Protrusion Rate Research Articles

Re-track: Software to analyze the retraction and protrusion velocities of neurites, filopodia and other structures

We have developed new software, Re-track, that will quantify the rates of retraction and protrusion of structures emanating from the central core of a cell, such as neurites or filopodia. Re-Track, uses time-lapse images of cells in TIFF format and calculates the velocity of retraction or protrusion of a selected structure. The software uses a flexible moving boundary and has the ability to correct this boundary throughout analysis. Re-Track is fast, platform independent, and user friendly, and it can be used to follow biological events such as changes in neuronal connections, tip-growing cells such as moss, adaptive migration of cells, and similar behavior in non-biological systems.

The impact of sphenoid sinus pneumatization type on the protrusion and dehiscence of the adjacent neurovascular structures: A prospective MDCT imaging study

Sphenoid sinus (SS) pneumatization is greatly variable. Precise determination of the pattern of SS pneumatization and its relationship to the vital neurovascular structures will provide a more comprehensive understanding of the regional SS anatomy to avoid the risk of potentially serious neural and vascular injury. We aimed to study the relationships among SS pneumatization type, extent of pneumatization, and protrusion/dehiscence of adjacent neurovascular structures using the multi-detector computed tomography. One hundred seventy healthy patients were prospectively examined by multi-detector computed tomography of the head. The type of pneumatization of the SS was determined. The protrusion/dehiscence of the internal carotid artery, optic nerve, vidian nerve, and maxillary nerve into the SS cavity was evaluated. The relationships between the SS pneumatization type and the extent of protrusion/dehiscence of the examined variables were statistically analyzed. We had 70 patients with no protrusion/dehiscence, 37 patients with unilateral protrusion/dehiscence, and 63 patients with bilateral protrusion/dehiscence. The presellar type was dominant (61.4%) in the group with no protrusion/dehiscence, the sellar type was dominant (56.8%) in the group with unilateral protrusion/dehiscence, and the postsellar type was dominant (55.6%) in the group with bilateral protrusion/dehiscence. The protrusion rates of the internal carotid artery, optic nerve, vidian nerve, and maxillary nerve in the postsellar type were 50.6%, 52.3%, 56.9%, and 54.1%, respectively (p< 0.001). The prevalence of protrusion/dehiscence of the neurovascular structures is significantly related to the type of SS and increases as the degree of SS pneumatization increases.

Efficient Front-Rear Coupling in Neutrophil Chemotaxis by Dynamic Myosin II Localization

Efficient chemotaxis requires rapid coordination between different parts of the cell in response to changing directional cues. Here, we investigate the mechanism of front-rear coordination in chemotactic neutrophils. We find that changes in the protrusion rate at the cell front are instantaneously coupled to changes in retraction at the cell rear, while myosin II accumulation at the rear exhibits a reproducible 9-15-s lag. In turning cells, myosin II exhibits dynamic side-to-side relocalization at the cell rear in response to turning of the leading edge and facilitates efficient turning by rapidly re-orienting the rear. These manifestations of front-rear coupling can be explained by a simple quantitative model incorporating reversible actin-myosin interactions with a rearward-flowing actin network. Finally, the system can be tuned by the degree of myosin regulatory light chain (MRLC) phosphorylation, which appears to be set in an optimal range to balance persistence of movement and turning ability.

Influence of cross-linking and retrograde flow on formation and dynamics of lamellipodium.

The forces that arise from the actin cortex play a crucial role in determining the membrane deformation. These include protrusive forces due to actin polymerization, pulling forces due to transient attachment of actin filaments to the membrane, retrograde flow powered by contraction of actomyosin network, and adhesion to the extracellular matrix. Here we present a theoretical model for membrane deformation resulting from the feedback between the membrane shape and the forces acting on the membrane. We model the membrane as a series of beads connected by springs and determine the final steady-state shape of the membrane arising from the interplay between pushing/pulling forces of the actin network and the resisting membrane tension. We specifically investigate the effect of the gel dynamics on the spatio-temporal deformation of the membrane until a stable lamellipodium is formed. We show that the retrograde flow and the cross-linking velocity play an essential role in the final elongation of the membrane. Interestingly, in the simulations where motor-induced contractility is switched off, reduced retrograde flow results in an increase in the rate and amplitude of membrane protrusion. These simulations are consistent with experimental observations that report an enhancement in protrusion efficiency as myosin II molecular motors are inhibited.

