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PubMed1968

On the role of microtubules in movement and alignment of nuclei in virus-induced syncytia.

Infection of baby hamster kidney (BHK21-F) cells with the parainfluenza virus SV5 causes rapid and extensive cell fusion. Time-lapse cinematography shows that when cells fuse, their nuclei migrate straight to the center of the syncytium at rates of 1-2 micro/min. Nuclei are often arranged in long, tightly packed, parallel rows in syncytia derived from the fibroblastic BHK21-F cells. Polarization microscopy shows birefringent material between and parallel to these rows of nuclei, and electron microscopy shows bundles of cytoplasmic microtubules, approximately 250 A in diameter, and filaments, approximately 80 A in diameter, parallel to and between the rows of nuclei. Colchicine treatment causes disappearance of microtubules from BHK21-F cells and an apparent increase in the number of 80-A filaments. Although colchicine-treated, SV5-infected cells fuse, their nuclei do not migrate or form rows but remain randomly scattered through the syncytial cytoplasm. Incubation at 4 degrees C does not disrupt microtubules in BHK21-F cells. Rows of nuclei have been isolated from SV5-induced syncytia, and the nuclei in them have been found to be intimately associated with microtubules but not with other cytoplasmic structures. These results suggest that microtubules demarcate cytoplasmic channels through which nuclei migrate and that they may also be involved in the mechanism of nuclear movement.

Animals↗
PubMed1994

A confocal microscopy study of anticytoskeletal antibody activity in patients with connective tissue disease.

The significance of the presence of antibodies to cytoskeleton proteins in patients with connective tissue diseases is not clear, as there is a high level of these antibodies in healthy controls. In an attempt to improve the visualization of the immunofluorescence binding pattern of autoantibodies to cytoskeletal structures in cultured fibroblasts, we have used confocal microscopy. Of the 256 serum samples tested, 155 (61%) WERE reactive with cytoplasmic structures. These reactive samples could be divided into seven patterns of binding, as determined by double-blind examination of single-section confocal images. While confirming the results of previous immunofluorescence studies which have shown that autoantibodies that bind to filamentous structures in the cytoplasm of cultured cells are common in patients with connective tissue diseases, we were able to identify three patterns of cytoskeletal binding which may be useful as an adjunct to other tests for the diagnosis of some connective tissue diseases, in particular systemic sclerosis (scleroderma) and rheumatoid arthritis/Sjogren's syndrome. None of the seven patterns was exclusive to a particular disease. We conclude that confocal microscopy may be of limited use as an adjunct to other serological assays in the diagnosis of some forms of connective tissue disease.

Actin Cytoskeleton↗
PubMed1983

Application of optical diffractometry in studies of cell fine structure. Comparison of arterial smooth muscle cells in contractile and synthetic state.

Arterial smooth muscle cells in contractile and synthetic state were analyzed by optical diffractometry. Cell sections (80-90 nm) were photographed in an electron microscope and diffraction patterns of the plates (negatives) were produced using a helium-neon laser. Radial and angular distributions of light intensity in the diffractograms were measured and digitized using an electronic detector plate consisting of ring- and wedge-shaped photosensitive elements; radial distributions provide information about size of structures and distances between them and angular distributions about spatial orientation of structures in the images. Micrographs of nuclei and cytoplasm were analyzed separately (40-50 plates in each group). Computerized statistical analysis of radial distributions of light intensity showed that the nuclear chromatin pattern differed between cells in contractile and synthetic state. The probability that the observed difference could have arisen purely by chance was less than 10(-5). Computer-aided classification to the a priori known cell group was correct in 96.5% of the cases. Analysis of radial distributions of light intensity similarly showed marked differences in cytoplasmic structure between cells in contractile state (dominated by bundles of myofilaments) and synthetic state (dominated by cisternae of rough endoplasmic reticulum). The probability that the observed difference could have arisen purely by chance was less than 10(-5). Computer-aided classification to the a priori known cell group was correct in 92.0% of the cases. In contrast, analysis of angular distributions of light intensity did not indicate any statistically significant differences between contractile and synthetic state cells. A likely reason is that both myofilaments and cisternae of rough endoplasmic reticulum were arranged in parallel. The results demonstrate that optical diffractometry is a useful method for image analysis in studies of cell fine structure. It provides information about size and orientation of structures with poorly defined shape and is particularly well suited for studies on cell differentiation and effects of pharmacological and other experimental treatments on cell fine structure. It represents an alternative and a complement to stereology for quantitative and objective evaluation of morphological data.

