Damien Cuvelier

External forces control mitotic spindle positioning

The response of cells to forces is essential for tissue morphogenesis and homeostasis. This response has been extensively investigated in interphase cells, but it remains unclear how forces affect dividing cells. We used a combination of micro-manipulation tools on human dividing cells to address the role of physical parameters of the micro-environment in controlling the cell division axis, a key element of tissue morphogenesis. We found that forces applied on the cell body direct spindle orientation during mitosis. We further show that external constraints induce a polarization of dynamic subcortical actin structures that correlate with spindle movements. We propose that cells divide according to cues provided by their mechanical micro-environment, aligning daughter cells with the external force field.

Bursting of sensitive polymersomes induced by curling.

Polymersomes, which are stable and robust vesicles made of block copolymer amphiphiles, are good candidates for drug carriers or micro/nanoreactors. Polymer chemistry enables almost unlimited molecular design of responsive polymersomes whose degradation upon environmental changes has been used for the slow release of active species. Here, we propose a strategy to remotely trigger instantaneous polymersome bursting. We have designed asymmetric polymer vesicles, in which only one leaflet is composed of responsive polymers. In particular, this approach has been successfully achieved by using a UV-sensitive liquid-crystalline copolymer. We study experimentally and theoretically this bursting mechanism and show that it results from a spontaneous curvature of the membrane induced by the remote stimulus. The versatility of this mechanism should broaden the range of applications of polymersomes in fields such as drug delivery, cosmetics and material chemistry.

Spreading dynamics and wetting transition of cellular aggregates.

We study the spreading of spheroidal aggregates of cells, expressing a tunable level of E-cadherin molecules, on glass substrates decorated with mixed fibronectin and polyethylene glycol. We observe the contact area by optical interferometry and the profile by side-view microscopy. We find a universal law of aggregate spreading at short times, which we interpret through an analogy with the spreading of viscoelastic droplets. At long times, we observe either partial wetting or complete wetting, with a precursor film of cells spreading around the aggregate with two possible states. In strongly cohesive aggregates this film is a cellular monolayer in the liquid state, whereas in weakly cohesive aggregates, cells escape from the aggregate, forming a 2D gas. The escape of isolated cells is a physical mechanism that appears also to be present in the progression of a noninvasive tumor into a metastatic malignant carcinoma, known as the epithelial-mesenchymal transition.

Micropatterned "adherent/repellent" glass surfaces for studying the spreading kinetics of individual red blood cells onto protein-decorated substrates

We report in this paper two simple and effective methods to decorate glass surfaces that enable protein micropatterning and subsequent spatially controlled adhesion of cells. The first method combines simultaneously the potentialities of two existing techniques, namely microcontact printing (μCP) and microfluidic networks (μFN) to achieve dual protein patterning in a single step. The second method is mainly based on the well-known property of poly(ethylene glycol) (PEG) to resist against protein adsorption. Both approaches were used to produce heterogeneous surfaces on which micron-size or submicronic streptavidin-coated lines alternate with cell-repellent areas. We first describe the implementation of the two methods and discuss the main pitfalls to avoid. Then, using these templates, we have monitored the kinetics of attachment of individual biotinylated (i.e. attractant towards streptavidin) red blood cells by directly measuring the propagation velocity of the adhesion front. Depending on the surface density of biotin, we found two distinct regimes, in agreement with existing theoretical models.

Robust method for high-throughput surface patterning of deformable substrates.

We describe a simple and robust method for high-throughput surface patterning of deformable substrates such as silicone rubber films covered with a thin layer of protein and cell antifouling hydrogel (PLL-g-PEG). The irradiation with deep UV (<200 nm) of PLL-g-PEG-coated rubber substrates through a synthetic quartz photomask created micropatterns over a large area of the substrate. Incubation with proteins resulted in stable patterns with high feature resolution. RPE1 cells seeded on fibronectin patterns were constrained for days even after stretching. We also propose the crossbow feature as an interesting example allowing the stretching of normalized stress fibers.

Mimicking cell/extracellular matrix adhesion with lipid membranes and solid substrates: requirements, pitfalls and proposals

The interest in physical approaches to the study of cell adhesion has generated numerous recent works on the development of substrates mimicking the extracellular matrix and the use of giant synthetic liposomes, commonly considered as basic models of living cells. The use of well-characterized bioactive substrates and artificial cells should allow us to gain new insight into the cell–extracellular matrix interactions, provided that their biomimetic relevance has been really proved. The aim of this paper is to define some minimal requirements for effective biomimetic features and to propose simple adhesion assays. We show, for instance, that immobilization of specific ligands is sometimes not sufficient t oe nsure specific adhesion of cells expressing the corresponding receptors. B yi nvestigating comparatively the adhesive behaviour of decorated erythrocytes and vesicles, we also discuss the potentialities and limitations of synthetic vesicles as test cells.

Nanofluidics in cellular tubes under oscillatory extension

Membrane nanotubes or tethers extruded from cells exhibit dynamic features that are believed to exhibit viscoelastic rheological properties. We have performed typical microrheology experiments on tethers pulled from red blood cells by measuring the force response to small oscillatory extensions or compressions. Our data, supported by a simple theoretical model, show that the force response does not reflect any intrinsic viscoelastic properties of the tethers themselves, but instead is dominated by the drainage of the internal cellular fluid into and out of the oscillating nanoconduit over a frequency-dependent penetration depth. The simplicity of tether rheology suggests its usage as a probe for measuring the local viscosity of the cytosol near the plasma membrane.

Intracellular transport: from physics to ... biology.

Considerable effort over the past three decades has allowed the identification of the protein families that control the cellular machinery responsible for intracellular transport within eukaryotic cells. These proteins are estimated to represent about 10–20% of the human “proteome.” The complexity of intracellular transport makes useful the development of model membranes. We describe here experimental systems based on lipid giant unilamellar vesicles (GUVs), which are attached to kinesin molecules. These systems give rise to thin membrane tubes and to complex tubular networks when incubated in vitro with microtubules and ATP. This type of assay, which mimics key events occurring during intracellular transport, allows physicists and biologists to understand how the unique mechanical properties of lipid membranes could be involved in the budding process, the sorting of cargo proteins and lipids, and the separation of the buds from a donor membrane.