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Traction force microscopy of migrating normal and H-ras transformed 3T3 fibroblasts.

Munevar, S; Wang , Y; Dembo, M
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /04/2001 EN
Relevância na Pesquisa
66.32%
Mechanical interactions between cell and substrate are involved in vital cellular functions from migration to signal transduction. A newly developed technique, traction force microscopy, makes it possible to visualize the dynamic characteristics of mechanical forces exerted by fibroblasts, including the magnitude, direction, and shear. In the present study such analysis is applied to migrating normal and transformed 3T3 cells. For normal cells, the lamellipodium provides almost all the forces for forward locomotion. A zone of high shear separates the lamellipodium from the cell body, suggesting that they are mechanically distinct entities. Timing and distribution of tractions at the leading edge bear no apparent relationship to local protrusive activities. However, changes in the pattern of traction forces often precede changes in the direction of migration. These observations suggest a frontal towing mechanism for cell migration, where dynamic traction forces at the leading edge actively pull the cell body forward. For H-ras transformed cells, pockets of weak, transient traction scatter among small pseudopods and appear to act against one another. The shear pattern suggests multiple disorganized mechanical domains. The weak, poorly coordinated traction forces...

High Resolution Traction Force Microscopy Based on Experimental and Computational Advances

Sabass, Benedikt; Gardel, Margaret L.; Waterman, Clare M.; Schwarz, Ulrich S.
Fonte: The Biophysical Society Publicador: The Biophysical Society
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
66.38%
Cell adhesion and migration crucially depend on the transmission of actomyosin-generated forces through sites of focal adhesion to the extracellular matrix. Here we report experimental and computational advances in improving the resolution and reliability of traction force microscopy. First, we introduce the use of two differently colored nanobeads as fiducial markers in polyacrylamide gels and explain how the displacement field can be computationally extracted from the fluorescence data. Second, we present different improvements regarding standard methods for force reconstruction from the displacement field, which are the boundary element method, Fourier-transform traction cytometry, and traction reconstruction with point forces. Using extensive data simulation, we show that the spatial resolution of the boundary element method can be improved considerably by splitting the elastic field into near, intermediate, and far field. Fourier-transform traction cytometry requires considerably less computer time, but can achieve a comparable resolution only when combined with Wiener filtering or appropriate regularization schemes. Both methods tend to underestimate forces, especially at small adhesion sites. Traction reconstruction with point forces does not suffer from this limitation...

Imaging in-plane and normal stresses near an interface crack using traction force microscopy

Xu, Ye; Engl, Wilfried C.; Jerison, Elizabeth R.; Wallenstein, Kevin J.; Hyland, Callen; Wilen, Larry A.; Dufresne, Eric R.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
66.14%
Colloidal coatings, such as paint, are all around us. However, we know little about the mechanics of the film-forming process because the composition and properties of drying coatings vary dramatically in space and time. To surmount this challenge, we extend traction force microscopy to quantify the spatial distribution of all three components of the stress at the interface of two materials. We apply this approach to image stress near the tip of a propagating interface crack in a drying colloidal coating and extract the stress intensity factor.

Three-Dimensional Traction Force Microscopy: A New Tool for Quantifying Cell-Matrix Interactions

Franck, Christian; Maskarinec, Stacey A.; Tirrell, David A.; Ravichandran, Guruswami
Fonte: Public Library of Science Publicador: Public Library of Science
Tipo: Artigo de Revista Científica
Publicado em 29/03/2011 EN
Relevância na Pesquisa
66.39%
The interactions between biochemical processes and mechanical signaling play important roles during various cellular processes such as wound healing, embryogenesis, metastasis, and cell migration. While traditional traction force measurements have provided quantitative information about cell matrix interactions in two dimensions, recent studies have shown significant differences in the behavior and morphology of cells when placed in three-dimensional environments. Hence new quantitative experimental techniques are needed to accurately determine cell traction forces in three dimensions. Recently, two approaches both based on laser scanning confocal microscopy have emerged to address this need. This study highlights the details, implementation and advantages of such a three-dimensional imaging methodology with the capability to compute cellular traction forces dynamically during cell migration and locomotion. An application of this newly developed three-dimensional traction force microscopy (3D TFM) technique to single cell migration studies of 3T3 fibroblasts is presented to show that this methodology offers a new quantitative vantage point to investigate the three-dimensional nature of cell-ECM interactions.