Animals↗
PubMed1981

Ultrastructural changes in the liver of patients with chronic non-A, non-B hepatitis.

Electron microscopy examination of liver biopsies from 8 patients with chronic non-A, non-B hepatitis revealed ultrastructural changes similar to those previously described in chimpanzees with experimentally induced acute non-A, non-B hepatitis. These changes consisted of intranuclear clusters of electron-dense, 15-27-nm particles that were detected in five out of the eight patients and of circular cytoplasmic structures that were present in seven cases. Other cytoplasmic abnormalities found in our patients related to the presence of curved membranes apparently developing from apposition of two cisternae of endoplasmic reticulum. In contrast with what has been reported in infected chimpanzees, the nuclear and cytoplasmic changes were not mutually exclusive in our patients, but coexisted in four of them.

Adult↗
PubMed2004

Increased importin-beta-dependent nuclear import of the actin modulating protein CapG promotes cell invasion.

CapG (gCap39) is a ubiquitous gelsolin-family actin modulating protein involved in cell signalling, receptor-mediated membrane ruffling, phagocytosis and motility. CapG is the only gelsolin-related actin binding protein that localizes constitutively to both nucleus and cytoplasm. Structurally related proteins like severin and fragmin are cytoplasmic because they contain a nuclear export sequence that is absent in CapG. Increased CapG expression has been reported in some cancers but a causal role for CapG in tumour development, including invasion and metastasis, has not been explored. We show that moderate expression of green fluorescent protein-tagged CapG (CapG-EGFP) in epithelial cells induces invasion into collagen type I and precultured chick heart fragments. Nuclear export sequence-tagged CapG-EGFP fails to induce invasion, whereas point mutations in the nuclear export sequence permitting nuclear re-entry restore cellular invasion. Nuclear import of CapG is energy-dependent and requires the cytosolic receptor importin beta but not importin alpha. Nuclear CapG does not possess intrinsic transactivation activity but suppresses VP16 transactivation of a luciferase reporter gene in a dose-dependent manner. Furthermore, invasion requires signalling through the Ras-phosphoinositide 3-kinase pathway and Cdc42 or RhoA, but not Rac1. We show for the first time active nuclear import of an actin binding protein, and our findings point to a role for nuclear CapG in eliciting invasion, possibly through interfering with the cellular transcription machinery.

Actins↗
PubMed1982

Heterogeneity of the early G2 phase-arrested cells; cytophotometric, interferometric, and EM morphometric studies.

Cells of antheridial filaments in Chara vulgaris L. blocked at the early G2 phase by a 5 day dark treatment were studied cytologically in order to determine causes of their heterogeneous response to light-induced mitotic activation. Cell size distribution patterns in dark inhibited populations were shown to be nearly consistent with those found within the control populations, comprising all the stages of interphase; cells which resume divisions in response to photoinduction attain, however, sizes characteristic of mitosis occurring in natural light conditions. Cytophotometric determinations of nuclei in cells given dark treatment indicated a complete homogeneity of populations in respect of replicated DNA contents. As evidence by nucleus-cytoplasm surface relations, interferometric measurements of their dry mass contents, and EM stereometric studies, the heterogeneity of early G2 phase arrested populations results from an imbalance between nuclear DNA-division cycle, which is continued up to the early stage of the G2 period, and the growth cycle, which in turn, due to its pronounced inhibition gives rise to the deficit of cytoplasm. Since no shortages in total volumes of structural cytoplasm components were found the number and volume of mitochondria were shown even largely increased, it is assumed that asynchrony during light-induced mitotic reactivation originates from quantitative differences of constituents within the area of ground cytoplasm.

Cell Nucleus↗
PubMed1976

[Localization in cells of Neisseria perflava and Klebsiella pneumoniae of antigens in common (cross-reacting) with tissues of the human bronchopulmonary apparatus].

