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PubMed2005

[Scanning probe microscopy for the study of morphological parameters of neutrophilic granulocytes].

Morphological parameters of human blood neutrophilic granulocytes were studied by scanning probe microscopy. The basic morphological characteristics of the living (unfixed) and fixed cells were compared. It was found that using the method described, the dimensions of the cells changed substantially and significantly depending upon the mode of fixation applied. The methanol fixation allowed to demonstrate the internal structure of a cell, though it sharply deformed the object of study. After glutaraldehyde fixation cells possessed the morphological characteristics similar to those of the living cells, but the diameter and height of the fixed cells were again different from those of living cells. The resolution of a scanning probe microscope was higher when the fixed cells were studied, permitting the visualization of a detailed cell structure, including the nucleus and cytoplasmic granules (in particular, when methanol fixation was used). However, this method of study is unreliable for the determination of cell dimensions (diameter and height).

Cell Size↗
PubMed1988

Intracellular colocalization of variant surface glycoprotein and transferrin-gold in Trypanosoma brucei.

Endocytosis and intracellular transport has been studied in the bloodstream forms of Trypanosoma brucei by light and electron microscopy, using colloidal gold coupled to bovine transferrin (transferrin-gold). The endocytosed transferrin-gold, visualized by silver intensification for light microscopy, was present in vesicular structures between the cell nucleus and flagellar pocket of the organism. At the ultrastructural level, transferrin-gold was present after a 10-min incubation in the flagellar pocket, coated vesicles, cisternal networks, and lysosomelike structures. Endocytosis and intracellular processing of T. brucei variable surface glycoprotein (VSG) was studied using two preparations of affinity-purified rabbit IgG directed against different parts of the VSG. One preparation of IgG was directed against the cross-reacting determinant (CRD): a complex glycolipid side chain covalently linked to the COOH-terminus of the VSG molecule. The other was directed against determinants on the rest of the VSG molecule. When the two IgG preparations were used on thawed, thin cryosections of trypanosomes that had been incubated in transferrin-gold before fixation, the organelles involved with transferrin-gold endocytosis labeled with both antibodies, as well as many vesicular, tubular, and vacuolar structures that did not contain endocytosed transferrin-gold. Both antibodies also labeled the cell surface. In double-labeling experiments both antibodies were closely associated except that IgG directed against the VSG molecule labeled all the cisternae of the Golgi apparatus, whereas anti-CRD IgG was shown to label only half of the Golgi apparatus. Evidence for sorting of VSG molecules from endocytosed transferrin-gold was found. Double-labeling experiments also showed some tubular profiles which labeled on one side with anti-CRD IgG and on the other side with anti-VSG IgG, suggesting a possible segregation of parts of the VSG molecule.

Animals↗
PubMed1995

Novel nuclear ribonucleoprotein structural components in the dormouse adrenal cortex during hibernation.

Adrenocortical cell nuclei of the dormouse Muscardinus avellanarius were investigated by electron microscopic immunocytochemistry in hibernating, arousing and euthermic individuals. While the basic structural constituents of the cell nucleus did not significantly modify in the three groups, novel structural components were found in nuclei of hibernating dormice. Lattice-like bodies (LBs), clustered granules (CGs), fibrogranular material (FGM) and granules associated with bundles of nucleoplasmic fibrils (NF) all contained ribonucleoproteins (RNPs), as shown by labeling with anti-snRNP (small nuclear RNP), anti-m3G-capped RNA and anti-hnRNP (heterogeneous nuclear RNP) antibodies. Moreover, the FGM also showed immunoreactivity for the proliferation associated nuclear antigen (PANA) and the non-snRNP splicing factor SC-35. All these nuclear structural components disappeared early during arousal and were not found in euthermic animals. These novel RNP-containing structures, which have not been observed in other tissues investigated so far in the same animal model, could represent storage and/or processing sites for pre-mRNA during the extreme metabolic condition of hibernation, to be quickly released upon arousal. NFs, which had been sometimes found devoid of associated granules in nuclei of brown adipose tissue from hi-bernating dormice, were present in much higher amounts in adrenocortical cell nuclei; they do not contain RNPs and their role remains to be elucidated. The possible roles of these structures are discussed in the frame of current knowledge of morpho-functional relationships in the cell nucleus.

Adrenal Cortex↗
PubMed2001

Effect of retinoic acid on the actin cytoskeleton in HL-60 cells.

