Isotropic Spreading Research Articles

Особенности турбулентности в электромагнитном перемешивателе жидкого металла

In the research work, conduction velocity sensors were successfully used to study the characteristics of turbulent flows arising in a liquid metal under the action of a rotating magnetic field in a vertical cylinder. At a certain magnitude of electromagnetic influence, on the graphs showing spectral energy of velocity pulsations there is observed a slope of ‘-5/3’, indicating developed homogeneous isotropic turbulence and spreading in a frequency range of up to two decades. The level of velocity pulsations varies from 4 to 10 % of the azimuthal rotation velocity. It is shown that the identified peaks in the velocity pulsation spectra correspond to the frequency of the liquid metal rotation around the cylinder axis. A nonlinear graph of the dependence of the liquid metal rotation frequency on the current frequency in the coils with a maximum near 50-75 Hz was obtained.

A unique role for clathrin light chain A in cell spreading and migration.

Clathrin heavy chain is the structural component of the clathrin triskelion, but unique functions for the two distinct and highly conserved clathrin light chains (CLCa and CLCb, also known as CLTA and CLTB, respectively) have been elusive. Here, we show that following detachment and replating, CLCa is uniquely responsible for promoting efficient cell spreading and migration. Selective depletion of CLCa, but not of CLCb, reduced the initial phase of isotropic spreading of HeLa, H1299 and HEK293 cells by 60-80% compared to siRNA controls, and wound closure and motility by ∼50%. Surface levels of β1-integrins were unaffected by CLCa depletion. However, CLCa was required for effective targeting of FAK (also known as PTK2) and paxillin to the adherent surface of spreading cells, for integrin-mediated activation of Src, FAK and paxillin, and for maturation of focal adhesions, but not their microtubule-based turnover. Depletion of CLCa also blocked the interaction of clathrin with the nucleation-promoting factor WAVE complex, and altered actin distribution. Furthermore, preferential recruitment of CLCa to budding protrusions was also observed. These results comprise the first identification of CLCa-specific functions, with implications for normal and neoplastic integrin-based signaling and cell migration.

Trabecular Architecture Determines Impulse Propagation Through the Early Embryonic Mouse Heart.

Most embryonic ventricular cardiomyocytes are quite uniform, in contrast to the adult heart, where the specialized ventricular conduction system is molecularly and functionally distinct from the working myocardium. We thus hypothesized that the preferential conduction pathway within the embryonic ventricle could be dictated by trabecular geometry. Mouse embryonic hearts of the Nkx2.5:eGFP strain between ED9.5 and ED14.5 were cleared and imaged whole mount by confocal microscopy, and reconstructed in 3D at 3.4 μm isotropic voxel size. The local orientation of the trabeculae, responsible for the anisotropic spreading of the signal, was characterized using spatially homogenized tensors (3 × 3 matrices) calculated from the trabecular skeleton. Activation maps were simulated assuming constant speed of spreading along the trabeculae. The results were compared with experimentally obtained epicardial activation maps generated by optical mapping with a voltage-sensitive dye. Simulated impulse propagation starting from the top of interventricular septum revealed the first epicardial breakthrough at the interventricular grove, similar to experimentally obtained activation maps. Likewise, ectopic activation from the left ventricular base perpendicular to dominant trabecular orientation resulted in isotropic and slower impulse spreading on the ventricular surface in both simulated and experimental conditions. We conclude that in the embryonic pre-septation heart, the geometry of the A-V connections and trabecular network is sufficient to explain impulse propagation and ventricular activation patterns.

