Pharmacological activation of PIEZO1 in human red blood cells prevents Plasmodium falciparum invasion.

An inherited gain-of-function variant (E756del) in the mechanosensitive cationic channel PIEZO1 was shown to confer a significant protection against severe malaria. Here, we demonstrate in vitro that human red blood cell (RBC) infection by Plasmodium falciparum is prevented by the pharmacological activation of PIEZO1. Yoda1 causes an increase in intracellular calcium associated with rapid echinocytosis that inhibits RBC invasion, without affecting parasite intraerythrocytic growth, division or egress. Notably, Yoda1 treatment significantly decreases merozoite attachment and subsequent RBC deformation. Intracellular Na+/K+ imbalance is unrelated to the mechanism of protection, although delayed RBC dehydration observed in the standard parasite culture medium RPMI/albumax further enhances the resistance to malaria conferred by Yoda1. The chemically unrelated Jedi2 PIEZO1 activator similarly causes echinocytosis and RBC dehydration associated with resistance to malaria invasion. Spiky outward membrane projections are anticipated to reduce the effective surface area required for both merozoite attachment and internalization upon pharmacological activation of PIEZO1. Globally, our findings indicate that the loss of the typical biconcave discoid shape of RBCs, together with an altered optimal surface to volume ratio, induced by PIEZO1 pharmacological activation prevent efficient P. falciparum invasion.

Speech can produce jet-like transport relevant to asymptomatic spreading of virus.

Many scientific reports document that asymptomatic and presymptomatic individuals contribute to the spread of COVID-19, probably during conversations in social interactions. Droplet emission occurs during speech, yet few studies document the flow to provide the transport mechanism. This lack of understanding prevents informed public health guidance for risk reduction and mitigation strategies, e.g., the "6-foot rule." Here we analyze flows during breathing and speaking, including phonetic features, using orders-of-magnitude estimates, numerical simulations, and laboratory experiments. We document the spatiotemporal structure of the expelled airflow. Phonetic characteristics of plosive sounds like "P" lead to enhanced directed transport, including jet-like flows that entrain the surrounding air. We highlight three distinct temporal scaling laws for the transport distance of exhaled material including 1) transport over a short distance (<0.5 m) in a fraction of a second, with large angular variations due to the complexity of speech; 2) a longer distance, ∼1 m, where directed transport is driven by individual vortical puffs corresponding to plosive sounds; and 3) a distance out to about 2 m, or even farther, where sequential plosives in a sentence, corresponding effectively to a train of puffs, create conical, jet-like flows. The latter dictates the long-time transport in a conversation. We believe that this work will inform thinking about the role of ventilation, aerosol transport in disease transmission for humans and other animals, and yield a better understanding of linguistic aerodynamics, i.e., aerophonetics.

High-Resolution Photonic Force Microscopy Based on Sharp Nanofabricated Tips.

Although near-field imaging techniques reach sub-nanometer resolution on rigid samples, it remains extremely challenging to image soft interfaces, such as biological membranes, due to the deformations induced by the probe. In photonic force microscopy, optical tweezers are used to manipulate and measure the scanning probe, allowing imaging of soft materials without force-induced artifacts. However, the size of the optically trapped probe still limits the maximum resolution. Here, we show a novel and simple nanofabrication protocol to massively produce optically trappable quartz particles which mimic the sharp tips of atomic force microscopy. Imaging rigid nanostructures with our tips, we resolve features smaller than 80 nm. Scanning the membrane of living malaria-infected red blood cells reveals, with no visible artifacts, submicron features termed knobs, related to the parasite activity. The use of nanoengineered particles in photonic force microscopy opens the way to imaging soft samples at high resolution.

Red cells' dynamic morphologies govern blood shear thinning under microcirculatory flow conditions.

