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At least 1,099 records · Page 61Linked to original sources
PubMed1998

Diisopropylglutathione ester protects A549 cells from the cytotoxic effects of sulphur mustard.

The A549 cell line was used to assess the ability of diisopropylglutathione (DIPE) to protect against a 100 microM challenge dose of sulphur mustard (HD) using gentian violet (GV), thiazolyl blue (MTT) and neutral red (NR) assays as indicators of cell culture viability. As part of a continuing study of the efficacy of protective nucleophiles as candidate treatments for HD poisoning, several different combinations of protectant and HD were used to determine the optimal means of protecting A549 cells from the effects of HD. It was found that DIPE (4 mM) could protect cells against the effects of HD though for optimal effect, DIPE had to be present at the time of HD challenge. Cultures protected with DIPE were up to 2.9-fold more viable than HD exposed cells 48 h after HD challenge when using the GV, MTT and NR assays to assess viability. Observations by phase contrast microscopy of GV stained cultures confirmed these findings. Pretreating A549 cultures with DIPE for 1 h followed by its removal prior to HD challenge did maintain cell viability, though at a relatively low level (only up to 1.4-fold more viable than HD only exposed cells). DIPE was also able to protect HD exposed A549 cultures when added to cell cultures at intervals of up to 12 to 15 min after the initial HD exposure, though viability tended to decrease over this period, so that at 1 h, addition of DIPE did not maintain the viability of the cultures. This is the first such report of the anti-HD protectant properties of DIPE in A549 cells. It is concluded that the protection observed against HD is probably largely due to extracellular inactivation of HD by DIPE.

Cell Line↗
PubMed2005

Comparison of urine contamination rates using three different methods of collection: clean-catch, cotton wool pad and urine bag.

Collecting uncontaminated urine specimens from infants is difficult. Commonly, an adhesive urinecollecting bag is used, which is uncomfortable. This study determined bacterial contamination rates using three methods of urine collection sequentially on the same infant (without known urinary tract infection)-clean-catch, cotton wool (sanitary) pad and urine bag. The study was undertaken in children under 3 years of age in the Institute of Maternal and Child Health of Pernambuco (IMIP), Recife, Brazil. Urine samples were analysed using phase contrast microscopy and routine culture. Culture of bacteria at any level was interpreted as a contaminated urine specimen. Cultures with > 10(5) colony-forming units/ml of one species by all three collection methods were regarded as true urinary tract infection and these children were excluded. Altogether, 534 urine samples from 191 patients were analysed. Median age was 2 months (1 day-36 months) and 124 (65%) were boys. Twelve children (6.3%) were considered to have true urinary tract infection, three were indeterminate and in 16 one or more samples were missing and all were excluded from analysis. There were more missing samples using the clean-catch method (12%) than when using the bag (4%) or pad (4%). Seventy-six of 160 (47.5%) children had evidence of bacterial contamination. Clean-catch specimens showed the least contamination (14.7%) and rates were similar between pads (29%) and bags (26.6%) (kappa = 0.40). Urine contamination rates were similar for sanitary pads and urine bags and significantly higher than for clean-catch (p<0.01). However, pads were a simple, non-invasive and comfortable alternative to bags.

Child, Preschool↗
PubMed1986

In vitro degradation of bone particles by human monocytes is decreased with the depletion of the vitamin K-dependent bone protein from the matrix.

The bone vitamin K-dependent protein osteocalcin has been suggested to play a role in bone resorption. By administering sodium warfarin to rats, it is possible to inhibit the vitamin K-dependent addition of the their gamma-carboxyglutamic acid residues to osteocalcin. This results in reduced amounts of osteocalcin bone, probably because devoid of the calcium-binding Gla residues, the protein no longer accumulates in bone. Preparations of bone obtained from rats treated with sodium warfarin for 6 weeks contained only 0.2% of normal levels of osteocalcin and were 90% reduced in the concentration of Gla. This bone could not be degraded by human monocytes in vitro as well as control bone (only 54% of control; P less than 0.003). Defects in the movement of cells to the bone were documented by phase contrast microscopy. Only 60% as many monocytes attached to the osteocalcin-depleted bone as to control bone in an in vitro attachment assay. These effects do not appear to be related to direct cellular toxicity. The degradation of bone in this in vitro system appears to be dependent on the osteocalcin content in matrix. This may result from defective movement of cells to bone and/or attachment to the bone.