Dynamic microtubules drive fibroblast spreading

ABSTRACTWhen cells with a mesenchymal type of motility come into contact with an adhesive substrate they adhere and start spreading by the formation of lamellipodia. Using a label-free approach and virtual synchronization approach we analyzed spreading in fibroblasts and cancer cells. In all cell lines spreading is a non-linear process undergoing isotropic or anisotropic modes with first fast (5–20 min) and then slow (30–120 min) phases. In the first 10 min cell area increases 2–4 times, while the absolute rate of initial spreading decreases 2–8 times. Fast spreading depends on actin polymerization and dynamic microtubules. Inhibition of microtubule growth was sufficient for a slowdown of initial spreading. Inhibition of myosin II in the presence of stable microtubules restored fast spreading. Inhibition of actin polymerization or complete depolymerization of microtubules slowed down fast spreading. However, in these cases inhibition of myosin II only partially restored spreading kinetics. We conclude that rapid growth of microtubules towards cell margins at the first stage of cell spreading temporarily inhibits phosphorylation of myosin II and is essential for the fast isotropic spreading. Comparison of the fibroblasts with cancer cells shows that fast spreading in different cell types shares similar kinetics and mechanisms, and strongly depends on dynamic microtubules.

Forced Spreading of Aqueous Solutions on Zwitterionic Sulfobetaine Surfaces for Rapid Evaporation and Solute Separation.

Solute separation of aqueous mixtures is mainly dominated by water vaporization. The evaporation rate of an aqueous drop grows with increasing the liquid-gas interfacial area. The spontaneous spreading behavior of a water droplet on a total wetting surface provides huge liquid-gas interfacial area per unit volume; however, it is halted by the self-pinning phenomenon upon addition of nonvolatile solutes. In this work, it is shown that the solute-induced self-pinning can be overcome by gravity, leading to anisotropic spreading much faster than isotropic spreading. The evaporation rate of anisotropic spreading on a zwitterionic sulfobetaine surface is 25 times larger as that on a poly(methyl methacrylate) surface. Dramatic enhancement of evaporation is demonstrated by simultaneous formation of fog atop liquid film. During anisotropic spreading, the solutes are quickly precipitated out within 30 s, showing the rapid solute-water separation. After repeated spreading process for the dye-containing solution, the mean concentration of the collection is doubled, revealing the concentration efficiency as high as 100%. Gravity-enhanced spreading on total wetting surfaces at room temperature is easy to scale-up with less energy consumption, and thus it has great potentials for the applications of solute separation and concentration.

Synergetic effect of laser patterning and micro coining for controlled lubricant propagation

In this study, the anisotropic spreading behavior of Poly-(alpha)-olefin oil (kinematic viscosity of 7.8 cSt at 100 °C) on stainless steel samples (AISI 403) having periodic, channel-like structures produced by hot micro-coining (periodicity of 400 μm and depth of 40 μm) as well as multi-scale structures (coining and laser patterning) was investigated. These results were compared to the behavior of periodic channels fabricated by direct laser interference patterning (periodicity of 5 μm and depth of 1 μm). The spreading behavior of a droplet (3 μl) was studied for a polished reference as well as for all modified surfaces and recorded by a digital light microscope. From this study, it can be concluded that the polished reference leads to an isotropic spreading behavior resulting from the stochastic surface roughness without any preferential orientation whereas all structured samples induce an anisotropic spreading behavior but with different degrees of anisotropy. The observed behavior can be well correlated with pinning induced by the grooves thus hindering the droplet propagation perpendicular to the grooves and the generation of capillary forces which favor the droplet movement along the grooves. It could be proved that the structural depth is a very desicive parameter with regard to the resulting spreading behavior. The multi-scale surface combining large structural depths and the steeper pattern geometry of the micro-coined surface with much smaller grooves of the laser-structure shows the largest anisotropic spreading behavior due to a stronger pinning and increased capillary forces.

Kindlin-2 cooperates with talin to activate integrins and induces cell spreading by directly binding paxillin.

