Mitotic apparatus

The in vivo micronucleus test is used for the detection of damage to chromosomes as well as the mitotic apparatus in bone marrow or peripheral blood cells of rodents. The assay system has been well standardized.14-17 The basic features of the test system are (1) the effect of the test chemical is observed in anucleated polychromatic erythrocytes (PCEs) (2) PCEs have a relatively short lifespan, so that any micronuclei they contain must have been generated as a result of recently induced chromosome damage (3) micronuclei are readily identifiable and their distribution is well defined and (4) the frequency of induced micronuclei in PCEs is dependent on sampling times. 

Until the past decade, the cytoplasm was widely considered to be structurally unorganized with the main division of labor at the organellar level. Certainly, relatively little was known about the nature of the cyto-skeleton (with the notable exception of the mitotic apparatus and striated muscle), and the dynamics of cytoplasmic behavior were conceptualized vaguely in terms of sol-gel transitions without a sound molecular foundation. Substantial improvements in electron, light, and fluorescence microscopy, as well as the isolation of discrete protein components of the cytoskeleton, have led the way to a much better appreciation of the structural organization of the cytoplasm. Indeed, the lacelike network of thin filaments, intermediate filaments, and microtubules in nonmuscle cells is as familiar today as the organelles identified... 

Microtubules in the cytoskeleton and mitotic apparatus are also in a state of dynamic equilibrium and flux with unpolymerized tubulin, and tubulin appears to be an excellent example of the proteins which Pauling (1953) postulated to exist as globular protomers or as insoluble, fibrous, supramolecular structures akin to unpolymerized and polymeric hemoglobin S. The current view of the microtubule cytoskeleton in nondividing celb comes from the development of tubulin-specific antibodies for indirect immunofluorescent localization of microtubules (Fuller et al., 1975 Weber et al., 1975). The general structural features of such cyto-... 

This theory clearly predicts that the shape of the polymer length distribution curve determines the shape of the time course of depolymerization. For example Kristofferson et al. (1980) were able to show that apparent first-order depolymerization kinetics arise from length distributions which are nearly exponential. It should also be noted that the above theory helps one to gain a better feeling for the time course of cytoskeleton or mitotic apparatus disassembly upon cooling cells to temperatures which destabilize microtubules and effect unidirectional depolymerization. Likewise, the linear depolymerization kinetic model could be applied to the disassembly of bacterial flagella, muscle and nonmuscle F-actin, tobacco mosaic virus, hemoglobin S fibers, and other linear polymers to elucidate important rate parameters and to test the sufficiency of the end-wise depolymerization assumption in such cases. 

The test is used for the detection of cytogenetic damage to the chromosomes or the mitotic apparatus of erythroblasts by analysis of erythrocytes for formation of micronuclei (small nuclei, separate from and additional to the main nuclei of cells, produced during the telophase of mitosis (meiosis) by lagging chromosome fragments or whole chromosomes). When a bone marrow erythroblast develops into a polychromatic erythrocyte (immature erythrocyte), the main nucleus is extruded any micronucleus that has been formed may remain behind in the otherwise anucleated cytoplasm. 

Positive results in the mammalian in vivo erythrocyte micronucleus test indicate that a substance produces micronuclei in the immature erythrocytes of the test species, which are the result of chromosomal damage or damage to the mitotic apparatus in the erythroblasts of the test species. 

The principal cytoskeletal proteins in non-muscle cells are actin, tubulin, and the components of intermediate filaments. Actin can exist either as monomers ( G-actin ) or polymerized into 70 A diameter double filament ( F-actin ). Polymerized actin usually is localized at the margins of the cells, linked by other proteins to the cell membrane. In contrast, tubulin forms hollow filaments, approximately 250 A in diameter, that are distributed within a cell in association, generally, with cell organelles. Stabilized microtubule structures are found in the flagella and cilia of eucaryotic cells however, in other instances - examples being the mitotic apparatus and the cytoskeletal elements arising in directed cell locomotion - the microtubules are temporal entities. Intermediate filaments, which are composed of keratin-like proteins, are approximately 100 A thick and form stable structural elements that impart rigidity, for example, to nerve axons and epithelial cells. 

The hallmark and molecular characteristic of progression is karyotype instability. This is due to disruption of mitotic apparatus, alteration of telomere function, DNA hypomethylation, chromosome translocations and recombination, gene amplification, and gene transposition. There is also alteration of mismatch repair genes in some cancers. 

Cyclic uptake and release of Ca2+ from the extracellular medium occur during mitosis in Physarum pofycephalum, and correlate with specific structural and kinetic events in the mitotic nuclei.442 The membrane system in the mitotic apparatus in Haemantkus endosperm cells functions in the localized release of Ca2+, so regulating the events of mitosis.443 It is known that calcium exerts effects on the stability of spindle microtubules. An alternative view is that free magnesium concentration acts as the fundamental regulator of the cell cycle.444 Tubulin polymerization depends on the presence of magnesium and the absence of calcium, and control of the Ca2+/Mg2+ ratio is relevant to spindle assembly. 

Many phytotoxins are mitotic inhibitors. However, the effects are seldom due to direct effects on the mitotic apparatus. Onion (Allium cepa L.) seeds are germinated in the presence of the natural products under a 14-h photoperiod.6 Root tips are prepared according to Armbruster et al.,2 and mitotic analysis is performed with a compound microscrope on 1000 cells per slide (three slides per treatment).6 9137 In addition to the proportions of cells in each stage of mitosis, the number of abnormal mitotic figures should be determined. 

NuMA Nuclear mitotic apparatus protein SP-H antigen... 

Borisy GG, Taylor EW. The mechanism of action of colchicine Colchicine binding to sea urchin eggs and the mitotic apparatus. 

An ELISA for the measurement of an NMP called nuclear mitotic apparatus protein in urine sample has been approved by the U.S. FDA for the management of patients with TCC of the urinary tract. The test is called NMP22 and is manufactured by Matritech, Inc. (Newton, Mass.)... 

Figure 34.1 Motion within cells. This high-voltage electron micrograph shows the mitotic apparatus in a metaphase mammalian cell. The targe cylindrical objects are chromosomes, and the threadlike structures stretched across the center are microtubules—tracks for the molecular motors that move chromosomes. Many processes, including chromosome segregation in mitosis, depend on the action of molecular-motor proteins.. [Courtesy of Dr. J. R. McIntosh.]...

Purpose. The purpose of the in vivo micronucleus test is to evaluate the potential of the test substance to cause chromosomal damage (clastogenicity) or damage to the mitotic apparatus (aneugenicity) by the analysis of micronuclei in erythrocytes sampled in bone marrow and/or in peripheral blood of experimental animals (usually rodents) (Schmid 1975 Heddle 1973 Heddle and Salamone 1981). 

Findings from studies discussed in Chapter 20 show that the association of microtubules with the endoplasmic reticulum and other membrane-bounded organelles may be critical to the location and organization of these organelles within the cell. For instance, if microtubules are destroyed by drugs such as nocodazole or colcemid, the ER loses its networklike organization. Microtubules are also critical to the formation of the mitotic apparatus—the elaborate, transient structure that captures and subsequently separates replicated chromosomes in cell division. 

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