RESUMEN
The physical properties of lipid bilayers comprising the cell membrane occupy the current spotlight of membrane biology. Their traditional representation as a passive 2D fluid has gradually been abandoned in favor of a more complex picture: an anisotropic time-dependent viscoelastic biphasic material, capable of transmitting or attenuating mechanical forces that regulate biological processes. In establishing new models, quantitative experiments are necessary when attempting to develop suitable techniques for dynamic measurements. Here, we map both the elastic and viscous properties of the model system 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) lipid bilayers using multifrequency atomic force microscopy (AFM), namely amplitude modulation-frequency modulation (AM-FM) AFM imaging in an aqueous environment. Furthermore, we investigate the effect of cholesterol (Chol) on the DPPC bilayer in concentrations from 0 to 60%. The AM-AFM quantitative maps demonstrate that at low Chol concentrations, the lipid bilayer displays a distinct phase separation and is elastic, whereas at higher Chol concentration, the bilayer appears homogenous and exhibits both elastic and viscous properties. At low-Chol contents, the Estorage modulus (elastic) dominates. As the Chol insertions increases, higher energy is dissipated; and although the bilayer stiffens (increase in Estorage), the viscous component dominates (Eloss). Our results provide evidence that the lipid bilayer exhibits both elastic and viscous properties that are modulated by the presence of Chol, which may affect the propagation (elastic) or attenuation (viscous) of mechanical signals across the cell membrane.
Asunto(s)
Membrana Celular/química , Colesterol/metabolismo , Membrana Dobles de Lípidos/química , Membrana Celular/metabolismo , Elasticidad , Membrana Dobles de Lípidos/metabolismo , Microscopía de Fuerza Atómica , Modelos Biológicos , ViscosidadRESUMEN
OBJECTIVE: The treatment of acne presents a major clinical and dermatological challenge. Investigating the nanomechanical properties of the microcomedone precursor lesions using atomic force microscopy (AFM) may prove beneficial in understanding their softening, dissolution and prevention. Although the exact biochemical mechanism of NaSal on microcomedones is not fully understood at present, it appears to exhibit a significant exfoliation effect on the skin via corneodesmosome dissolution. METHODS: Therefore, to support this exploration, sodium salicylate (NaSal), a common ingredient employed in skin care products, is applied ex vivo to microcomedones,collected by nose strip adhesive tape, and their nanomechanical properties are assessed using AFM. Although the exact biochemical mechanism of NaSal on microcomedones is not fully understood at present, it appears to exhibit a significant exfoliation effect on the skin via corneodesmosome dissolution. RESULTS: Herein, our findings demonstrate that when microcomedones are treated with 2% NaSal, samples appeared significantly more compliant ('softer') ((1.3 ± 0.62) MPa) when compared to their pre-treated measurements ((7.2 ± 3.6) MPa; p = 0.038). Furthermore, elastic modulus maps showed that after 2% NaSal treatment, areas in the microcomedone appeared softer and swollen in some, but not in all areas, further proving the valuable impact of 2% NaSal solution in altering the biomechanical properties and morphologies in microcomedones. CONCLUSION: Our results are the first of their kind to provide qualitative and quantitative mechanobiological evidence that 2% NaSal decreases the elastic modulus of microcomedones. Therefore, this study provides evidence that NaSal can be beneficial as an active ingredient in topical treatments aimed at targeting microcomedones.
OBJECTIF: Le traitement de l'acné présente un défi clinique et dermatologique majeur. L'étude des propriétés nanomécaniques des lésions précurseurs en tant que microcomédons à l'aide de la microscopie à force atomique (AFM) peut s'avérer bénéfique pour comprendre leur ramollissement, leur dissolution et leur prévention. MÉTHODES: Par conséquent, pour soutenir cette exploration, le salicylate de sodium (NaSal), un ingrédient couramment utilisé dans les produits de soins de la peau, est appliqué ex vivo aux microcomédons et leurs propriétés nanomécaniques sont évaluées à l'aide de l'AFM. Bien que le mécanisme biochimique exact du NaSal sur les microcomédons ne soit pas entièrement compris à l'heure actuelle, il semble présenter un effet exfoliant significatif sur la peau via la dissolution des cornéodesmosomes. RÉSULTATS: Ici, nos résultats démontrent que lorsque les microcomédons sont traités avec 2% de NaSal, les échantillons semblaient significativement plus conformes ("plus doux") ((1.3 ± 0.62) MPa) par rapport à leurs mesures pré-traitées ((7.2 ± 3.6) MPa ; P = 0,03826). De plus, les cartes du module d'élasticité ont montré qu'après un traitement à 2 % de NaSal, les zones du microcomédon semblaient plus molles et gonflées dans certaines zones, mais pas dans toutes, prouvant ainsi l'impact précieux d'une solution de NaSal à 2 % dans la modification des propriétés biomécaniques et de la morphologie des microcomédons. CONCLUSION: Nos résultats sont les premiers du genre à fournir des preuves mécanobiologiques qualitatives et quantitatives que 2% de NaSal diminue le module d'élasticité des microcomédons. Par conséquent, cette étude fournit des preuves que NaSal peut être bénéfique en tant qu'ingrédient actif dans les traitements topiques visant à cibler les microcomédons.