Effects of Opsonin Density on Phagocytic Behavior of Human Neutrophils

Although many components of the molecular machinery driving phagocytosis in immune cells have been identified, a detailed understanding of the underlying biophysical mechanisms remains elusive. Here, we investigate phagocytic protrusion dynamics in human neutrophils by monitoring mechanical processes and signaling events during phagocytosis of targets coated with varying densities of rabbit immunoglobulin G (IgG). We examine effects of the opsonin density in two complementary experiments: phagocytosis of IgG-coated polystyrene microspheres, and frustrated phagocytic cell spreading on IgG-coated glass coverslips. We facilitate interactions between neutrophils and microspheres using two opposing glass micropipettes to hold and manipulate the cell and target particle; this also allows us to quantify cell morphology and approximate cortical tension during phagocytosis. To analyze frustrated phagocytosis, we image the area of contact between spreading cells and the coverslip with high resolution using reflection interference contrast microscopy (RICM). The frustrated phagocytosis assay revealed that the overall speed of contact-area growth is largely independent of IgG density. However, cells are more likely to spread and reach a larger maximum contact area on higher densities of IgG. Monitoring also the cytosolic calcium concentration, we found that the timing of calcium surges during frustrated phagocytosis depends on IgG density – at lower densities, the cells reach larger contact areas before exhibiting global calcium bursts. Overall, these findings indicate that the speed of cell spreading is determined by the rate of actin protrusion rather than the density of adhesive contacts, while the number of ligands available for binding to Fcγ-receptors impacts both calcium signaling dynamics and the likelihood and extent of cell spreading. This suggests that initial actin polymerization is largely independent of calcium signaling, while later stages of phagocytosis mechanics are regulated by elevated global calcium levels.

NADPH-oxidase-derived ROS alterscell migration by modulating adhesions dynamics.

Cell migration requires the coordinated activation of structural and signalling molecules, such as the RhoGTPase Rac1. It is known that the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex assembly, which generates reactive oxygen species (ROS) at the cell membrane, also relies on Rac1 activation, indicating a possible effect of ROS during cell migration. In this study, we evaluated the effect of NADPH-oxidase-derived ROS on the migration process. Using time-lapse videos of CHO.K1 cells plated on fibronectin (2μg/ml) or collagen (5μg/cm2 ), we observed that depletion of ROS by N-acetyl-cysteine (NAC, 10mM), an unspecific antioxidant, or diphenyliodonium (DPI, 10μM), a NADPH-oxidase inhibitor, induced a ∼50% decrease in migration speed and severely impacted migration directionality. Then, we analysed the effects of NADPH oxidase on three migratory events: protrusion rate, adhesion process and signalling pathways related to cell migration. DPI induced an increase of ∼3 protrusion/cell, which were 2× faster but had a ∼50% retraction when compared with control. By pull-down assay, we observed no changes on Rac1 activation, indicating that ROS-mediated effects were related to downstream molecules, such as adhesion-related molecules. A reduction of the adhesion marker FAK-Y397 levels in cells treated with NAC and DPI was observed. In order to analyse adhesion dynamics, CHO.K1 cells transfected with paxillin-GFP analysed with total internal reflectance fluorescence (TIRF) indicated that DPI (5μM) induced larger adhesions when compared with control. These results indicate that the local generation of NADPH-oxidase-derived ROS can modulate cell migration due to changes on adhesion dynamics and signalling. This study highlights the physiological requirement of ROS for cell migration and the potential use of these molecules as targets to modulate the cell migration process at different diseases.