Animals↗
PubMed1993

Absence of triglyceride accumulation in lipoprotein lipase-deficient human monocyte-macrophages incubated with human very low density lipoprotein.

Lipoprotein lipase, a lipolytic enzyme essential for normal hydrolysis of triglycerides in very low density lipoprotein (VLDL) and chylomicrons, is found in several cell types, including macrophages. The role of lipoprotein lipase in mediating the uptake of normal VLDL triglycerides into human cultured monocyte-derived macrophages was studied using macrophage cells from a functionally lipoprotein lipase-deficient patient and macrophages of cells from a normal subject. After incubation with VLDL, massive accumulation of phase refractile (lipid) inclusions were noted by phase contrast microscopy within the normal, but not within the lipoprotein lipase-deficient, macrophages. Chemical determinations of intracellular lipid confirmed massive triglyceride accumulation within normal macrophages, but not in lipoprotein lipase-deficient macrophages. VLDL-derived cholesterol did not accumulate in either cell. These results confirm an additional role of lipoprotein lipase, that of mediating triglyceride accumulation into macrophages from normal human VLDL. Human monocyte-macrophages genetically deficient in a functional lipoprotein lipase will be useful to determine the role of lipoprotein lipase in macrophage accumulation of lipid from other forms of triglyceride-carrying lipoproteins, including hypertriglyceridemic VLDL, beta-VLDL, and chylomicrons.

Adult↗
PubMed1995

Prostaglandin E2 alters human orbital fibroblast shape through a mechanism involving the generation of cyclic adenosine monophosphate.

Orbital fibroblasts from patients with Graves' ophthalmopathy, when treated with prostaglandin E2 (PGE2), become stellate and develop prominent cellular processes. In this paper, we describe results of studies designed to characterize the action of PGE2 on orbital fibroblast shape changes in vitro. Orbital and dermal fibroblasts were incubated with PGE2, one of several prostanoid analogues, 8-br-cAMP or forskolin and were then visualized by phase-contrast microscopy. Other studies involved seeding cells in special chambers equipped with electrodes for cell sensing using electric cell-substrate impedance sensing (ECIS) to detect changes in shape. PGE2 (10(-7) mol/L) elicited a rapid and dramatic alteration in the shape of orbital fibroblasts but not those derived from the skin. Cells became stellate and developed prominent cytoplasmic processes that extended out from the central area containing the cell nucleus. The effects were stereoselective in that a number of structurally related compounds, including Sulprostone, PGI2, PGF2 alpha, thromboxane A2, thromboxane B2, and 11 deoxy,16,16 dimethyl PGE2 failed to elicit a similar shape change. Butaprost (10(-5) mol/L), a specific EP2 agonist, elicited a similar shape-change as that observed with PGE2. 16,16-dimethyl PGE2, a nonselective agonist, could mimic the action of PGE2. The effect of PGE2 was apparent at 10(-8) mol/L, maximal at a concentration of 10(-7) mol/L and took 4-8 hr to evolve completely. Cycloheximide (10 micrograms/mL) and actinomycin D (1 micrograms/mL) failed to block the shape change. The morphologic change could be reproduced by addition of 8-br-cAMP (3 mmol/L) and by forskolin (5 mumol/L). Moreover, PGE2 and Butaprost treatment elicited in orbital cultures a massive increase in endogenous cAMP production while analogues not affecting cell shape failed to influence cyclic nucleotide generation. Three strains of orbital fibroblasts from patients with Graves' ophthalmopathy and three from normal orbits were tested and all responded to PGE2 (10(-7) mol/L). Four strains of dermal fibroblasts failed to respond to PGE2. The changes in orbital fibroblast morphology were accompanied by a marked decrease in monolayer impedance as assessed by electric cell-substrate impedance sensing. The earliest effects were apparent within 30 min using this sensitive technique. The widely recognized roles of PGE2 and related compounds in the mediation of the inflammatory response make our current findings in orbital fibroblasts of potential importance to the pathogenesis of Graves' ophthalmopathy.

