Tutorial: fluorescence lifetime microscopy of membrane mechanosensitive Flipper probes.

Measuring forces within living cells remains a technical challenge. In this Tutorial, we cover the development of hydrophobic mechanosensing fluorescent probes called Flippers, whose fluorescence lifetime depends on lipid packing. Flipper probes can therefore be used as reporters for membrane tension via the measurement of changes in their fluorescence lifetime.

Apical dehydration impairs the cystic fibrosis airway epithelium barrier via a 1-integrin/YAP1 pathway.

Defective hydration of airway surface mucosa is associated with lung infection in cystic fibrosis (CF), partly caused by disruption of the epithelial barrier integrity. Although rehydration of the CF airway surface liquid (ASL) alleviates epithelium vulnerability to infection by junctional protein expression, the mechanisms linking ASL to barrier integrity are unknown. We show here the strong degradation of YAP1 and TAZ proteins in well-polarized CF human airway epithelial cells (HAECs), a process that was prevented by ASL rehydration.

Curvature in Biological Systems: Its Quantification, Emergence, and Implications across the Scales.

Surface curvature both emerges from, and influences the behavior of, living objects at length scales ranging from cell membranes to single cells to tissues and organs. The relevance of surface curvature in biology is supported by numerous experimental and theoretical investigations in recent years. In this review, first, a brief introduction to the key ideas of surface curvature in the context of biological systems is given and the challenges that arise when measuring surface curvature are discussed.

Epithelial cells adapt to curvature induction via transient active osmotic swelling.

Generation of tissue curvature is essential to morphogenesis. However, how cells adapt to changing curvature is still unknown because tools to dynamically control curvature in vitro are lacking. Here, we developed self-rolling substrates to study how flat epithelial cell monolayers adapt to a rapid anisotropic change of curvature.

Poly-l-lysine/Laminin Surface Coating Reverses Glial Cell Mechanosensitivity on Stiffness-Patterned Hydrogels.

Brain tissues demonstrate heterogeneous mechanical properties, which evolve with aging and pathologies. The observation in these tissues of smooth to sharp rigidity gradients raises the question of brain cell responses to both different values of rigidity and their spatial variations, in dependence on the surface chemistry they are exposed to. Here, we used recent techniques of hydrogel photopolymerization to achieve stiffness texturing down to micrometer resolution in polyacrylamide hydrogels.

Cells on Hydrogels with Micron-Scaled Stiffness Patterns Demonstrate Local Stiffness Sensing.

Cell rigidity sensing-a basic cellular process allowing cells to adapt to mechanical cues-involves cell capabilities exerting force on the extracellular environment. In vivo, cells are exposed to multi-scaled heterogeneities in the mechanical properties of the surroundings. Here, we investigate whether cells are able to sense micron-scaled stiffness textures by measuring the forces they transmit to the extracellular matrix.

Modulation of designer biomimetic matrices for optimized differentiated intestinal epithelial cultures.

The field of intestinal biology is thirstily searching for different culture methods that complement the limitations of organoids, particularly the lack of a differentiated intestinal compartment. While being recognized as an important milestone for basic and translational biological studies, many primary cultures of intestinal epithelium (IE) rely on empirical trials using hydrogels of various stiffness, whose mechanical impact on epithelial organization remains vague until now. Here, we report the development of hydrogel scaffolds with a range of elasticities and their influence on IE expansion, organization, and differentiation.

Bending toward differentiation.

Both biochemical and mechanical signals coordinate all processes at the origin of the formation of functional organs, including tissue folding, cell shape, and differentiation. In this issue of Developmental Cell, Blonski et al. establish a direct consequence of epithelial monolayer folding on nuclear shape and gene expression.

Laser-Assisted Strain Engineering of Thin Elastomer Films to Form Variable Wavy Substrates for Cell Culture.

Endothelial and epithelial cells usually grow on a curved environment, at the surface of organs, which many techniques have tried to reproduce. Here a simple method is proposed to control curvature of the substrate. Prestrained thin elastomer films are treated by infrared laser irradiation in order to rigidify the surface of the film.

Curvature-dependent constraints drive remodeling of epithelia.

Epithelial tissues function as barriers that separate the organism from the environment. They usually have highly curved shapes, such as tubules or cysts. However, the processes by which the geometry of the environment and the cell's mechanical properties set the epithelium shape are not yet known.

A fluorescent membrane tension probe.

Cells and organelles are delimited by lipid bilayers in which high deformability is essential to many cell processes, including motility, endocytosis and cell division. Membrane tension is therefore a major regulator of the cell processes that remodel membranes, albeit one that is very hard to measure in vivo. Here we show that a planarizable push-pull fluorescent probe called FliptR (fluorescent lipid tension reporter) can monitor changes in membrane tension by changing its fluorescence lifetime as a function of the twist between its fluorescent groups.

Geometrical Determinants of Neuronal Actin Waves.

Hippocampal neurons produce in their early stages of growth propagative, actin-rich dynamical structures called actin waves. The directional motion of actin waves from the soma to the tip of neuronal extensions has been associated with net forward growth, and ultimately with the specification of neurites into axon and dendrites. Here, geometrical cues are used to control actin wave dynamics by constraining neurons on adhesive stripes of various widths.

Tuning the adhesive geometry of neurons: length and polarity control.

Neurons acquire their functional and morphological axo-dendritic polarity by extending, from competing minor processes (neurites), one long axon among numerous dendrites. We employed complementary sets of micropatterns built from 2 and 6 μm wide stripes of various lengths to constrain hippocampal neuron shapes. Using these geometries, we have (i) limited the number of neuronal extensions to obtain a minimal in vitro system of bipolar neurons and (ii) controlled the neurite width during growth by the generation of a progressive cell shape asymmetry on either side of the cellular body.