Integrins require an activation step prior to ligand binding and signaling. How talin and kindlin contribute to these events in non-hematopoietic cells is poorly understood. Here we report that fibroblasts lacking either talin or kindlin failed to activate β1 integrins, adhere to fibronectin (FN) or maintain their integrins in a high affinity conformation induced by Mn(2+). Despite compromised integrin activation and adhesion, Mn(2+) enabled talin- but not kindlin-deficient cells to initiate spreading on FN. This isotropic spreading was induced by the ability of kindlin to directly bind paxillin, which in turn bound focal adhesion kinase (FAK) resulting in FAK activation and the formation of lamellipodia. Our findings show that talin and kindlin cooperatively activate integrins leading to FN binding and adhesion, and that kindlin subsequently assembles an essential signaling node at newly formed adhesion sites in a talin-independent manner.

Recombinant Phage Coated 1D Al2O3 Nanostructures for Controlling the Adhesion and Proliferation of Endothelial Cells.

A novel synthesis of a nanostructured cell adhesive surface is investigated for future stent developments. One-dimensional (1D) Al2O3 nanostructures were prepared by chemical vapor deposition of a single source precursor. Afterwards, recombinant filamentous bacteriophages which display a short binding motif with a cell adhesive peptide (RGD) on p3 and p8 proteins were immobilized on these 1D Al2O3 nanostructures by a simple dip-coating process to study the cellular response of human endothelial EA hy.926. While the cell density decreased on as-deposited 1D Al2O3 nanostructures, we observed enhanced cell proliferation and cell-cell interaction on recombinant phage overcoated 1D Al2O3 nanostructures. The recombinant phage overcoating also supports an isotropic cell spreading rather than elongated cell morphology as we observed on as-deposited Al2O3 1D nanostructures.

Experimental and numerical insights into isotropic spreading and deterministic dewetting of dielectrowetted films.

Dielectrowetting effects of surface wrinkling, isotropic vs anisotropic spreading, electrode geometry, and deterministic dewetting are presented both experimentally and by 3D numerical modeling. The numerical results are generated by COMSOL in conjunction with the phase-field and electrohydrodynamic methods, including comparisons to experimental data. The dynamic behavior of the two-phase system has been accurately characterized on both the macro- and microscopic level. This work provides a deeper theoretical insight into the operating physics of dielectrowetting superspreading devices.

Microtubules suppress blebbing and stimulate lamella extension in spreading fibroblasts

We compared spreading of Vero fibroblasts when microtubules were depolymerized or stabilized. After initial attachment cells start blebbing that continues for different time and abruptly transfers into spreading. After spreading initiation, most cells spread in an anisotropic manner through stochastic formation of lamellipodia. A second mode was rapid, isotropic spreading via formation of circular lamellum that occurs in 15% of cells. The rate of spreading was maximal at the beginning and decreased during the first hour according to logarithmic law. After 60 min many cells formed stable efges and started migrating on the substrate. However, cell area slowly continued to increase. Actin bundles are formed 20 min after cell attachment and they first run along cell boundary. This system disassembles within 20-40 min and is substituted with stress fibers crossing the cell. In the isotropically spread cells no actin bunbles are seen. Microtubules in the spreading cells enter into large blebs and all nascent lamella and later form radial array. When MTs has been depolymerized or stabilized blebbing started before cells attached to the substrate and continue much longer than in control cells. In both cases the initial rate of spreading decrease several fold, and remains constant for many hours. After 24 h the mean area occupied by cells with altered MT system was the same as in control. Alteration of MT system had moderate effect on actin system--formation of actin cables started at the same time as in control (within 20 min upon cell attachment), however, they grew even in cells undergoing prolonged blebbing. Actin cables running along cell margin were similar to tat in control cells, but they did not disappear up to 1 h. When stabilized, microtubules form chaotic array: they do not enter blebs and in spread cells run parallel to the cell margin at a distance of 3-5 microm. We conclude that dynamic microtubules speed up completion of blebbing and promote early stages of fibroblasts spreading.

