scholarly journals THE DISTRIBUTION OF SPINDLE MICROTUBULES DURING MITOSIS IN CULTURED HUMAN CELLS

1971 ◽  
Vol 49 (2) ◽  
pp. 468-497 ◽  
Author(s):  
J. Richard McIntosh ◽  
Story C. Landis
Keyword(s):  
Electron Microscopy ◽  
Hela Cells ◽  
Metaphase Plate ◽  
Human Cells ◽  
Late Anaphase ◽  
Monolayer Cultures ◽  
Spindle Axis ◽  
Early Anaphase ◽  
Cultured Human Cells ◽  
Spindle Microtubules

WI-38 and HeLa cells in mitosis have been selected from fixed monolayer cultures and serially sectioned for electron microscopy. Sections perpendicular to the spindle axis permit counting of the number of microtubules at each position on the spindle axis and hence the preparation of tubule distribution profiles. Errors intrinsic to this method are discussed. The changes in the tubule distributions from one mitotic stage to another provide evidence concerning the behavior of the spindle tubules during mitosis. The ratio of the number of tubules passing the chromosomes on the metaphase plate to the maximum number in each half spindle is about 1/2. This ratio changes little in early anaphase, and then decreases in late anaphase at about the same time that a zone of increased tubule number develops at the middle of the interzone. The region where the stem bodies form contains about 3/2 the number of tubules seen elsewhere in the interzone. This ratio is almost constant as the mid-body forms in telophase and then increases to 2/1 in early interphase before the final stages of cytokinesis occur.

scholarly journals ULTRASTRUCTURAL ANALYSIS OF MITOTIC SPINDLE ELONGATION IN MAMMALIAN CELLS IN VITRO

1971 ◽  
Vol 50 (2) ◽  
pp. 416-431 ◽  
Author(s):  
B. R. Brinkley ◽  
Joiner Cartwright
Keyword(s):  
Mitotic Spindle ◽  
Mammalian Cells ◽  
Chinese Hamster ◽  
Spindle Axis ◽  
Hamster Strain ◽  
Early Anaphase ◽  
Spindle Microtubules ◽  
Spindle Elongation ◽  
Elongation Process

The mitotic spindle of many mammalian cells undergoes an abrupt elongation at anaphase. In both cultured rat kangaroo (strain PtK1) and Chinese hamster (strain Don-C) fibroblasts, the distance from pole to pole at metaphase doubles during anaphase and telophase. In order to determine the organization and distribution of spindle microtubules during the elongation process, cells were fixed and flat embedded in Epon 812. Selected cells were photographed with the phase-contrast microscope and then serially sectioned perpendicular to the major spindle axis. Microtubule profiles were counted in selected sections, and the number was plotted with respect to position along the spindle axis. Interpretation of the distribution profiles indicated that not all interpolar microtubules extended from pole to pole. It is estimated that 55–70% of the interpolar microtubules are overlapped at the cell equator while 30–45% extend across the equator into both half spindles. This arrangement appeared to persist from early anaphase (before elongation) until telophase after the elongation process. Although sliding or shearing of microtubules may occur in the spindle, such appears not to be the mechanism by which the spindle elongates in anaphase. Instead, our data support the hypothesis that spindle elongation occurs by growth of prepositioned microtubules which "push" the poles apart.

Comparative study of hydrophilic and hydrophobic ionic liquids for observing cultured human cells by scanning electron microscopy

2011 ◽  
Vol 74 (12) ◽  
pp. 1104-1108 ◽  
Author(s):  
Yasuhito Ishigaki ◽  
Yuka Nakamura ◽  
Teruaki Takehara ◽  
Takayuki Kurihara ◽  
Hironori Koga ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Ionic Liquids ◽  
Comparative Study ◽  
Human Cells ◽  
Cultured Human Cells ◽  
Hydrophobic Ionic Liquids ◽  
Scanning Electron

scholarly journals Organization of spindle microtubules in Ochromonas danica.

1980 ◽  
Vol 87 (3) ◽  
pp. 531-545 ◽  
Author(s):  
D H Tippit ◽  
L Pillus ◽  
J Pickett-Heaps
Keyword(s):  
Average Length ◽  
Numerical Data ◽  
Tracking Errors ◽  
Mitotic Apparatus ◽  
Ochromonas Danica ◽  
Spindle Poles ◽  
Late Anaphase ◽  
Early Anaphase ◽  
Spindle Microtubules ◽  
Spindle Elongation

