Chen, Christopher S.

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Now showing 1 - 9 of 9
  • Publication
    E-cadherin engagement stimulates proliferation via Rac1
    (2006-05-08) Liu, Wendy F; Nelson, Celeste M; Pirone, Dana M; Chen, Christopher S.
    E-cadherin has been linked to the suppression of tumor growth and the inhibition of cell proliferation in culture. We observed that progressively decreasing the seeding density of normal rat kidney-52E (NRK- 52E) or MCF-10A epithelial cells from confluence, indeed, released cells from growth arrest. Unexpectedly, a further decrease in seeding density so that cells were isolated from neighboring cells decreased proliferation. Experiments using microengineered substrates showed that E-cadherin engagement stimulated the peak in proliferation at intermediate seeding densities, and that the proliferation arrest at high densities did not involve E-cadherin, but rather resulted from a crowding-dependent decrease in cell spreading against the underlying substrate. Rac1 activity, which was induced by E-cadherin engagement specifically at intermediate seeding densities, was required for the cadherin-stimulated proliferation, and the control of Rac1 activation by E-cadherin was mediated by p120- catenin. Together, these findings demonstrate a stimulatory role for E-cadherin in proliferative regulation, and identify a simple mechanism by which cell–cell contact may trigger or inhibit epithelial cell proliferation in different settings.
  • Publication
    Separate but not equal: Differential mechanical roles for myosin isoforms
    (2007-05-01) Chen, Christopher S.
    Cells undergo many structural-mechanical changes as an inextricable component of cellular motility, cytokinesis, and changes in cell shape. The mere act of receptor-mediated adhesion to extracellular matrix involves massive changes in cytoskeletal organization, spreading, and flattening of cells against the matrix, and the generation of traction forces through the contractile activity of cells. The primary force-generating proteins involved in these mechanical processes are thought to be the nonmuscle myosin II's (NMM-II). Of the three different nonmuscle myosin II isoforms that have been identified, two (NMM-IIA and NMM-IIB) are found almost ubiquitously in higher organisms. Yet, it has remained unclear whether these molecules play redundant, overlapping, or distinct roles in the varying mechanical functions of cells.
  • Publication
    Nanotechnology for Cell–Substrate Interactions
    (2006-01-01) Sniadecki, Nathan J; Desai, Ravi A; Ruiz, Sami Alom; Chen, Christopher S.
    In the pursuit to understand the interaction between cells and their underlying substrates, the life sciences are beginning to incorporate micro- and nanotechnology-based tools to probe and measure cells. The development of these tools portends endless possibilities for new insights into the fundamental relationships between cells and their surrounding microenvironment that underlie the physiology of human tissue. Here, we review techniques and tools that have been used to study how a cell responds to the physical factors in its environment. We also discuss unanswered questions that could be addressed by these approaches to better elucidate the molecular processes and mechanical forces that dominate the interactions between cells and their physical scaffolds.
  • Publication
    Magnetic microposts as an approach to apply forces to living cells
    (2007-07-14) Sniadecki, Nathan J; Anguelouch, Alexandre; Yang, Michael T; Lamb, Corinne M; Liu, Zhijun; Kirschner, Stuart B; Liu, Yaohua; Reich, Daniel H; Chen, Christopher S.
    Cells respond to mechanical forces whether applied externally or generated internally via the cytoskeleton. To study the cellular response to forces separately, we applied external forces to cells via microfabricated magnetic posts containing cobalt nanowires interspersed among an array of elastomeric posts, which acted as independent sensors to cellular traction forces. A magnetic field induced torque in the nanowires, which deflected the magnetic posts and imparted force to individual adhesions of cells attached to the array. Using this system, we examined the cellular reaction to applied forces and found that applying a step force led to an increase in local focal adhesion size at the site of application but not at nearby nonmagnetic posts. Focal adhesion recruitment was enhanced further when cells were subjected to multiple force actuations within the same time interval. Recording the traction forces in response to such force stimulation revealed two responses: a sudden loss in contractility that occurred within the first minute of stimulation or a gradual decay in contractility over several minutes. For both types of responses, the subcellular distribution of loss in traction forces was not confined to locations near the actuated micropost, nor uniformly across the whole cell, but instead occurred at discrete locations along the cell periphery. Together, these data reveal an important dynamic biological relationship between external and internal forces and demonstrate the utility of this microfabricated system to explore this interaction. Supporting materials: http://www.pnas.org/cgi/content/full/0611613104/DC1
  • Publication
    Emergence of Patterned Stem Cell Differentiation Within Multicellular Structures
    (2008-11-01) Ruiz, Sami Alom; Chen, Christopher S.
    The ability of stem cells to differentiate into specified lineages in the appropriate locations is vital to morphogenesis and adult tissue regeneration. Although soluble signals are important regulators of patterned differentiation, here we show that gradients of mechanical forces can also drive patterning of lineages. In the presence of soluble factors permitting osteogenic and adipogenic differentiation, human mesenchymal stem cells at the edge of multicellular islands differentiate into the osteogenic lineage, whereas those in the center became adipocytes. Interestingly, changing the shape of the multicellular sheet modulated the locations of osteogenic versus adipogenic differentiation. Measuring traction forces revealed gradients of stress that preceded and mirrored the patterns of differentiation, where regions of high stress resulted in osteogenesis, whereas stem cells in regions of low stress differentiated to adipocytes. Inhibiting cytoskeletal tension suppressed the relative degree of osteogenesis versus adipogenesis, and this spatial patterning of differentiation was also present in three-dimensional multicellular clusters. These findings demonstrate a role for mechanical forces in linking multicellular organization to spatial differentials of cell differentiation, and they represent an important guiding principle in tissue patterning that could be exploited in stem cell-based therapies.
