Dynamics of anisotropic colloidal systems: What to choose, DLS, DDM or XPCS?

Investigation of the dynamics of colloids in bulk can be hindered by issues such as multiple scattering and sample opacity. These challenges are exacerbated when dealing with inorganic materials. In this study, we employed a model system of Akaganeite colloidal rods to assess three leading dynamics measurement techniques: 3D-(depolarized) dynamic light scattering (3D-(D)DLS), polarized-differential dynamic microscopy (P-DDM), and x-ray photon correlation spectroscopy (XPCS). Our analysis revealed that the translational and rotational diffusion coefficients captured by these methods show a remarkable alignment. Additionally, by examining the q-ranges and maximum volume fractions for each approach, we offer insights into the best technique for investigating the dynamics of anisotropic systems at the colloidal scale.

Tissue fluidification promotes a cGAS-STING cytosolic DNA response in invasive breast cancer.

The process in which locally confined epithelial malignancies progressively evolve into invasive cancers is often promoted by unjamming, a phase transition from a solid-like to a liquid-like state, which occurs in various tissues. Whether this tissue-level mechanical transition impacts phenotypes during carcinoma progression remains unclear. Here we report that the large fluctuations in cell density that accompany unjamming result in repeated mechanical deformations of cells and nuclei. This triggers a cellular mechano-protective mechanism involving an increase in nuclear size and rigidity, heterochromatin redistribution and remodelling of the perinuclear actin architecture into actin rings. The chronic strains and stresses associated with unjamming together with the reduction of Lamin B1 levels eventually result in DNA damage and nuclear envelope ruptures, with the release of cytosolic DNA that activates a cGAS-STING (cyclic GMP-AMP synthase-signalling adaptor stimulator of interferon genes)-dependent cytosolic DNA response gene program. This mechanically driven transcriptional rewiring ultimately alters the cell state, with the emergence of malignant traits, including epithelial-to-mesenchymal plasticity phenotypes and chemoresistance in invasive breast carcinoma.

Echo speckle imaging of dynamic processes in soft materials.

We present a laser-speckle imaging technique, termed Echo speckle imaging (ESI), that quantifies the local dynamics in biological tissue and soft materials with a noise level around or below 10% of the measured signal without affecting the spatial resolution. We achieve this through an unconventional speckle beam illumination that creates changing, statistically independent illumination conditions and substantially increases the measurement accuracy. Control experiments for dynamically homogeneous and heterogeneous soft materials and tissue phantoms illustrate the performance of the method. We show that this approach enables us to precision-monitor purely dynamic heterogeneities in turbid soft media with a lateral resolution of 100 µm and better.

Non-invasive measurement of nuclear relative stiffness from quantitative analysis of microscopy data.

The connection between the properties of a cell tissue and those of the single constituent cells remains to be elucidated. At the purely mechanical level, the degree of rigidity of different cellular components, such as the nucleus and the cytoplasm, modulates the interplay between the cell inner processes and the external environment, while simultaneously mediating the mechanical interactions between neighboring cells. Being able to quantify the correlation between single-cell and tissue properties would improve our mechanobiological understanding of cell tissues. Here we develop a methodology to quantitatively extract a set of structural and motility parameters from the analysis of time-lapse movies of nuclei belonging to jammed and flocking cell monolayers. We then study in detail the correlation between the dynamical state of the tissue and the deformation of the nuclei. We observe that the nuclear deformation rate linearly correlates with the local divergence of the velocity field, which leads to a non-invasive estimate of the elastic modulus of the nucleus relative to the one of the cytoplasm. We also find that nuclei belonging to flocking monolayers, subjected to larger mechanical perturbations, are about two time stiffer than nuclei belonging to dynamically arrested monolayers, in agreement with atomic force microscopy results. Our results demonstrate a non-invasive route to the determination of nuclear relative stiffness for cells in a monolayer.

Multiscale heterogeneous dynamics in two-dimensional glassy colloids.

