マルチ スケール キノウ ヲ ユウスル コウソク ジドウ マイクロ マニピュレーション システム

Ebubekir Avci, Chanh-Nghiem Nguyen, Kenichi Ohara, Yasushi Mae, Tatsuo Arai, Analysis and suppression of residual vibration in microhand for high-speed single-cell manipulation, International Journal of Mechatronics and Automation, 2013-Vol.3, No.2, pp.110-11

Collective motion of cells: from experiments to models

Swarming or collective motion of living entities is one of the most common and spectacular manifestations of living systems having been extensively studied in recent years. A number of general principles have been established. The interactions at the level of cells are quite different from those among individual animals therefore the study of collective motion of cells is likely to reveal some specific important features which are overviewed in this paper. In addition to presenting the most appealing results from the quickly growing related literature we also deliver a critical discussion of the emerging picture and summarize our present understanding of collective motion at the cellular level. Collective motion of cells plays an essential role in a number of experimental and real-life situations. In most cases the coordinated motion is a helpful aspect of the given phenomenon and results in making a related process more efficient (e.g., embryogenesis or wound healing), while in the case of tumor cell invasion it appears to speed up the progression of the disease. In these mechanisms cells both have to be motile and adhere to one another, the adherence feature being the most specific to this sort of collective behavior. One of the central aims of this review is both presenting the related experimental observations and treating them in the light of a few basic computational models so as to make an interpretation of the phenomena at a quantitative level as well.Comment: 24 pages, 25 figures, 13 reference video link

Single-molecule experiments in biological physics: methods and applications

I review single-molecule experiments (SME) in biological physics. Recent technological developments have provided the tools to design and build scientific instruments of high enough sensitivity and precision to manipulate and visualize individual molecules and measure microscopic forces. Using SME it is possible to: manipulate molecules one at a time and measure distributions describing molecular properties; characterize the kinetics of biomolecular reactions and; detect molecular intermediates. SME provide the additional information about thermodynamics and kinetics of biomolecular processes. This complements information obtained in traditional bulk assays. In SME it is also possible to measure small energies and detect large Brownian deviations in biomolecular reactions, thereby offering new methods and systems to scrutinize the basic foundations of statistical mechanics. This review is written at a very introductory level emphasizing the importance of SME to scientists interested in knowing the common playground of ideas and the interdisciplinary topics accessible by these techniques. The review discusses SME from an experimental perspective, first exposing the most common experimental methodologies and later presenting various molecular systems where such techniques have been applied. I briefly discuss experimental techniques such as atomic-force microscopy (AFM), laser optical tweezers (LOT), magnetic tweezers (MT), biomembrane force probe (BFP) and single-molecule fluorescence (SMF). I then present several applications of SME to the study of nucleic acids (DNA, RNA and DNA condensation), proteins (protein-protein interactions, protein folding and molecular motors). Finally, I discuss applications of SME to the study of the nonequilibrium thermodynamics of small systems and the experimental verification of fluctuation theorems. I conclude with a discussion of open questions and future perspectives.Comment: Latex, 60 pages, 12 figures, Topical Review for J. Phys. C (Cond. Matt

Workshop on "Robotic assembly of 3D MEMS".

Proceedings of a workshop proposed in IEEE IROS'2007.The increase of MEMS' functionalities often requires the integration of various technologies used for mechanical, optical and electronic subsystems in order to achieve a unique system. These different technologies have usually process incompatibilities and the whole microsystem can not be obtained monolithically and then requires microassembly steps. Microassembly of MEMS based on micrometric components is one of the most promising approaches to achieve high-performance MEMS. Moreover, microassembly also permits to develop suitable MEMS packaging as well as 3D components although microfabrication technologies are usually able to create 2D and "2.5D" components. The study of microassembly methods is consequently a high stake for MEMS technologies growth. Two approaches are currently developped for microassembly: self-assembly and robotic microassembly. In the first one, the assembly is highly parallel but the efficiency and the flexibility still stay low. The robotic approach has the potential to reach precise and reliable assembly with high flexibility. The proposed workshop focuses on this second approach and will take a bearing of the corresponding microrobotic issues. Beyond the microfabrication technologies, performing MEMS microassembly requires, micromanipulation strategies, microworld dynamics and attachment technologies. The design and the fabrication of the microrobot end-effectors as well as the assembled micro-parts require the use of microfabrication technologies. Moreover new micromanipulation strategies are necessary to handle and position micro-parts with sufficiently high accuracy during assembly. The dynamic behaviour of micrometric objects has also to be studied and controlled. Finally, after positioning the micro-part, attachment technologies are necessary