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Acné Vulgar/tratamiento farmacológico , Fármacos Dermatológicos/química , Salicilato de Sodio/química , Administración Tópica , Módulo de Elasticidad , Voluntarios Sanos , Humanos , Microscopía de Fuerza Atómica , Piel/efectos de los fármacosRESUMEN
Insulin growth factor 1 (IGF1) is a major anabolic signal that is essential during skeletal development, cellular adhesion and migration. Recent transcriptomic studies have shown that there is an upregulation in IGF1 expression in calvarial osteoblasts derived from patients with single-suture craniosynostosis (SSC). Upregulation of the IGF1 signaling pathway is known to induce increased expression of a set of osteogenic markers that previously have been shown to be correlated with contractility and migration. Although the IGF1 signaling pathway has been implicated in SSC, a correlation between IGF1, contractility and migration has not yet been investigated. Here, we examined the effect of IGF1 activation in inducing cellular contractility and migration in SSC osteoblasts using micropost arrays and time-lapse microscopy. We observed that the contractile forces and migration speeds of SSC osteoblasts correlated with IGF1 expression. Moreover, both contractility and migration of SSC osteoblasts were directly affected by the interaction of IGF1 with IGF1 receptor (IGF1R). Our results suggest that IGF1 activity can provide valuable insight for phenotype-genotype correlation in SSC osteoblasts and might provide a target for therapeutic intervention.
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Factor I del Crecimiento Similar a la Insulina/metabolismo , Transducción de Señal/fisiología , Adhesión Celular/fisiología , Movimiento Celular/fisiología , Células Cultivadas , Humanos , Osteoblastos/metabolismo , Osteoblastos/fisiología , Receptor IGF Tipo 1/metabolismoRESUMEN
The mechanical properties of living cells are highly regulated by remodeling dynamics of the cytoarchitecture, and are linked to a wide variety of physiological and pathological processes. Microtubules (MT) and actomyosin contractility are both involved in regulating focal adhesion (FA) size and cortical elasticity in living cells. Although several studies have examined the effects of MT depolymerization or actomyosin activation on biological processes, very few have investigated the influence of both on the mechanical properties, FA assembly, and spreading of fibroblast cells. Here, we examine how activation of both processes modulates cortical elasticity as a function of time. Enhancement of contractility (calyculin A treatment) or the depolymerization of MTs (nocodazole treatment) individually caused a time-dependent increase in FA size, decrease in cell height and an increase in cortical elasticity. Surprisingly, sequentially stimulating both processes led to a decrease in cortical elasticity, loss of intact FAs and a concomitant increase in cell height. Our results demonstrate that loss of MTs disables the ability of fibroblast cells to maintain increased contractility and cortical elasticity upon activation of myosin-II. We speculate that in the absence of an intact MT network, a large amount of contractile tension is transmitted directly to FA sites resulting in their disassembly. This implies that tension-mediated FA growth may have an upper bound, beyond which disassembly takes place. The interplay between cytoskeletal remodeling and actomyosin contractility modulates FA size and cell height, leading to dynamic time-dependent changes in the cortical elasticity of fibroblast cells.