Interfacial competitive adsorption of different amphipathicity emulsifiers and milk protein affect fat crystallization, physical properties, and morphology of frozen aerated emulsion

This study investigated interfacial competitive adsorption of different emulsifiers, including lipophilic [sucrose stearate 370 (S370); mono- and di-acylglycerols (MDG)] and hydrophilic [sucrose stearate 1670 (S1670) and Tween 20 (TW20)] emulsifiers with milk proteins and its influence on fat coalescence behavior in frozen aerated emulsions (10% w/w fat). Lipophilic emulsifiers accelerated fat crystallization, promoting interface heterogeneous nucleation and partial coalescence of fat droplets. In contrast, hydrophilic emulsifiers favored large fat crystal formation with a reduced rate and capacity of fat crystal protrusion. The combination of surfactants (0.5% w/w) with milk proteins (5% w/w) produced concerted effects on the stability of emulsions. The lipophilic surfactant-protein mixture also enabled strong aeration activity (overrun) and a short melting starting time. Moreover, a remarkable array of bee wax-like network was observed in such lipophilic emulsifier-stabilized frozen whipped emulsions with a similar effect seen between S370 and MDG. However, for whipped emulsions produced with hydrophilic emulsifiers, especially with TW20, fewer and irregularly-shaped air bubbles were notable, contrary to the round, slightly arching shape with glossy surface in frozen whipped emulsions with lipophilic emulsifiers. Overall, lipophilic emulsifiers were superior to hydrophilic emulsifiers in producing the well-textured frozen aerated emulsions.

On the relation between filament density, force generation, and protrusion rate in mesenchymal cell motility.

Lamellipodia are flat membrane protrusions formed during mesenchymal motion. Polymerization at the leading edge assembles the actin filament network and generates protrusion force. How this force is supported by the network and how the assembly rate is shared between protrusion and network retrograde flow determines the protrusion rate. We use mathematical modeling to understand experiments changing the F-actin density in lamellipodia of B16-F1 melanoma cells by modulation of Arp2/3 complex activity or knockout of the formins FMNL2 and FMNL3. Cells respond to a reduction of density with a decrease of protrusion velocity, an increase in the ratio of force to filament number, but constant network assembly rate. The relation between protrusion force and tension gradient in the F-actin network and the density dependency of friction, elasticity, and viscosity of the network explain the experimental observations. The formins act as filament nucleators and elongators with differential rates. Modulation of their activity suggests an effect on network assembly rate. Contrary to these expectations, the effect of changes in elongator composition is much weaker than the consequences of the density change. We conclude that the force acting on the leading edge membrane is the force required to drive F-actin network retrograde flow.

Crawling migration under chemical signalling: A stochastic model

Cell migration is an essential and complex mechanism involved in many physiological processes such as tissue formation or tumor invasion. External and internal cues are known to guide cell motion by regulating cytoskeleton dynamics. Here, we approach cell‐crawling migration under a large‐scale assumption so that it reduces to a particle in motion. However, we describe the cell displacement as the result of its internal activity: the cell velocity is a function of the (discrete) protrusive forces exerted by filopodia on the substrate. Cell polarization ability is modeled in the feedback that the cell motion exerts on the protrusion rates: faster cells form preferentially protrusions in the direction of motion. In addition, we incorporate a gradient in attractive chemical signal, which may vary in time. We perform numerical simulations on this model, showing that the balance between cells self‐polarized internal machinery and signal sensing leads to nontrivial behaviors. Finally, by using the mathematical framework of structured population processes previously developed to study population dynamics, we study rigorously the mathematical model, and we derive some of its fundamental properties.

Logistic regression analysis on the outcome predictive factors of ruptured lumbar disc herniation

To investigate the outcome predictive factors of ruptured lumbar disc herniation after conservative treatment. From June 2009 to June 2016, 147 patients with ruptured lumbar intervertebral disc herniation were treated with conservative treatment in the orthopedics department of Suzhou Traditional Chinese Medicine Hospital for clinical efficacy and MRI follow-up. Multivariate Logistic regression analysis(Stepwise regression method)was used to analyze the relationship between the 11 categorical variables and absorptivity of protrusions: sex(X1), age(X2), course of disease(X3) , the rate of protrusion(X4), the Komori type(X5), the MSU type(X6), the Iwabuchi type(X7), the Pfirrmann grade(X8), the Modic change on adjacent vertebrae(X9), spinal canal morphology(X10), the Schizas types of cauda equina sedimentation sign(X11). A total of 64 cases of prominent reabsorption among all cases followed-up (absorption rate>=30%), accounting for 43.5%. The reabsorption of protrusions is more likely to occur in patients with a duration of less than 1 year(P=0.006), MSU type 3 (P=0.001), Iwabuchi type 1 or 5 (P=0.000), the Schizas type of cauda equina sedimentation sign A or B(P=0.004). Regression equation Y=-10.363+1.916X3+1.446X4-1.445X5+2.070X6+4.679X7+1.125X9+1.023X10+2.223X11. Such factors as age, gender, Pfirrmann classification and spinal canal morphology had no significant effect on reabsorption of protrusions. Ruptured lumbar disc herniation can be reabsorbed after nonoperative treatment. And the reabsorption of protrusions is more likely to occur in patients with a duration of less than 1 year, MSU type 3, Iwabuchi type 1 or 5, the Schizas type of cauda equina sedimentation sign A or B, which can be used as the key reference factors for predicting the outcome of the projections.

