Instability of pleomorphic tubulin paracrystals artificially induced by Vinca alkaloids in tissue-cultured cells

This review outlines the establishment, properties, and use of two lines of cultured Nb rat lymphoma cells. The cultured cells have retained important properties of the cancers of origin, such as dependency on prolactin for growth and a high sensitivity to antineoplastic Vinca alkaloids. The cultures have been useful for defining the hormonal dependency of the lymphomas in the animal and for studying the progression of the lymphomas from hormonal dependency to autonomy. A new, specific and highly sensitive in vitro bioassay for lactogenic hormones has been developed from one of the cultures. The use of the lymphoma cell cultures has revealed unsuspected pharmacological differences between the closely related, clinically useful antineoplastic Vinca alkaloids, vinblastine and vincristine. The lymphoma cell cultures are also useful tools for studying biochemical, cell cycle related events which follow the mitogenic stimulation of cells by a polypeptide growth factor.

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Immunoelectron microscopic localization of the 210,000-mol wt microtubule-associated protein in cultured cells of primates.

The Journal of Cell Biology ◽

10.1083/jcb.91.2.438 ◽

1981 ◽

Vol 91 (2) ◽

pp. 438-445 ◽

Cited By ~ 19

Author(s):

M De Brabander ◽

J C Bulinski ◽

G Geuens ◽

J De Mey ◽

G G Borisy

Keyword(s):

Molecular Weight ◽

Intermediate Filaments ◽

Sodium Azide ◽

Cultured Cells ◽

Entire Length ◽

Ultrastructural Immunocytochemistry ◽

Microscopic Localization ◽

Protein Map ◽

Tubulin Paracrystals

Results from ultrastructural immunocytochemistry on glutaraldehyde-fixed cells confirmed and extended findings previously obtained with immunofluorescence. A microtubule-associated protein (MAP) of 210,000 molecular weight was shown to be specifically associated with all cytoplasmic and mitotic microtubules along their entire length in primate cells. Specific labeling with the anti-MAP antibody could not be detected on any other subcellular structures, notably the centrosomes, kinetochores, microfilaments, and intermediate filaments. Treatment with the microtubule-disrupting drug, nocodazole, induced diffusion of the MAP throughout the cytoplasm. During repolymerization of microtubules following disassembly by nocodazole, the association of the MAP with the microtubules was intermediate and complete. When cells were treated with vinblastine, the tubulin paracrystals formed were heavily stained by the antibody. Neither sodium azide nor taxol affected the association of the MAP with microtubules.

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Ultrastructural Correlations between Clonal Human Tumor Colonies and in Vivo Neoplasms

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100053711 ◽

1982 ◽

Vol 40 ◽

pp. 210-211

Author(s):

M.J. Murphy ◽

R.R. Price ◽

J.C. Sloman

Keyword(s):

Cultured Cells ◽

Human Tumor ◽

Cell Type ◽

Tumor Mass ◽

Initial Cell ◽

Cloning Assay ◽

Human Tumor Cloning ◽

The Relationship

The in vitro human tumor cloning assay originally described by Salmon and Hamburger has been applied recently to the investigation of differential anti-tumor drug sensitivities over a broad range of human neoplasms. A major problem in the acceptance of this technique has been the question of the relationship between the cultured cells and the original patient tumor, i.e., whether the colonies that develop derive from the neoplasm or from some other cell type within the initial cell population. A study of the ultrastructural morphology of the cultured cells vs. patient tumor has therefore been undertaken to resolve this question. Direct correlation was assured by division of a common tumor mass at surgical resection, one biopsy being fixed for TEM studies, the second being rapidly transported to the laboratory for culture.