Asunto(s)
Membrana Celular/fisiología , Proteínas Contráctiles/fisiología , Elasticidad/fisiología , Microtúbulos/fisiología , Animales , Adhesión Celular/efectos de los fármacos , Adhesión Celular/fisiología , Membrana Celular/ultraestructura , Proliferación Celular/efectos de los fármacos , Citoesqueleto/efectos de los fármacos , Citoesqueleto/fisiología , Citoesqueleto/ultraestructura , Elasticidad/efectos de los fármacos , Adhesiones Focales/efectos de los fármacos , Adhesiones Focales/fisiología , Adhesiones Focales/ultraestructura , Ratones , Microtúbulos/ultraestructura , Contracción Muscular/fisiología , Células 3T3 NIH , Nocodazol/farmacología , Moduladores de Tubulina/farmacologíaRESUMEN
Growing evidence suggests that critical cellular processes are profoundly influenced by the cross talk between extracellular nanomechanical forces and the material properties of the cellular microenvironment. Although many studies have examined either the effect of nanomechanical forces or the material properties of the microenvironment on biological processes, few have investigated the influence of both. Here, we performed simultaneous atomic force microscopy and traction force microscopy to demonstrate that muscle precursor cells (myoblasts) rapidly generate a significant increase in traction when stimulated with a local 10 nN force. Cells were cultured and nanomechanically stimulated on hydrogel substrates with controllable local elastic moduli varying from ~16-89 kPa, as confirmed with atomic force microscopy. Importantly, cellular traction dynamics in response to nanomechanical stimulation only occurred on substrates that were similar to the elasticity of working muscle tissue (~64-89 kPa) as opposed to substrates mimicking resting tissue (~16-51 kPa). The traction response was also transient, occurring within 30 s, and dissipating by 60 s, during constant nanomechanical stimulation. The observed biophysical dynamics are very much dependent on rho-kinase and myosin-II activity and likely contribute to the physiology of these cells. Our results demonstrate the fundamental ability of cells to integrate nanoscale information in the cellular microenvironment, such as nanomechanical forces and substrate mechanics, during the process of mechanotransduction.
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Materiales Biocompatibles/química , Elasticidad , Mecanotransducción Celular , Mioblastos/citología , Animales , Línea Celular , Módulo de Elasticidad , Hidrogeles/química , Ratones , Microscopía de Fuerza Atómica/métodos , Mioblastos/metabolismo , Miosinas/antagonistas & inhibidores , Miosinas/metabolismo , Estrés Mecánico , Quinasas Asociadas a rho/antagonistas & inhibidores , Quinasas Asociadas a rho/metabolismoRESUMEN
We show that surface swelling is the first step in the interaction of a single femtosecond laser pulse with PMMA. This is followed by perforation of the swollen structure and material ejection. The size of the swelling and the perforated hole increases with pulse energy. After certain energy the swelling disappears and the interaction is dominated by the ablated hole. This behaviour is independent of laser polarization. The threshold energy at which the hole size coincides with size of swelling is 1.5 times that of the threshold for surface swelling. 2D molecular dynamics simulations show surface swelling at low pulse energies along with void formation below the surface within the interaction region. Simulations show that at higher energies, the voids coalesce and grow, and the interaction is dominated by material ejection.
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Rayos Láser , Modelos Químicos , Polimetil Metacrilato/química , Polimetil Metacrilato/efectos de la radiación , Simulación por Computador , Ensayo de Materiales , Dosis de Radiación , Propiedades de Superficie/efectos de la radiaciónRESUMEN
Annually, an estimated seven million deaths are linked to exposure to airborne pollutants. Despite extensive epidemiological evidence supporting clear associations between poor air quality and a range of short- and long-term health effects, there are considerable gaps in our understanding of the specific mechanisms by which pollutant exposure induces adverse biological responses at the cellular and tissue levels. The development of more complex, predictive, in vitro respiratory models, including two- and three-dimensional cell cultures, spheroids, organoids and tissue cultures, along with more realistic aerosol exposure systems, offers new opportunities to investigate the cytotoxic effects of airborne particulates under controlled laboratory conditions. Parallel advances in high-resolution microscopy have resulted in a range of in vitro imaging tools capable of visualizing and analysing biological systems across unprecedented scales of length, time and complexity. This article considers state-of-the-art in vitro respiratory models and aerosol exposure systems and how they can be interrogated using high-resolution microscopy techniques to investigate cell-pollutant interactions, from the uptake and trafficking of particles to structural and functional modification of subcellular organelles and cells. These data can provide a mechanistic basis from which to advance our understanding of the health effects of airborne particulate pollution and develop improved mitigation measures.