The role of the cytoskeleton in cellular force generation in 2D and 3D environments

Kraning-Rush, Casey M.; Carey, Shawn P.; Califano, Joseph P.; Smith, Brooke N.; Reinhart-King, Cynthia A.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
46.34%
To adhere and migrate, cells generate forces through the cytoskeleton that are transmitted to the surrounding matrix. While cellular force generation has been studied on 2D substrates, less is known about cytoskeletal-mediated traction forces of cells embedded in more in vivo-like 3D matrices. Recent studies have revealed important differences between the cytoskeletal structure, adhesion, and migration of cells in 2D and 3D. Because the cytoskeleton mediates force, we sought to directly compare the role of the cytoskeleton in modulating cell force in 2D and 3D. MDA-MB-231 cells were treated with agents that perturbed actin, microtubules, or myosin, and analyzed for changes in cytoskeletal organization and force generation in both 2D and 3D. To quantify traction stresses in 2D, Traction Force Microscopy was used; in 3D, force was assessed based on single cell-mediated collagen fibril reorganization imaged using Confocal Reflectance Microscopy. Interestingly, even though previous studies have observed differences in cell behaviors like migration in 2D and 3D, our data indicate that forces generated on 2D substrates correlate with forces within 3D matrices. Disruption of actin, myosin or microtubules in either 2D or 3D microenvironments disrupts cell-generated force. These data suggest that despite differences in cytoskeletal organization in 2D and 3D...

Microtubule depolymerization induces traction force increase through two distinct pathways

Rape, Andrew; Guo, Wei-hui; Wang, Yu-li
Fonte: Company of Biologists Publicador: Company of Biologists
Tipo: Artigo de Revista Científica
Publicado em 15/12/2011 EN
Relevância na Pesquisa
56.35%
Traction forces increase after microtubule depolymerization; however, the signaling mechanisms underlying this, in particular the dependence upon myosin II, remain unclear. We investigated the mechanism of traction force increase after nocodazole-induced microtubule depolymerization by applying traction force microscopy to cells cultured on micropatterned polyacrylamide hydrogels to obtain samples of homogeneous shape and size. Control cells and cells treated with a focal adhesion kinase (FAK) inhibitor showed similar increases in traction forces, indicating that the response is independent of FAK. Surprisingly, pharmacological inhibition of myosin II did not prevent the increase of residual traction forces upon nocodazole treatment. This increase was abolished upon pharmacological inhibition of FAK. These results suggest two distinct pathways for the regulation of traction forces. First, microtubule depolymerization activates a myosin-II-dependent mechanism through a FAK-independent pathway. Second, microtubule depolymerization also enhances traction forces through a myosin-II-independent, FAK-regulated pathway. Traction forces are therefore regulated by a complex network of complementary signals and force-generating mechanisms.

Improved throughput traction microscopy reveals pivotal role for matrix stiffness in fibroblast contractility and TGF-β responsiveness