Cells, Cultured↗
PubMed1992

Embryonic stem cell-derived cystic embryoid bodies form vascular channels: an in vitro model of blood vessel development.

Murine embryonic stem cells can differentiate in vitro to form cystic embryoid bodies (CEB) that contain different structures and cell types. The blood islands are one such structure that consist of immature hematopoietic cells surrounded by endothelial cells, the first identifiable vascular cells. CEBs differentiated in vitro developed blood islands initially, and subsequently these blood islands matured to form vascular channels containing hematopoietic cells. Phase contrast microscopy demonstrated the presence of channels in mature CEBs grown in suspension culture, and high resolution light and electron microscopy showed that the cells lining these channels were endothelial cells. The channels appeared less organized than the vasculature of the mature yolk sac. The hematopoietic cells were occasionally seen 'flowing' through the CEB channels, although their numbers were reduced relative to the yolk sac. Analysis of primary CEB cultures showed the presence of cells with two characteristics of endothelial cells: approximately 30% of the cells labelled with fluorescent acetylated low density lipoprotein and a small number of cells were positive for von Willebrand's factor by immunostaining. Thus we conclude that a primitive vasculature forms in CEBs differentiated in vitro, and that not only primary differentiation of endothelial cells but also some aspects of vascular maturation are intrinsic to this cell culture system. CEBs are therefore a useful model for the study of developmental blood vessel formation.

Animals↗
PubMed1992

Wound closure in foetal rat skin.

Foetal rat skin rapidly closes an open wound in organ culture and in vivo, this possibly being unique to organs still in the morphogenetic stage. In the present study, examination was made of morphological changes in foetal rat skin during closure of open wounds inflicted at day 16 of gestation. Phase-contrast microscopy of open-wounded skin cultured in vitro indicated inward spreading of the peripheral skin to be responsible for wound closure. Wound closure in vitro was inhibited by cytochalasin B (10 micrograms/ml), not by hydroxyurea (2 mM), indicating prenatal wound closure to be mediated by regulation of the microfilament system rather than cell proliferation. During wound closure in vitro and in vivo, light and scanning electron microscopy of the peripheral skin showed cells in the periderm, the outermost layer of the foetal epidermis, to elongate centripetally and en masse, whereas the shape of underlying epidermal cells not to change. Numerous spindle-shaped cells and fibrous matrices in the mesenchyme were redistributed, becoming oriented along the wound edge. Following isolation of the mesenchyme and epidermis by treatment with Dispase and separate culturing, the capacity for wound closure in vitro was found to be retained only by the mesenchyme. Cellular activity within the mesenchyme, rather than in the epidermis, would thus appear essential to wound closure in foetal rat.

Animals↗
PubMed1985

Rhizobium attachment to clover roots.

The adhesion of rhizobia to surfaces of clover roots was examined by an indirect plate-counting assay and phase-contrast microscopy. The number of Rhizobium trifolii cells attached to clover root segments increased in approximately linear fashion during the first hour of incubation, but did not change appreciably thereafter. The addition of 30 mM-2-deoxy-D-glucose, which effectively inhibits binding of clover root lectin, did not promote the release of previously attached bacteria nor inhibit subsequent attachment to either root segments or root hairs. Rhizobia of several heterologous species attached to clover roots in numbers comparable to those of strains of R. trifolii, the homologous species. These results indicate that rhizobia have effective mechanisms of adhesion to non-host roots and that clover lectin contributed little or nothing to attachment under the conditions examined.

