scholarly journals Histochemical and Molecular Characterization of Spongiosal Cells in Native Tissue, Two- and Three-Dimensional Cultures of Rat Aortic Valve

2016 ◽  
Vol 2016 ◽  
pp. 1-8
Author(s):  
Vibudha Nanduri ◽  
Beheita Moein ◽  
Avinash Raj Thatipalli ◽  
Gopal Pande ◽  
Anasuya Ganguly
Keyword(s):  
Aortic Valve ◽  
Cell Cultures ◽  
Three Dimensional ◽  
Cellular Organization ◽  
Collagen Gels ◽  
Cytochemical Staining ◽  
Cell Lineages ◽  
Native Tissue ◽  
Immunofluorescence Studies

The histocytochemical and molecular analysis of cells that constitute the aortic valve (AV) of the rat heart was done in this study. We have focussed on the identity of cells in the spongiosal layer of the valve by immunofluorescence studies using lineage specific markers and cytochemical staining. We have established two-dimensional (2D) cultures of cells from isolated rat AV leaflets and maintained endothelial and interstitial valvular cells (IVC) over a period of six to eight weeks. Using “passage 0” cells from 2D valvular cultures, we could reconstruct the three-dimensional (3D) valvular tissue in collagen gels that showed very similar cellular organization and marker expression profile, as that of the native tissue. Lineage specific markers in the native tissue and cell cultures were studied by Reverse Transcriptase-PCR and immunofluorescence for VCAM-I, α-SMA, collagen I, CD71, collagen II, and E-cadherin markers. This is the first report on the identification of cell lineages in the spongiosal layer of AV and the successful reconstruction of 3D valvular tissue from primary cell cultures of AV.

scholarly journals Identification and partial characterization of three low-molecular-weight collagenous polypeptides synthesized by chondrocytes cultured within collagen gels in the absence and in the presence of fibronectin

1983 ◽  
Vol 211 (2) ◽  
pp. 417-426 ◽  
Author(s):  
G J Gibson ◽  
C M Kielty ◽  
C Garner ◽  
S L Schor ◽  
M E Grant
Keyword(s):  
Molecular Weight ◽  
Peptide Mapping ◽  
Three Dimensional ◽  
Low Molecular Weight ◽  
Collagen Gels ◽  
Serum Supplement ◽  
Electrophoretic Mobilities ◽  
Reducing Conditions ◽  
Interstitial Collagens

Culture of chick-embryo sternal-cartilage chondrocytes within three-dimensional collagen gels promotes the synthesis of three low-molecular-weight collagenous polypeptides. The proportions of these novel collagens synthesized and released into the medium are markedly influenced by the presence or the absence of fibronectin in the serum supplement. Chondrocytes cultured on plastic dishes appear to synthesize only small amounts of these low-molecular-weight species. The three species (designated G, H and J) were characterized with respect to the proportion of [14C]proline incorporated into each polypeptide occurring as hydroxy[14C]proline and with respect to their susceptibilities to bacterial collagenase. On the basis of their electrophoretic mobilities under reducing conditions, the G, H and J polypeptides were calculated to have Mr 59 000, 69 000 and 84 000 respectively. Chymotrypsin digestion converted the G collagen into a species containing polypeptides of Mr 45 000, whereas the H and J polypeptides yielded a single band of Mr 53 000. The H and J polypeptides were found to occur as disulphide-linked aggregates, as was the chymotrypsin-digestion product. Peptide ‘mapping’ has shown that G, H and J polypeptides show no common identity and are distinct from the known interstitial collagens. Native G collagen was digested by human collagenase to discrete products, whereas H and J chains were not cleaved under identical conditions.

Scanning Electron Microscopy and Electron Microprobe Analysis of Malignant Cell Cultures

1968 ◽  
Vol 26 ◽  
pp. 164-165
Author(s):  
R. I. Johnsson-Hegyeli ◽  
A. F. Hegyeli ◽  
D. K. Landstrom ◽  
W. C. Lane
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Cell Cultures ◽  
Electron Microprobe ◽  
Microprobe Analysis ◽  
Electron Microprobe Analysis ◽  
Three Dimensional ◽  
Malignant Cell ◽  
Interference Microscopy ◽  
Scanning Electron

Last year we reported on the use of reflected light interference microscopy (RLIM) for the direct color photography of the surfaces of living normal and malignant cell cultures without the use of replicas, fixatives, or stains. The surface topography of living cells was found to follow underlying cellular structures such as nuceloli, nuclear membranes, and cytoplasmic organelles, making possible the study of their three-dimensional relationships in time. The technique makes possible the direct examination of cells grown on opaque as well as transparent surfaces. The successful in situ electron microprobe analysis of the elemental composition and distribution within single tissue culture cells was also reported.This paper deals with the parallel and combined use of scanning electron microscopy (SEM) and the two previous techniques in a study of living and fixed cancer cells. All three studies can be carried out consecutively on the same experimental specimens without disturbing the cells or their structural relationships to each other and the surface on which they are grown. KB carcinoma cells were grown on glass coverslips in closed Leighto tubes as previously described. The cultures were photographed alive by means of RLIM, then fixed with a fixative modified from Sabatini, et al (1963).