  • Publication
    An Inhibitory Role for FAK in Regulating Proliferation: A Link Between Limited Adhesion and RhoA-ROCK Signaling
    (2006-07-17) Pirone, Dana M; Liu, Wendy F; Ruiz, Sami Alom; Gao, Lin; Raghavan, Srivatsan; Lemmon, Christopher A; Romer, Lewis H; Chen, Christopher S.
    Focal adhesion kinase (FAK) transduces cell adhesion to the extracellular matrix into proliferative signals. We show that FAK overexpression induced proliferation in endothelial cells, which are normally growth arrested by limited adhesion. Interestingly, displacement of FAK from adhesions by using a FAK−/− cell line or by expressing the C-terminal fragment FRNK also caused an escape of adhesion-regulated growth arrest, suggesting dual positive and negative roles for FAK in growth regulation. Expressing kinase-dead FAK-Y397F in FAK−/− cells prevented uncontrolled growth, demonstrating the antiproliferative function of inactive FAK. Unlike FAK overexpression–induced growth, loss of growth control in FAK−/− or FRNK-expressing cells increased RhoA activity, cytoskeletal tension, and focal adhesion formation. ROCK inhibition rescued adhesion-dependent growth control in these cells, and expression of constitutively active RhoA or ROCK dysregulated growth. These findings demonstrate the ability of FAK to suppress and promote growth, and underscore the importance of multiple mechanisms, even from one molecule, to control cell proliferation.
  • Publication
    Mechanical Tugging Force Regulates the Size of Cell-Cell Junctions
    (2010-03-31) Cohen, Daniel M.; Liu, Zhijun; Yang, Michael T.; Tan, John L.; Sniadecki, Nathan J.; Chen, Christopher S.; Ruiz, Sami Alom; Nelson, Celeste M.
    Actomyosin contractility affects cellular organization within tissues in part through the generation of mechanical forces at sites of cell–matrix and cell–cell contact. While increased mechanical loading at cell–matrix adhesions results in focal adhesion growth, whether forces drive changes in the size of cell–cell adhesions remains an open question. To investigate the responsiveness of adherens junctions (AJ) to force, we adapted a system of microfabricated force sensors to quantitatively report cell–cell tugging force and AJ size. We observed that AJ size was modulated by endothelial cell–cell tugging forces: AJs and tugging force grew or decayed with myosin activation or inhibition, respectively. Myosin-dependent regulation of AJs operated in concert with a Rac1, and this coordinated regulation was illustrated by showing that the effects of vascular permeability agents (S1P, thrombin) on junctional stability were reversed by changing the extent to which these agents coupled to the Rac and myosin-dependent pathways. Furthermore, direct application of mechanical tugging force, rather than myosin activity per se, was sufficient to trigger AJ growth. These findings demonstrate that the dynamic coordination of mechanical forces and cell–cell adhesive interactions likely is critical to the maintenance of multicellular integrity and highlight the need for new approaches to study tugging forces.
  • Publication
    Assembly of multicellular constructs and microarrays of cells using magnetic nanowires
    (2005-06-01) Tanase, Monica; Felton, Edward J.; Gray, Darren S.; Hultgren, Anne; Chen, Christopher S.; Reich, Daniel H.
    An approach is described for controlling the spatial organization of mammalian cells using ferromagnetic nanowires in conjunction with patterned micromagnet arrays. The nanowires are fabricated by electrodeposition in nanoporous templates, which allows for precise control of their size and magnetic properties. The high aspect ratio and large remanent magnetization of the nanowires enable suspensions of cells bound to Ni nanowires to be controlled with low magnetic fields. This was used to produce one- and two-dimensional field-tuned patterning of suspended 3T3 mouse fibroblasts. Self-assembled one-dimensional chains of cells were obtained through manipulation of the wires' dipolar interactions. Ordered patterns of individual cells in two dimensions were formed through trapping onto magnetic microarrays of ellipsoidal permalloy micromagnets. Cell chains were formed on the arrays by varying the spacing between the micromagnets or the strength of fluid flow over the arrays. The positioning of cells on the array was further controlled by varying the direction of an external magnetic field. These results demonstrate the possibility of using magnetic nanowires to organize cells.
  • Publication
    Magnetic microposts for mechanical stimulation of biological cells: Fabrication, characterization, and analysis
    (2008-04-16) Sniadecki, Nathan J; Lamb, Corinne M; Liu, Yaohua; Chen, Christopher S.; Reich, Daniel H
    Cells use force as a mechanical signal to sense and respond to their microenvironment. Understanding how mechanical forces affect living cells requires the development of tool sets that can apply nanoscale forces and also measure cellular traction forces. However, there has been a lack of techniques that integrate actuation and sensing components to study force as a mechanical signal. Here, we describe a system that uses an array of elastomeric microposts to apply external forces to cells through cobalt nanowires embedded inside the microposts. We first biochemically treat the posts’ surfaces to restrict cell adhesion to the posts' tips. Then by applying a uniform magnetic field (B < 0.3 T), we induce magnetic torque on the nanowires that is transmitted to a cell's adhesion site as an external force. We have achieved external forces of up to 45 nN, which is in the upper range of current nanoscale force-probing techniques. Nonmagnetic microposts, similarly prepared but without nanowires, surround the magnetic microposts and are used to measure the traction forces and changes in cell mechanics. We record the magnitude and direction of the external force and the traction forces by optically measuring the deflection of the microposts, which linearly deflect as cantilever springs. With this approach, we can measure traction forces before and after force stimulation in order to monitor cellular response to forces. We present the fabrication methods, magnetic force characterization, and image analysis techniques used to achieve the measurements.