A study was made of localization in the cells of Neisseria perflava and Klebsiella pneumoniae of antigens common with the tissues of the human broncho-pulmonary apparatus. A study was made of cross reactions of some structures of Neisseria perflava and Klebsiella pneumoniae cells (capsule, walls, cytoplasmic structure fraction, hyaloplasm) in the complement fixation test with antilung sera. Antigens cross reacting with the antilung sera were localized not only in the surface structures (walls) of bacterial cells, but also in the "deep" components (cytoplasm fraction rich in ribonucleoproteids) of the microbe.

Antigens↗
PubMed1999

SNARE membrane trafficking dynamics in vivo.

The ER/Golgi soluble NSF attachment protein receptor (SNARE) membrin, rsec22b, and rbet1 are enriched in approximately 1-micrometer cytoplasmic structures that lie very close to the ER. These appear to be ER exit sites since secretory cargo concentrates in and exits from these structures. rsec22b and rbet1 fused to fluorescent proteins are enriched at approximately 1-micrometer ER exit sites that remained more or less stationary, but periodically emitted streaks of fluorescence that traveled generally in the direction of the Golgi complex. These exit sites were reused and subsequent tubules or streams of vesicles followed similar trajectories. Fluorescent membrin- enriched approximately 1-micrometer peripheral structures were more mobile and appeared to translocate through the cytoplasm back and forth, between the periphery and the Golgi area. These mobile structures could serve to collect secretory cargo by fusing with ER-derived vesicles and ferrying the cargo to the Golgi. The post-Golgi SNAREs, syntaxin 6 and syntaxin 13, when fused to fluorescent proteins each displayed characteristic patterns of movement. However, syntaxin 13 was the only SNARE whose life cycle appeared to involve interactions with the plasma membrane. These studies reveal the in vivo spatiotemporal dynamics of SNARE proteins and provide new insight into their roles in membrane trafficking.

Animals↗
PubMed1975

Observation of Kolmer's crystalloid outside the retina. Presence in the corneal endothelium in various conditions.

A structure similar to Kolmer's crystalloid, an organelle found in the horizontal cells of the retina, was observed in endothelial cells of the cornea from four patients (one case of hyperparathyroidism and three cases of corneal dystrophy). Based on a morphological analysis it is concluded that the structures in the present cases are identical with the cytoplasmic structure of the horizontal cell, and that the membranous tubes probably constitute a special form of rough surfaced endoplasmic reticulum, eventually involved in transport and/or a specialized protein synthesis.

Cell Membrane↗
PubMed1993

The gamma subunit of the B cell antigen-receptor complex is a C-terminally truncated product of the B29 gene.

The predominant Ag-receptor complex of B cells consists of mIgM or mIgD noncovalently associated with glycosylated heterodimers of Ig-alpha and Ig-beta or Ig-alpha and Ig-gamma. Upon B cell stimulation the associated proteins are phosphorylated, giving rise to pp32/33 (alpha), pp37 (beta), and pp34 (previously designated gamma). Ig-alpha and Ig-beta contain extended cytoplasmic structure (61 and 48 amino acids, respectively) and associate with cytoplasmic effectors indicating that they are directly involved in signal transduction. Here we report analysis of the structural relationship of mIgM- and mIgD-associated Ig-beta and Ig-gamma chains from mice. N-terminal sequence, immunoblotting, and physicochemical analyses show that both Ig-beta and Ig-gamma are products of the B cell-specific B29 gene and demonstrate that the 37-kDa Ig-beta protein is the full length predicted product of the B29 gene. The Ig-associated protein that migrates in the 34-kDa range is actually two distinct species. The minor species is a phosphorylatable and underglycosylated form of full length Ig-beta, and the major species is a C-terminally truncated form of B29, which we now designate Ig-gamma. This conclusion is based on the observations that Ig-gamma is composed of a core protein which is 3 to 4 kDa smaller than deglycosylated Ig-beta, it is not phosphorylated, unlike Ig-beta, and it does not react with an antiserum raised against a peptide of the seven C-terminal amino acids of B29. Based on these findings we estimate that Ig-gamma is truncated by about 30 to 36 amino acid residues and hypothesize that the most 3' B29 exon, which encodes the 32 C-terminal residues, may not be expressed in Ig-gamma. All of the documented B29 products are found in association with both mIgM and mIgD. Interestingly, Ig-gamma is found in intermediate and low density splenic B cells, but is not detectable in resting B cells. This raises the possibility that it may confer some distinct signaling function on the Ag receptors of these cells.

