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Rotation Tracking and Adhesion Footprinting Reveal Asymmetric Rolling Adhesion Mechanism
Monday, February 29, 2016 Thus, ring tension has little effect on the constriction rate, which is set by the intrinsic rate of synthesis by the cell’s septum synthesis machinery. 1506-Pos Board B483 Multi-scale Computational Model of Epithelial Cell Proliferation and Mechanics Ali Nematbakhsh1, Pavel Brodskiy2, Zhiliang Xu1, Jeremiah J. Zartman2, Mark Alber1. 1 Department of Applied and Computational Mathematics and Statistics, University of Notre Dame, Notre Dame, IN, USA, 2Department of Chemical and Biomolecular Engineering, University of Notre Dame, Notre Dame, IN, USA. Epithelia are sheets of tightly adherent, asymmetrically organized cells that provide a protective barrier for organs. The close association of epithelial cells through physical interactions and active adhesion leads to tightly packed cellular networks. Several computational models have been developed to simulate the dynamics of cell growth, division and neighboring cell-cell interactions and shape changes, which drive tissue morphogenesis. However, cells are typically approximated as polygons to simplify computation effort, resulting in poor descriptions of many morphogenetic processes including mitotic rounding. We will describe newly developed cell-based subcellular element (SCE) computational model implemented on high performance Graphical Processing Units (GPUs) cluster. The model represents mechanical interaction of both the internal cytoplasm and outer membrane with subcellular nodes based on potentials energy functions. Model simulations are shown to match cell and tissue properties of the developing Drosophila wing disc including the overall distributions of cell neighbor numbers. The model does not require ad hoc rules to simulate cell neighbor changes and predicts that 4-edge vertices and T1 transitions can result from mitotic cell rounding. High speed computational framework is flexible and can be extended to study the apical dynamics of wound healing and epithelial-mesenchymal transitions, where epithelial cells strongly deviate from polygonal network geometries. 1507-Pos Board B484 Intact Immunotaxis Comprises an Intricate Spatiotemporal Hierarchy of Distinct Chemotactic Processes - A New Paradigm Volkmar Heinrich. Biomedical Engineering, University of California Davis, Davis, CA, USA. A close look at cytokine-directed chemotaxis reveals an intriguing mechanistic dilemma. If cytokine gradients provided the only directional cue for chemotaxing immune cells, these cells would ultimately target the cytokine-producing cells of their own host. Such friendly fire may well be an inherent part of an indiscriminate mop-up duty of immune cells in the inflammatory response. However, it would preclude these cells from partaking in any pathogenspecific defenses requiring pre-contact recognition of invaders. Evidently, effective immune-cell recruitment requires additional directional guidance via chemical signals that emanate from the surfaces of microbes rather than host cells. To detect minuscule amounts of chemoattractant produced at the surfaces of bacteria, fungi, and surrogate particles, we have developed a single-cell, pure-chemotaxis assay that employs human neutrophils as ultrasensitive biodetectors. We use micropipette manipulation and/or optical tweezers to maneuver target particles into the proximity of non-adherent, initially quiescent neutrophils. Microscopic inspection of the resulting neutrophil morphology provides a clear readout of the chemotactic activity of the cells. In all cases where neutrophils extended chemotactic protrusions toward nearby targets, this cell response required serum. For bacterial targets such as Salmonella Typhimurium and Escherichia coli, and for zymosan particles as well as fungal particles of Coccidioides posadasii and Candida albicans, this type of microbe-guided chemotaxis was predominantly mediated by complement, in particular the anaphylatoxin C5a. The pre-contact neutrophil response to these targets was generally vigorous but short range, consistent with our analytical solution of a theoretical diffusion-reaction problem that takes into account the deactivation of anaphylatoxins by carboxypeptidases. Together, these findings establish complement-mediated chemotaxis as a universal homing mechanism by which chemotaxing immune cells implement a last-minute course correction toward bacterial, fungal, and model pathogens. 1508-Pos Board B485 Modeling the Effects of Focal Adhesion Size Restriction on Cell Shape during Spreading Magdalena Stolarska1, Kara Huyett1, Aravind Rammohan2. 1 Mathematics, University of St. Thomas, St. Paul, MN, USA, 2Mathematics, Corning, Inc., Corning, NY, USA.
