Edouard Hannezo

Glass-like dynamics of collective cell migration

Collective cell migration in tissues occurs throughout embryonic development, during wound healing, and in cancerous tumor invasion, yet most detailed knowledge of cell migration comes from single-cell studies. As single cells migrate, the shape of the cell body fluctuates dramatically through cyclic processes of extension, adhesion, and retraction, accompanied by erratic changes in migration direction. Within confluent cell layers, such subcellular motions must be coupled between neighbors, yet the influence of these subcellular motions on collective migration is not known. Here we study motion within a confluent epithelial cell sheet, simultaneously measuring collective migration and subcellular motions, covering a broad range of length scales, time scales, and cell densities. At large length scales and time scales collective migration slows as cell density rises, yet the fastest cells move in large, multicell groups whose scale grows with increasing cell density. This behavior has an intriguing analogy to dynamic heterogeneities found in particulate systems as they become more crowded and approach a glass transition. In addition we find a diminishing self-diffusivity of short-wavelength motions within the cell layer, and growing peaks in the vibrational density of states associated with cooperative cell-shape fluctuations. Both of these observations are also intriguingly reminiscent of a glass transition. Thus, these results provide a broad and suggestive analogy between cell motion within a confluent layer and the dynamics of supercooled colloidal and molecular fluids approaching a glass transition.

Notch Lineages and Activity in Intestinal Stem Cells Determined by a New Set of Knock-In Mice

The conserved role of Notch signaling in controlling intestinal cell fate specification and homeostasis has been extensively studied. Nevertheless, the precise identity of the cells in which Notch signaling is active and the role of different Notch receptor paralogues in the intestine remain ambiguous, due to the lack of reliable tools to investigate Notch expression and function in vivo. We generated a new series of transgenic mice that allowed us, by lineage analysis, to formally prove that Notch1 and Notch2 are specifically expressed in crypt stem cells. In addition, a novel Notch reporter mouse, Hes1-EmGFPSAT, demonstrated exclusive Notch activity in crypt stem cells and absorptive progenitors. This roster of knock-in and reporter mice represents a valuable resource to functionally explore the Notch pathway in vivo in virtually all tissues.

Alignment of cellular motility forces with tissue flow as a mechanism for efficient wound healing

Recent experiments have shown that spreading epithelial sheets exhibit a long-range coordination of motility forces that leads to a buildup of tension in the tissue, which may enhance cell division and the speed of wound healing. Furthermore, the edges of these epithelial sheets commonly show finger-like protrusions whereas the bulk often displays spontaneous swirls of motile cells. To explain these experimental observations, we propose a simple flocking-type mechanism, in which cells tend to align their motility forces with their velocity. Implementing this idea in a mechanical tissue simulation, the proposed model gives rise to efficient spreading and can explain the experimentally observed long-range alignment of motility forces in highly disordered patterns, as well as the buildup of tensile stress throughout the tissue. Our model also qualitatively reproduces the dependence of swirl size and swirl velocity on cell density reported in experiments and exhibits an undulation instability at the edge of the spreading tissue commonly observed in vivo. Finally, we study the dependence of colony spreading speed on important physical and biological parameters and derive simple scaling relations that show that coordination of motility forces leads to an improvement of the wound healing process for realistic tissue parameters.

Instabilities of monolayered epithelia: shape and structure of villi and crypts.

We study theoretically the shapes of a dividing epithelial monolayer of cells lying on top of an elastic stroma. The negative tension created by cell division provokes a buckling instability at a finite wave vector leading to the formation of periodic arrays of villi and crypts. The instability is similar to the buckling of a metallic plate under compression. We use the results to rationalize the various structures of the intestinal lining observed in vivo. Taking into account the coupling between cell division and local curvature, we obtain different patterns of villi and crypts, which could explain the different morphologies of the small intestine and the colon.