Amino Acid Sequence↗
PubMedDate not supplied

Change in ultrastructure of apical cells in wheat shoots in ontogenesis.

Electron microscopy and morphometric studies were made of apical meristem cells of a shoot of wheat (Triticum aestivum L., variety Saratovskaya-29) at various phases of ontogenesis: vegetative, prefloral, and beginning of flowering (from seed embryo to the beginning of formation of the head). It was demonstrated that during this period there are changes in cytoplasmic structures, their number, proportions, and internal structures. Changes were disclosed which were linked to the hereditary course of growth and differentiation of the apex, that is, changes that were evidently ontogenetic (dynamics of lipid drops, of the Golgi apparatus, endoplasmic reticulum, and to some degree, the structure of plastids and mitochondria). It was demonstrated that in the course of the vegatative phase, changes in activity of the apex meristem and its various zones are connected with comparatively small quantitative changes in content of basic organelle cytoplasm, and their structural transformations are more significant. The activation of the apex in the pre-floral phase is linked to an increase in formation of membrane systems, increase in the functional activity of organelles, and increase in the formation of ribosomes and their density in the cytoplasm which provides conditions for a transition of the apex to generative differentiation.

Microscopy, Electron↗
PubMed1981

The mast cell: IV. An ultrastructural and autoradiographic study of the distribution and maturation of peritoneal mast cells in the rat.

Ultrastructural studies of peritoneal mast cells in adult and young rats showed that results are comparable to those obtained by light microscopic and histochemical studies as far as their maturation and distribution are concerned. It was also evident that mast cells possess cytoplasmic structures common to other cells including microtubules and microfilaments in addition to their distinctive cytoplasmic granules. Granules appear to develop from the vicinity of the Golgi apparatus as progranules in a structured sequence to full maturity rather than from mitochondria or from the cytoplasm de novo. In newborn and young rats mast cells are mostly immature and exhibit frequent mitotic activity. Autoradiographic studies showed that young Stages 1 and 2 peritoneal mast cells are primarily responsible for mitotic and therefore proliferative activity. It was concluded that the final density of mast cells is achieved in the early stages by proliferation and possibly by heteroplastic differentiation of precursor cells.

Animals↗
PubMed1963

The fine structure of Rhodospirillum rubrum.

The fine structure of Rhodospirillum rubrum grown under a series of defined conditions has been examined in thin sections prepared by the methods of Ryter and Kellenberger. In cells grown anaerobically at different light intensities, the abundance of 500 A membrane-bounded vesicles in the cytoplasm is inversely related to light intensity, and directly related to cellular chlorophyll content. When the chlorophyll content of the cell is low, the vesicles are exclusively peripheral in location; they extend more deeply into the cytoplasm when the chlorophyll content is high. Typical vesicles also occur, though rarely, in cells grown aerobically in the dark, which have a negligible chlorophyll content. When synthesis of the photosynthetic pigment system is induced in a population of aerobically grown cells by incubation under semianaerobic conditions in the dark, the vesicles become increasingly abundant with increasing cellular chlorophyll content, and the cells eventually acquire the cytoplasmic structure that is characteristic of cells growing anaerobically at a high light intensity. Poststaining with lead hydroxide reveals that the membranes surrounding the 500 A vesicles are indistinguishable in structure from the cytoplasmic membrane, and continuous with it in some areas of the sections. The bearing of these observations on current notions concerning the organization of the bacterial photosynthetic apparatus is discussed.

Cell Membrane↗
PubMed2003

Ultrastructural study of Entamoeba invadens encystation and excystation.

In the life cycle of Entamoeba species, the cyst and all the processes associated to it have been poorly studied. Entamoeba invadens, a serpent's parasite, has been commonly accepted as a model for the study of encystation and excystation. Here we analyzed through scanning and transmission electron microscopy the in vitro morphological differentiation of both processes. During encystation, the formation of an irregular net of fibrillar material on the surface of precysts was observed. In thin sections of cryofixed and cryosubstituted specimens, abundant vacuoles containing a microfibrillar material of similar appearance to the structural components of the cyst wall were found in the cytoplasm. Assays with a calcofluor probe on cryosections of encysting trophozoites and precysts showed the presence of fluorescent circular cytoplasmic structures. In the cyst stage, the fluorescence was located on the surface. During excystation, the detachment of the metacyst from the cyst wall was observed through scanning electron microscopy. Metacysts endocyting amorphous material which may correspond to cyst wall residues were commonly found. By transmission electron microscopy the formation of a crescent-shaped space between the plasma membrane and the cyst wall was observed. Abundant small electrondense bodies were found in the cytoplasm. Many of them were in close apposition to the plasma membrane and frequently some of them were seen projecting towards this newly formed space. Our results suggest that the microfibrillar content of the vacuoles corresponds to the cyst wall material, that the electrondense bodies may be involved in the excystation process, and that part of the cyst wall residues may be endocyted by the parasite.