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Focal adhesions (FAs) are known to exhibit mechanosensitive properties. Changes in FA size, distribution, and dynamics is leveraged in controlling various signaling networks that enable a cell to demonstrate mechanochemical sensitivity to its environment. Here, we present a two-dimensional model and finite element simulations of a cell interacting with a deformable substrate through FA complexes. The cell is treated as a hypoelastic material that is allowed to undergo active deformation representative of spreading and localized actomyosin contractions, and the substrate is modeled as linearly elastic. FA complexes are modeled by collections of linear springs that can form and break dynamically. Our aim is to understand how controlling the size of FAs, either via disassembly by microtubules or by ligand patterning, affects cellular responses. We model microtubule induced FA disassembly by systematically removing FA springs from regions closest to the cell nucleus. Alternately we also examine control of FA size by ligand patterning using reaction-diffusion equations describing the interchange between bound and unbound integrins, which accounts for the conservation of the total number of integrins within the cell. We verify our model of the coupling between integrin activation and actomyosin contractions by demonstrating that stress fibers form between adhesive patches as reported in the experimental work of The´ry et al [1]. In addition, we compare the effects of the proposed two models for controlling FA evolution on intracellular stresses, substrate displacement patterns, FA distribution, and cell shape for different substrate stiffnesses and ligand patterns. [1] M. The´ry, A. Pe´pin, E. Dressaire,Y. Chen, and M. Bornens (2006). Cell Distribution of Stress Fibers in Response to the Geometry of the Adhesive Geometry, Cell Motility and the Cytoskeleton, 63, pp. 341-355. 1509-Pos Board B486 Rotation Tracking and Adhesion Footprinting Reveal Asymmetric Rolling Adhesion Mechanism Isaac T.S. Li1,2, Taekjip Ha3,4, Yann R. Chemla1. 1 Department of Physics and Centre for Physics of Living Cells, University of Illinois Urbana-Champaign, Urbana, IL, USA, 2Department of Chemistry, University of British Columbia, Kelowna, BC, Canada, 3Department of Biophysics and Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD, USA, 4Howard Hughes Medical Institute, Baltimore, MD, USA. Rolling adhesion is the behaviour that leukocytes and circulating tumour cells exhibit as they passively roll along blood vessel walls under flow. It plays a critical role in capturing cells in the blood, guiding them toward inflammation sites, and activating cell signalling pathways to enable their subsequent transmigration. Rolling adhesion is mediated by catch-bond-like interactions between selectins expressed on endothelial cells lining blood vessels and P-selectin glycoprotein ligand-1 (PSGL-1) found at microvilli tips of leukocytes. Despite our understanding of individual components of this process, how the molecular details of adhesion bonds scale to cell-surface adhesion and rolling behaviour remains poorly understood. Here, we developed a method that maps the functional adhesion sites and their strength on a leukocyte surface. The method relies on tracking the rotational angle of a single rolling cell, which confers advantages over standard methods that track the centreof-mass alone. Constructing the adhesion map from the instantaneous angular velocity reveals that the adhesion profile along the rolling circumference is inhomogeneous. We corroborated these findings by obtaining a footprint of molecular adhesion events using DNA-based molecular force probes. Our results reveal that adhesion at the functional level is not uniformly distributed over the leukocyte surface as previously assumed, but is instead patchy. 1510-Pos Board B487 Cell-Substrate Interaction Determines Cellular Volume and Shape Jiaxiang Tao, Sean Sun. Johns Hopkins University, Baltimore, MD, USA. Multiple experimental results have shown eukaryotic cells are able to respond to its mechanical environment. Such responds are not only crucial during cell migration, polarization and tissue formation, but also determining cellular volume and shape. In this study, we show a simple mechanical force balance, coupled with previously-purposed chemical model on Rho GTPase activation, is able to predict the cellular shape when cells spreading on substrates with different sizes and/or stiffness, indicating the importance of mechanical forces that regulates different cell activities, including myosin activities, cellular volume, as well as traction stresses between cell and substrate. Moreover, if cell is placed in the growth medium, such mechanical signal may trigger cell division. With previously developed FRET pair, we are also able to observe RhoA activity during cell spreading experimentally, which is a crucial prove to our purposed model.