Marinković, Aleksandar; Mih, Justin D.; Park, Jin-Ah; Liu, Fei; Tschumperlin, Daniel J.
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
46.35%
Lung fibroblast functions such as matrix remodeling and activation of latent transforming growth factor-β1 (TGF-β1) are associated with expression of the myofibroblast phenotype and are directly linked to fibroblast capacity to generate force and deform the extracellular matrix. However, the study of fibroblast force-generating capacities through methods such as traction force microscopy is hindered by low throughput and time-consuming procedures. In this study, we improved at the detail level methods for higher-throughput traction measurements on polyacrylamide hydrogels using gel-surface-bound fluorescent beads to permit autofocusing and automated displacement mapping, and transduction of fibroblasts with a fluorescent label to streamline cell boundary identification. Together these advances substantially improve the throughput of traction microscopy and allow us to efficiently compute the forces exerted by lung fibroblasts on substrates spanning the stiffness range present in normal and fibrotic lung tissue. Our results reveal that lung fibroblasts dramatically alter the forces they transmit to the extracellular matrix as its stiffness changes, with very low forces generated on matrices as compliant as normal lung tissue. Moreover...

Multidimensional traction force microscopy reveals out-of-plane rotational moments about focal adhesions

Legant, Wesley R.; Choi, Colin K.; Miller, Jordan S.; Shao, Lin; Gao, Liang; Betzig, Eric; Chen, Christopher S.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
66.31%
Recent methods have revealed that cells on planar substrates exert both shear (in-plane) and normal (out-of-plane) tractions against the extracellular matrix (ECM). However, the location and origin of the normal tractions with respect to the adhesive and cytoskeletal elements of cells have not been elucidated. We developed a high-spatiotemporal-resolution, multidimensional (2.5D) traction force microscopy to measure and model the full 3D nature of cellular forces on planar 2D surfaces. We show that shear tractions are centered under elongated focal adhesions whereas upward and downward normal tractions are detected on distal (toward the cell edge) and proximal (toward the cell body) ends of adhesions, respectively. Together, these forces produce significant rotational moments about focal adhesions in both protruding and retracting peripheral regions. Temporal 2.5D traction force microscopy analysis of migrating and spreading cells shows that these rotational moments are highly dynamic, propagating outward with the leading edge of the cell. Finally, we developed a finite element model to examine how rotational moments could be generated about focal adhesions in a thin lamella. Our model suggests that rotational moments can be generated largely via shear lag transfer to the underlying ECM from actomyosin contractility applied at the intracellular surface of a rigid adhesion of finite thickness. Together...

Remanent cell traction force in renal vascular smooth muscle cells induced by integrin-mediated mechanotransduction

Balasubramanian, Lavanya; Lo, Chun-Min; Sham, James S. K.; Yip, Kay-Pong
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
56.41%
It was previously demonstrated in isolated renal vascular smooth muscle cells (VSMCs) that integrin-mediated mechanotransduction triggers intracellular Ca2+ mobilization, which is the hallmark of myogenic response in VSMCs. To test directly whether integrin-mediated mechanotransduction results in the myogenic response-like behavior in renal VSMCs, cell traction force microscopy was used to monitor cell traction force when the cells were pulled with fibronectin-coated or low density lipoprotein (LDL)-coated paramagnetic beads. LDL-coated beads were used as a control for nonintegrin-mediated mechanotransduction. Pulling with LDL-coated beads increased the cell traction force by 61 ± 12% (9 cells), which returned to the prepull level after the pulling process was terminated. Pulling with noncoated beads had a minimal increase in the cell traction force (12 ± 9%, 8 cells). Pulling with fibronectin-coated beads increased the cell traction force by 56 ± 20% (7 cells). However, the cell traction force was still elevated by 23 ± 14% after the pulling process was terminated. This behavior is analogous to the changes of vascular resistance in pressure-induced myogenic response, in which vascular resistance remains elevated after myogenic constriction. Fibronectin is a native ligand for α5β1-integrins in VSMCs. Similar remanent cell traction force was found when cells were pulled with beads coated with β1-integrin antibody (Ha2/5). Activation of β1-integrin with soluble antibody also triggered variations of cell traction force and Ca2+ mobilization...

