Confinement and substrate topology strongly affect the behavior of cell populations and, in particular, their collective migration. In vitro experiments dealing with these aspects require strategies of surface patterning that remain effective over long times (typically several days) and ways to control the surface topology in three dimensions. Here, we describe protocols addressing these two aspects. High-resolution patterning of a robust cell-repellent coating is achieved by etching the coating through a photoresist mask patterned directly on the coated surface. Out-of-plane curvature can be controlled using glass wires or corrugated "wavy" surfaces.

Mathematical models have been widely used to describe the collective movement of bacteria by chemotaxis. In particular, bacterial concentration waves traveling in a narrow channel have been experimentally observed and can be precisely described thanks to a mathematical model at the macroscopic scale. Such model was derived in [1] using a kinetic model based on an accurate description of the mesoscopic run-and-tumble process. We extend this approach to study the behavior of the interaction between two populations of E. Coli. Separately, each population travels with its own speed in the channel. When put together, a synchronization of the speed of the traveling pulses can be observed. We show that this synchronization depends on the fraction of the fast population. Our approach is based on mathematical analysis of a macroscopic model of partial differential equations. Numerical simulations in comparison with experimental observations show qualitative agreement.

For the first time the study at various length scales of E. coli proliferation modes within Si(HIPE) inorganic macrocellular foams is proposed. Both qualitatively and semi-quantitatively, bacterial proliferation within the foam is not homogeneous and is directly linked to the random distribution of Si(HIPE) macroscopic cells. When inoculated in a nutrient loaded Si(HIPE), the bacterial growth is enhanced within the Si(HIPE) matrices compared to the surrounding LB media. The bacterial growth kinetics tends to be faster and the concentration at saturation is roughly 100% times higher. In the case of a Si(HIPE) host free of nutrients, bacterial motion occurs as an infiltration wave; the peak of this propagation wave moves at a constant speed of 88 μm h-1, while bacterial concentrations within the Si(HIPE) host reach levels far above the ones reached in the presence of nutrients, suggesting a real synergetic relationship between the bacterial colony guests and the Si(HIPE) host. When a nutrient reservoir is present at the opposite position from which bacteria were inoculated, bacterial proliferation is associated with a coalescence process between the growing colonies that were rapidly established within the first hours. When the Si(HIPE) host was fully colonized we found out a specific distance between adjacent colonies of 5 and 15 μm in good correspondence with the repartition of the wall to wall distances of the Si(HIPE) macroscopic cells, meaning that bacterial repartition once colonization occurs is optimum. These results show that Si(HIPE) foams are outstanding candidates for strengthened bacterial proliferation without motion restriction imposed by conventional silica gels.

A microfluidic device is a powerful tool to manipulate in a controlled manner at spatiotemporal scales for biological systems. Here, we describe a simple diffusion-based assay to generate and measure the effect of biochemical perturbations within the cytoplasm of cell-free extracts from Xenopus eggs. Our approach comprises a microliter reservoir and a model cytoplasm that are separated by a synthetic membrane containing sub-micrometric pores through which small molecules and recombinant proteins can diffuse. We have used this system to examine the perturbation of elements of the mitotic spindle, which is a microtubule-based bipolar structure involved in the segregation of the replicated genome to daughter cells during cell division. First, we used the small molecule inhibitor monastrol to target kinesin-5, a molecular motor that maintains the microtubule spindle bipolarity. Next, we explored the dynamics of the mitotic spindle by monitoring the exchange between unpolymerized and polymerized tubulin within microtubule fibers. These results show that a simple diffusion-based system can generate biochemical perturbations directly within a cell-free cytoplasm based on Xenopus egg extracts at the time scale of minutes. Our assay is therefore suitable for monitoring the dynamics of supramolecular assemblies within cell-free extracts in response to perturbations. This strategy opens up broad perspectives including phenotype screening or mechanistic studies of biological assembly processes and could be applied to other cell-free extracts such as those derived from mammalian or bacterial cells.

Nat. Cell Biol. 16, 217–223 (2014); published online 23 February 2014; corrected after print 26 February 2014 In the version of this Article originally published there were some errors: The second citation of reference 1 in the first sentence of the main text should have been of reference 2 ('...or by the release of a physical barrier2.

Migration collective : un partage des tâches entre cellules leaders et coordination supracellulaire Myriam Reffay1,2, Maria-Carla Parrini3, Olivier Cochet-Escartin1, Benoit Ladoux2,4, Axel Buguin1, Sylvie Coscoy1, Francois Amblard1, Jacques Camonis3, Pascal Silberzan1 1 Laboratoire physico-chimie Curie, UMR 168, Institut Curie, centre de recherche, CNRS, UPMC, Paris, F-75248, France ; 2 Laboratoire matiere et systemes complexes UMR 7057, Universite Paris Diderot, CNRS, Paris, F-75251, France ; 3 groupe analyse des reseaux de transduction, U830, Institut Curie, centre de recherche, Inserm, Paris, F-75248, France. 4 Adresses actuelles : Institut Jacques Monod (IJM), CNRS UMR 7592 et Universite Paris Diderot, Paris F75205, France ; Mechanobiology Institute, National University of Singapore 117411, Singapore. myriam.reffay@univ-paris-diderot.fr

It is now well established that topological microstructures play a key role in the physical properties of surfaces. Stimulus-induced variations of topological microstructure should therefore lead to a change in the physical properties of microstructured responsive surfaces. In this paper, we demonstrate that roughness changes alter the wetting properties of responsive organic surfaces. Oriented nematic liquid crystalline elastomers (LCEs) are used to construct the microstructured surfaces via a replica molding technique. The topological microstructure of the surfaces covered with micropillars changes with temperature, due to the reversible contraction of the LCE pillars along the long axis at the nematic-to-isotropic phase transition. This is directly observed for the first time under environmental scanning electron microscopy (E-SEM). A high boiling point liquid, glycerol, is used to continuously monitor the contact angle change with temperature. The glycerol contact angle of the microstructured surfaces covered with small pillars decreases from 118° at room temperature to 80° at 140 °C, corresponding to a transition from Cassie state to Wenzel state.