Matrix elasticity regulates mesenchymal stem cell chemotaxis.

Efficient homing of human mesenchymal stem cells (hMSCs) is likely to be dictated by a combination of physical and chemical factors present in the microenvironment. However, crosstalk between the physical and chemical cues remains incompletely understood. Here, we address this question by probing the efficiency of epidermal growth factor (EGF)-induced hMSC chemotaxis on substrates of varying stiffness (3, 30 and 600 kPa) inside a polydimethylsiloxane(PDMS) microfluidic device. Chemotactic speed was found to be the sum of a stiffness-dependent component and a chemokine concentration-dependent component. While the stiffness-dependent component scaled inversely with stiffness, the chemotactic component was independent of stiffness. Faster chemotaxis on the softest 3 kPa substrates is attributed to a combination of weaker adhesions and higher protrusion rate. While chemotaxis was mildly sensitive to contractility inhibitors, suppression of chemotaxis upon actin depolymerization demonstrates the role of actin-mediated protrusions in driving chemotaxis. In addition to highlighting the collective influence of physical and chemical cues in chemotactic migration, our results suggest that hMSC homing is more efficient on softer substrates.

A Rac1-FMNL2 signaling module affects cell-cell contact formation independent of Cdc42 and membrane protrusions.

De novo formation of epithelial cell-cell contacts relies on actin-based protrusions as well as tightly controlled turnover of junctional actin once cells encounter each other and adhesion complexes assemble. The specific contributions of individual actin regulators on either protrusion formation or junctional actin turnover remain largely unexplored. Based on our previous findings of Formin-like 2 (FMNL2)-mediated control of junctional actin dynamics, we investigated its potential role in membrane protrusions and impact on newly forming epithelial contacts. CRISPR/Cas9-mediated loss of FMNL2 in human MCF10A cells combined with optogenetic control of Rac1 activity confirmed its critical function in the establishment of intercellular contacts. While lamellipodial protrusion rates remained unaffected, FMNL2 knockout cells were characterized by impaired filopodia formation similar to depletion of the Rho GTPase Cdc42. Silencing of Cdc42, however, failed to affect FMNL2-mediated contact formation. Hence, we propose a cell-cell contact-specific and Rac1-mediated function of FMNL2 entirely independent of Cdc42. Consistent with this, direct visualizations of native epithelial junction formation revealed a striking and specifically Rac1- and not Cdc42-dependent recruitment of FMNL2 to newly forming junctions as well as established cell-cell contacts within epithelial sheets.

How Filaments Density Impacts Force Generation and Protrusion Rate of Lamellipodium in Motile Cells

How Filaments Density Impacts Force Generation and Protrusion Rate of Lamellipodium in Motile Cells

A Stochastic Model For Protrusion Activity

In this work we approach cell migration under a large-scale assumption, so that the system reduces to a particle in motion. Unlike classical particle models, the cell displacement results from its internal activity: the cell velocity is a function of the (discrete) protrusive forces exerted by filopodia on the substrate. Cell polarisation ability is modeled in the feedback that the cell motion exerts on the protrusion rates: faster cells form preferentially protrusions in the direction of motion. By using the mathematical framework of structured population processes previously developed to study population dynamics [4], we introduce rigorously the mathematical model and we derive some of its fundamental properties. We perform numerical simulations on this model showing that different types of trajectories may be obtained: Brownian-like, persistent, or intermittent when the cell switches between both previous regimes. We find back the trajectories usually described in the literature for cell migration.

A free-boundary model of a motile cell explains turning behavior.