The cytoskeleton, in addition to its structural and kinetic functions, is also involved in modulating signal transfer in cell proliferation, differentiation and death. In some myeloid leukemic cell lines, the process of cell differentiation accompanied by apoptosis, can be induced by all-trans retinoic acid (ATRA). In this report, we describe the morphological changes in actin cytoskeleton, taking place during apoptosis in cells of the human leukemic HL-60 cell line. By using fluorescent microscopy, the morphology of microfilaments and the proportion of apoptotic cells in the cell populations untreated or treated with 10(-6) M ATRA were detected. Interphase HL-60 cells showed aggregations of short, thick microfilament bundles in the region between the plasma membrane and the nucleus. In comparison with both interphase and mitotic cells, the cells with apoptotic nuclear fragmentation showed a different organisation of the actin cytoskeleton. The following types of F-actin structures were observed: (i) Cells with a high number of large dots/patches of F-actin under the plasma membrane. These dots might be localised only in the part of the cell or occurred under the whole plasma membrane. This arrangement was often associated with a diffuse signal for F-actin. (ii) Cells with 3D-network of F-actin fibres through the cytoplasm between remnants of the cell nucleus. This 3D-structure probably played an important active role in the process of apoptotic bodies formation. (iii) Cells without any detectable signal for F-actin or cells with only a very low F-actin signal. Both of these showed typical apoptotic collapse of chromatin. It is concluded that the actin cytoskeleton is a dynamic structure actively involved in the executive phase of the process of apoptosis. It is suggested that the rearrangement of the microfilament network and its subsequent degradation are necessary for the main morphological changes of apoptotic cells, i.e., plasma membrane blebbing and apoptotic bodies formation.

Actins↗
PubMed1995

A1 catecholamine cell group: fine structure and synaptic input from the nucleus of the solitary tract.

Preembedding immunoperoxidase staining methods were used to characterize tyrosine hydroxylase-immunoreactive (TH-ir) elements in the caudal ventrolateral medulla, and to determine the extent to which neurons of the A1 cell group are directly innervated by projections of the nucleus of the solitary tract (NTS). TH-ir neurons in the A1 region were medium-sized and multipolar. They possessed rounded nuclei with infrequent invaginations, well-developed Golgi apparati, high cytoplasmic densities of mitochondria, and a low to moderate tendency for rough endoplasmic reticulum (RER) to align in parallel stacks. A1 cell bodies were commonly juxtaposed to TH-positive and TH-negative neurons, myelinated profiles, glia and/or vascular elements, but close membrane appositions were only seen with glial elements. Synaptic input to A1 neurons was predominantly asymmetric, provided virtually exclusively by non-TH-ir terminals, and directed principally to dendritic shafts; A1 somata are relatively sparsely innervated. In a second experiment, silver-intensified immunogold localization of TH-ir was combined with immunoperoxidase labeling for anterogradely transported Phaseolus vulgaris-leucoagglutinin (PHA-L), following tracer injections in the caudal aspect of the medial division of the NTS. These experiments revealed a small proportion of PHA-L-labeled axon terminals that made asymmetric contacts with dendritic shafts of TH-ir neurons. These results suggest that the fine structure and synaptic input of A1 neurons are somewhat distinct from that of rostrally situated C1 catecholamine cells. In addition, while they document a direct NTS-A1 projection that may participate in the interoceptive control of vasopressin secretion, the bulk of ventrolaterally directed projections from the caudomedial NTS contact noncatecholaminergic elements in the A1 region, some of which may correspond to so-called depressor neurons implicated in the baroreflex control of sympathetic outflow and vasopressin secretion.

Animals↗
PubMedDate not supplied

Fibrosarcoma mimicking plasmacytoma or carcinoma: an ultrastructural study of 4 cases.

Because the fibroblast has a remarkable capability of phenotypic modulations, reflected in both morphologic and immunohistochemical (IHC) changes, ultrastructural studies are mandatory to identify the variants of fibroblasts. Myofibroblasts or histiofibroblasts are such examples, demonstrating chimeric ultrastructural features of fibroblastic cells in common with smooth muscle cells or with histiocytes, respectively. The presence of epithelioid fibroblastic cells sharing morphologic features with epithelial or plasma cells has not been yet characterized. The authors identified 4 cases of fibrosarcomas (FS) characterized by an unusual phenotype and associated with peculiar ultrastructural findings. The electron microscopic (EM) findings were correlated with the histologic appearance and immunoprofile. All tumors were located in the extremities, 3 in soft tissues and 1 in the bone. By light microscopy 2 cases were composed predominantly by round uniform cells with a striking plasmacytoid appearance. One case mimicked carcinoma, composed predominantly by epithelioid cells and scattered giant tumor cells. The fourth case showed a mixture of plasmacytoid-like and epithelioid cells. By IHC, tumor cells were positive for vimentin and in 2 cases also for epithelial membrane antigen. Kappa/lambda light chain and cytokeratins markers were negative. By EM all 4 tumors showed in addition to classic features of fibroblasts, unusual epithelial-type features, such as secretory granules of "neurosecretory-type" (3 cases), rudimentary cell junctions (3 cases), microvilli (2 cases), and lumen-like structures (1 case). One plasmacytoid-type tumor showed finely granular extracellular deposits. The study describe 4 examples of fibrosarcomas with unusual features at the ultrastructural level, which are associated microscopically with a peculiar phenotype, mimicking plasmacytoma or carcinoma. These findings broaden the spectrum of fibroblastic cell variants in neoplasia.