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Maytansine-Induced Mitotic Arrest in Human Lymphoblasts

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100079747 ◽

1977 ◽

Vol 35 ◽

pp. 460-461

Author(s):

Tai-Te Chao ◽

John Sullivan ◽

Awtar Krishan

Keyword(s):

Electron Microscopy ◽

Cell Cycle ◽

Transmission Electron Microscopy ◽

Tubulin Polymerization ◽

Vinca Alkaloids ◽

Antimitotic Activity ◽

Transmission Electron ◽

Flow Microfluorometry ◽

Brain Tubulin

Maytansine, a novel ansa macrolide (1), has potent anti-tumor and antimitotic activity (2, 3). It blocks cell cycle traverse in mitosis with resultant accumulation of metaphase cells (4). Inhibition of brain tubulin polymerization in vitro by maytansine has also been reported (3). The C-mitotic effect of this drug is similar to that of the well known Vinca- alkaloids, vinblastine and vincristine. This study was carried out to examine the effects of maytansine on the cell cycle traverse and the fine struc- I ture of human lymphoblasts.Log-phase cultures of CCRF-CEM human lymphoblasts were exposed to maytansine concentrations from 10-6 M to 10-10 M for 18 hrs. Aliquots of cells were removed for cell cycle analysis by flow microfluorometry (FMF) (5) and also processed for transmission electron microscopy (TEM). FMF analysis of cells treated with 10-8 M maytansine showed a reduction in the number of G1 cells and a corresponding build-up of cells with G2/M DNA content.

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Electron Probe Microanalysis of Frozen Hydrated Kidneys, Isolated Cells, and Cultured Cells

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100054595 ◽

1982 ◽

Vol 40 ◽

pp. 402-403 ◽

Cited By ~ 1

Author(s):

Claude Lechene

Keyword(s):

Liquid Nitrogen ◽

Electron Probe Microanalysis ◽

Electron Probe ◽

Cultured Cells ◽

Liquid Nitrogen Temperature ◽

Elemental Content ◽

Isolated Cells ◽

Renal Tubular Cells ◽

Diamond Saw ◽

Saw Blade

Electron probe microanalysis of frozen hydrated kidneysThe goal of the method is to measure on the same preparation the chemical elemental content of the renal luminal tubular fluid and of the surrounding renal tubular cells. The following method has been developed. Rat kidneys are quenched in solid nitrogen. They are trimmed under liquid nitrogen and mounted in a copper holder using a conductive medium. Under liquid nitrogen, a flat surface is exposed by sawing with a diamond saw blade at constant speed and constant pressure using a custom-built cryosaw. Transfer into the electron probe column (Cameca, MBX) is made using a simple transfer device maintaining the sample under liquid nitrogen in an interlock chamber mounted on the electron probe column. After the liquid nitrogen is evaporated by creating a vacuum, the sample is pushed into the special stage of the instrument. The sample is maintained at close to liquid nitrogen temperature by circulation of liquid nitrogen in the special stage.

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The Infection of Cells by Bullet-Shaped Viruses

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100063779 ◽

1969 ◽

Vol 27 ◽

pp. 372-373

Author(s):

Frederick A. Murphy ◽

Alyne K. Harrison ◽

Sylvia G. Whitfield

Keyword(s):

Electron Microscopy ◽

Physical Properties ◽

Host Cell ◽

Thin Section ◽

Cultured Cells ◽

Cell Infection ◽

Thin Section Electron Microscopy ◽

Section Electron ◽

Section Electron Microscopy

The bullet-shaped viruses are currently classified together on the basis of similarities in virion morphology and physical properties. Biologically and ecologically the member viruses are extremely diverse. In searching for further bases for making comparisons of these agents, the nature of host cell infection, both in vivo and in cultured cells, has been explored by thin-section electron microscopy.

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Ultrastructure of two neoplasms in culture compared with original biopsies

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100110623 ◽

1984 ◽

Vol 42 ◽

pp. 50-51

Author(s):

Joseph M. Harb ◽

James T. Casper ◽

Vlcki Piaskowski

Keyword(s):

Electron Microscopy ◽

Tissue Culture ◽

Biopsy Specimen ◽

Cultured Cells ◽

Light And Electron Microscopy ◽

Human Tumor ◽

Soft Agar ◽

Human Tumor Cells ◽

Morphological Distinction ◽

Tumor Types

The application of tissue culture and the newer methodologies of direct cloning and colony formation of human tumor cells in soft agar hold promise as valuable modalities for a variety of diagnostic studies, which include morphological distinction between tumor types by electron microscopy (EM). We present here two cases in which cells in culture expressed distinct morphological features not apparent in the original biopsy specimen. Evaluation of the original biopsies by light and electron microscopy indicated both neoplasms to be undifferentiated sarcomas. Colonies of cells propagated in soft agar displayed features of rhabdomyoblasts in one case, and cultured cells of the second biopsy expressed features of Ewing's sarcoma.