Blood viscosity decreases with shear stress, a property essential for an efficient perfusion of the vascular tree. Shear thinning is intimately related to the dynamics and mutual interactions of RBCs, the major component of blood. Because of the lack of knowledge about the behavior of RBCs under physiological conditions, the link between RBC dynamics and blood rheology remains unsettled. We performed experiments and simulations in microcirculatory flow conditions of viscosity, shear rates, and volume fractions, and our study reveals rich RBC dynamics that govern shear thinning. In contrast to the current paradigm, which assumes that RBCs align steadily around the flow direction while their membranes and cytoplasm circulate, we show that RBCs successively tumble, roll, deform into rolling stomatocytes, and, finally, adopt highly deformed polylobed shapes for increasing shear stresses, even for semidilute volume fractions of the microcirculation. Our results suggest that any pathological change in plasma composition, RBC cytosol viscosity, or membrane mechanical properties will affect the onset of these morphological transitions and should play a central role in pathological blood rheology and flow behavior.

Flow-Induced Transitions of Red Blood Cell Shapes under Shear.

A recent study of red blood cells (RBCs) in shear flow [Lanotte et al., Proc. Natl. Acad. Sci. U.S.A. 113, 13289 (2016)PNASA60027-842410.1073/pnas.1608074113] has demonstrated that RBCs first tumble, then roll, transit to a rolling and tumbling stomatocyte, and finally attain polylobed shapes with increasing shear rate, when the viscosity contrast between cytosol and blood plasma is large enough. Using two different simulation techniques, we construct a state diagram of RBC shapes and dynamics in shear flow as a function of shear rate and viscosity contrast, which is also supported by microfluidic experiments. Furthermore, we illustrate the importance of RBC shear elasticity for its dynamics in flow and show that two different kinds of membrane buckling trigger the transition between subsequent RBC states.

Sinking a Granular Raft.

We report experiments that yield new insights on the behavior of granular rafts at an oil-water interface. We show that these particle aggregates can float or sink depending on dimensionless parameters taking into account the particle densities and size and the densities of the two fluids. We characterize the raft shape and stability and propose a model to predict its shape and maximum length to remain afloat. Finally we find that wrinkles and folds appear along the raft due to compression by its own weight, which can trigger destabilization. These features are characteristics of an elastic instability, which we discuss, including the limitations of our model.

Dynamics of colloid accumulation under flow over porous obstacles.

The accumulation of colloidal particles to build dense structures from dilute suspensions may follow distinct routes. The mechanical, structural and geometrical properties of these structures depend on local hydrodynamics and colloidal interactions. Using model suspensions flowing into microfabricated porous obstacles, we investigate this interplay by tuning both the flow pattern and the ionic strength. We observe the formation of a large diversity of shapes, and demonstrate that growing structures in turn influence the local velocity pattern, favouring particle deposition either locally or over a wide front. We also show that these structures are labile, stabilised by the flow pushing on them, in low ionic strength conditions, or cohesive, in a gel-like state, at higher ionic strength. The interplay between aggregate cohesion and erosion thus selects preferential growth modes and therefore dictates the final shape of the structure.

The dynamics of interacting folds under biaxial compressive stresses.

The gradual in-plane compression of a solid film bonded to a soft substrate can lead to surface wrinkling and even to the formation of a network of folds for sufficiently high strain. An understanding of how these folds initiate, propagate, and interact with each other is still lacking. In a previous study, we developed an experimental system to observe the wrinkle-to-fold transition of layered elastic materials under biaxial compressive stresses. Here we focus on the dynamic interaction of a pair of propagating folds under biaxial compression. We find experimentally that their behavior is mediated through their tips and depends on the separation of the tips and their angle of interception. When the angle is lower than 45°, the two folds either form a unique fold by the coalescence of their tips when close enough, or bend their trajectories to intersect each other and form a lenticular region in analogy with cracks. When the angle is higher then 45°, the folds simply intersect and form a T-like junction. We rationalize this behavior by conducting numerical simulations to visualize the stress field around the two tips and find that the initial geometric position of the tips primarily determines the final state of the folds.

