37 research outputs found
Rigidity Matching between Cells and the Extracellular Matrix Leads to the Stabilization of Cardiac Conduction
Biomechanical dynamic interactions between cells and the extracellular environment dynamically regulate physiological tissue behavior in living organisms, such as that seen in tissue maintenance and remodeling. In this study, the substrate-induced modulation of synchronized beating in cultured cardiomyocyte tissue was systematically characterized on elasticity-tunable substrates to elucidate the effect of biomechanical coupling. We found that myocardial conduction is significantly promoted when the rigidity of the cell culture environment matches that of the cardiac cells (4 kiloPascals). The stability of spontaneous target wave activity and calcium transient alternans in high frequency-paced tissue were both enhanced when the cell substrate and cell tissue showed the same rigidity. By adapting a simple theoretical model, we reproduced the experimental trend on the rigidity matching for the synchronized excitation. We conclude that rigidity matching in cell-to-substrate interactions critically improves cardiomyocyte-tissue synchronization, suggesting that mechanical coupling plays an essential role in the dynamic activity of the beating heart
Protein-protein interactions of the hyperthermophilic archaeon Pyrococcus horikoshii OT3
BACKGROUND: Although 2,061 proteins of Pyrococcus horikoshii OT3, a hyperthermophilic archaeon, have been predicted from the recently completed genome sequence, the majority of proteins show no similarity to those from other organisms and are thus hypothetical proteins of unknown function. Because most proteins operate as parts of complexes to regulate biological processes, we systematically analyzed protein-protein interactions in Pyrococcus using the mammalian two-hybrid system to determine the function of the hypothetical proteins. RESULTS: We examined 960 soluble proteins from Pyrococcus and selected 107 interactions based on luciferase reporter activity, which was then evaluated using a computational approach to assess the reliability of the interactions. We also analyzed the expression of the assay samples by western blot, and a few interactions by in vitro pull-down assays. We identified 11 hetero-interactions that we considered to be located at the same operon, as observed in Helicobacter pylori. We annotated and classified proteins in the selected interactions according to their orthologous proteins. Many enzyme proteins showed self-interactions, similar to those seen in other organisms. CONCLUSION: We found 13 unannotated proteins that interacted with annotated proteins; this information is useful for predicting the functions of the hypothetical Pyrococcus proteins from the annotations of their interacting partners. Among the heterogeneous interactions, proteins were more likely to interact with proteins within the same ortholog class than with proteins of different classes. The analysis described here can provide global insights into the biological features of the protein-protein interactions in P. horikoshii
Manipulation of Cell Movement by Designing Microelasticity Gradient Field of Cell Culture Substrate
Relationship between apical membrane elasticity and stress fiber organization in fibroblasts analyzed by fluorescence and atomuc force microscopy
To investigate the relationship between cellular microelasticity and the structural features of cytoskeletons (CSKs), a microindentation test for apical cell membranes and observation of the spatio-distribution of actin CSKs of fibroblasts were performed by fluorescence and atomic force microscopy (FM/AFM). The indentation depths of apical cell membranes were measured from AFM force–indentation (f–i) curves under equal final loads and mapped two-dimensionally to show the relative distribution of local microelasticity on cell membranes. Intracellular spatial distribution of actin CSKs was visualized fluorescently by high Z-resolution cross-sectional observation of a cell on which indentation mapping analysis had been performed in advance. Structural features of stress fibers (SFs) were observed as three typical patterns of dense SF, sparse SF and sparser SF cell groups, which were quantitated using the degree of orientation in apical SFs (ASFs) that had been defined using two-dimensional Fourier analysis. In indentation depth maps, the upper nuclear region was markedly softer than the pseudopodium region. The mean indentation depth of the upper nuclear region decreased with increased SF density in whole cells and the degree of orientation of ASF, although the pseudopodium region did not exhibit such a trend. The apical membrane of adhered cells was found to tend to stiffen with the increase in both density and degree of orientation of SFs
Adhesion Forces of the Blood Plasma Proteins on Self-Assembled Monolayer Surfaces of Alkanethiolates with Different Functional Groups Measured by an Atomic Force Microscope
Distribution of Young’s moduli in gels with different unit sizes and peak elasticities.
<p>The width of each unit in series A and B was 100 µm and 120 µm, respectively. Series 1, 2, and 3 had a peak elasticity of ca. 100 kPa, 300 kPa, and 500 kPa, respectively. The ratio of ascending:descending elasticity gradients in each unit was 1:2, i.e., the X-position of the peak of elasticity was located at ca. 30 µm and 40 µm in series A and B, respectively. The conditions used to prepare each of the gels are described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0078067#pone-0078067-t001" target="_blank">Table 1</a>.</p
Rectified Cell Migration on Saw-Like Micro-Elastically Patterned Hydrogels with Asymmetric Gradient Ratchet Teeth
<div><p>To control cell motility is one of the essential technologies for biomedical engineering. To establish a methodology of the surface design of elastic substrate to control the long-range cell movements, here we report a sophisticated cell culture hydrogel with a micro-elastically patterned surface that allows long-range durotaxis. This hydrogel has a saw-like pattern with asymmetric gradient ratchet teeth, and rectifies random cell movements. Durotaxis only occurs at boundaries in which the gradient strength of elasticity is above a threshold level. Consequently, in gels with unit teeth patterns, durotaxis should only occur at the sides of the teeth in which the gradient strength of elasticity is above this threshold level. Therefore, such gels are expected to support the long-range biased movement of cells via a mechanism similar to the Feynman-Smoluchowski ratchet, i.e., rectified cell migration. The present study verifies this working hypothesis by using photolithographic microelasticity patterning of photocurable gelatin gels. Gels in which each teeth unit was 100–120 µm wide with a ratio of ascending:descending elasticity gradient of 1:2 and a peak elasticity of ca. 100 kPa supported the efficient rectified migration of 3T3 fibroblast cells. In addition, long-range cell migration was most efficient when soft lanes were introduced perpendicular to the saw-like patterns. This study demonstrates that asymmetric elasticity gradient patterning of cell culture gels is a versatile means of manipulating cell motility.</p> </div
Long-range cell migration on gels with saw-like asymmetric elasticity patterns and perpendicular soft lanes (PSLs).
<p>The conditions used to prepare the gels are described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0078067#pone-0078067-t001" target="_blank">Table 1</a>. (a) Phase contrast microscopy image of the gel. Sale bar: 100 µm. (b) Distribution of Young’s modulus in a single unit of the gel. Unit size: 90 µm. Elasticity peak: ca. 500 kPa. (c) Trajectories of 3T3 fibroblasts cultured on gels with PSLs. Cells were observed by time-lapse microscopy for 24 hr and images were acquired at 15 min intervals. (d) Time-course of ensemble averaged trajectories with standard deviation in the presence (red) and absence (blue) of PSLs. n=30 cells.</p
Schematic diagram showing how cell adhesive hydrogels with saw-like asymmetric elasticity patterns are fabricated.
<p>In the first step, a soft base gel is prepared by irradiation of a photocurable sol of styrenated gelatin with visible light. In the second step, the base gel is photo-irradiated through slits in a photomask for a defined period of time. The gel is continuously displaced in a perpendicular direction to the photomask at a defined speed using a computer-assisted X-Y movement stage. The duration of irradiation is set so that the StG sol surface is irradiated asymmetrically.</p