Animals↗
PubMed1981

The relationship between the nuage and the chromatid body during spermatogenesis in the rat.

Cytoplasmic structures ultrastructurally similar to the nuage are present in the cytoplasm of all spermatogenic cells in adult rats. The nuage is a discrete organelle which should not be confused with the chromatoid body. In step 7-8 spermatids transient contact is established between the nuage by the chromatoid body. This indicates a very specific recognition of the nuage by the chromatoid body. It is suggested that the nuage and the chromatoid body are separate cell organelles the functions of which are somehow related to each other.

Animals↗
PubMed1992

Monoclonal antiphospholipid antibody reactivity against human placental trophoblast.

Naturally occurring antibodies against the negatively charged phospholipids cardiolipin (CL) and phosphatidylserine (PS) have been associated with recurrent pregnancy loss. One prevalent hypothesis proposes that antiphospholipid antibody (aPL) mediated pathophysiology is through increased placental thrombosis. In this study we investigated the reactivity of three mouse monoclonal aPLs with term and 26 week human placental preparations. Each monoclonal antibody reacted differently with CL and PS; 3SB9b reacted with PS (CL-/PS+), D11A4 reacted with CL (CL+/PS-) and BA3B5C4 reacted with both CL and PS (CL+/PS+). 3SB9b reacted strongly with the syncytiotrophoblastic layer of both formalin fixed and frozen placental tissue. Sporadic reactivity was observed against the cytotrophoblastic layer. BA3B5C4 reacted strongly and specifically with cytotrophoblastic cells. D11A4 had only weak reactivity in the subtrophoblastic stromal region of the placenta in frozen sections. aPL staining was also observed against extravillous cytotrophoblast. BA3B5C4 stained cytoplasmic structures, whereas 3SB9b stained the plasma membrane region with little cytoplasmic staining. These data suggest that the trophoblastic layer is reactive with aPLs and may potentially be directly damaged through mechanisms unrelated to thrombosis. In addition, the trophoblastic layer directly in contact with the maternal circulation is most reactive with aPLs that are PS+ rather than CL+. The differential reactivity of 3SB9b and BA3B5C4 suggests that the antigenic conformation involving PS on the cytotrophoblast is altered concurrent with fusion into the syncytium.

Antibodies, Monoclonal↗
PubMed1990

Cytoplasmic dynein is localized to kinetochores during mitosis.

Recent evidence suggests that the force for poleward movement of chromosomes during mitosis is generated at or close to the kinetochores. Chromosome movement depends on motion relative to microtubules, but the identities of the motors remain uncertain. One candidate for a mitotic motor is dynein, a large multimeric enzyme which can move along microtubules toward their slow growing end. Dyneins were originally found in axonemes of cilia and flagella where they power microtubule sliding. Recently, cytoplasmic dyneins have also been found, and specific antibodies have been raised against them. The cellular localization of dynein has previously been studied with several antibodies raised against flagellar dynein, but the relevance of these data to the distribution of cytoplasmic dynein is not known. Antibodies raised against cytoplasmic dyneins have shown localization of dynein antigens to the mitotic spindles in Caenorhabditis elegans embryos (Lye et al., personal communication) and punctate cytoplasmic structures in Dictyostelium amoebae. Using antibodies that recognize subunits of cytoplasmic dyneins, we show here that during mitosis, cytoplasmic dynein antigens concentrate near the kinetochores, centrosomes and spindle fibres of HeLa and PtK1 cells, whereas at interphase they are distributed throughout the cytoplasm. This is consistent with the hypothesis that cytoplasmic dynein is a mitotic motor.

Adenosine Triphosphatases↗