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This study compared the surface properties and rising velocities of pristine and weathered plastic production pellets, to evaluate impacts of environmental conditions. Rising velocities were measured for 140 weathered pellets collected from a Spanish beach and compared with pristine low-density polyethylene, high-density polyethylene and polypropylene pellets. A subset of 49 weathered pellets were analysed by Fourier-transform infrared spectroscopy (FTIR), with all found to be polyethylene. Experimental rising velocities for the weathered pellets varied widely, from (2.36 ± 0.01) cm s-1 to (10.56 ± 0.26) cm s-1, with a mean value of (5.79 ± 0.06) cm s-1. Theoretical rising velocities were consistently higher than experimental velocities for all pellet types: on average 136% of experimental values for weathered pellets. This discrepancy was more distinct for less spherical pellets, which were often more weathered. Flatter pellets often oscillated as they rose, which explains at least some of this finding. Atomic force microscopy (AFM) analysis revealed that the roughness of the pristine and weathered pellets was (59 ± 11) nm, and (74 ± 26) nm respectively. X-ray photoelectron spectroscopy (XPS) analysis showed that the proportion of surface oxidised carbon species were 2.3% and 4.0% of the total carbon signal for a pristine and a weathered pellet, respectively; consistent with photochemical reactions changing the surface chemistry of weathered pellets. As determined by density column, weathered pellets had slightly lower experimental densities than pristine pellets. Overall, this study illustrates why it is important that modelling studies on the environmental fate and/or movements of microplastics validate or correct predictions using experimental data.
RESUMEN
The role of hyaluronan (HA) in periodontal healing has been speculated via its interaction with the CD44 receptor. While HA-CD44 interactions have previously been implicated in numerous cell types; effect and mechanism of exogenous HA on periodontal ligament (PDL) cells is less clear. Herein, we examine the effect of exogenous HA on contractility and migration in human and murine PDL cells using arrays of microposts and time-lapse microscopy. Our findings observed HA-treated human PDL cells as more contractile and less migratory than untreated cells. Moreover, the effect of HA on contractility and focal adhesion area was abrogated when PDL cells were treated with Y27632, an inhibitor of rho-dependent kinase, but not when these cells were treated with ML-7, an inhibitor of myosin light chain kinase. Our results provide insight into the mechanobiology of PDL cells, which may contribute towards the development of therapeutic strategies for periodontal healing and tissue regeneration.
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Adhesión Celular , Movimiento Celular , Receptores de Hialuranos/fisiología , Ácido Hialurónico/metabolismo , Ligamento Periodontal/fisiología , Adolescente , Adulto , Animales , Proliferación Celular , Células Cultivadas , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ligamento Periodontal/citología , Fosforilación , Transducción de Señal , Adulto Joven , Quinasas Asociadas a rho/metabolismoRESUMEN
The current understanding of the role of the cell membrane is in a state of flux. Recent experiments show that conventional models, considering only electrophysiological properties of a passive membrane, are incomplete. The neuronal membrane is an active structure with mechanical properties that modulate electrophysiology. Protein transport, lipid bilayer phase, membrane pressure and stiffness can all influence membrane capacitance and action potential propagation. A mounting body of evidence indicates that neuronal mechanics and electrophysiology are coupled, and together shape the membrane potential in tight coordination with other physical properties. In this review, we summarise recent updates concerning electrophysiological-mechanical coupling in neuronal function. In particular, we aim at making the link with two relevant yet often disconnected fields with strong clinical potential: the use of mechanical vibrations-ultrasound-to alter the electrophysiogical state of neurons, e.g., in neuromodulation, and the theories attempting to explain the action of general anaesthetics. STATEMENT OF SIGNIFICANCE: General anaesthetics revolutionised medical practice; now an apparently unrelated technique, ultrasound neuromodulation-aimed at controlling neuronal activity by means of ultrasound-is poised to achieve a similar level of impact. While both technologies are known to alter the electrophysiology of neurons, the way they achieve it is still largely unknown. In this review, we argue that in order to explain their mechanisms/effects, the neuronal membrane must be considered as a coupled mechano-electrophysiological system that consists of multiple physical processes occurring concurrently and collaboratively, as opposed to sequentially and independently. In this framework the behaviour of the cell membrane is not the result of stereotypical mechanisms in isolation but instead emerges from the integrative behaviour of a complexly coupled multiphysics system.
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Potenciales de Acción , Anestesia General , Encéfalo/fisiopatología , Membrana Celular , Modelos Neurológicos , Neuronas , Ondas Ultrasónicas , Animales , HumanosRESUMEN
Craniosynostosis is the premature fusion of the calvarial sutures that is associated with a number of physical and intellectual disabilities spanning from pediatric to adult years. Over the past two decades, techniques in molecular genetics and more recently, advances in high-throughput DNA sequencing have been used to examine the underlying pathogenesis of this disease. To date, mutations in 57 genes have been identified as causing craniosynostosis and the number of newly discovered genes is growing rapidly as a result of the advances in genomic technologies. While contributions from both genetic and environmental factors in this disease are increasingly apparent, there remains a gap in knowledge that bridges the clinical characteristics and genetic markers of craniosynostosis with their signaling pathways and mechanotransduction processes. By linking genotype to phenotype, outlining the role of cell mechanics may further uncover the specific mechanotransduction pathways underlying craniosynostosis. Here, we present a brief overview of the recent findings in craniofacial genetics and cell mechanics, discussing how this information together with animal models is advancing our understanding of craniofacial development.