Theory of epithelial sheet morphology in three dimensions

Significance Epithelia are the tissue layers that line organs throughout the body. Their complex movements and extensive reorganization have been widely studied as a model system of embryo development. Epithelial cells have been theoretically described using physical models close to those used for soap bubbles and foams. Nevertheless, although morphogenesis is intrinsically three-dimensional, previous works have mostly considered a two-dimensional planar geometry. In this paper, we provide a theoretical three-dimensional description of epithelial sheets, which describes within a single framework many developmental transitions, such as the formation of cavities or cellular tubes of a given size. We provide simple scaling laws that could be verified experimentally, for each of these transitions. Morphogenesis during embryo development requires the coordination of mechanical forces to generate the macroscopic shapes of organs. We propose a minimal theoretical model, based on cell adhesion and actomyosin contractility, which describes the various shapes of epithelial cells and the bending and buckling of epithelial sheets, as well as the relative stability of cellular tubes and spheres. We show that, to understand these processes, a full 3D description of the cells is needed, but that simple scaling laws can still be derived. The morphologies observed in vivo can be understood as stable points of mechanical equations and the transitions between them are either continuous or discontinuous. We then focus on epithelial sheet bending, a ubiquitous morphogenetic process. We calculate the curvature of an epithelium as a function of actin belt tension as well as of cell–cell and and cell–substrate tension. The model allows for a comparison of the relative stabilities of spherical or cylindrical cellular structures (acini or tubes). Finally, we propose a unique type of buckling instability of epithelia, driven by a flattening of individual cell shapes, and discuss experimental tests to verify our predictions.

Physics of active jamming during collective cellular motion in a monolayer

Significance Collective cell motion is very important in many biological processes such as wound healing, embryogenesis, or cancer progression. Nevertheless, it is not clear which parameters control the transition from freely moving single cells to collective jammed motion. In this article, we uncover complex dynamics as a cell monolayer ages, where cell motion is shown to gradually slow down with time, while the distance over which cell displacements are correlated first increases drastically and then decreases. This change of behavior is not controlled by cell density but rather by a maturation of the cell−cell and cell−substrate contacts. By comparing experiments, analytic model, and detailed particle-based simulations, we shed light on this biological amorphous solidification process. Although collective cell motion plays an important role, for example during wound healing, embryogenesis, or cancer progression, the fundamental rules governing this motion are still not well understood, in particular at high cell density. We study here the motion of human bronchial epithelial cells within a monolayer, over long times. We observe that, as the monolayer ages, the cells slow down monotonously, while the velocity correlation length first increases as the cells slow down but eventually decreases at the slowest motions. By comparing experiments, analytic model, and detailed particle-based simulations, we shed light on this biological amorphous solidification process, demonstrating that the observed dynamics can be explained as a consequence of the combined maturation and strengthening of cell−cell and cell−substrate adhesions. Surprisingly, the increase of cell surface density due to proliferation is only secondary in this process. This analysis is confirmed with two other cell types. The very general relations between the mean cell velocity and velocity correlation lengths, which apply for aggregates of self-propelled particles, as well as motile cells, can possibly be used to discriminate between various parameter changes in vivo, from noninvasive microscopy data.

Identity and dynamics of mammary stem cells during branching morphogenesis

During puberty, the mouse mammary gland develops into a highly branched epithelial network. Owing to the absence of exclusive stem cell markers, the location, multiplicity, dynamics and fate of mammary stem cells (MaSCs), which drive branching morphogenesis, are unknown. Here we show that morphogenesis is driven by proliferative terminal end buds that terminate or bifurcate with near equal probability, in a stochastic and time-invariant manner, leading to a heterogeneous epithelial network. We show that the majority of terminal end bud cells function as highly proliferative, lineage-committed MaSCs that are heterogeneous in their expression profile and short-term contribution to ductal extension. Yet, through cell rearrangements during terminal end bud bifurcation, each MaSC is able to contribute actively to long-term growth. Our study shows that the behaviour of MaSCs is not directly linked to a single expression profile. Instead, morphogenesis relies upon lineage-restricted heterogeneous MaSC populations that function as single equipotent pools in the long term.

