Reconstruction Of Nuclei Research Articles

Three-dimensional reconstruction of cell nuclei, internalized quantum dots and sites of lipid peroxidation

BackgroundThe purpose of the study was to develop and illustrate three-dimensional (3-D) reconstruction of nuclei and intracellular lipid peroxidation in cells exposed to oxidative stress induced by quantum dots. Programmed cell death is characterized by multiple biochemical and morphological changes in different organelles, including nuclei, mitochondria and lysosomes. It is the dynamics of the spatio-temporal changes in the signalling and morphological adaptations which will ultimately determine the 'shape' and fate of the cell.ResultsWe present new approaches to the 3-D reconstruction of organelle morphology and biochemical changes in confocal live-cell images. We demonstrate the 3-D shapes of nuclei, the 3-D intracellular distributions of QDs and the accompanying lipid-membrane peroxidation, and provide methods for quantification.ConclusionThis study provides an approach to 3-D organelle and nanoparticle visualization in the context of cell death; however, this approach is also applicable more generally to investigating changes in organelle morphology in response to therapeutic interventions, stressful stimuli and internalized nanoparticles. Moreover, the approach provides quantitative data for such changes, which will help us to better integrate compartmentalization of subcellular events and to link morphological and biochemical changes with physiological outcomes.

Determination of the mean caliper diameter of lung nuclei by a method which is independent of shape assumptions.

When determining the number of nuclei (therefore cells) present in a known volume of tissue it is necessary to determine the overall mean caliper diameter of nuclei for each cell type studied. Most previous methods for determining the mean caliper diameters (D) of of cell nuclei were strongly dependetn upon assumptions of the shape of the particles in question (Greeley et al., 1978). A computer program has now been designed to calculate D from three-dimensional reconstructions of nuclei in a manner that is independent of all shape assumptions. The Ds of ten capillary endothelial cell nuclei from normal rat lungs have been determined by this shape-independent method. The resulting overall mean caliper diameter for this group of ten nuclei varied only slightly from that previously estimated for them by a method which assumes them to be triaxial ellipsoids. In a similar comparison, the Ds for ten normal human lung endothelial cell nuclei were determined by both methods. The overall mean caliper diameter determined by the shape-independent method was 10% larger than that estimated by the shape-dependent method. This suggests that human lung endothelial nuclei vary significantly from the shape assumption (triaxial ellipsoid) employed in the shape-dependent method, and the shape-independent method was considered to yield a more accurate measurement of D.

Synaptonemal complexes in the mosquitoCulex quinquefasciatus

The presence of synaptonemal complexes between the paired homologous chromosomes at meiotic prophase is a prerequisite for meiotic crossing over, and it may be important for the regular disjunctions of the chromosomes at meiosis I (Moses, 1968; Westergaard and von Wettstein, 1972; Gillies, 1975). Reconstructions of nuclei during zygotene and pachytene have shown that the ends of the synaptonemal complexes in many organisms are attached to the nuclear membrane, often in a polarized fashion (Moens, 1969; Rasmussen, 1976); such a bouquet arrangement of the chromosomes is found in Culex.Materials and MethodsOvaries from Culex quinquefasciatus were fixed in glutaraldehyde, followed by 0s04, and embedded in Epon. The synaptonemal complexes were reconstructed from serial sections.Results and DiscussionCulex has 3 pairs of very long metacentric or slightly submetacentric chromosomes which during pachytene loop around the nucleus several times (Fig. 1). The centromeric regions are fused, and the synaptonemal complexes do not continue through the structure.