Nano handling and measurement of biological cells in culture

A thesis submitted to the University of Bedfordshire in partial fulfillment of the requirements for the degree of Doctor of PhilosophyThis thesis systematically investigates the nano handling and measurement techniques for biological cells in culture and studies the techniques to realize innovative and multi-functional applications in biomedicine. Among them, the technique based on AFM is able to visualize and quantify the dynamics of organic cells in culture on the nano scale. Especially, the cellular shear adhesion force on the various locations of biological cells was firstly accurately measured in the research of the cell-substrate interaction in terms of biophysical perspective. The innovative findings are conductive to study the cell-cell adhesion, the cell-matrix adhesion which is related to the cell morphology structure, function, deformation ability and adhesion of cells and better understand the cellular dynamic behaviors. Herein, a new liquid-AFM probe unit and an increment PID control algorithm were implemented suitable for scanning the cell samples under the air conditions and the liquid environments. The influence between the surface of sample and the probe, and the damage of probe during the sample scanning were reduced. The proposed system is useful for the nano handling and measurement of living cells. Besides, Besides, to overcome the limitations of liquid-AFMs, the multiple optical tweezers were developed to integrate with the liquid-AFM. The technique based on laser interference is able to characterize the optical trap stiffness and the escape velocity, especially to realize the capture and sorting of multiple cells by a polarization-controlled periodic laser interference. It can trap and move hundreds of cells without physical contact, and has broad application prospects in cytology. Herein, a new experimental method integrated with the positioning analysis in the Z direction was used to improve the fluid force method for the calibration and characterize the mechanical forces exerted on optical traps and living cells. Moreover, a sensitive and highly efficient polarization-controlled three-beam interference set-up was developed for the capture and sorting of multiple cells. By controlling the polarization angles of the beams, various intensity distributions and different sizes of dots were obtained. Subsequently, we have experimentally observed multiple optical tweezers and the sorting of cells with different polarization angles, which are in accordance with the theoretical analysis

Biomaterial-Mediated Reprogramming of the Wound Interface to Enhance Meniscal Repair

Endogenous repair of fibrous connective tissues is limited, and there exist few successful strategies to improve healing after injury. As such, new methods that advance repair by enhancing cell migration to the wound interface, extracellular matrix (ECM) production, and tissue integration would represent a marked clinical advance. Using the adult meniscus as a test platform, we hypothesized that ECM density and stiffness increase throughout tissue maturation, and that these age-related changes present biophysical barriers to interstitial cell migration during wound healing. We further posited that modulating the matrix could remove these impediments, enabling endogenous cells to reach the injury site. To test our hypotheses, we compared the microenvironment of fetal and adult meniscal ECM via atomic force microscopy (AFM) indentation and second harmonic generation (SHG) imaging of the collagenous matrix. We also explored interstitial cell mobility through fetal and adult native tissue environments using a three-dimensional ex vivo system. We further investigated strategies that might expedite cell migration, including enzymatic degradation of the ECM with collagenase to reduce matrix stiffness and increase porosity. To restrict these biological manipulations to the wound interface, we fabricated a delivery system in which selected biofactors were stored inside composite electrospun nanofibrous scaffolds and released upon hydration. The ability for bioactive scaffolds to enhance the cellularity and integration of meniscal injuries was evaluated in vivo using tissue explants in a subcutaneous implantation model, as well as an orthotopic meniscal injury model. Our findings suggest that matrix stiffness, density, and organization increase with meniscal development at the expense of cell mobility. Our results also indicate that partial digestion of the wound interface with collagenase improves repair by creating a more compliant and porous microenvironment that facilitates cell migration. Furthermore, when scaffolds containing collagenase-releasing fibers were placed inside meniscal defects, enzymatic digestion was localized and resulted in improved cellular colonization and closure of the wound site, similar to treatment with aqueous collagenase. This innovative approach of targeted delivery may aid the many patients that exhibit meniscal tears by promoting integrative repair, thereby circumventing the pathologic consequences of partial meniscus removal, and may find widespread application in the treatment of injuries to a variety of dense connective tissues