Cell Adhesion↗
PubMed1978

Intracellular motility of mitochondria: role of the inner compartment in migration and shape changes of mitochondria in XTH-cells.

Mitochondrial movements have been followed by phase-contrast microscopy in living XTH-cells (Xenopus laevis tadpole-heart cells) in tissue culture. The same organelles have been viewed subsequently in electron micrographs. Locomotion of mitochondria proceeds at velocities up to 100 micrometer/min. Formation of branches of mitochondria and other shape changes may occur with the same speed. Mitochondrial motility can be classified into 4 types: (I) Alternating extension and contraction at the two ends of rod-shaped mitochondria. (2) Lateral branching. (3) Alternate stretching and contraction of arbitrary parts of a mitochondrion amounting to a kind of peristaltic action. (4) Transverse wave propagation along the organelle. Types I to 3 can be reduced to the same underlying principle; they cause locomotion. Formation of mitochondrial extensions is due to elongation of cristae. The observations are discussed in terms of 4 specific proposals. (I) Intracellular locomotion of mitochondria is caused by local enlargements and contractions of the organelles. (2) The shape changes are correlated with alterations in the arrangement of the cristae. (3) Such arrangements are not associated with overall swelling or shrinkage of the mitochondrion; they are local features. (4) Estimates of the time required for rearrangement of the inner compartment amount to less than 0.3 s for single crista arrangements during the fastest shape changes, and less than 1-3 s during slower alterations. This high velocity is in good accord with the hypothesis of energy conservation by conformational events during oxidative phosphorylation.

Animals↗
PubMed1983

Behaviour and structure of the leading lamella in moving fibroblasts. I. Occurrence and centripetal movement of arc-shaped microfilament bundles beneath the dorsal cell surface.

Arc-shaped bundles of microfilaments are frequently found beneath the dorsal surface of the leading lamella of chick embryo fibroblasts. These structures, called arcs, form parallel to, and 2-10 micron from the leading edge of the lamella and they then move centripetally through the lamella and disappear in front of the cell nucleus. Arcs move centripetally at a mean speed of 1.33 (+/- 0.08 S.E.) micron min-1 relative to the substratum. Arcs are specifically associated with cells that are actively protrusive. They are common in fanshaped fibroblasts and in those cells that are newly spreading on a substratum. Arcs are absent from fibroblasts that have only small lamellae or that are polygonal. The correlation between are formation and protrusive activity is also clearly shown by the locomotory behaviour of fan-shaped cells. When protrusion of the lamella is increased by tail detachment, fibroblasts often develop numerous arcs, which chase each other backwards through the lamella. Conversely, arc formation ceases during contact inhibition of locomotion. In cells with large convex-edged lamellae there is a dorsal submembraneous sheath of microfilaments. All of the filaments comprising the sheath are oriented parallel to the margin of the lamella. Arcs are regions of the sheath where the microfilaments are more densely packed and hence visible by phase-contrast microscopy. There is no obvious relationship between the dorsally situated arcs and microfilament sheath, and the stress fibres that are associated with the ventral cell surface. The similarities between the movement and behaviour of arcs and the centripetal transport of particles on the surface of the lamella suggest a role for the microfilament sheath in the movement of particles.

Animals↗
PubMed1985

Mutual desmosome formation between all binary combinations of human, bovine, canine, avian and amphibian cells: desmosome formation is not tissue- or species-specific.

