scholarly journals Tendon Development Requires Regulation of Cell Condensation and Cell Shape via Cadherin-11-Mediated Cell-Cell Junctions

2007 ◽  
Vol 27 (17) ◽  
pp. 6218-6228 ◽  
Author(s):  
Susan H. Richardson ◽  
Tobias Starborg ◽  
Yinhui Lu ◽  
Sally M. Humphries ◽  
Roger S. Meadows ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Plasma Membrane ◽  
Cell Shape ◽  
Plasma Membranes ◽  
Three Dimensional ◽  
Collagen Fibrils ◽  
Cell Junctions ◽  
Membrane Channels ◽  
Dimensional Reconstruction ◽  
Cell Cell

ABSTRACT The ability of tendon to transmit forces from muscle to bone is directly attributable to an extracellular matrix (ECM) containing parallel bundles of collagen fibrils. Although the biosynthesis of collagen is well characterized, how cells deposit the fibrils in regular parallel arrays is not understood. Here we show that cells in the tendon mesenchyme are nearly cylindrical and are aligned side by side and end to end along the proximal-distal axis of the limb. Using three-dimensional reconstruction electron microscopy, we show that the cells have deep channels in their plasma membranes and contain bundles of parallel fibrils that are contiguous from one cell to another along the tendon axis. A combination of electron microscopy, microarray analysis, and immunofluorescence suggested that the cells are held together by cadherin-11-containing cell-cell junctions. Using a combination of RNA interference and electron microscopy, we showed that knockdown of cadherin-11 resulted in cell separation, loss of plasma membrane channels, and misalignment of the collagen fibrils in the ECM. Our results show that tendon formation in the developing limb requires precise regulation of cell shape via cadherin-11-mediated cell-cell junctions and coaxial alignment of plasma membrane channels in longitudinally stacked cells.

Conformational characterization of nucleosome structure by Electron Microscopy

1993 ◽  
Vol 51 ◽  
pp. 194-195
Author(s):  
Gregory J. Czarnota
Keyword(s):  
Electron Microscopy ◽  
Structural Changes ◽  
Physiological State ◽  
Gene Activation ◽  
Three Dimensional ◽  
Macromolecular Complex ◽  
Ultrastructural Studies ◽  
Ionic Environment ◽  
Nucleosome Structure ◽  
Dimensional Reconstruction

Chromatin structure at the fundamental level of the nucleosome is important in vital cellular processes. Recent biochemical and genetic analyses show that nucleosome structure and structural changes are very active participants in gene expression, facilitating or inhibiting transcription and reflecting the physiological state of the cell. Structural states and transitions for this macromolecular complex, composed of DNA wound about a heterotypic octamer of variously modified histone proteins, have been measured by physico-chemical techniques and by enzyme-accessibility and are recognized to occur with various post-translational modifications, gene activation, transformation and with ionic-environment. In spite of studies which indicate various forms of nucleosome structure, all current x-ray and neutron diffraction studies have consistently resulted in only one structure, suggestive of a static conformation. In contrast, two-dimensional electron microscopy studies and three-dimensional reconstruction techniques have yielded different structures. These fundamental differences between EM and other ultrastructural studies have created a long standing quandary, which I have addressed and resolved using spectroscopic electron microscopy and statistical analyses of nucleosome images in a study of nucleosome structure with ionic environment.

Three-Dimensional Reconstruction of Native Androctonus Australis Hemocyanin

1990 ◽  
Vol 48 (1) ◽  
pp. 452-453
Author(s):  
Nicolas Boisset ◽  
Jean-Christophe Taveau ◽  
Jean Lamy ◽  
Terence Wagenknecht ◽  
Michael Radermacher ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Solid Body ◽  
Body Surface ◽  
Three Dimensional ◽  
Staining Technique ◽  
Three Dimensional Reconstruction ◽  
Dimensional Reconstruction ◽  
View Reconstruction ◽  
Surface Representations ◽  
Top View

Hemocyanin, the respiratory pigment of the scorpion Androctonus australis is composed of 24 kidney shaped subunits. A model of architecture supported by many indirect arguments has been deduced from electron microscopy (EM) and immuno-EM. To ascertain, the disposition of the subunits within the oligomer, the 24mer was submitted to three-dimensional reconstruction by the method of single-exposure random-conical tilt series.A sample of native hemocyanin, prepared with the double layer negative staining technique, was observed by transmisson electron microscopy under low-dose conditions. Six 3D-reconstructions were carried out indenpendently from top, side and 45°views. The results are composed of solid-body surface representations, and slices extracted from the reconstruction volume.The main two characters of the molecule previously reported by Van Heel and Frank, were constantly found in the solid-body surface representations. These features are the presence of two different faces called flip and flop and a rocking of the molecule around an axis passing through diagonnally opposed hexamers. Furthermore, in the solid-body surface of the top view reconstruction, the positions and orientations of the bridges connecting the half molecules were found in excellent agreement with those predicted by the model.

scholarly journals Spatial and temporal relationships between cadherins and PECAM-1 in cell-cell junctions of human endothelial cells.