The entire framework of microtubules (MTs) in the mitotic apparatus of Ochromonas danica is reconstructed (except at the spindle poles) from transverse serial sections. Eleven spindles were sectioned and used for numerical data, but only four were reconstructed: a metaphase, an early anaphase, a late anaphase, and telophase. Four major classes of MTs are observed: (a) free MTs (MTs not attached to either pole); (b) interdigitated MTs (MTs attached to one pole which laterally associate with MTs from the opposite pole); (c) polar MTs (MTs attached to one pole); (d) kinetochore MTs (kMTs). Pole-to-pole MTs are rare and may be caused by tracking errors. During anaphase, the kMTs, free MTs, and polar MTs shorten until most disappear, while interdigitated MTs lengthen. In the four reconstructed spindles, the number of MTs decreases between early anaphase and telophase from 881 to 285, while their average length increases from 1.66 to 4.98 micron. The total length of all the MTs in the spindle (placed end to end) remains at 1.42 +/- 0.04 mm between these stages. At late anaphase and telophase the spindle is comprised mainly of groups of interdigitated MTs. Such MTs from opposite poles form a region of overlap in the middle of the spindle. During spindle elongation (separation of the poles), the length of the overlap region does not decrease. These results are compatible with theories that suggest that MTs directly provide the force that elongates the spindle, either by MT polymerization alone or by MT sliding with concomitant MT polymerization.

scholarly journals Biotin-tubulin incorporates into kinetochore fiber microtubules during early but not late anaphase.

1989 ◽  
Vol 109 (5) ◽  
pp. 2257-2265 ◽  
Author(s):  
P Wadsworth ◽  
E Shelden ◽  
G Rupp ◽  
C L Rieder
Keyword(s):  
Electron Microscopy ◽  
Dynamic Behavior ◽  
Electron Microscopic ◽  
Late Anaphase ◽  
Early Anaphase

The dynamic behavior of kinetochore fiber microtubules has been examined in PtK1 cells during anaphase of mitosis. Cells in anaphase were injected with biotin-tubulin and, at various intervals after injection, fixed for light or electron microscopic immunolocalization of biotin-tubulin-containing microtubules. When cells in early to mid anaphase were injected with biotin-tubulin and fixed 1-2 min later, fluorescence was observed throughout the spindle, including the region of the kinetochore fibers. Electron microscopy of early to mid anaphase cells, after processing with immunogold methods, revealed both labeled and unlabeled microtubules in the kinetochore fibers; some labeled microtubules contacted the kinetochores. When late anaphase cells were injected with biotin-tubulin, and fixed a few minutes later, little fluorescence was observed in the kinetochore fibers. Electron microscopy confirmed that kinetochore fibers in late anaphase cells were refractory to tubulin incorporation. The results of these experiments demonstrate that the kinetochore fiber incorporates new microtubules during early anaphase but that this incorporation ceases in mid to late anaphase. Thus, microtubule turnover within the kinetochore fiber does not abruptly cease at the onset of anaphase and anaphase kinetochore fiber microtubules are more dynamic than previously suspected.

scholarly journals NITROUS OXIDE: EFFECTS ON THE MITOTIC APPARATUS AND CHROMOSOME MOVEMENT IN HELA CELLS

1973 ◽  
Vol 58 (1) ◽  
pp. 96-106 ◽  
Author(s):  
B. R. Brinkley ◽  
Potu N. Rao
Keyword(s):  
Nitrous Oxide ◽  
Hela Cells ◽  
Metaphase Plate ◽  
Microtubule Assembly ◽  
Chromosome Movement ◽  
Spindle Microtubule ◽  
Mitotic Apparatus ◽  
Ultrastructural Studies ◽  
Multipolar Spindles ◽  
Spindle Microtubules

When HeLa cells were grown in the presence of nitrous oxide (N2O) under pressure (80 lb/in2) mitosis was inhibited and the chromosomes displayed a typical colchicine metaphase (c-metaphase) configuration when examined by light microscopy. When the cells were returned to a 37°C incubator, mitosis was resumed and the cells entered G1 synchronously. Ultrastructural studies of N2O-blocked cells revealed a bipolar spindle with centriole pairs at each pole. Both chromosomal and interpolar (pole-to-pole) microtubules were also present. Thus, N2O, unlike most c-mitotic agents, appeared to have little or no effect upon spindle microtubule assembly. However, the failure of chromo somes to become properly aligned onto the metaphase plate indicated an impairment in normal prometaphase movement. The alignment of spindle microtubules was frequently atypical with some chromosomal microtubules extending from kinetochores to the poles, while others extended out at acute angles from the spindle axis. These ultrastructural studies indicated that N2O blocked cells at a stage in mitosis more advanced than that produced by Colcemid or other c-mitotic agents. Like Colcemid, however, prolonged arrest in mitosis with N2O led to an increased incidence of multipolar spindles.