Our previous work has suggested that the molecular components of desmosomes are highly conserved between different tissues and different vertebrate species. In order to determine whether the adhesion recognition mechanism of desmosomes is also conserved we have examined the specificity of desmosome formation between different epithelial cell types by co-culturing binary combinations of cells from different species and from epidermal and non-epidermal origin. The following cell types were used: human (HeLa, cervical carcinoma), bovine (Madin Darby bovine kidney, MDBK), canine (Madin Darby canine kidney, MDCK), avian (chick embryonic corneal epithelium) and amphibian (Rana pipiens, adult corneal epithelium). Different cells in co-culture were identified on the basis of at least one of the following criteria: (1) morphology by phase-contrast microscopy; (2) presence or absence of staining of cytokeratin with monoclonal antibody LE61; (3) morphology at the electron microscope level. Mutual desmosome formation between different cell types was assessed using fluorescent antibody staining with anti-desmoplakin antibodies and confirmed using electron microscopy. We have found that mutual desmosome formation occurred between all binary combinations of human, bovine, canine, avian and amphibian cells. Thus there is complete non-selectivity of desmosome formation between five different epithelial cell types from three vertebrate classes. Our results suggest that desmosome formation is not tissue- or species-specific and that the mechanism for intercellular binding involved in desmosomal adhesion is highly conserved.

Animals↗
PubMed1988

Distribution of pericentriolar material in multipolar spindles induced by colcemid treatment in Chinese hamster ovary cells.

Mitotic Chinese hamster ovary cells were obtained by treatment with microtubule drugs under various conditions, and the shape of spindles was analysed by phase-contrast microscopy of isolated spindles, and by indirect immunofluorescence staining of whole mitotic cells with anti-tubulin antibody. Bipolarity of spindles was maintained after treatment with 0.05 microM of colcemid for 3.5 h, but increased exposure to higher concentrations (0.32 microM) and for longer durations (5.5 h) led to a marked rise in multipolar spindles. Nocodazole treatment, on the other hand, failed to show a multiplicity of spindle poles even at 3.3 microM. Each pole of a multipolar spindle was associated with pericentriolar material, as shown by staining with an autoimmune serum specific for pericentriolar material. The number of locations with free pericentriolar material capable of polymerizing microtubules in vitro also increased with increasing numbers of spindle poles, suggesting that dispersion of the pericentriolar material resulted in the production of many microtubule-nucleating sites in multipolar spindles. The different efficiencies of recovery from different drugs, which have been known to be quite variable, may be partly due to the different extent of dispersion of the pericentriolar material.

Animals↗
PubMed1994

A simple method for the assessment of macrophage scavenger receptor-ligand interaction: adherence of erythrocytes coated with oxidized low density lipoprotein and modified albumin to macrophages.

A simple method is described for the assessment of the binding of macrophage scavenger receptor and its ligands such as oxidized low density lipoprotein (ox-LDL) and maleylated bovine serum albumin (m-BSA). In this method, the binding of ox-LDL or m-BSA to macrophages is observed as the adherence of erythrocytes precoated with ox-LDL or m-BSA under phase-contrast microscopy. Erythrocytes were coated with native LDL or ox-LDL simply by incubating mouse erythrocytes with LDL. The ox-LDL-coated erythrocytes attached to the monolayer of mouse peritoneal macrophages without phagocytosis at 37 degrees C, while native LDL-coated erythrocytes did not. The extent of the adherence of the ox-LDL-coated erythrocytes to the macrophages was conveniently expressed as the proportion of the macrophages binding the coated erythrocytes. An optimal concentration of ox-LDL for erythrocyte coating giving maximum erythrocyte adherence to macrophages was 10 micrograms/ml as protein at an erythrocyte concentration of 2%. The binding of the ox-LDL-coated erythrocytes to the macrophages could be inhibited by ligands for scavenger receptor, and reflected the extent of LDL oxidation. Mouse erythrocytes were successfully coated with m-BSA using chromium(III) ion as adsorbent. The m-BSA-coated erythrocytes attached to the macrophages, while native BSA-coated erythrocytes did not. The adherence was inhibitable with ligands for scavenger receptor, and reflected the extent of maleylation. The method would be useful particularly for measurement of the ligand-binding activity of the receptor, and the receptor-binding activity of the ligands.