Mathematical Modeling of the Impact of Actin and Keratin Filaments on Keratinocyte Cell Spreading

Keratin intermediate filaments (IFs) form cross-linked arrays to fulfill their structural support function in epithelial cells and tissues subjected to external stress. How the cross-linking of keratin IFs impacts the morphology and differentiation of keratinocytes in the epidermis and related surface epithelia remains an open question. Experimental measurements have established that keratinocyte spreading area is inversely correlated to the extent of keratin IF bundling in two-dimensional culture. In an effort to quantitatively explain this relationship, we developed a mathematical model in which isotropic cell spreading is considered as a first approximation. Relevant physical properties such as actin protrusion, adhesion events, and the corresponding response of lamellum formation at the cell periphery are included in this model. Through optimization with experimental data that relate time-dependent changes in keratinocyte surface area during spreading, our simulation results confirm the notion that the organization and mechanical properties of cross-linked keratin filaments affect cell spreading; in addition, our results provide details of the kinetics of this effect. These in silico findings provide further support for the notion that differentiation-related changes in the density and intracellular organization of keratin IFs affect tissue architecture in epidermis and related stratified epithelia.

The E3 Ubiquitin-Ligase HACE1 Catalyzes the Ubiquitylation of Active Rac1

Rac1 small GTPase controls essential aspects of cell biology and is a direct target of numerous bacterial virulence factors. The CNF1 toxin of pathogenic Escherichia coli addresses Rac1 to ubiquitin-proteasome system (UPS). We report the essential role of the tumor suppressor HACE1, a HECT-domain containing E3 ubiquitin-ligase, in the targeting of Rac1 to UPS. HACE1 binds preferentially GTP-bound Rac1 and catalyzes its polyubiquitylation. HACE1 expression increases the ubiquitylation of Rac1, when the GTPase is activated by point mutations or by the GEF-domain of Dbl. RNAi-mediated depletion of HACE1 blocks the ubiquitylation of active Rac1 and increases GTP-bound Rac1 cellular levels. HACE1 antagonizes cell isotropic spreading, a hallmark of Rac1 activation, and is required for endothelial cell monolayer invasion by bacteria. Together, these data establish the role of the HACE1 E3 ubiquitin-ligase in controlling Rac1 ubiquitylation and activity.

Membrane Feedback Controls Signaling Regulation of Cell Spreading

Cell motility and spreading are regulated by signaling from the integrin receptors. Interaction between the integrin receptors and substrates such as fibronectin triggers the activation of downstream signaling pathways, resulting in the activation of actin regulating proteins such as Arp2/3, gelsolin and profilin. We have developed an integrated model of cell spreading regulated by integrin signaling network to understand the role of signaling during fast isotropic spreading, when fibroblasts spread on fibronectin coated slides. The three dimensional stochastic spreading model was developed using differential geometry techniques and is coupled with a deterministic model for integrin signaling regulation. We find that cell spreading is a robust process and depends on signaling only for the initiation of spreading but not for maintaining the spreading dynamics. Our model further predicts that signaling dynamics in the absence of Cdc42 and WASP reduce the spreading rate to a small extent but do not affect the shape evolution of the spreading cell. These predictions were verified by experiments conducted with dominant negative Cdc42 cells and wiskostatin effects on cell spreading. Computational analyses predicted that the spreading shape evolution is controlled by the physical properties of the plasma membrane such as membrane surface load and membrane bending ridigity. Simulations from our model identified that changing these properties affects the spreading dynamics, in particular the shape evolution. In contrast, changing information flow through the cell signaling network has little effect. Overall isotropic fast spreading of fibroblasts on fibronectin-coated surfaces depends strongly on the biophysical properties of the plasma membrane and is robust to changes in the signaling dynamics.