Characterization of photosystem II with the atomic-force microscope

1992 ◽  
Vol 50 (2) ◽  
pp. 1146-1147
Author(s):  
Kathleen M. Marr ◽  
Mary K. Lyon
Keyword(s):  
Photosystem Ii ◽  
Atomic Force Microscope ◽  
Three Dimensional ◽  
Biologically Active ◽  
Atomic Force ◽  
Freeze Etching ◽  
Image Averaging ◽  
Oxygen Evolving ◽  
Averaging Techniques

Photosystem II (PSII) is different from all other reaction centers in that it splits water to evolve oxygen and hydrogen ions. This unique ability to evolve oxygen is partly due to three oxygen evolving polypeptides (OEPs) associated with the PSII complex. Freeze etching on grana derived insideout membranes revealed that the OEPs contribute to the observed tetrameric nature of the PSIl particle; when the OEPs are removed, a distinct dimer emerges. Thus, the surface of the PSII complex changes dramatically upon removal of these polypeptides. The atomic force microscope (AFM) is ideal for examining surface topography. The instrument provides a topographical view of individual PSII complexes, giving relatively high resolution three-dimensional information without image averaging techniques. In addition, the use of a fluid cell allows a biologically active sample to be maintained under fully hydrated and physiologically buffered conditions. The OEPs associated with PSII may be sequentially removed, thereby changing the surface of the complex by one polypeptide at a time.

convergent-beam diffraction from surfaces

1987 ◽  
Vol 45 ◽  
pp. 30-33
Author(s):  
J. A. Eades ◽  
A. E. Smith ◽  
D. F. Lynch
Keyword(s):  
Reciprocal Lattice ◽  
Three Dimensional ◽  
Grazing Incidence ◽  
Three Dimensions ◽  
Plane Figure ◽  
Transmission Electron ◽  
Convergent Beam ◽  
Beam Patterns ◽  
Beam Diffraction

It is quite simple (in the transmission electron microscope) to obtain convergent-beam patterns from the surface of a bulk crystal. The beam is focussed onto the surface at near grazing incidence (figure 1) and if the surface is flat the appropriate pattern is obtained in the diffraction plane (figure 2). Such patterns are potentially valuable for the characterization of surfaces just as normal convergent-beam patterns are valuable for the characterization of crystals.There are, however, several important ways in which reflection diffraction from surfaces differs from the more familiar electron diffraction in transmission.GeometryIn reflection diffraction, because of the surface, it is not possible to describe the specimen as periodic in three dimensions, nor is it possible to associate diffraction with a conventional three-dimensional reciprocal lattice.

Construction of plastic contact deformation maps on ceramics: a case study on aluminum nitride

1996 ◽  
Vol 54 ◽  
pp. 636-637
Author(s):  
D. L. Callahan
Keyword(s):  
Plastic Deformation ◽  
Aluminum Nitride ◽  
Mechanical Response ◽  
Three Dimensional ◽  
Bright Field Image ◽  
Perfectly Plastic ◽  
Contrast Analysis ◽  
Deformation Patterns

Modern polishing, precision machining and microindentation techniques allow the processing and mechanical characterization of ceramics at nanometric scales and within entirely plastic deformation regimes. The mechanical response of most ceramics to such highly constrained contact is not predictable from macroscopic properties and the microstructural deformation patterns have proven difficult to characterize by the application of any individual technique. In this study, TEM techniques of contrast analysis and CBED are combined with stereographic analysis to construct a three-dimensional microstructure deformation map of the surface of a perfectly plastic microindentation on macroscopically brittle aluminum nitride.The bright field image in Figure 1 shows a lg Vickers microindentation contained within a single AlN grain far from any boundaries. High densities of dislocations are evident, particularly near facet edges but are not individually resolvable. The prominent bend contours also indicate the severity of plastic deformation. Figure 2 is a selected area diffraction pattern covering the entire indentation area.

Three-dimensional image analysis and visualization in confocal light microscopy

1993 ◽  
Vol 51 ◽  
pp. 158-159
Author(s):  
J. K. Samarabandu ◽  
R. Acharya ◽  
D. R. Pareddy ◽  
P. C. Cheng
Keyword(s):  
Depth Perception ◽  
Computer Graphic ◽  
Three Dimensional ◽  
Cellular Organization ◽  
Data Set ◽  
Computational Burden ◽  
Interactive Operation ◽  
Cell Organization ◽  
Confocal Images ◽  
3D Digitization

In the study of cell organization in a maize meristem, direct viewing of confocal optical sections in 3D (by means of 3D projection of the volumetric data set, Figure 1) becomes very difficult and confusing because of the large number of nucleus involved. Numerical description of the cellular organization (e.g. position, size and orientation of each structure) and computer graphic presentation are some of the solutions to effectively study the structure of such a complex system. An attempt at data-reduction by means of manually contouring cell nucleus in 3D was reported (Summers et al., 1990). Apart from being labour intensive, this 3D digitization technique suffers from the inaccuracies of manual 3D tracing related to the depth perception of the operator. However, it does demonstrate that reducing stack of confocal images to a 3D graphic representation helps to visualize and analyze complex tissues (Figure 2). This procedure also significantly reduce computational burden in an interactive operation.

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