scholarly journals Differences in Virus-Induced Cell Morphology and in Virus Maturation between MVA and Other Strains (WR, Ankara, and NYCBH) of Vaccinia Virus in Infected Human Cells

2003 ◽  
Vol 77 (19) ◽  
pp. 10606-10622 ◽  
Author(s):  
Juan Carlos Gallego-Gómez ◽  
Cristina Risco ◽  
Dolores Rodríguez ◽  
Pilar Cabezas ◽  
Susana Guerra ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
New York ◽  
Vaccinia Virus ◽  
Hela Cells ◽  
Morphological Changes ◽  
Human Cells ◽  
The Other ◽  
Transmission Electron Microscopy Analysis ◽  
Modified Vaccinia Virus Ankara ◽  
Electron Microscopy Analysis

ABSTRACT Live recombinants based on attenuated modified vaccinia virus Ankara (MVA) are potential vaccine candidates against a broad spectrum of diseases and tumors. To better understand the efficacy of MVA as a human vaccine, we analyzed by confocal and electron microscopy approaches MVA-induced morphological changes and morphogenetic stages during infection of human HeLa cells in comparison to other strains of vaccinia virus (VV): the wild-type Western Reserve (WR), Ankara, and the New York City Board of Health (NYCBH) strains. Confocal microscopy studies revealed that MVA infection alters the cytoskeleton producing elongated cells (bipolar), which do not form the characteristic actin tails. Few virions are detected in the projections connecting neighboring cells. In contrast, cells infected with the WR, Ankara, and NYCBH strains exhibit a stellated (multipolar) or rounded morphology with actin tails. A detailed transmission electron microscopy analysis of HeLa cells infected with MVA showed important differences in fine ultrastructure and amounts of the viral intermediates compared to cells infected with the other VV strains. In HeLa cells infected with MVA, the most abundant viral forms are intracellular immature virus, with few intermediates reaching the intracellular mature virus (IMV) form, at various stages of maturation, which exhibit a more rounded shape than IMVs from cells infected with the other VV strains. The “IMVs” from MVA-infected cells have an abnormal internal structure (“atypical” viruses) with potential alterations in the core-envelope interactions and are unable to significantly acquire the additional double envelope to render intracellular envelope virus. The presence of potential cell-associated envelope virus is very scarce. Our findings revealed that MVA in human cells promotes characteristic morphological changes to the cells and is able to reach the IMV stage, but these virions were not structurally normal and the subsequent steps in the morphogenetic pathway are blocked.

scholarly journals Mitochondrial nucleoid morphology and respiratory function are altered in Drp1-deficient HeLa cells

2019 ◽  
Vol 167 (3) ◽  
pp. 287-294 ◽  
Author(s):  
Azusa Ota ◽  
Takaya Ishihara ◽  
Naotada Ishihara
Keyword(s):  
Hela Cells ◽  
Copy Number ◽  
Respiratory Function ◽  
Respiratory Activity ◽  
Mitochondrial Fission ◽  
Fluorescent Microscopy ◽  
Human Cells ◽  
Mtdna Copy Number ◽  
Consumption Rates ◽  
Cultured Human Cells

Abstract Mitochondria are dynamic organelles that frequently divide and fuse with each other. The dynamin-related GTPase protein Drp1 has a key role in mitochondrial fission. To analyse the physiological roles of Drp1 in cultured human cells, we analysed Drp1-deficient HeLa cells established by genome editing using CRISPR/Cas9. Under fluorescent microscopy, not only mitochondria were elongated but their DNA (mtDNA) nucleoids were extremely enlarged in bulb-like mitochondrial structures (‘mito-bulbs’) in the Drp1-deficient HeLa cells. We further found that respiratory activity, as measured by oxygen consumption rates, was severely repressed in Drp1-deficient HeLa cells and that this was reversible by the co-repression of mitochondrial fusion factors. Although mtDNA copy number was not affected, several respiratory subunits were repressed in Drp1-deficient HeLa cells. These results suggest that mitochondrial fission is required for the maintenance of active respiratory activity and the morphology of mtDNA nucleoids in human cells.

scholarly journals Interpolar spindle microtubules in PTK cells.