Finite Element Analysis of Traction Force Microscopy: Influence of Cell Mechanics, Adhesion, and Morphology

Zielinski, Rachel; Mihai, Cosmin; Kniss, Douglas; Ghadiali, Samir N.
Fonte: American Society of Mechanical Engineers Publicador: American Society of Mechanical Engineers
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
66.42%
The interactions between adherent cells and their extracellular matrix (ECM) have been shown to play an important role in many biological processes, such as wound healing, morphogenesis, differentiation, and cell migration. Cells attach to the ECM at focal adhesion sites and transmit contractile forces to the substrate via cytoskeletal actin stress fibers. This contraction results in traction stresses within the substrate/ECM. Traction force microscopy (TFM) is an experimental technique used to quantify the contractile forces generated by adherent cells. In TFM, cells are seeded on a flexible substrate and displacements of the substrate caused by cell contraction are tracked and converted to a traction stress field. The magnitude of these traction stresses are normally used as a surrogate measure of internal cell contractile force or contractility. We hypothesize that in addition to contractile force, other biomechanical properties including cell stiffness, adhesion energy density, and cell morphology may affect the traction stresses measured by TFM. In this study, we developed finite element models of the 2D and 3D TFM techniques to investigate how changes in several biomechanical properties alter the traction stresses measured by TFM. We independently varied cell stiffness...

Improved-Throughput Traction Microscopy Based on Fluorescence Micropattern for Manual Microscopy

Liu, Kai; Yuan, Yuan; Huang, Jianyong; Wei, Qiong; Pang, Mingshu; Xiong, Chunyang; Fang, Jing
Fonte: Public Library of Science Publicador: Public Library of Science
Tipo: Artigo de Revista Científica
Publicado em 01/08/2013 EN
Relevância na Pesquisa
46.43%
Traction force microscopy (TFM) is a quantitative technique for measuring cellular traction force, which is important in understanding cellular mechanotransduction processes. Traditional TFM has a significant limitation in that it has a low measurement throughput, commonly one per TFM dish, due to a lack of cell position information. To obtain enough cellular traction force data, an onerous workload is required including numerous TFM dish preparations and heavy cell-seeding activities, creating further difficulty in achieving identical experimental conditions among batches. In this paper, we present an improved-throughput TFM method using the well-developed microcontact printing technique and chemical modifications of linking microbeads to the gel surface to address these limitations. Chemically linking the microbeads to the gel surface has no significant influence on cell proliferation, morphology, cytoskeleton, and adhesion. Multiple pairs of force loaded and null force fluorescence images can be easily acquired by means of manual microscope with the aid of a fluorescence micropattern made by microcontact printing. Furthermore, keeping the micropattern separate from cells by using gels effectively eliminates the potential negative effect of the micropattern on the cells. This novel design greatly improves the analysis throughput of traditional TFM from one to at least twenty cells per petri dish without losing unique advantages...

High Resolution, Large Deformation 3D Traction Force Microscopy

Toyjanova, Jennet; Bar-Kochba, Eyal; López-Fagundo, Cristina; Reichner, Jonathan; Hoffman-Kim, Diane; Franck, Christian
Fonte: Public Library of Science Publicador: Public Library of Science
Tipo: Artigo de Revista Científica
Publicado em 16/04/2014 EN
Relevância na Pesquisa
66.31%
Traction Force Microscopy (TFM) is a powerful approach for quantifying cell-material interactions that over the last two decades has contributed significantly to our understanding of cellular mechanosensing and mechanotransduction. In addition, recent advances in three-dimensional (3D) imaging and traction force analysis (3D TFM) have highlighted the significance of the third dimension in influencing various cellular processes. Yet irrespective of dimensionality, almost all TFM approaches have relied on a linear elastic theory framework to calculate cell surface tractions. Here we present a new high resolution 3D TFM algorithm which utilizes a large deformation formulation to quantify cellular displacement fields with unprecedented resolution. The results feature some of the first experimental evidence that cells are indeed capable of exerting large material deformations, which require the formulation of a new theoretical TFM framework to accurately calculate the traction forces. Based on our previous 3D TFM technique, we reformulate our approach to accurately account for large material deformation and quantitatively contrast and compare both linear and large deformation frameworks as a function of the applied cell deformation. Particular attention is paid in estimating the accuracy penalty associated with utilizing a traditional linear elastic approach in the presence of large deformation gradients.