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Uultrastructure of Microtubules

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100094528 ◽

1972 ◽

Vol 30 ◽

pp. 252-253

Author(s):

Ariaki Nagayama

Keyword(s):

Fine Structure ◽

Cultured Cells ◽

Present Report ◽

Confluent Monolayer ◽

Purification Method ◽

Vinca Rosea ◽

L Cells ◽

Antimitotic Activity ◽

Monolayer Cultures ◽

Rapid Purification

Vinblastine(Vb) or vincristine, alkaloid derived from Vinca rosea is known for its antimitotic activity by regrouping of microtubules into paracrystalline form within the cells. A rapid purification method of vinblastine-induced microtubular paracrystals(PC) has provided us with a fresh and pure microtubular material demonstrating the presence of a labile ATPase associated with the PC. The present report is concerned with the fine structure of purified microtubules of mammalian cultured cells.Confluent monolayer cultures of L cells were incubated for 20hrs with 10-5 M Vb (donated from Shionogi Seiyaku & Co., Osaka, Japan).

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DNA nick end labeling: a reliable marker for apoptosis?

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100141184 ◽

1995 ◽

Vol 53 ◽

pp. 962-963

Author(s):

Anne F. Bushnell ◽

Sarah Webster ◽

Lynn S. Perlmutter

Keyword(s):

Cell Death ◽

Cultured Cells ◽

Apoptotic Cell ◽

Morphological Characteristics ◽

Detection Methods ◽

Tissue Preparation ◽

Preparation Methods ◽

Dna Laddering ◽

Disease States ◽

Reliable Marker

Apoptosis, or programmed cell death, is an important mechanism in development and in diverse disease states. The morphological characteristics of apoptosis were first identified using the electron microscope. Since then, DNA laddering on agarose gels was found to correlate well with apoptotic cell death in cultured cells of dissimilar origins. Recently numerous DNA nick end labeling methods have been developed in an attempt to visualize, at the light microscopic level, the apoptotic cells responsible for DNA laddering.The present studies were designed to compare various tissue processing techniques and staining methods to assess the occurrence of apoptosis in post mortem tissue from Alzheimer's diseased (AD) and control human brains by DNA nick end labeling methods. Three tissue preparation methods and two commercial DNA nick end labeling kits were evaluated: the Apoptag kit from Oncor and the Biotin-21 dUTP 3' end labeling kit from Clontech. The detection methods of the two kits differed in that the Oncor kit used digoxigenin dUTP and anti-digoxigenin-peroxidase and the Clontech used biotinylated dUTP and avidinperoxidase. Both used 3-3' diaminobenzidine (DAB) for final color development.

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Intralysosomal Accumulation of Colloidal Gold-Low Density Lipoprotein Conjugates in Chloroquine-Treated Fibroblasts

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010011951x ◽

1985 ◽

Vol 43 ◽

pp. 546-547 ◽

Cited By ~ 1

Author(s):

Dean A. Handley ◽

Cynthia M. Arbeeny ◽

Larry D. Witte

Keyword(s):

Colloidal Gold ◽

Cultured Cells ◽

Low Density Lipoprotein ◽

Density Lipoprotein ◽

Coated Vesicle ◽

Low Density ◽

Cholesterol Esters ◽

Cultured Fibroblasts ◽

Surface Receptors ◽

Ldl Particles

Low density lipoproteins (LDL) are the major cholesterol carrying particles in the blood. Using cultured cells, it has been shown that LDL particles interact with specific surface receptors and are internalized via a coated pit-coated vesicle pathway for lysosomal catabolism. This (Pathway has been visualized using LDL labeled to ferritin or colloidal gold. It is now recognized that certain lysomotropic agents, such as chloroquine, inhibit lysosomal enzymes that degrade protein and cholesterol esters. By interrupting cholesterol ester hydrolysis, chloroquine treatment results in lysosomal accumulation of cholesterol esters from internalized LDL. Using LDL conjugated to colloidal gold, we have examined the ultrastructural effects of chloroquine on lipoprotein uptake by normal cultured fibroblasts.

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