Under a constant flow velocity V, a flexible polymer (in steady state) is expected to display three types of conformation: 1) unperturbed at low V; 2) "trumpet": partly stretched, with certain scaling laws (F. Brochard-Wyart, Europhys. Lett., 23 (1993) 105) at intermediate V; 3) "stem and flower" with a completely stretched portion (the stem) at high V. We discuss regime 3) here. We also consider transients where (starting from regime 3)) the flow is abruptly suppressed and the resulting decay law for the stem is of the Rouse type.

Inspired by the lotus leaf, scientists have developed many superhydrophobic surfaces, some of which show remarkable switching between hydrophobic and hydrophilic state under external stimuli. However, the switch usually relies on the change of chemical properties rather than on the modification of the topographic structure of the surface. In this paper, the roughness‐change‐related switchable wetting properties of microstructured responsive surfaces made of nematic liquid crystalline elastomers (LCEs) is reported. First, various carbonate LC monomers and side‐on LCEs are synthesized with low nematic‐to‐isotropic transition temperature, TNI. Then, LCEs prepared from 3″‐vinylcarbonyloxypropyl 2,5‐di(4′‐octyloxybenzoyloxy)benzoate monomer, with TNI of 76 °C and contraction of 34% are used to construct a surface covered with micropillar arrays by using a replica molding technique. The contraction of the micropillars induces a reversible roughness change of the microstructured surface. Water contact angle of this microstructured surface changed with temperature, indicating a successful approach at building a surface with switchable wetting properties.

The bacterium Escherichia coli in suspension in a liquid medium swims by a succession of runs and tumbles, effectively describing a random walk. The tumbles randomize incompletely, i.e. with a directional persistence, the orientation taken by the bacterium. Here, we model these tumbles by an active rotational diffusion process characterized by a diffusion coefficient and a diffusion time. In homogeneous media, this description accounts well for the experimental reorientations. In shallow gradients of nutrients, tumbles are still described by a unique rotational diffusion coefficient. Together with an increase in the run length, these tumbles significantly contribute to the net chemotactic drift via a modulation of their duration as a function of the direction of the preceding run. Finally, we discuss the limits of this model in propagating concentration waves characterized by steep gradients. In that case, the effective rotational diffusion coefficient itself varies with the direction of the preceding run. We propose that this effect is related to the number of flagella involved in the reorientation process.

Characterizing the migration of a population of cells remains laborious and somewhat subjective. Advances in genetics and robotics allow researchers to perform many experiments in parallel, but analyzing the large sets of data remains a bottleneck. Here we describe a rapid, fully automated correlation-based method for cell migration analysis, compatible with standard video microscopy. This method allows for the computation of quantitative migration parameters via an extensive dynamic mapping of cell displacements.

Collective cell migration is often characterized by the spontaneous onset of multicellular protrusions (known as fingers) led by a single leader cell. Working with epithelial Madin-Darby canine kidney monolayers we show that cells within the fingers, as compared with the epithelium, are well oriented and polarized along the main finger direction, which suggests that these cells actively migrate. The cell orientation and polarity decrease continuously from the tip toward the epithelium over a penetration distance of typically two finger lengths. Furthermore, laser photoablation experiments at various locations along these fingers demonstrate that the cells in the fingers are submitted to a tensile stress whose value is larger close to the tip. From a dynamical point of view, cells entering a finger gradually polarize on timescales that depend upon their particular initial position. Selective laser nanosurgery of the leader lamellipodium shows not only that these structures need a leader to progress, but that this leader itself is the consequence of a prior self-organization of the cells forming the finger. These results highlight the complex interplay between the collective orientation within the fingers and the mechanical action of the leader.

Thermophoresis, the Soret effect, depletes a high concentration of a polyethylene glycol polymer solution from the hot region and builds a concentration gradient. In such a solution, solutes of small concentration experience thermophoresis and polyethylene glycol concentration-dependent restoring forces. We report that by using focused laser heating and varying the polyethylene glycol concentration one observes geometrical localizations of solutes like DNA and RNA into patterns such as a ring. For DNA up to 5.6 kbp, the ring size decreases following a behavior analogous to a gel electrophoresis separation. Above 5.6 kbp, the ring diameter increases with the DNA length. Mixtures of DNA and RNA can be separated as well as different RNA lengths. Separation of colloids is also observed. The experiments might be relevant for the separation of small RNA ribozymes in an early stage of life.

Chemotactic bacteria are known to collectively migrate towards sources of attractants. In confined convectionless geometries, concentration “waves” of swimming Escherichia coli can form and propagate through a self-organized process involving hundreds of thousands of these microorganisms. These waves are observed in particular in microcapillaries or microchannels; they result from the interaction between individual chemotactic bacteria and the macroscopic chemical gradients dynamically generated by the migrating population. By studying individual trajectories within the propagating wave, we show that, not only the mean run length is longer in the direction of propagation, but also that the directional persistence is larger compared to the opposite direction. This modulation of the reorientations significantly improves the efficiency of the collective migration. Moreover, these two quantities are spatially modulated along the concentration profile. We recover quantitatively these microscopic and macroscopic observations with a dedicated kinetic model.