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Migration of a fibroblast along a collagen fiber can be regarded as cell locomotion in one-dimension (1D). In this process, a cell protrudes forward, forms a new adhesion, produces traction forces, and releases its rear adhesion in order to advance itself along a path. However, how a cell coordinates its adhesion formation, traction forces, and rear release in 1D migration is unclear. Here, we studied fibroblasts migrating along a line of microposts. We found that when the front of a cell protruded onto a new micropost, the traction force produced at its front increased steadily, but did so without a temporal correlation in the force at its rear. Instead, the force at the front coordinated with a decrease in force at the micropost behind the front. A similar correlation in traction forces also occurred at the rear of a cell, where a decrease in force due to adhesion detachment corresponded to an increase in force at the micropost ahead of the rear. Analysis with a bio-chemo-mechanical model for traction forces and adhesion dynamics indicated that the observed relationship between traction forces at the front and back of a cell is possible only when cellular elasticity is lower than the elasticity of the cellular environment.
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Movimiento Celular , Fibroblastos/citología , Animales , Fenómenos Biomecánicos , Simulación por Computador , Elasticidad , Ratones , Células 3T3 NIH , Factores de TiempoRESUMEN
Focal adhesions play a fundamental role in force sensing, which influences a variety of cellular processes and functions, particularly migration and the cell cycle. They consist of large macromolecular assemblies of proteins that associate with integrins, in order to serve as anchor points between the cell and the extracellular matrix. These dynamic regions act as a hub for sensing and transmission of mechanical cues between cells and their surrounding microenvironments. A number of techniques have been used to study focal adhesions, including optical microscopy, substrate micropatterning techniques, and tools which can directly manipulate cells, such as the atomic force microscope. Mechanical stimulation of cells leads to changes in cell contractility, stress fiber remodeling, and focal adhesion position and size; several of the responses explored in this chapter.
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Señales (Psicología) , Adhesiones Focales/metabolismo , Animales , Elasticidad , Humanos , Microscopía de Fuerza AtómicaRESUMEN
There are numerous approaches for producing natural and synthetic 3D scaffolds that support the proliferation of mammalian cells. 3D scaffolds better represent the natural cellular microenvironment and have many potential applications in vitro and in vivo. Here, we demonstrate that 3D cellulose scaffolds produced by decellularizing apple hypanthium tissue can be employed for in vitro 3D culture of NIH3T3 fibroblasts, mouse C2C12 muscle myoblasts and human HeLa epithelial cells. We show that these cells can adhere, invade and proliferate in the cellulose scaffolds. In addition, biochemical functionalization or chemical cross-linking can be employed to control the surface biochemistry and/or mechanical properties of the scaffold. The cells retain high viability even after 12 continuous weeks of culture and can achieve cell densities comparable with other natural and synthetic scaffold materials. Apple derived cellulose scaffolds are easily produced, inexpensive and originate from a renewable source. Taken together, these results demonstrate that naturally derived cellulose scaffolds offer a complementary approach to existing techniques for the in vitro culture of mammalian cells in a 3D environment.
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Técnicas de Cultivo de Célula/métodos , Celulosa/análisis , Malus/química , Andamios del Tejido/química , Animales , Flores/química , Técnica del Anticuerpo Fluorescente , Células HeLa , Humanos , Ratones , Microscopía de Fuerza Atómica , Microscopía Electrónica de Rastreo , Mioblastos , Células 3T3 NIHRESUMEN
High-pass filtering is required for the removal of background field inhomogeneities in magnetic resonance phase images. This high-pass filtering smooths across boundaries between areas with large differences in phase. The most prominent boundary is the surface of the brain where areas with large phase values inside the brain are located close to areas outside the brain where the phase is, on average, zero. Cortical areas, which are of great interest in brain MRI, are therefore often degraded by high-pass filtering. Here, we propose the use of the bilateral filter for the high-pass filtering step. The bilateral filter is essentially a Gaussian filter that stops smoothing at boundaries. We show that the bilateral filter improves image quality at the brain's surface, without sacrificing contrast within the brain.