extracts. PPP2R1A requirement was abolished by NHSL1 depletion. PPP2R1A mutations found in tumors impaired WSC binding and migration regulation, suggesting that this novel function of PPP2R1A is critical for its tumor suppressor activity.

Throughout their life, plant root are submitted to mechanical stresses due to pressure exerted by the soil. So far, few studies addressed root cell deformation and calcium signaling elicited by soil compression. In this study, we designed a microchip inspired by pneumatic microvalve concept in order to deliver a lateral pressure to the root of a plant expressing the RGECO1-mTurquoise calcium reporter. Lateral pressure applied on the root induced a moderate elastic deformation of root cortical cells and elicited a multicomponent calcium signal at the onset of the pressure pulse, followed by a second one at the release of the pressure. This indicates that straining rather than stressing of tissues is relevant to trigger the calcium signal. The calcium elevation was restricted to the tissue under pressure and did not propagate. Additionally, the calcium signals exhibited a remarkable attenuation upon repetitive stimulations. A microvalve concept mimicking lateral soil pressure was developed. Non-damaging lateral compression of the root induces an elastic deformation of cortical cells. A multicomponent calcium signal is elicited at the onset of a pressure pulse and upon release of the pressure. Straining rather than stressing of tissues is relevant to trigger the calcium signal. The calcium signal is localized at the tissue under pressure and does not propagate. Calcium signals exhibit a remarkable attenuation upon repetitive stimulations.

To accomplish their critical task of removing infected cells and fighting pathogens, leukocytes activate by forming specialized interfaces with other cells. Using an innovative micropipette rheometer, we show in three different cell types that when stimulated by microbeads mimicking target cells, leukocytes become up to ten times stiffer and more viscous. These mechanical changes initiate within seconds after contact and evolve rapidly over minutes. Remarkably, leukocyte elastic and viscous properties evolve in parallel, preserving a well-defined ratio that constitutes a mechanical signature specific to each cell type. The current results indicate that simultaneously tracking both elastic and viscous properties during an active cell process provides a new way to investigate cell mechanical processes. Our findings also suggest that dynamic immuno-mechanical measurements provide an identifier of leukocyte type and an indicator of the cell’s state of activation.

To accomplish their critical task of removing infected cells and fighting pathogens, leukocytes activate by forming specialized interfaces with other cells. Using an innovative micropipette rheometer, we show in three different cell types that when stimulated by microbeads mimicking target cells, leukocytes become up to ten times stiffer and more viscous. These mechanical changes initiate within seconds after contact and evolve rapidly over minutes. Remarkably, leukocyte elastic and viscous properties evolve in parallel, preserving a well-defined ratio that constitutes a mechanical signature specific to each cell type. The current results indicate that simultaneously tracking both elastic and viscous properties during an active cell process provides a new way to investigate cell mechanical processes. Our findings also suggest that dynamic immuno-mechanical measurements provide an identifier of leuko-cyte type and an indicator of the cell's state of activation.