Signaling Network Triggers and Membrane Physical Properties Control the Actin Cytoskeleton-Driven Isotropic Phase of Cell Spreading

Cell spreading is regulated by signaling from the integrin receptors that activate intracellular signaling pathways to control actin filament regulatory proteins. We developed a hybrid model of whole-cell spreading in which we modeled the integrin signaling network as ordinary differential equations in multiple compartments, and cell spreading as a three-dimensional stochastic model. The computed activity of the signaling network, represented as time-dependent activity levels of the actin filament regulatory proteins, is used to drive the filament dynamics. We analyzed the hybrid model to understand the role of signaling during the isotropic phase of fibroblasts spreading on fibronectin-coated surfaces. Simulations showed that the isotropic phase of spreading depends on integrin signaling to initiate spreading but not to maintain the spreading dynamics. Simulations predicted that signal flow in the absence of Cdc42 or WASP would reduce the spreading rate but would not affect the shape evolution of the spreading cell. These predictions were verified experimentally. Computational analyses showed that the rate of spreading and the evolution of cell shape are largely controlled by the membrane surface load and membrane bending rigidity, and changing information flow through the integrin signaling network has little effect. Overall, the plasma membrane acts as a damper such that only ∼5% of the actin dynamics capability is needed for isotropic spreading. Thus, the biophysical properties of the plasma membrane can condense varying levels of signaling network activities into a single cohesive macroscopic cellular behavior.

Interplay of RhoA and Motility in the Programmed Spreading of Daughter Cells Postmitosis

Upon cortical retraction in mitosis, mammalian cells have a dramatically decreased physical association with their environment. Hence, mechanisms that prevent mitotic detachment and ensure appropriate positioning of the resulting daughter cells are critical for effective tissue morphogenesis and repair, and are the subject of this study. We find that, unlike low-motility cells, highly motile cells spread isotropically upon division and do not typically reoccupy their mother-cell footprint, and often even disseminate their mitotic cells. To elucidate these different motility-based phenotypes, we investigated their partial recapitulation and rescue using defined molecular perturbations. We show that activated RhoA is localized at the mitotic cell cortex, and Rho-associated kinase inhibition increases the degree of reoccupation of the mother-cell outline in highly motile cells. Conversely, we show that induction of motility in low-motility cells by RasV12 overexpression results in increased isotropic daughter-cell spreading. We thus propose that a balance between cortical retraction forces, which depend in part on RhoA activation, and substrate adhesion forces, which diminish with increasing motility rates, governs the integrity of mitotic actin retraction fibers and influences subsequent daughter-cell spreading. This balance of forces during mitosis has implications for cancer metastasis.

Cell spreading as a hydrodynamic process.

Many cell types have the ability to move themselves by crawling on extra-cellular matrices. Although cell motility is governed by actin and myosin filament assembly, the pattern of the movement follows the physical properties of the network ensemble average. The first step of motility, cell spreading on matrix substrates, involves a transition from round cells in suspension to polarized cells on substrates. Here we show that the spreading dynamics on 2D surfaces can be described as a hydrodynamic process. In particular, we show that the transition from isotropic spreading at early time to anisotropic spreading is reminiscent of the fingering instability observed in many spreading fluids. During cell spreading, the main driving force is the polymerization of actin filaments that push the membrane forward. From the equilibrium between the membrane force and the cytoskeleton, we derive a first order expression of the polymerization stress that reproduces the observed behavior. Our model also allows an interpretation of the effects of pharmacological agents altering the polymerization of actin. In particular we describe the influence of Cytochalasin D on the nucleation of the fingering instability.