Model-based Traction Force Microscopy Reveals Differential Tension in Cellular Actin Bundles

Soiné, Jérôme R. D.; Brand, Christoph A.; Stricker, Jonathan; Oakes, Patrick W.; Gardel, Margaret L.; Schwarz, Ulrich S.
Fonte: Public Library of Science Publicador: Public Library of Science
Tipo: Artigo de Revista Científica
Publicado em 06/03/2015 EN
Relevância na Pesquisa
66.31%
Adherent cells use forces at the cell-substrate interface to sense and respond to the physical properties of their environment. These cell forces can be measured with traction force microscopy which inverts the equations of elasticity theory to calculate them from the deformations of soft polymer substrates. We introduce a new type of traction force microscopy that in contrast to traditional methods uses additional image data for cytoskeleton and adhesion structures and a biophysical model to improve the robustness of the inverse procedure and abolishes the need for regularization. We use this method to demonstrate that ventral stress fibers of U2OS-cells are typically under higher mechanical tension than dorsal stress fibers or transverse arcs.

A Novel Cell Traction Force Microscopy to Study Multi-Cellular System

Tang, Xin; Tofangchi, Alireza; Anand, Sandeep V.; Saif, Taher A.
Fonte: Public Library of Science Publicador: Public Library of Science
Tipo: Artigo de Revista Científica
Publicado em 05/06/2014 EN
Relevância na Pesquisa
66.41%
Traction forces exerted by adherent cells on their microenvironment can mediate many critical cellular functions. Accurate quantification of these forces is essential for mechanistic understanding of mechanotransduction. However, most existing methods of quantifying cellular forces are limited to single cells in isolation, whereas most physiological processes are inherently multi-cellular in nature where cell-cell and cell-microenvironment interactions determine the emergent properties of cell clusters. In the present study, a robust finite-element-method-based cell traction force microscopy technique is developed to estimate the traction forces produced by multiple isolated cells as well as cell clusters on soft substrates. The method accounts for the finite thickness of the substrate. Hence, cell cluster size can be larger than substrate thickness. The method allows computing the traction field from the substrate displacements within the cells' and clusters' boundaries. The displacement data outside these boundaries are not necessary. The utility of the method is demonstrated by computing the traction generated by multiple monkey kidney fibroblasts (MKF) and human colon cancerous (HCT-8) cells in close proximity, as well as by large clusters. It is found that cells act as individual contractile groups within clusters for generating traction. There may be multiple of such groups in the cluster...

Three-Dimensional Quantification of Cellular Traction Forces and Mechanosensing of Thin Substrata by Fourier Traction Force Microscopy

del Álamo, Juan C.; Meili, Ruedi; Álvarez-González, Begoña; Alonso-Latorre, Baldomero; Bastounis, Effie; Firtel, Richard; Lasheras, Juan C.
Fonte: Public Library of Science Publicador: Public Library of Science
Tipo: Artigo de Revista Científica
Publicado em 04/09/2013 EN
Relevância na Pesquisa
66.43%
We introduce a novel three-dimensional (3D) traction force microscopy (TFM) method motivated by the recent discovery that cells adhering on plane surfaces exert both in-plane and out-of-plane traction stresses. We measure the 3D deformation of the substratum on a thin layer near its surface, and input this information into an exact analytical solution of the elastic equilibrium equation. These operations are performed in the Fourier domain with high computational efficiency, allowing to obtain the 3D traction stresses from raw microscopy images virtually in real time. We also characterize the error of previous two-dimensional (2D) TFM methods that neglect the out-of-plane component of the traction stresses. This analysis reveals that, under certain combinations of experimental parameters (cell size, substratums' thickness and Poisson's ratio), the accuracy of 2D TFM methods is minimally affected by neglecting the out-of-plane component of the traction stresses. Finally, we consider the cell's mechanosensing of substratum thickness by 3D traction stresses, finding that, when cells adhere on thin substrata, their out-of-plane traction stresses can reach four times deeper into the substratum than their in-plane traction stresses. It is also found that the substratum stiffness sensed by applying out-of-plane traction stresses may be up to 10 times larger than the stiffness sensed by applying in-plane traction stresses.