1993 ◽  
Vol 123 (6) ◽  
pp. 1475-1489 ◽  
Author(s):  
D N Mastronarde ◽  
K L McDonald ◽  
R Ding ◽  
J R McIntosh
Keyword(s):  
Electron Microscopy ◽  
Cross Section ◽  
Specific Interaction ◽  
Spindle Pole ◽  
Serial Sections ◽  
Spindle Poles ◽  
Late Anaphase ◽  
Computer Image Processing ◽  
Spindle Microtubules ◽  
Anaphase B

Spindle microtubules (MTs) in PtK1 cells, fixed at stages from metaphase to telophase, have been reconstructed using serial sections, electron microscopy, and computer image processing. We have studied the class of MTs that form an interdigitating system connecting the two spindle poles (interpolar MTs or ipMTs) and their relationship to the spindle MTs that attach to kinetochores (kMTs). Viewed in cross section, the ipMTs cluster with antiparallel near neighbors throughout mitosis; this bundling becomes much more pronounced as anaphase proceeds. While the minus ends of most kMTs are near the poles, those of the ipMTs are spread over half of the spindle length, with at least 50% lying > 1.5 microns from the poles. Longitudinal views of the ipMT bundles demonstrate a major rearrangement of their plus ends between mid- and late anaphase B. However, the minus ends of these MTs do not move appreciably farther from the spindle midplane, suggesting that sliding of these MTs contributes little to anaphase B. The minus ends of ipMTs are markedly clustered in the bundles of kMTs throughout anaphase A. These ends lie close to kMTs much more frequently than would be expected by chance, suggesting a specific interaction. As sister kinetochores separate and kMTs shorten, the minus ends of the kMTs remain associated with the spindle poles, but the minus ends of many ipMTs are released from the kMT bundles, allowing the spindle pole and the kMTs to move away from the ipMTs as the spindle elongates.

scholarly journals Nucleoplasmic organization of small nuclear ribonucleoproteins in cultured human cells.

1993 ◽  
Vol 121 (4) ◽  
pp. 715-727 ◽  
Author(s):  
A G Matera ◽  
D C Ward
Keyword(s):  
Hela Cells ◽  
Human Cells ◽  
Complete Agreement ◽  
Antisense Probe ◽  
Coiled Bodies ◽  
Highly Sensitive ◽  
U2 Snrnp ◽  
Cultured Human Cells ◽  
Small Nuclear Ribonucleoproteins

The organization of eight small nuclear ribonucleoproteins (the U1, U2, U4, U5, and U6 RNAs previously studied by others and three additional snRNAs, U11, U12, and 7SK) has been investigated in cultured human cells by fluorescence in situ hybridization with antisense DNA and 2'-O-Me RNA oligonucleotides. Using highly sensitive digital imaging microscopy we demonstrate that all of these snRNAs are widespread throughout the nucleoplasm, but they are excluded from the nucleoli. In addition, the U2, U4, U5, U6, and U12 snRNAs are concentrated in discrete nuclear foci, known as coiled bodies, but U1 and 7SK are not. In addition to coiled bodies, a classic speckled pattern was observed in the nucleoplasm of monolayer-grown HeLa cells, whereas suspension-grown HeLa cells revealed a more diffuse nucleoplasmic labeling. Immunofluorescence staining using various snRNP-specific antisera shows complete agreement with that of their antisense snRNA oligonucleotide counterparts. Although U2 RNA is concentrated in coiled bodies, quantitation of the fluorescence signals from the U2 antisense probe reveals that the bulk of the U2 snRNP is located in the nucleoplasm. Furthermore, simultaneous visualization of the U2 snRNAs and the tandemly repeated U2 genes demonstrates that coiled bodies are not the sites of U2 transcription.

Platinum Compounds as Stains for Electron Microscopy

1974 ◽  
Vol 32 ◽  
pp. 230-231
Author(s):  
S. K. Aggarwal ◽  
P. McAllister ◽  
R. W. Wagner ◽  
B. Rosenberg
Keyword(s):  
Electron Microscopy ◽  
Hela Cells ◽  
Uranyl Acetate ◽  
Sarcoma 180 ◽  
Platinum Compounds ◽  
Kb Cells ◽  
En Bloc ◽  
Human Lymphoma ◽  
Ultrathin Sections ◽  
Blue Coloration

Uranyl acetate has been used as an electron stain for en bloc staining as well as for staining ultrathin sections in conjunction with various lead stains (Fig. 1). Present studies reveal that various platinum compounds also show promise as electron stains. Certain platinum compounds have been shown to be effective anti-tumor agents. Of particular interest are the compounds with either uracil or thymine as one of the ligands (cis-Pt(II)-uracil; cis-Pt(II)-thymine). These compounds are amorphous, highly soluble in water and often exhibit an intense blue coloration. These compounds show enough electron density to be used as stains for electron microscopy. Most of the studies are based on various cell lines (human AV, cells, human lymphoma cells, KB cells, Sarcoma-180 ascites cells, chick fibroblasts and HeLa cells) while studies on tissue blocks are in progress.

Sign in / Sign up

Export Citation Format

Share Document