Clusters of red blood cells in microcapillary flow: hydrodynamic versus macromolecule induced interaction.

We present experiments on RBCs that flow through micro-capillaries under physiological conditions. The strong flow-shape coupling of these deformable objects leads to a rich variety of cluster formation. We show that the RBC clusters form as a subtle imbrication between hydrodynamic interactions and adhesion forces because of plasma proteins, mimicked by the polymer dextran. Clusters form along the capillaries and macromolecule-induced adhesion contributes to their stability. However, at high yet physiological flow velocities, shear stresses overcome part of the adhesion forces, and cluster stabilization due to hydrodynamics becomes stronger. For the case of pure hydrodynamic interaction, cell-to-cell distances have a pronounced bimodal distribution. Our 2D-numerical simulations on vesicles capture the transition between adhesive and non-adhesive clusters at different flow velocities.

Microfluidic study of enhanced deposition of sickle cells at acute corners.

Sickle cell anemia is a blood disorder, known to affect the microcirculation and is characterized by painful vaso-occlusive crises in deep tissues. During the last three decades, many scenarios based on the enhanced adhesive properties of the membrane of sickle red blood cells have been proposed, all related to a final decrease in vessels lumen by cells accumulation on the vascular walls. Up to now, none of these scenarios considered the possible role played by the geometry of the flow on deposition. The question of the exact locations of occlusive events at the microcirculatory scale remains open. Here, using microfluidic devices where both geometry and oxygen levels can be controlled, we show that the flow of a suspension of sickle red blood cells around an acute corner of a triangular pillar or of a bifurcation, leads to the enhanced deposition and aggregation of cells. Thanks to our devices, we follow the growth of these aggregates in time and show that their length does not depend on oxygenation levels; instead, we find that their morphology changes dramatically to filamentous structures when using autologous plasma as a suspending fluid. We finally discuss the possible role played by such aggregates in vaso-occlusive events.

A simple model to understand the effect of membrane shear elasticity and stress-free shape on the motion of red blood cells in shear flow.

An analytical model was proposed by Keller and Skalak in 1982 to understand the motion of red blood cells in shear flow. The cell was described as a fluid ellipsoid of fixed shape. This model was extended in 2007 to introduce shear elasticity of the red blood cell membrane. Here, this model is further extended to take into account that the cell discoid shape physiologically observed is not a stress-free shape. The model shows that spheroid stress-free shapes allow us to fit the experimental data with the values of shear elasticity typical to that found with micropipette and optical tweezer experiments. In the range of moderate shear rates (for which RBCs keep their discoid shape) this model enables us to quantitatively determine (i) an effective cell viscosity, which combines membrane and hemoglobin viscosities and (ii) an effective shear modulus of the membrane that combines the shear modulus and the stress-free shape. This model can also be used to determine RBC mechanical parameters not only in the tanktreading regime when cells are suspended in medium of high viscosity but also in the tumbling regime characteristic of cells suspended in media of low viscosity. In this regime, a transition is predicted between a rigid-like tumbling motion and a fluid-like tumbling motion above a critical shear rate, which is directly related to the mechanical parameters of the cell.

Curling and rolling dynamics of naturally curved ribbons.

When a straight rod is bent and suddenly released on one end, a burst of dispersive flexural waves propagates down the material as predicted by linear beam theories. However, we show that for ribbons with a longitudinal natural radius of curvature a0, geometrical constraints lead to strain localization which controls the dynamics. This localized region of deformation selects a specific curling deformation front which travels down the ribbon when initially flattened and released. Performing experiments on different ribbons, in air and in water, we show that initially, on length scales on the order of a0, the curling front moves as a power law of time with an exponent ranging from 0.5 to 2 for increasing values of the ribbons' width. At longer time scales, the material wraps itself at a constant speed Vr into a roll of radius R ≠ a0. The relationship between Vr and R is calculated by a balance between kinetic, elastic and gravitational energy and both internal and external powers dissipated. When gravity and drag are negligible, we observe that a0/R reaches a limiting value of 0.48 that we predict by solving the Elastica on the curled ribbon considering the centrifugal forces due to rotation. The solution we propose represents a solitary traveling curvature wave which is reminiscent to propagating instabilities in mechanics.