To understand shapes and movements of cells undergoing lamellipodial motility, we systematically explore minimal free-boundary models of actin-myosin contractility consisting of the force-balance and myosin transport equations. The models account for isotropic contraction proportional to myosin density, viscous stresses in the actin network, and constant-strength viscous-like adhesion. The contraction generates a spatially graded centripetal actin flow, which in turn reinforces the contraction via myosin redistribution and causes retraction of the lamellipodial boundary. Actin protrusion at the boundary counters the retraction, and the balance of the protrusion and retraction shapes the lamellipodium. The model analysis shows that initiation of motility critically depends on three dimensionless parameter combinations, which represent myosin-dependent contractility, a characteristic viscosity-adhesion length, and a rate of actin protrusion. When the contractility is sufficiently strong, cells break symmetry and move steadily along either straight or circular trajectories, and the motile behavior is sensitive to conditions at the cell boundary. Scanning of a model parameter space shows that the contractile mechanism of motility supports robust cell turning in conditions where short viscosity-adhesion lengths and fast protrusion cause an accumulation of myosin in a small region at the cell rear, destabilizing the axial symmetry of a moving cell.

Microstructure Evolution and Protrusion of Electroplated Cu-Filled Through-Silicon Vias Subjected to Thermal Cyclic Loading

Through-silicon vias (TSVs) have become an important technology for three-dimensional integrated circuit (3D IC) packaging. Protrusion of electroplated Cu-filled vias is a critical reliability issue for TSV technology. In this work, thermal cycling tests were carried out to identify how the microstructure affects protrusion during thermal cycling. Cu protrusion occurs when the loading temperature is higher than 149°C. During the first five thermal cycles, the grain size of Cu plays a dominant role in the protrusion behavior. Larger Cu grain size before thermal cycling results in greater Cu protrusion. With increasing thermal cycle number, the effect of the Cu grain size reduces and the microstrain begins to dominate the Cu protrusion behavior. Higher magnitude of microstrain within Cu results in greater protrusion increment during subsequent thermal cycles. When the thermal cycle number reaches 25, the protrusion rate of Cu slows down due to strain hardening. After 30 thermal cycles, the Cu protrusion stabilizes within the range of 1.92 μm to 2.09 μm.

Facial soft tissue changes during the pre-pubertal and pubertal growth phase: a mixed longitudinal laser-scanning study.

Facial soft tissues changes during growth roughly tend to mimic the underlying hard tissues, but not completely. The aim of this mixed longitudinal study was to assess facial growth among pre-pubertal and pubertal subjects without malocclusion using a non-invasive three-dimensional laser scanning system. Fifty-nine subjects (30 females and 29 males) aged at baseline 5.4-8.9 years with normal occlusion were clustered into the younger, older pre-pubertal, and pubertal groups according to age and the absence/presence of a standing height growth spurt. Three-dimensional facial images were obtained using laser scanners for five consecutive years. Several transversal, sagittal, and vertical parameters were assessed for between and within group comparisons. Significant overall changes of almost all parameters were seen within each group (P < 0.05) without any group differences (P > 0.05). The younger pre-pubertal group showed greater annual growth rates of lip prominence; both pre-pubertal groups showed greater rates in facial middle third height. The pubertal group showed greater annual rates in facial profile angle changes during the growth peak. A high standing height increment (7cm) was used as the threshold for subject allocation in the pubertal group. Soft tissue facial growth has generally similar amounts and rates irrespective of the pubertal growth spurt. Pre-pubertal subjects show greater annual rates of facial middle third height changes while pubertal subjects show greater annual rates of chin protrusion.