1994 ◽  
Vol 126 (1) ◽  
pp. 247-258 ◽  
Author(s):  
O Ayalon ◽  
H Sabanai ◽  
M G Lampugnani ◽  
E Dejana ◽  
B Geiger
Keyword(s):  
Endothelial Cells ◽  
Plasma Membrane ◽  
Vascular System ◽  
Adherens Junctions ◽  
Microscopic Analysis ◽  
Membrane Receptors ◽  
Temporal Organization ◽  
Cell Junctions ◽  
Short Treatment ◽  
Cell Cell

The integrity of the endothelial layer, which lines the entire cavity of the vascular system, depends on tight adhesion of the cells to the underlying basement membrane as well as to each other. It has been previously shown that such interactions occur via membrane receptors that determine the specificity, topology, and mechanical properties of the surface adhesion. Cell-cell junctions between endothelial cells, in culture and in situ, involve both Ca(2+)-dependent and -independent mechanisms that are mediated by distinct adhesion molecules. Ca(2+)-dependent cell-cell adhesion occurs mostly via members of the cadherin family, which locally anchor the microfilament system to the plasma membrane, in adherens junctions. Ca(2+)-independent adhesions were reported to mainly involve members of the Ig superfamily. In this study, we performed three-dimensional microscopic analysis of the relative subcellular distributions of these two endothelial intercellular adhesion systems. We show that cadherins are located at adjacent (usually more apical), yet clearly distinct domains of the lateral plasma membrane, compared to PECAM-1. Moreover, cadherins were first organized in adherens junctions within 2 h after seeding of endothelial cells, forming multiple lateral patches which developed into an extensive belt-like structure over a period of 24 h. PECAM-1 became associated with surface adhesions significantly later and became progressively associated with the cadherin-containing adhesions. Cadherins and PECAM-1 also differed in their detergent extractability, reflecting differences in their mode of association with the cytoskeleton. Moreover, the two adhesion systems could be differentially modulated since short treatment with the Ca2+ chelator EGTA, disrupted the cadherin junctions leaving PECAM-1 apparently intact. These results confirm that endothelial cells possess distinct intercellular contact mechanisms that differ in their spatial and temporal organization as well as in their functional properties.

scholarly journals Initial events during phagocytosis by macrophages viewed from outside and inside the cell: membrane-particle interactions and clathrin.

1982 ◽  
Vol 94 (3) ◽  
pp. 613-623 ◽  
Author(s):  
J Aggeler ◽  
Z Werb
Keyword(s):  
Electron Microscopy ◽  
Plasma Membrane ◽  
Plasma Membranes ◽  
Cell Attachment ◽  
High Resolution Electron Microscopy ◽  
Peritoneal Macrophages ◽  
Coated Pits ◽  
Thin Sections ◽  
Particle Uptake ◽  
Freeze Dried

The initial events during phagocytosis of latex beads by mouse peritoneal macrophages were visualized by high-resolution electron microscopy of platinum replicas of freeze-dried cells and by conventional thin-section electron microscopy of macrophages postfixed with 1% tannic acid. On the external surface of phagocytosing macrophages, all stages of particle uptake were seen, from early attachment to complete engulfment. Wherever the plasma membrane approached the bead surface, there was a 20-nm-wide gap bridged by narrow strands of material 12.4 nm in diameter. These strands were also seen in thin sections and in replicas of critical-point-dried and freeze-fractured macrophages. When cells were broken open and the plasma membrane was viewed from the inside, many nascent phagosomes had relatively smooth cytoplasmic surfaces with few associated cytoskeletal filaments. However, up to one-half of the phagosomes that were still close to the cell surface after a short phagocytic pulse (2-5 min) had large flat or spherical areas of clathrin basketwork on their membranes, and both smooth and clathrin-coated vesicles were seen fusing with or budding off from them. Clathrin-coated pits and vesicles were also abundant elsewhere on the plasma membranes of phagocytosing and control macrophages, but large flat clathrin patches similar to those on nascent phagosomes were observed only on the attached basal plasma membrane surfaces. These resulted suggest that phagocytosis shares features not only with cell attachment and spreading but also with receptor-mediated pinocytosis.

scholarly journals Immunofluorescence and electron microscopy of the cytoplasmic surface of the human erythrocyte membrane and its interaction with Sendai virus.