Multiscale approach to link red blood cell dynamics, shear viscosity, and ATP release.

RBCs are known to release ATP, which acts as a signaling molecule to cause dilation of blood vessels. A reduction in the release of ATP from RBCs has been linked to diseases such as type II diabetes and cystic fibrosis. Furthermore, reduced deformation of RBCs has been correlated with myocardial infarction and coronary heart disease. Because ATP release has been linked to cell deformation, we undertook a multiscale approach to understand the links between single RBC dynamics, ATP release, and macroscopic viscosity all at physiological shear rates. Our experimental approach included microfluidics, ATP measurements using a bioluminescent reaction, and rheology. Using microfluidics technology with high-speed imaging, we visualize the deformation and dynamics of single cells, which are known to undergo motions such as tumbling, swinging, tanktreading, and deformation. We report that shear thinning is not due to cellular deformation as previously believed, but rather it is due to the tumbling-to-tanktreading transition. In addition, our results indicate that ATP release is constant at shear stresses below a threshold (3 Pa), whereas above the threshold ATP release is increased and accompanied by large cellular deformations. Finally, performing experiments with well-known inhibitors, we show that the Pannexin 1 hemichannel is the main avenue for ATP release both above and below the threshold, whereas, the cystic fibrosis transmembrane conductance regulator only contributes to deformation-dependent ATP release above the stress threshold.

Spatiotemporal dynamics of the proton motive force on single bacterial cells.

Electrochemical gradients across biological membranes are vital for cellular bioenergetics. In bacteria, the proton motive force (PMF) drives essential processes like adenosine triphosphate production and motility. Traditionally viewed as temporally and spatially stable, recent research reveals a dynamic PMF behavior at both single-cell and community levels. Moreover, the observed lateral segregation of respiratory complexes could suggest a spatial heterogeneity of the PMF. Using a light-activated proton pump and detecting the activity of the bacterial flagellar motor, we perturb and probe the PMF of single cells. Spatially homogeneous PMF perturbations reveal millisecond-scale temporal dynamics and an asymmetrical capacitive response. Localized perturbations show a rapid lateral PMF homogenization, faster than proton diffusion, akin to the electrotonic potential spread observed in passive neurons, explained by cable theory. These observations imply a global coupling between PMF sources and consumers along the membrane, precluding sustained PMF spatial heterogeneity but allowing for rapid temporal changes.

A novel mechanism for egress of malarial parasites from red blood cells.

The culminating step of the intraerythrocytic development of Plasmodium falciparum, the causative agent of malaria, is the spectacular release of multiple invasive merozoites on rupture of the infected erythrocyte membrane. This work reports for the first time that the whole process, taking place in time scales as short as 400 milliseconds, is the result of an elastic instability of the infected erythrocyte membrane. Using high-speed differential interference contrast (DIC) video microscopy and epifluorescence, we demonstrate that the release occurs in 3 main steps after osmotic swelling of the infected erythrocyte: a pore opens in ~ 100 milliseconds, ejecting 1-2 merozoites, an outward curling of the erythrocyte membrane is then observed, ending with a fast eversion of the infected erythrocyte membrane, pushing the parasites forward. It is noteworthy that this last step shows slight differences when infected erythrocytes are adhering. We rationalize our observations by considering that during the parasite development, the infected erythrocyte membrane acquires a spontaneous curvature and we present a subsequent model describing the dynamics of the curling rim. Our results show that sequential erythrocyte membrane curling and eversion is necessary for the parasite efficient angular dispersion and might be biologically essential for fast and numerous invasions of new erythrocytes.

Capillary force on a micrometric sphere trapped at a fluid interface exhibiting arbitrary curvature gradients.