Free Form Deformation–Based Image Registration Improves Accuracy of Traction Force Microscopy

Jorge-Peñas, Alvaro; Izquierdo-Alvarez, Alicia; Aguilar-Cuenca, Rocio; Vicente-Manzanares, Miguel; Garcia-Aznar, José Manuel; Van Oosterwyck, Hans; de-Juan-Pardo, Elena M.; Ortiz-de-Solorzano, Carlos; Muñoz-Barrutia, Arrate
Fonte: Public Library of Science Publicador: Public Library of Science
Tipo: Artigo de Revista Científica
Publicado em 07/12/2015 EN
Relevância na Pesquisa
66.22%
Traction Force Microscopy (TFM) is a widespread method used to recover cellular tractions from the deformation that they cause in their surrounding substrate. Particle Image Velocimetry (PIV) is commonly used to quantify the substrate’s deformations, due to its simplicity and efficiency. However, PIV relies on a block-matching scheme that easily underestimates the deformations. This is especially relevant in the case of large, locally non-uniform deformations as those usually found in the vicinity of a cell’s adhesions to the substrate. To overcome these limitations, we formulate the calculation of the deformation of the substrate in TFM as a non-rigid image registration process that warps the image of the unstressed material to match the image of the stressed one. In particular, we propose to use a B-spline -based Free Form Deformation (FFD) algorithm that uses a connected deformable mesh to model a wide range of flexible deformations caused by cellular tractions. Our FFD approach is validated in 3D fields using synthetic (simulated) data as well as with experimental data obtained using isolated endothelial cells lying on a deformable, polyacrylamide substrate. Our results show that FFD outperforms PIV providing a deformation field that allows a better recovery of the magnitude and orientation of tractions. Together...

Numeric reconstruction of 2D cellular actomyosin network from substrate displacement

Nishitani,Wagner Shin; Carbonari,Ronny Calixto; Alencar,Adriano Mesquita
Fonte: Sociedade Brasileira de Engenharia Biomédica Publicador: Sociedade Brasileira de Engenharia Biomédica
Tipo: Artigo de Revista Científica Formato: text/html
Publicado em 01/01/2015 EN
Relevância na Pesquisa
66.14%
Introduction: One of the fundamental structural elements of the cell is the cytoskeleton. Along with myosin, actin microfilaments are responsible for cellular contractions, and their organization may be related to pathological changes in myocardial tissue. Due to the complexity of factors involved, numerical modeling of the cytoskeleton has the potential to contribute to a better understanding of mechanical cues in cellular activities. In this work, a systematic method was developed for the reconstruction of an actomyosin topology based on the displacement exerted by the cell on a flexible substrate. It is an inverse problem which could be considered a phenomenological approach to traction force microscopy (TFM). Methods An actomyosin distribution was found with a topology optimization method (TOM), varying the material density and angle of contraction of each element of the actomyosin domain. The routine was implemented with a linear material model for the bidimensional actomyosin elements and tridimensional substrate. The topology generated minimizes the nodal displacement squared differences between the generated topology and experimental displacement fields obtained by TFM. The structure resulting from TOM was compared to the actin structures observed experimentally with a GFP-attached actin marker. Results The optimized topology reproduced the main features of the experimental actin and its squared displacement differences were 11.24 µm2...