On approaching the glass transition, a dense colloid exhibits a dramatic slowdown with minute structural changes. Most microscopy experiments directly follow the motion of individual particles in real space, whereas scattering experiments typically probe the collective dynamics in reciprocal space at variable wavevector q. Multiscale studies of glassy dynamics are experimentally demanding and, thus, seldom performed. By using two-dimensional hard-sphere colloids at various area fractions ϕ, we show here that Differential Dynamic Microscopy (DDM) can be effectively used to measure the collective dynamics of a glassy colloid in a range of q within a single experiment. As ϕ is increased, the single decay of the intermediate scattering functions is progressively replaced by a more complex relaxation that we fit to a sum of two stretched-exponential decays. The slowest process, corresponding to the long-time particle escapes from caging, has a characteristic time τs = 1/(DLq2) with diffusion coefficient DL∼(ϕc-ϕ)2.8, and ϕc ≃ 0.81. The fast process exhibits, instead, a non-Brownian scaling of the characteristic time τf(q) and a relative amplitude a(q) that monotonically increases with q. Despite the non-Brownian nature of τf(q), we succeed in estimating the short-time diffusion coefficient Dcage, whose ϕ-dependence is practically negligible compared to the one of DL. Finally, we extend DDM to measure the q-dependent dynamical susceptibility χ4(q, t), a powerful yet hard-to-access multiscale indicator of dynamical heterogeneities. Our results show that DDM is a convenient tool to study the dynamics of colloidal glasses over a broad range of time and length scales.

Bistability of Dielectrically Anisotropic Nematic Crystals and the Adaptation of Endothelial Collectives to Stress Fields.

Endothelial monolayers physiologically adapt to flow and flow-induced wall shear stress, attaining ordered configurations in which elongation, orientation, and polarization are coherently organized over many cells. Here, with the flow direction unchanged, a peculiar bi-stable (along the flow direction or perpendicular to it) cell alignment is observed, emerging as a function of the flow intensity alone, while cell polarization is purely instructed by flow directionality. Driven by the experimental findings, the parallelism between endothelia is delineated under a flow field and the transition of dual-frequency nematic liquid crystals under an external oscillatory electric field. The resulting physical model reproduces the two stable configurations and the energy landscape of the corresponding system transitions. In addition, it reveals the existence of a disordered, metastable state emerging upon system perturbation. This intermediate state, experimentally demonstrated in endothelial monolayers, is shown to expose the cellular system to a weakening of cell-to-cell junctions to the detriment of the monolayer integrity. The flow-adaptation of monolayers composed of healthy and senescent endothelia is successfully predicted by the model with adjustable nematic parameters. These results may help to understand the maladaptive response of in vivo endothelial tissues to disturbed hemodynamics and the progressive functional decay of senescent endothelia.

Deformation profiles and microscopic dynamics of complex fluids during oscillatory shear experiments.

Oscillatory shear tests are widely used in rheology to characterize the linear and non-linear mechanical response of complex fluids, including the yielding transition. There is an increasing urge to acquire detailed knowledge of the deformation field that is effectively present across the sample during these tests; at the same time, there is mounting evidence that the macroscopic rheological response depends on the elusive microscopic behavior of the material constituents. Here we employ a strain-controlled shear-cell with transparent walls to visualize and quantify the dynamics of tracers embedded in various cyclically sheared complex fluids, ranging from almost-ideal elastic to yield stress fluids. For each sample, we use image correlation processing to measure the macroscopic deformation field, and echo-differential dynamic microscopy to probe the microscopic irreversible sample dynamics in reciprocal space; finally, we devise a simple scheme to spatially map the rearrangements in the sheared sample, once again without tracking the tracers. For the yield stress sample, we obtain a wave-vector dependent characterization of shear-induced diffusion across the yielding transition, which is accompanied by a three-order-of-magnitude speed-up of the dynamics and by a transition from localized, intermittent rearrangements to a more spatially homogeneous and temporally uniform activity. Our tracking free approach is intrinsically multi-scale, can successfully discriminate between different types of dynamics, and can be automated to minimize user intervention. Applications are many, as it can be translated to other imaging modes, including fluorescence, and can be used with sub-resolution tracers and even without tracers, for samples that provide intrinsic optical contrast.

Hecw controls oogenesis and neuronal homeostasis by promoting the liquid state of ribonucleoprotein particles.

Specialised ribonucleoprotein (RNP) granules are a hallmark of polarized cells, like neurons and germ cells. Among their main functions is the spatial and temporal modulation of the activity of specific mRNA transcripts that allow specification of primary embryonic axes. While RNPs composition and role are well established, their regulation is poorly defined. Here, we demonstrate that Hecw, a newly identified Drosophila ubiquitin ligase, is a key modulator of RNPs in oogenesis and neurons. Hecw depletion leads to the formation of enlarged granules that transition from a liquid to a gel-like state. Loss of Hecw activity results in defective oogenesis, premature aging and climbing defects associated with neuronal loss. At the molecular level, reduced ubiquitination of the Fmrp impairs its translational repressor activity, resulting in altered Orb expression in nurse cells and Profilin in neurons.