1979 ◽  
Vol 83 (2) ◽  
pp. 338-347 ◽  
Author(s):  
M Büechi ◽  
T Bächi
Keyword(s):  
Electron Microscopy ◽  
Plasma Membrane ◽  
Plasma Membranes ◽  
Sendai Virus ◽  
Light And Electron Microscopy ◽  
Distal Portion ◽  
Double Labeling ◽  
Virus Particles ◽  
Viral Antigens ◽  
Cytoplasmic Surface

A method was developed for directly observing the inner surfaces of plasma membranes by light and electron microscopy. Human erythrocytes were attached to cover slips (glass or mica) treated with aminopropylsilane and glutaraldehyde, and then disrupted by direct application of a jet of buffer, which removed the distal portion of the cells, thus exposing the cytoplasmic surface (PS) of the flattened membranes. Antispectrin antibodies and Sendai virus particles were employed as sensitive markers for, respectively, the PS and the external surface (ES) of the membrane; their localization by immunofluorescence or electron microscopy demonstrated that the major asymmetrical features of the plasma membrane were preserved. The fusion of Sendai virus particles with cells was investigated using double-labeling immunofluorescence techniques. Virus adsorbed to the ES of cells at 4 degrees C was not accessible to fluorescein-labeled antibodies applied from the PS side. After incubation at 37 degrees C, viral antigens could be detected at the PS. These antigens, however, remained localized and did not diffuse from the site of attachment, as is usually seen in viral antigens accessible on the ES. They may therefore represent internal viral antigens not incorporated into the plasma membrane as a result of virus-cell fusion.

Grid Computing in 3D Electron Microscopy Reconstruction

2012 ◽  
pp. 881-898
Author(s):  
J.R. Bilbao-Castro ◽  
I. García ◽  
J.J. Fernández
Keyword(s):  
Electron Microscopy ◽  
Grid Computing ◽  
Three Dimensional ◽  
Dimensional Electron ◽  
Reconstruction Process ◽  
Computing Paradigm ◽  
Structural Conformation ◽  
Reconstructed Volume ◽  
Dimensional Reconstruction ◽  
Computational Resources

Three-dimensional electron microscopy allows scientists to study biological specimens and to understand how they behave and interact with each other depending on their structural conformation. Electron microscopy projections of the specimens are taken from different angles and are processed to obtain a virtual three-dimensional reconstruction for further studies. Nevertheless, the whole reconstruction process, which is composed of many different subtasks from the microscope to the reconstructed volume, is not straightforward nor cheap in terms of computational costs. Different computing paradigms have been applied in order to overcome such high costs. While classic parallel computing using mainframes and clusters of workstations is usually enough for average requirements, there are some tasks which would fit better into a different computing paradigm – such as grid computing. Such tasks can be split up into a myriad of subtasks, which can then be run independently using as many computational resources as are available. This chapter explores two of these tasks present in a typical three-dimensional electron microscopy reconstruction process. In addition, important aspects like fault-tolerance are widely covered; given that the distributed nature of a grid infrastructure makes it inherently unstable and difficult to predict.

Grid Computing in 3D Electron Microscopy Reconstruction

2011 ◽  
pp. 392-409
Author(s):  
J.R. Bilbao Castro ◽  
I. Garcia Fernandez ◽  
J. Fernandez
Keyword(s):  
Electron Microscopy ◽  
Grid Computing ◽  
Three Dimensional ◽  
Dimensional Electron ◽  
Reconstruction Process ◽  
Computing Paradigm ◽  
Structural Conformation ◽  
Reconstructed Volume ◽  
Dimensional Reconstruction ◽  
Computational Resources

Three-dimensional electron microscopy allows scientists to study biological specimens and to understand how they behave and interact with each other depending on their structural conformation. Electron microscopy projections of the specimens are taken from different angles and are processed to obtain a virtual three-dimensional reconstruction for further studies. Nevertheless, the whole reconstruction process, which is composed of many different subtasks from the microscope to the reconstructed volume, is not straightforward nor cheap in terms of computational costs. Different computing paradigms have been applied in order to overcome such high costs. While classic parallel computing using mainframes and clusters of workstations is usually enough for average requirements, there are some tasks which would fit better into a different computing paradigm – such as grid computing. Such tasks can be split up into a myriad of subtasks, which can then be run independently using as many computational resources as are available. This chapter explores two of these tasks present in a typical three-dimensional electron microscopy reconstruction process. In addition, important aspects like fault-tolerance are widely covered; given that the distributed nature of a grid infrastructure makes it inherently unstable and difficult to predict.

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