Medial Protrusio Technique in Cementless Total Hip Arthroplasty for Developmental Dysplasia of the Hip: A Prospective 6- to 9-Year Follow-Up of 43 Consecutive Patients

BackgroundThe medial protrusio technique may be used during total hip arthroplasty (THA) on patients with developmental dysplasia. However, studies have yet to determine whether a cementless cup can be sufficiently stable to withstand loading forces. This study aimed to assess the clinical and radiographic outcomes of this technique. Furthermore, we sought to determine the relationship between the rate of medial protrusion and the incidence of cup loosening. MethodsThirty-nine patients (43 hips) underwent cementless THA between April 2006 and March 2009 by using the medial protrusio technique. These patients participated in a 6- to 9-year follow-up. Their clinical and radiographic data were gathered prospectively. ResultsThe average Harris Hip Score improved from 43.1 ± 15.4 points preoperatively to 91.9 ± 12.8 points at the final follow-up (P < .001). The mean height of hip center and the distance of hip center medialization were 2.4 ± 0.6 and 2.5 ± 0.9 cm, respectively. The rate of medial protrusion and the rate of cup coverage were 42.1 ± 12.4% and 96.8 ± 5.1%, respectively. The rate of medial protrusion ranged from 18.3% to 58.3% in 38 hips (group A) and from 61.3% to 68.9% in 5 hips (group B). None of the cups in group A loosened or failed, 2 failures occurred in group B (0% vs 40.0%; P = .011). ConclusionsDevelopmental dysplasia was treated through THA using the medial protrusio technique, which easily achieves a sufficient superolateral host bony coverage of the cup and promotes socket reconstruction at the true acetabulum. The rate of medial protrusion of <60% may be necessary to obtain excellent clinical and radiographic midterm results.

Three-dimensional matrix fiber alignment modulates cell migration and MT1-MMP utility by spatially and temporally directing protrusions.

Multiple attributes of the three-dimensional (3D) extracellular matrix (ECM) have been independently implicated as regulators of cell motility, including pore size, crosslink density, structural organization, and stiffness. However, these parameters cannot be independently varied within a complex 3D ECM protein network. We present an integrated, quantitative study of these parameters across a broad range of complex matrix configurations using self-assembling 3D collagen and show how each parameter relates to the others and to cell motility. Increasing collagen density resulted in a decrease and then an increase in both pore size and fiber alignment, which both correlated significantly with cell motility but not bulk matrix stiffness within the range tested. However, using the crosslinking enzyme Transglutaminase II to alter microstructure independently of density revealed that motility is most significantly predicted by fiber alignment. Cellular protrusion rate, protrusion orientation, speed of migration, and invasion distance showed coupled biphasic responses to increasing collagen density not predicted by 2D models or by stiffness, but instead by fiber alignment. The requirement of matrix metalloproteinase (MMP) activity was also observed to depend on microstructure, and a threshold of MMP utility was identified. Our results suggest that fiber topography guides protrusions and thereby MMP activity and motility.

Anatomical variants of surgically important landmarks in the sphenoid sinus: a radiologic study in Southeast Asian patients.

Failure of a surgeon to understand the local variations of the anatomical landmarks of the sphenoid sinus is a potential risk factor to cause damage to the optic nerve (ON) or internal carotid artery (ICA) that lies on the walls of the sphenoid sinus. The aim of this study was to identify the anatomical variants of the sphenoid sinus and its related surrounding structures among the Southeast Asian (SEA) population, based on computed tomography (CT) scans. This cross-sectional study analyzed 300 CT scans of the brain, paranasal sinuses (PNS), and head and neck (H&N) at a tertiary referral centre in Malaysia utilizing the Osirix software. The images were reconstructed into 1 mm cuts on bone window. Demographic details and scan findings were documented in a standardized data collection sheet. The rates of ON dehiscence, ICA dehiscence and ICA protrusion in the SEA population were 7.0, 3.0 and 10.0 %, respectively. The rate of ON protrusion was 2.3 %. There was no statistically significant relationship (p > 0.05) noted on Chi-square test, between anterior clinoid process (ACP) pneumatization and ON protrusion. The rate of Onodi cells in our population was 14.3 %. The average vertical distance of the ostia from the roof of the posterior choanae was 1.42 cm (±0.32). The horizontal distance of the ostia from the anterior end of the superior turbinate was 1.58 cm (±0.41) and the oblique distance of the ostia from the anterior nasal spine was 5.35 cm (±0.48). Independent t tests showed that there is a statistically significant difference between the means of each of these parameters (p < 0.001) and their international averages. The rate of ON protrusion is lower in the SEA population, whereas the rates of ON dehiscence, ICA dehiscence and ICA protrusion fall within the range of international averages. In our population, ACP pneumatization is not related to ON protrusion. The distance of the ostia from given landmarks was significantly shorter than in other studies.