Adult↗
PubMed1995

Nuclear matrix and the cell cycle.

The facts that the nuclear matrix represents a structural framework of the cell nucleus and that nuclear events, such as DNA replication, transcription, and DNA repair, are associated with this skeletal structure suggest that its components are subject to cell cycle-regulatory mechanisms. Cell cycle regulation has been shown for nuclear lamina assembly and disassembly during mitosis and chromatin reorganization. Little attention has so far been paid to internal nuclear matrix proteins and matrix-associated proteins with respect to the cell cycle. This survey attempts to summarize available data and presents experimental evidence that important metabolic functions of the nucleus are regulated by the transient, cell cycle-dependent attachment of enzymes and regulatory proteins to the nuclear matrix. Results on thymidine kinase and RNA polymerase during the synchronous cell cycle of Physarum polycephalum demonstrate that reversible binding to the nuclear matrix represents an additional level of regulation for nuclear processes.

Animals↗
PubMed2006

The nucleolus: a model for the organization of nuclear functions.

Nucleoli are the prominent contrasted structures of the cell nucleus. In the nucleolus, ribosomal RNAs (rRNAs) are synthesized, processed and assembled with ribosomal proteins. The size and organization of the nucleolus are directly related to ribosome production. The organization of the nucleolus reveals the functional compartmentation of the nucleolar machineries that depends on nucleolar activity. When this activity is blocked, disrupted or impossible, the nucleolar proteins have the capacity to interact independently of the processing activity. In addition, nucleoli are dynamic structures in which nucleolar proteins rapidly associate and dissociate with nucleolar components in continuous exchanges with the nucleoplasm. At the time of nucleolar assembly, the processing machineries are recruited in a regulated manner in time and space, controlled by different kinases and form intermediate structures, the prenucleolar bodies. The participation of stable pre-rRNAs in nucleolar assembly was demonstrated after mitosis and during development but this is an intriguing observation since the role of these pre-rRNAs is presently unknown. A brief report on the nucleolus and diseases is proposed as well as of nucleolar functions different from ribosome biogenesis.

Cell Cycle↗
PubMed2004

The functional architecture of the nucleus as analysed by ultrastructural cytochemistry.

Ultrastructural cytochemistry has been, for many years now, a major tool for investigating structure-function relationships in the cell nucleus. It has been essential in approaching the roles which different nuclear structural constituents can play in nuclear functions. This article briefly summarises transmission electron microscopic studies aimed at characterising in situ nuclear architectural domains and their involvement in main nuclear functions, such as DNA replication, hnRNA transcription and pre-mRNA processing. It discusses the importance of ultrastructural cytochemistry in high resolution analyses of intranuclear distribution of chromatin domains and their topological relationships with other structural interphase nuclear constituents. It puts forward the central role of the perichromatin region as a functional nuclear domain. Finally, it attempts to critically evaluate some future applications of ultrastructural investigations of the nucleus and stresses the importance of combining them with light microscopic analyses of living cells.

Animals↗
PubMedDate not supplied

[Structural changes in the chromatin during the cell differentiation of the erythroid series].

Data about the changes in the molecular organization of the cell nucleus on different levels of its structure during erythroid cell differentiation are presented. Changes are observed in DNA methylation and secondary structure. Protein composition of the chromatin and the length of the nucleosomal repeat change. Data about the peculiarities of the chromatin structure of erythroid-specific genes and their association to the nuclear skeleton are presented as well.

Alpha-Globulins↗
PubMed1993

Distribution of [241Am] in hepato-pancreatic nuclear proteins of the lobster (Homarus gammarus).

Recent studies on the bio-accumulation of transuranic elements in marine invertebrates have demonstrated an intracellular concentration of radionuclides in the digestive gland. The present investigation aims to establish the distribution of [241Am] within the different structures of the cell nucleus. Lobsters were experimentally contaminated through feeding and then killed. The nuclei of hepato-pancreatic cells were isolated, purified and dissociated. The different protein categories were separated in media of increasing ionic strength. The distribution of americium in this material yielded results showing a preferential fixation onto the structural proteins of the nuclear matrix. The radiological consequences of such an affinity are discussed in terms of the life-times of the ligands concerned.

Americium↗