Defining the clonal dynamics leading to mouse skin tumour initiation

The changes in cell dynamics after oncogenic mutation that lead to the development of tumours are currently unknown. Here, using skin epidermis as a model, we assessed the effect of oncogenic hedgehog signalling in distinct cell populations and their capacity to induce basal cell carcinoma, the most frequent cancer in humans. We found that only stem cells, and not progenitors, initiated tumour formation upon oncogenic hedgehog signalling. This difference was due to the hierarchical organization of tumour growth in oncogene-targeted stem cells, characterized by an increase in symmetric self-renewing divisions and a higher p53-dependent resistance to apoptosis, leading to rapid clonal expansion and progression into invasive tumours. Our work reveals that the capacity of oncogene-targeted cells to induce tumour formation is dependent not only on their long-term survival and expansion, but also on the specific clonal dynamics of the cancer cell of origin.

Cortical instability drives periodic supracellular actin pattern formation in epithelial tubes

Significance Robust pattern formation is a ubiquitous question of developmental biology. In 1952, Turing proposed in a seminal paper that this could be achieved by a hydrodynamical instability of two diffusing species reacting with each other, but direct experimental evidence of how a mechanism is implemented biologically is still lacking. In this paper, we show by a combination of experiment and theory that the actin cytoskeleton, one of the main force-producing mechanisms in biology, has the property to self-organize into regular supracellular patterns in vivo in Drosophila. We show that the wavelength of the pattern depends on the physical properties of the gel and can be modified experimentally. An essential question of morphogenesis is how patterns arise without preexisting positional information, as inspired by Turing. In the past few years, cytoskeletal flows in the cell cortex have been identified as a key mechanism of molecular patterning at the subcellular level. Theoretical and in vitro studies have suggested that biological polymers such as actomyosin gels have the property to self-organize, but the applicability of this concept in an in vivo setting remains unclear. Here, we report that the regular spacing pattern of supracellular actin rings in the Drosophila tracheal tubule is governed by a self-organizing principle. We propose a simple biophysical model where pattern formation arises from the interplay of myosin contractility and actin turnover. We validate the hypotheses of the model using photobleaching experiments and report that the formation of actin rings is contractility dependent. Moreover, genetic and pharmacological perturbations of the physical properties of the actomyosin gel modify the spacing of the pattern, as the model predicted. In addition, our model posited a role of cortical friction in stabilizing the spacing pattern of actin rings. Consistently, genetic depletion of apical extracellular matrix caused strikingly dynamic movements of actin rings, mirroring our model prediction of a transition from steady to chaotic actin patterns at low cortical friction. Our results therefore demonstrate quantitatively that a hydrodynamical instability of the actin cortex can trigger regular pattern formation and drive morphogenesis in an in vivo setting.

Balance between Apical Membrane Growth and Luminal Matrix Resistance Determines Epithelial Tubule Shape

The morphological stability of biological tubes is crucial for the efficient circulation of fluids and gases. Failure of this stability causes irregularly shaped tubes found in multiple pathological conditions. Here, we report that Drosophila mutants of the ESCRT III component Shrub/Vps32 exhibit a strikingly elongated sinusoidal tube phenotype. This is caused by excessive apical membrane synthesis accompanied by the ectopic accumulation and overactivation of Crumbs in swollen endosomes. Furthermore, we demonstrate that the apical extracellular matrix (aECM) of the tracheal tube is a viscoelastic material coupled with the apical membrane. We present a simple mechanical model in which aECM elasticity, apical membrane growth, and their interaction are three vital parameters determining the stability of biological tubes. Our findings demonstrate a mechanical role for the extracellular matrix and suggest that the interaction of the apical membrane and an elastic aECM determines the final morphology of biological tubes independent of cell shape.