We report theoretical predictions and measurements of the capillary force acting on a spherical colloid smaller than the capillary length that is placed on a curved fluid interface of arbitrary shape. By coupling direct imaging and interferometry, we are able to measure the in situ colloid contact angle and to correlate its position with respect to the interface curvature. Extremely tiny capillary forces down to femtonewtons can be measured with this method. Measurements agree well with a theory relating the capillary force to the gradient of Gaussian curvature and to the mean curvature of the interface prior to colloidal deposition. Numerical calculations corroborate these results.

Red blood cell membrane dynamics during malaria parasite egress.

Precisely how malaria parasites exit from infected red blood cells to further spread the disease remains poorly understood. It has been shown recently, however, that these parasites exploit the elasticity of the cell membrane to enable their egress. Based on this work, showing that parasites modify the membrane's spontaneous curvature, initiating pore opening and outward membrane curling, we develop a model of the dynamics of the red blood cell membrane leading to complete parasite egress. As a result of the three-dimensional, axisymmetric nature of the problem, we find that the membrane dynamics involve two modes of elastic-energy release: 1), at short times after pore opening, the free edge of the membrane curls into a toroidal rim attached to a membrane cap of roughly fixed radius; and 2), at longer times, the rim radius is fixed, and lipids in the cap flow into the rim. We compare our model with the experimental data of Abkarian and co-workers and obtain an estimate of the induced spontaneous curvature and the membrane viscosity, which control the timescale of parasite release. Finally, eversion of the membrane cap, which liberates the remaining parasites, is driven by the spontaneous curvature and is found to be associated with a breaking of the axisymmetry of the membrane.

Hierarchical folding of elastic membranes under biaxial compressive stress.

Mechanical instabilities that cause periodic wrinkling during compression of layered materials find applications in stretchable electronics and microfabrication, but can also limit an application's performance owing to delamination or cracking under loading and surface inhomogeneities during swelling. In particular, because of curvature localization, finite deformations can cause wrinkles to evolve into folds. The wrinkle-to-fold transition has been documented in several systems, mostly under uniaxial stress. However, the nucleation, the spatial structure and the dynamics of the invasion of folds in two-dimensional stress configurations remain elusive. Here, using a two-layer polymeric system under biaxial compressive stress, we show that a repetitive wrinkle-to-fold transition generates a hierarchical network of folds during reorganization of the stress field. The folds delineate individual domains, and each domain subdivides into smaller ones over multiple generations. By modifying the boundary conditions and geometry, we demonstrate control over the final network morphology. The ideas introduced here should find application in the many situations where stress impacts two-dimensional pattern formation.

Dynamic stiffening of the flagellar hook.

For many bacteria, motility stems from one or more flagella, each rotated by the bacterial flagellar motor, a powerful rotary molecular machine. The hook, a soft polymer at the base of each flagellum, acts as a universal joint, coupling rotation between the rigid membrane-spanning rotor and rigid flagellum. In multi-flagellated species, where thrust arises from a hydrodynamically coordinated flagellar bundle, hook flexibility is crucial, as flagella rotate significantly off-axis. However, consequently, the thrust applies a significant bending moment. Therefore, the hook must simultaneously be compliant to enable bundle formation yet rigid to withstand large hydrodynamical forces. Here, via high-resolution measurements and analysis of hook fluctuations under dynamical conditions, we elucidate how it fulfills this double functionality: the hook shows a dynamic increase in bending stiffness under increasing torsional stress. Such strain-stiffening allows the system to be flexible when needed yet reduce deformation under high loads, enabling high speed motility.

Colloid science: non-spherical bubbles.

Surface tension gives gas bubbles their perfect spherical shape by minimizing the surface area for a given volume. Here we show that gas bubbles and liquid drops can exist in stable, non-spherical shapes if the surface is covered, or 'armoured', with a close-packed monolayer of particles. When two spherical armoured bubbles are fused, jamming of the particles on the interface supports the unequal stresses that are necessary to stabilize a non-spherical shape.