307a
Focal adhesions (FAs) are known to exhibit mechanosensitive properties. Changes in FA size, distribution, and dynamics is leveraged in controlling various signaling networks that enable a cell to demonstrate mechanochemical sensitivity to its environment. Here, we present a two-dimensional model and finite element simulations of a cell interacting with a deformable substrate through FA complexes. The cell is treated as a hypoelastic material that is allowed to undergo active deformation representative of spreading and localized actomyosin contractions, and the substrate is modeled as linearly elastic. FA complexes are modeled by collections of linear springs that can form and break dynamically. Our aim is to understand how controlling the size of FAs, either via disassembly by microtubules or by ligand patterning, affects cellular responses. We model microtubule induced FA disassembly by systematically removing FA springs from regions closest to the cell nucleus. Alternately we also examine control of FA size by ligand patterning using reaction-diffusion equations describing the interchange between bound and unbound integrins, which accounts for the conservation of the total number of integrins within the cell. We verify our model of the coupling between integrin activation and actomyosin contractions by demonstrating that stress fibers form between adhesive patches as reported in the experimental work of The´ry et al [1]. In addition, we compare the effects of the proposed two models for controlling FA evolution on intracellular stresses, substrate displacement patterns, FA distribution, and cell shape for different substrate stiffnesses and ligand patterns. [1] M. The´ry, A. Pe´pin, E. Dressaire,Y. Chen, and M. Bornens (2006). Cell Distribution of Stress Fibers in Response to the Geometry of the Adhesive Geometry, Cell Motility and the Cytoskeleton, 63, pp. 341-355. 1509-Pos Board B486 Rotation Tracking and Adhesion Footprinting Reveal Asymmetric Rolling Adhesion Mechanism Isaac T.S. Li1,2, Taekjip Ha3,4, Yann R. Chemla1. 1 Department of Physics and Centre for Physics of Living Cells, University of Illinois Urbana-Champaign, Urbana, IL, USA, 2Department of Chemistry, University of British Columbia, Kelowna, BC, Canada, 3Department of Biophysics and Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD, USA, 4Howard Hughes Medical Institute, Baltimore, MD, USA. Rolling adhesion is the behaviour that leukocytes and circulating tumour cells exhibit as they passively roll along blood vessel walls under flow. It plays a critical role in capturing cells in the blood, guiding them toward inflammation sites, and activating cell signalling pathways to enable their subsequent transmigration. Rolling adhesion is mediated by catch-bond-like interactions between selectins expressed on endothelial cells lining blood vessels and P-selectin glycoprotein ligand-1 (PSGL-1) found at microvilli tips of leukocytes. Despite our understanding of individual components of this process, how the molecular details of adhesion bonds scale to cell-surface adhesion and rolling behaviour remains poorly understood. Here, we developed a method that maps the functional adhesion sites and their strength on a leukocyte surface. The method relies on tracking the rotational angle of a single rolling cell, which confers advantages over standard methods that track the centreof-mass alone. Constructing the adhesion map from the instantaneous angular velocity reveals that the adhesion profile along the rolling circumference is inhomogeneous. We corroborated these findings by obtaining a footprint of molecular adhesion events using DNA-based molecular force probes. Our results reveal that adhesion at the functional level is not uniformly distributed over the leukocyte surface as previously assumed, but is instead patchy. 1510-Pos Board B487 Cell-Substrate Interaction Determines Cellular Volume and Shape Jiaxiang Tao, Sean Sun. Johns Hopkins University, Baltimore, MD, USA. Multiple experimental results have shown eukaryotic cells are able to respond to its mechanical environment. Such responds are not only crucial during cell migration, polarization and tissue formation, but also determining cellular volume and shape. In this study, we show a simple mechanical force balance, coupled with previously-purposed chemical model on Rho GTPase activation, is able to predict the cellular shape when cells spreading on substrates with different sizes and/or stiffness, indicating the importance of mechanical forces that regulates different cell activities, including myosin activities, cellular volume, as well as traction stresses between cell and substrate. Moreover, if cell is placed in the growth medium, such mechanical signal may trigger cell division. With previously developed FRET pair, we are also able to observe RhoA activity during cell spreading experimentally, which is a crucial prove to our purposed model.