Mechanisms Controlling Cell Size and Shape during Isotropic Cell Spreading

Cell motility is important for many developmental and physiological processes. Motility arises from interactions between physical forces at the cell surface membrane and the biochemical reactions that control the actin cytoskeleton. To computationally analyze how these factors interact, we built a three-dimensional stochastic model of the experimentally observed isotropic spreading phase of mammalian fibroblasts. The multiscale model is composed at the microscopic levels of three actin filament remodeling reactions that occur stochastically in space and time, and these reactions are regulated by the membrane forces due to membrane surface resistance (load) and bending energy. The macroscopic output of the model (isotropic spreading of the whole cell) occurs due to the movement of the leading edge, resulting solely from membrane force-constrained biochemical reactions. Numerical simulations indicate that our model qualitatively captures the experimentally observed isotropic cell-spreading behavior. The model predicts that increasing the capping protein concentration will lead to a proportional decrease in the spread radius of the cell. This prediction was experimentally confirmed with the use of Cytochalasin D, which caps growing actin filaments. Similarly, the predicted effect of actin monomer concentration was experimentally verified by using Latrunculin A. Parameter variation analyses indicate that membrane physical forces control cell shape during spreading, whereas the biochemical reactions underlying actin cytoskeleton dynamics control cell size (i.e., the rate of spreading). Thus, during cell spreading, a balance between the biochemical and biophysical properties determines the cell size and shape. These mechanistic insights can provide a format for understanding how force and chemical signals together modulate cellular regulatory networks to control cell motility.

Cell spreading as a hydrodynamic process

Many cell types have the ability to move themselves by crawling on extra-cellular matrices. Although cell motility is governed by actin and myosin filament assembly, the pattern of the movement follows the physical properties of the network ensemble average. The first step of motility, cell spreading on matrix substrates, involves a transition from round cells in suspension to polarized cells on substrates. Here we show that the spreading dynamics on 2D surfaces can be described as a hydrodynamic process. In particular, we show that the transition from isotropic spreading at early time to anisotropic spreading is reminiscent of the fingering instability observed in many spreading fluids. During cell spreading, the main driving force is the polymerization of actin filaments that push the membrane forward. From the equilibrium between the membrane force and the cytoskeleton, we derive a first order expression of the polymerization stress that reproduces the observed behavior. Our model also allows an interpretation of the effects of pharmacological agents altering the polymerization of actin. In particular we describe the influence of Cytochalasin D on the nucleation of the fingering instability.

Modeling the Effects of Nearby Buildings on Inter-Floor Radio-Wave Propagation

Two buildings (A and B) have been modeled and analyzed with a 2D TE <sub xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">z</sub> implementation of the finite-difference time-domain (FDTD) algorithm in order to identify and characterize the mechanisms allowing signals to propagate between floors, specifically reflection and scattering from nearby buildings. Results have been extended to 2.5D by assuming isotropic spreading in the third dimension. In both scenarios considered, reflections from surrounding buildings are found to increase the average received power on adjacent floors-up to 9.7 dB and 32 dB for buildings A and B respectively. Measurements of the impulse response in Building A, made with a sliding correlator channel sounder, show a number of long-delay pulses, which can be attributed to specific reflection paths. Based on these findings, a simple two-component propagation model to predict the sector-average signal strengths is proposed and validated against measurements of the received power. The direct component is modeled as free space with a 22 dB/floor attenuation factor, and the reflected component is modeled as free space with reflection/transmission coefficients of 0.5. The RMS prediction error for this model is 3.2 dB.

Modelling the Effects of Nearby Buildings on Inter-Floor Radio-Wave Propagation

Two buildings (A and B) have been modeled and analyzed with a 2D TE <sub xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">z</sub> implementation of the finite-difference time-domain (FDTD) algorithm in order to identify and characterize the mechanisms allowing signals to propagate between floors, specifically reflection and scattering from nearby buildings. Results have been extended to 2.5D by assuming isotropic spreading in the third dimension. In both scenarios considered, reflections from surrounding buildings are found to increase the average received power on adjacent floors-up to 9.7 dB and 32 dB for buildings A and B respectively. Measurements of the impulse response in Building A, made with a sliding correlator channel sounder, show a number of long-delay pulses, which can be attributed to specific reflection paths. Based on these findings, a simple two-component propagation model to predict the sector-average signal strengths is proposed and validated against measurements of the received power. The direct component is modeled as free space with a 22 dB/floor attenuation factor, and the reflected component is modeled as free space with reflection/transmission coefficients of 0.5. The RMS prediction error for this model is 3.2 dB.