Probing roto-translational diffusion of small anisotropic colloidal particles with a bright-field microscope.

Soft and biological materials are often composed of elementary constituents exhibiting an incessant roto-translational motion at the microscopic scale. Tracking this motion with a bright-field microscope becomes increasingly challenging when the particle size becomes smaller than the microscope resolution, a case which is frequently encountered. Here we demonstrate squared-gradient differential dynamic microscopy (SG-DDM) as a tool to successfully use bright-field microscopy to extract the roto-translational dynamics of small anisotropic colloidal particles, whose rotational motion cannot be tracked accurately in direct space. We provide analytical justification and experimental demonstration of the method by successful application to an aqueous suspension of peanut-shaped particles.

Disentangling collective motion and local rearrangements in 2D and 3D cell assemblies.

The accurate quantification of cellular motility and of the structural changes occurring in multicellular aggregates is critical in describing and understanding key biological processes, such as wound repair, embryogenesis and cancer invasion. Current methods based on cell tracking or velocimetry either suffer from limited spatial resolution or are challenging and time-consuming, especially for three-dimensional (3D) cell assemblies. Here we propose a conceptually simple, robust and tracking-free approach for the quantification of the dynamical activity of cells via a two-step procedure. We first characterise the global features of the collective cell migration by registering the temporal stack of the acquired images. As a second step, a map of the local cell motility is obtained by performing a mean squared amplitude analysis of the intensity fluctuations occurring when two registered image frames acquired at different times are subtracted. We successfully apply our approach to cell monolayers undergoing a jamming transition, as well as to monolayers and 3D aggregates that exhibit a cooperative unjamming-via-flocking transition. Our approach is capable of disentangling very efficiently and of assessing accurately the global and local contributions to cell motility.

Multiple dynamic regimes in a coarsening foam.

Intermittent dynamics driven by internal stress imbalances in disordered systems is a fascinating yet poorly understood phenomenon. Here, we study it for a coarsening foam. By exploiting differential dynamic microscopy and particle tracking we determine the dynamical characteristics of the foam at different ages in reciprocal and direct space, respectively. At all wavevectors q investigated, the intermediate scattering function exhibits a compressed exponential decay. However, the access to unprecedentedly small values of q highlights the existence of two distinct regimes for the q-dependence of the foam relaxation rate Γ(q). At high q, Γ(q) ∼ q consistent with directionally-persistent and intermittent bubble displacements. At low q, we find the surprising scaling Γ(q) ∼ q δ , with δ = 1.6 ± 0.2. The analysis of the bubble displacement distribution in real space reveals the existence of a displacement cut-off of the order of the bubble diameter. Introducing such cut-off length in an existing model, describing stress-driven dynamics in disordered systems, fully accounts for the observed behavior in direct and reciprocal space.

Emergence of Multiscale Dynamics in Colloidal Gels.

To gain insight into the kinetics of colloidal gel evolution at low particle volume fractions ϕ, we utilize differential dynamic microscopy to investigate particle aggregation, geometric percolation, and the subsequent transition to nonergodic dynamics. We report the emergence of unexpectedly rich multiscale dynamics upon the onset of nonergodicity, which separates the wave vectors q into three different regimes. In the high-q domain, the gel exhibits ϕ-independent internal vibrations of fractal clusters. The intermediate-q domain is dominated by density fluctuations at the length scale of the clusters, as evidenced by the q independence of the relaxation time τ. In the low-q domain, the scaling of τ as q^{-3} suggests that the network appears homogeneous. The transitions between these three regimes introduce two characteristic length scales, distinct from the cluster size.

Functional transcription promoters at DNA double-strand breaks mediate RNA-driven phase separation of damage-response factors.

Damage-induced long non-coding RNAs (dilncRNA) synthesized at DNA double-strand breaks (DSBs) by RNA polymerase II are necessary for DNA-damage-response (DDR) focus formation. We demonstrate that induction of DSBs results in the assembly of functional promoters that include a complete RNA polymerase II preinitiation complex, MED1 and CDK9. Absence or inactivation of these factors causes a reduction in DDR foci both in vivo and in an in vitro system that reconstitutes DDR events on nucleosomes. We also show that dilncRNAs drive molecular crowding of DDR proteins, such as 53BP1, into foci that exhibit liquid-liquid phase-separation condensate properties. We propose that the assembly of DSB-induced transcriptional promoters drives RNA synthesis, which stimulates phase separation of DDR factors in the shape of foci.