Absorption↗
PubMed1986

Development of cultured cardiac cells and difference of their structures in vivo.

Some interesting features in the morphological study of rat cardiac cells were obtained by use of the light and phase-contrast microscopy. The histological approach and comparison of the ultrastructures of the cultured cardiac cells with uncultured cells were included in the studies as a control. Four main points are discussed: the process of single beating cells that eventually constitutes the cell cluster and network, the function and histological content of the network, the mechanisms of synchronized beating cells, the substance of the cell surface coat and intercellular connection.

Animals↗
PubMed1992

[Concentration and form of asbestos fibers in tap drinking water contaminated from a water supply pipe with asbestos-cement].

The identification and concentration of asbestos fibers in tap drinking water supplied in a central area of Akita Prefecture, Japan, were determined by phase-contrast microscopy and a scanning electron microscope equipped with an energy-dispersive X-ray microanalyzer. The following results were obtained. 1. Asbestos fibers were found in the tap water from two areas in which an asbestos-cement pipe was used for public water supply. The concentrations of asbestos fibers in the tap water were 2.7 x 10(4) to 27.0 x 10(4) fibers per liter of water in area A and 10.0 x 10(4) to 21.0 x 10(4) in area B. On the other hand, no asbestos fiber contamination was observed in tap water of area C, which shared a common water source with area A. A vinyl chloride pipe was used over the entire length of the water supply in route C. 2. Crocidolite was the predominant type of asbestos fiber detected in the tap water. Chrysotile and a mixture of chrysotile and amosite were also observed. 3. Almost all asbestos fibers detected in the tap water possessed the form of thick or sheaved fibers with lengths ranging from ca. 5 to 10 microns. Their shapes were very different from those of asbestos fibers found in the atmosphere. The typical form of the latter is short (ca. 1 micron in length) and needle-like. 4. It was suggested that the contamination of asbestos fibers in the tap water was caused by erosion and peeling off of the inner wall of the asbestos-cement pipe used as a conduit. In order to evaluate the safety of drinking water in Japan, an extensive survey on asbestos-fiber contamination in tap water is necessary.

Asbestos↗
PubMed1990

Tissue culture of feline normal mammary gland.

To study the growth and morphology of feline mammary epithelial cells in vitro, normal feline mammary tissues as well as cells collected from feline milk were cultured and examined using phase-contrast microscopy and immunohistochemistry. The method employed was a modification of Hiratsuka et al., and was found to be useful for the culture of dissociated feline mammary tissue. Small polygonal cells and large spindle-shaped cells that were positively stained with anti-keratin and anti-actin antibodies formed colonies with a pavement-like structure in culture of mid-pregnant mammary gland. Cultures of involuting mammary gland were composed mainly of small spindle-shaped cells, which were negative for keratin immunostaining and thought to be fibroblasts. Myofibroblasts, which participate in feline mammary carcinogenesis, and show keratin negative, actin positive, were not encountered in the present culture system of normal mammary gland. The cells from milk formed small colonies, but did not grow to reach confluent monolayer.

Animals↗
PubMed2005

Glioma expansion in collagen I matrices: analyzing collagen concentration-dependent growth and motility patterns.

We study the growth and invasion of glioblastoma multiforme (GBM) in three-dimensional collagen I matrices of varying collagen concentration. Phase-contrast microscopy studies of the entire GBM system show that invasiveness at early times is limited by available collagen fibers. At early times, high collagen concentration correlates with more effective invasion. Conversely, high collagen concentration correlates with inhibition in the growth of the central portion of GBM, the multicellular tumor spheroid. Analysis of confocal reflectance images of the collagen matrices quantifies how the collagen matrices differ as a function of concentration. Studying invasion on the length scale of individual invading cells with a combination of confocal and coherent anti-Stokes Raman scattering microscopy reveals that the invasive GBM cells rely heavily on cell-matrix interactions during invasion and remodeling.

Cell Line, Tumor↗