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  • Title: The measurement of induced genetic change in mammalian germ cells.
    Author: Adler ID, Pacchierotti F, Russo A.
    Journal: Methods Mol Biol; 2012; 817():335-75. PubMed ID: 22147580.
    Abstract:
    In vivo methods are described to detect clastogenic and aneugenic effects of chemical agents in male and female germ cells in vivo. The knowledge of stages of germ cell development and their duration for a given test animal is essential for these experiments. Commonly, mice or rats are employed. Structural chromosome aberrations can be analyzed microscopically in mitotic cell divisions of differentiating spermatogonia, zygotes, or early embryos as well as in first meiotic cell divisions of spermatocytes and oocytes. Numerical chromosome aberrations are scorable during second meiotic divisions of spermatocytes and oocytes. The micronucleus test is applicable to early round spermatids and to first cleavage embryos, and as in somatic cells, it assesses structural as well as numerical chromosome aberrations. In contrast to the somatic micronucleus assay, the timing of cell sampling determines whether the micronuclei scored in round spermatids were formed from structural or numerical aberrations, i.e. with short treatment-sampling intervals the micronuclei are formed by exposed meiotic divisions and represent induced non-disjunction. On the -contrary, after longer intervals of 12-14 days micronuclei are formed from induced unstable structural aberrations in differentiating spermatogonia or during the last round of DNA-synthesis in early spermatocytes. Furthermore, labelling with fluorescent DNA-probes can be used to confirm these theoretical expectations. The mouse sperm-FISH assay is totally based on scoring colour spots from individual chromosomes (e.g. X, Y, and 8) hybridized with specific DNA-probes. The most animal demanding assay described here is the dominant lethal test. It is commonly performed with treated male laboratory rodents and allows the determination of the most sensitive developmental stage of spermatogenesis to a particular chemical under test. Theoretically, unstable structural chromosome aberrations in sperm will lead to foetal deaths after fertilization at around the time of implantation in the uterus wall. These can be scored as deciduomata or early dead foetuses in the uterus wall of the females at mid-pregnancy. None of the tests described in this chapter provide data for a quantitative estimate of the genetic risk to progeny from exposed germ cells. The only tests on which such calculations can be based, the heritable translocation assay and the specific locus test, are so animal and time-consuming that they can no more be performed anywhere in the world and thus are not described here.
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