Unjamming overcomes kinetic and proliferation arrest in terminally differentiated cells and promotes collective motility of carcinoma.

During wound repair, branching morphogenesis and carcinoma dissemination, cellular rearrangements are fostered by a solid-to-liquid transition, known as unjamming. The biomolecular machinery behind unjamming and its pathophysiological relevance remain, however, unclear. Here, we study unjamming in a variety of normal and tumorigenic epithelial two-dimensional (2D) and 3D collectives. Biologically, the increased level of the small GTPase RAB5A sparks unjamming by promoting non-clathrin-dependent internalization of epidermal growth factor receptor that leads to hyperactivation of the kinase ERK1/2 and phosphorylation of the actin nucleator WAVE2. This cascade triggers collective motility effects with striking biophysical consequences. Specifically, unjamming in tumour spheroids is accompanied by persistent and coordinated rotations that progressively remodel the extracellular matrix, while simultaneously fluidizing cells at the periphery. This concurrent action results in collective invasion, supporting the concept that the endo-ERK1/2 pathway is a physicochemical switch to initiate collective invasion and dissemination of otherwise jammed carcinoma.

Flocking transitions in confluent tissues.

Collective cell migration in dense tissues underlies important biological processes, such as embryonic development, wound healing and cancer invasion. While many aspects of single cell movements are now well established, the mechanisms leading to displacements of cohesive cell groups are still poorly understood. To elucidate the emergence of collective migration in mechanosensitive cells, we examine a self-propelled Voronoi (SPV) model of confluent tissues with an orientational feedback that aligns a cell's polarization with its local migration velocity. While shape and motility are known to regulate a density-independent liquid-solid transition in tissues, we find that aligning interactions facilitate collective motion and promote solidification, with transitions that can be predicted by extending statistical physics tools such as effective temperature to this far-from-equilibrium system. In addition to accounting for recent experimental observations obtained with epithelial monolayers, our model predicts structural and dynamical signatures of flocking, which may serve as gateway to a more quantitative characterization of collective motility.

Dark field differential dynamic microscopy enables accurate characterization of the roto-translational dynamics of bacteria and colloidal clusters.

Micro- and nanoscale objects with anisotropic shape are key components of a variety of biological systems and inert complex materials, and represent fundamental building blocks of novel self-assembly strategies. The time scale of their thermal motion is set by their translational and rotational diffusion coefficients, whose measurement may become difficult for relatively large particles with small optical contrast. Here we show that dark field differential dynamic microscopy is the ideal tool for probing the roto-translational Brownian motion of anisotropic shaped particles. We demonstrate our approach by successful application to aqueous dispersions of non-motile bacteria and of colloidal aggregates of spherical particles.

Image windowing mitigates edge effects in Differential Dynamic Microscopy.

Differential Dynamic Microscopy (DDM) analyzes traditional real-space microscope images to extract information on sample dynamics in a way akin to light scattering, by decomposing each image in a sequence into Fourier modes, and evaluating their time correlation properties. DDM has been applied in a number of soft-matter and colloidal systems. However, objects observed to move out of the microscope's captured field of view, intersecting the edges of the acquired images, can introduce spurious but significant errors in the subsequent analysis. Here we show that application of a spatial windowing filter to images in a sequence before they enter the standard DDM analysis can reduce these artifacts substantially. Moreover, windowing can increase significantly the accessible range of wave vectors probed by DDM, and may further yield unexpected information, such as the size polydispersity of a colloidal suspension.

Active diffusion and advection in Drosophila oocytes result from the interplay of actin and microtubules.

Transport in cells occurs via a delicate interplay of passive and active processes, including diffusion, directed transport and advection. Despite progress in super-resolution microscopy, discriminating and quantifying these processes is a challenge, requiring tracking of rapidly moving, sub-diffraction objects in a crowded, noisy environment. Here we use differential dynamic microscopy with different contrast mechanisms to provide a thorough characterization of the dynamics in the Drosophila oocyte. We study the movement of vesicles and the elusive motion of a cytoplasmic F-actin mesh, a known regulator of cytoplasmic flows. We find that cytoplasmic motility constitutes a combination of directed motion and random diffusion. While advection is mainly attributed to microtubules, we find that active diffusion is driven by the actin cytoskeleton, although it is also enhanced by the flow. We also find that an important dynamic link exists between vesicles and cytoplasmic F-actin motion, as recently suggested in mouse oocytes.