Metaphase analysis

Metaphase analysis or microiuidc-us assay in rodent bone marrow... 

In vitro metaphase analysis (or mouse micronucleus test)... 

Toxicological properties Acute toxicity (by two routes of admission) Skin irritation Eye irritation Skin sensitization 28 days subacute toxicity Ames test In vitro metaphase analysis (or mouse micronucleus test)... 

Metaphase analysis of cultured mammalian cells and of treated animals. 

Rats and mice are generally used for in vivo studies, with the mouse being employed for bone marrow micronucleus analysis and the rat for metaphase analysis, but both can be used for either. Mice are cheaper and easier to handle than rats, and only a qualitative difference in response has been found between the species (Albanese et al., 1987). Chinese hamsters are also widely used for metaphase analysis because of their low diploid chromosome number of 22. However, there are few other historical toxicological data for this species. 

Metaphase Analysis. Metaphase analysis can be performed in any tissue with actively dividing cells, but bone marrow is the tissue most often examined. Cells are treated with a test compound and are arrested in metaphase by the administration of colcemid or colchicine at various sampling times after treatment. Preparations are examined for structural chromosome damage. Because the bone marrow has a good blood supply, the cells should be exposed to the test compound or its metabolites in the peripheral blood supply. Additionally, these cells are sensitive to S-dependent and S-independent mutagens (Topham et al., 1983). 

Positive results from the in vitro micronucleus test indicate that the test substance induces chromosome damage and/or damage to the cell division apparams, in cultured mammahan somatic cells. Immunochemical labehng (FISH fluorescence in sim hybridization) of kinetochores, or hybridization with general or chromosome specific centromeric/telomeric probes can provide useful information on the mechanism of micronucleus formation. Use of cytokinesis block facilitates the acquisition of the additional mechanistic information (e.g., chromosome nondisjunction) that can be obtained by FISH techniques. The micronucleus assay has a number of advantages over metaphase analysis performed to measure chromosome aberrations (see OECD TG 487 draft). 

Prenatal and postnatal development none Genetic toxicology human lymphocyte metaphase analysis, mammalian cell mutation assay, mouse micronucleus test Carcinogenicity none... 

Type of Study METAPHASE ANALYSIS Genetic end Point CHROMOSOME ABERRATIONS Species and Strain HAMSTER CHINESE Cell CHL FIBROBLAST... 

EUard, S. Toper, S. Stemp, G. Parry, E. M. Wilcox, R Parry, J. M. A comparison of conventional metaphase analysis of Giemsa-stained chromosomes with multi-color fluorescence in situ hybridization. Mutagenesis 1996, 11, 537—546. 

Damage induced in whole animals can be detected in in vivo chromosome assays in either somatic or germinal cells by examination of metaphases or the formation of micronuclei. The micronucleus test can also detect whole chromosome loss or aneuploidy in the absence of clastogenic activity and is considered comparable in sensitivity to chromosome analysis (Tsuchimoto and Matter, 1979). 

Uchiyama S, Kobayashi S, Takata H, Ishihara T, Hori N, Higashi T, Hayashihara K, Sone T, Higo D, Nirasawa T, et al (2005) Proteome analysis of human metaphase chromosomes. J Biol Chem 280 16994-17004... 

During mitosis, aU the DNA is highly condensed to allow separation of the sister chromatids. This is the only time in the ceE cycle when the chromosome structure is visible. Chromosome abnormalities may be assessed on mitotic chromosomes by karyotype analysis (metaphase chromosomes) and by banding techniques (prophase or prometaphase), which identify aneu-ploidy, translocations, deletions, inversions, and duplications. 

An analysis of different stages can be made by fixing cells at different times after treatment, when the cells appear at metaphase, where the chromosomes can be analyzed. With synchronous cells, treatment at different times after initiation of synchronization permits cells in different stages of the cycle to be treated. 

Structural and Numerical Alterations. The basic principles underlying in vivo cytogenetic assays are the same as those for in vitro assays, discussed above (Section 25.3.4b). The cell types that are available for the analysis of chromosome alterations are those that are cycling cell populations, such as bone marrow and spermatogonial cells. For acute exposures, analysis of metaphase cells at their first metaphase after S-phase treatment and analysis of all aberration types is appropriate for hazard evaluation. For chronic exposures, it is inappropriate to analyze unstable aberration types the informative approach is to analyze stable types (e.g., reciprocal translocations and inversions) using FISFI. 

Figure 5. FISH mapping of ISYNA1 gene. Each dot represents the double FISH signals detected on chromosome 19 and 4, after analysis of 100 metaphase chromosomal spreads. Chromosome 19p 13.1 harbors the expressed gene and chromosome 4pl5, the pseudogene.

Palekar LD, Eyre JF, Most BM, et al. 1987. Metaphase and anaphase analysis of V79 cells exposed to erionite, UICC chrysotile and UlCC crocidolite. Carcinogenesis 8 553-560. 

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