Traction force microscopy on soft elastic substrates: a guide to recent computational advances

Schwarz, Ulrich S.; Soine, Jerome R. D.
Fonte: Universidade Cornell Publicador: Universidade Cornell
Tipo: Artigo de Revista Científica
Publicado em 08/06/2015
Relevância na Pesquisa
66.26%
The measurement of cellular traction forces on soft elastic substrates has become a standard tool for many labs working on mechanobiology. Here we review the basic principles and different variants of this approach. In general, the extraction of the substrate displacement field from image data and the reconstruction procedure for the forces are closely linked to each other and limited by the presence of experimental noise. We discuss different strategies to reconstruct cellular forces as they follow from the foundations of elasticity theory, including two- versus three-dimensional, inverse versus direct and linear versus non-linear approaches. We also discuss how biophysical models can improve force reconstruction and comment on practical issues like substrate preparation, image processing and the availability of software for traction force microscopy.; Comment: Revtex, 29 pages, 3 PDF figures, 2 tables. BBA - Molecular Cell Research, online since 27 May 2015, special issue on mechanobiology

Three-Dimensional Quantification of Cellular Traction Forces and Mechanosensing of Thin Substrata by Fourier Traction Force Microscopy

del Alamo, Juan C; Meili, Ruedi; Alvarez-Gonzalez, Begoña; Alonso-Latorre, Baldomero; Bastounis, Effie; Firtel, Richard; Lasheras, Juan C
Fonte: Universidade Cornell Publicador: Universidade Cornell
Tipo: Artigo de Revista Científica
Publicado em 18/06/2013
Relevância na Pesquisa
66.43%
We introduce a novel three-dimensional (3D) traction force microscopy (TFM) method motivated by the recent discovery that cells adhering on plane surfaces exert both in-plane and out-of-plane traction stresses. We measure the 3D deformation of the substratum on a thin layer near its surface, and input this information into an exact analytical solution of the elastic equilibrium equation. These operations are performed in the Fourier domain with high computational efficiency, allowing to obtain the 3D traction stresses from raw microscopy images virtually in real time. We also characterize the error of previous two-dimensional (2D) TFM methods that neglect the out-of-plane component of the traction stresses. This analysis reveals that, under certain combinations of experimental parameters (\ie cell size, substratums' thickness and Poisson's ratio), the accuracy of 2D TFM methods is minimally affected by neglecting the out-of-plane component of the traction stresses. Finally, we consider the cell's mechanosensing of substratum thickness by 3D traction stresses, finding that, when cells adhere on thin substrata, their out-of-plane traction stresses can reach four times deeper into the substratum than their in-plane traction stresses. It is also found that the substratum stiffness sensed by applying out-of-plane traction stresses may be up to 10 times larger than the stiffness sensed by applying in-plane traction stresses.

Three-Dimensional Traction Force Microscopy: A New Tool for Quantifying Cell-Matrix Interactions

Franck, Christian; Maskarinec, Stacey A.; Tirrell, David A.; Ravichandran, Guruswami
Fonte: Public Library of Science Publicador: Public Library of Science
Tipo: Article; PeerReviewed Formato: application/pdf; image/tiff; image/tiff; image/tiff
Publicado em 29/03/2011
Relevância na Pesquisa
66.39%
The interactions between biochemical processes and mechanical signaling play important roles during various cellular processes such as wound healing, embryogenesis, metastasis, and cell migration. While traditional traction force measurements have provided quantitative information about cell matrix interactions in two dimensions, recent studies have shown significant differences in the behavior and morphology of cells when placed in three-dimensional environments. Hence new quantitative experimental techniques are needed to accurately determine cell traction forces in three dimensions. Recently, two approaches both based on laser scanning confocal microscopy have emerged to address this need. This study highlights the details, implementation and advantages of such a three-dimensional imaging methodology with the capability to compute cellular traction forces dynamically during cell migration and locomotion. An application of this newly developed three-dimensional traction force microscopy (3D TFM) technique to single cell migration studies of 3T3 fibroblasts is presented to show that this methodology offers a new quantitative vantage point to investigate the three-dimensional nature of cell-ECM interactions.