Mechanics of Fibroblast Migration: a Dissertation

Cell migration involves complex mechanical interactions between cells or between cells and the underlying substrate. Using a newly developed technique, traction force microscopy , I have been able to visualize the dynamic characteristics of mechanical forces exerted by migrating fibroblasts such as magnitude, direction, and shear. For NIH 3T3 fibroblasts, I found that the lamellipodium provides nearly all of the force necessary for cell migration. A high shear zone separates the lamellipodium from the remainder of the cell body, suggesting that they are mechanically distinct entities. The timing of the tractions at the leading edge, as well as the spatial distribution, bears no apparent relationship to concurrent local protrusive activities, yet changes in traction force patterns often precede changes in migration direction. In H-ras transformed cells I found isolated regions of weak, transient traction forces in pseudopods all along the cell that appeared to act against one another. The resulting shear pattern suggested that there were multiple disorganized mechanical domains. These results support a frontal towing model for cell migration where the dynamic traction forces at the leading edge served to actively pull the cell body forward. In H-ras transformed cells, the weak poorly coordinated traction forces coupled with weak cell substrate-adhesions were likely responsible for the abnormal motile behavior of these cells. To probe the mechanical interactions beneath various regions of migrating fibroblasts, a cell substrate inhibitor (GRGDTP peptide) was locally applied while imaging stress distribution on the substrate utilizing traction force microscopy. I found that both spontaneous and GRGDTP induced detachment of the trailing edge resulted in extensive cell shortening with no change in overall traction force magnitude or cell migration. Conversely, leading edge disruption resulted in a dramatic global loss of traction forces pnor to any significant cell shortening. These results suggested that fibroblasts transmit their contractile forces to the substrate through two distinct types of adhesions. Leading edge adhesions were unique in their ability to transmit active propulsive forces whereas trailing end adhesions created passive resistance during cell migration and readily redistributed their loads upon detachment. I have also investigated how fibroblasts regulate traction forces based on mechanical input. My results showed that stretching forces applied through the flexible substrate induced increases in both intracellular calcium concentration and traction forces in fibroblasts. Treatment with gadolinium, a well known stretch-activated ion channel inhibitor, was found to inhibit both traction forces and cell migration without inhibiting cellular spread morphology or protrusive activities. Gadolinium treatment also caused a pronounced decrease in vinculin and phosphotyrosine concentrations from focal adhesions. Local application of gadolinium to the trailing region had no detectable effect on overall traction forces or cell migration, whereas local application to the leading edge caused a global inhibition of traction forces and an inhibition of migration. These observations suggest that stretch activated entry of calcium ions in the frontal region serves to regulate the organization of focal adhesions and the output of mechanical forces. Together my experiments elucidate how fibroblasts exert mechanical forces to propel their movements, and how fibroblasts utilize mechanical input to regulate their movements

Possible origins of macroscopic left-right asymmetry in organisms

I consider the microscopic mechanisms by which a particular left-right (L/R) asymmetry is generated at the organism level from the microscopic handedness of cytoskeletal molecules. In light of a fundamental symmetry principle, the typical pattern-formation mechanisms of diffusion plus regulation cannot implement the "right-hand rule"; at the microscopic level, the cell's cytoskeleton of chiral filaments seems always to be involved, usually in collective states driven by polymerization forces or molecular motors. It seems particularly easy for handedness to emerge in a shear or rotation in the background of an effectively two-dimensional system, such as the cell membrane or a layer of cells, as this requires no pre-existing axis apart from the layer normal. I detail a scenario involving actin/myosin layers in snails and in C. elegans, and also one about the microtubule layer in plant cells. I also survey the other examples that I am aware of, such as the emergence of handedness such as the emergence of handedness in neurons, in eukaryote cell motility, and in non-flagellated bacteria.Comment: 42 pages, 6 figures, resubmitted to J. Stat. Phys. special issue. Major rewrite, rearranged sections/subsections, new Fig 3 + 6, new physics in Sec 2.4 and 3.4.1, added Sec 5 and subsections of Sec

Amoeboid Shape Dynamics on Flat and Topographically Modified Surfaces

I present an analysis of the shape dynamics of the amoeba Dictyostelium discoideum, a model system for the study of cellular migration. To better understand cellular migration in complicated 3-D environments, cell migration was studied on simple 3-D surfaces, such as cliffs and ridges. D. discoideum interact with surfaces without forming mature focal adhesion complexes. The cellular response to the surface topography was characterized by measuring and looking for patterns in cell shape. Dynamic cell shape is a measure of the interaction between the internal biochemical state of a cell and its external environment. For D. discoideum migrating on flat surfaces, waves of high boundary curvature were observed to travel from the cell front to the cell back. Curvature waves are also easily seen in cells that do not adhere to a surface, such as cells that are electrostatically repelled from the coverslip or cells that are extended over the edge of micro-fabricated cliffs. At the leading edge of adhered cells, these curvature waves are associated with protrusive activity, suggesting that protrusive motion can be thought of as a wave-like process. The wave-like character of protrusions provides a plausible mechanism for the ability of cells to swim in viscous fluids and to navigate complex 3-D topography. Patterning of focal adhesion complexes has previously been implicated in contact guidance (polarization or migration parallel to linear topographical structures). However, significant contact guidance is observed in D. discoideum, which lack focal adhesion complexes. Analyzing the migration of cells on nanogratings of ridges spaced various distances apart, ridges spaced about 1.5 micrometers apart were found to guide cells best. Contact guidance was modeled as an interaction between wave-like processes internal to the cell and the periodicity of the nanograting. The observed wavelength and speed of the oscillations that best couple to the surface are consistent with those of protrusive dynamics. Dynamic sensing via actin or protrusive dynamics might then play a role in contact guidance

Fabrication of biomedical devices through electro-fluid-dynamic-based techniques

Fabrication of biomedical devices through electro-fluid-dynamic-based technique