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In Vivo Studies
Published in Alexandra C. Miller, Depleted Uranium, 2006
David E. McClain, Alexandra C. Miller
Experiments with cultured cells have also demonstrated the capacity of uranium to induce genotoxic changes. (Lin 1991) showed that uranyl nitrate increased frequencies of micronuclei, sister chromatid exchange, and chromosomal aberrations in Chinese hamster ovary (CHO) cells. Assessments using a human cell model were again conducted by Miller et al., who used in vitro studies to demonstrate that DU was genotoxic by measuring the induction of sister chromatid exchanges, micronuclei, and dicentric chromosomes in the same human cell model used to study neoplastic transformation. Three examples of chromosomal damage measured in these studies are shown in Figure 1.3, including micronuclei, dicentric, and sister chromatid exchange. Studies examining the effect of DU exposure on the induction of these chromosomal changes revealed that DU (in both soluble and insoluble) form could induce an increase in genotoxic damage in comparison to control levels (Table 1.3). Furthermore, these were the first studies to demonstrate that DU could induce a radiation-specific marker, a dicentric chromosome. This finding led to additional studies to address the role of alpha particle radiation damage caused by DU exposure. The nonradioactive heavy metals like nickel which were genotoxic, causing sister chromatid exchanges and micronuclei formation, did not induce dicentric chromosomes; therefore, these data also suggest that DU causes genotoxic damage via a radioactive mechanism.
Chromosomal damage in occupationally exposed health professionals assessed by two cytogenetic methods
Published in Archives of Environmental & Occupational Health, 2023
Dita Kadlcikova, Petra Musilova, Hana Hradska, Miluse Vozdova, Marketa Petrovova, Marek Svoboda, Jiri Rubes
Prior to hybridization, the locations of DAPI (4′,6-diamidino-2-phenylindole) stained metaphase cells, potentially suitable for further analysis, were captured by an automatic metaphase finder system (Metafer, MetaSystems, Altlussheim, Germany) using Zeiss Axio Imager Z2 fluorescence microscope (Carl Zeiss Microimaging GmbH, Jena, Germany). The slides were denatured in 0.07 M NaOH (according to a protocol provided by MetaSystems for multi-color probe kits) and hybridized with denatured painting probes for chromosomes 1 (orange), 2 (green) and 4 (green; MetaSystems) and a pancentromeric probe (biotin, Cambio, Cambridge, United Kingdom). After overnight hybridization at 37 °C, the slides were washed in 0.4× SSC at 72 °C and the biotin labeled probe was detected by avidin conjugated to the near infrared fluorescent dye Cy5 (Invitrogen, Waltham, Massachusetts, USA). Chromosomes were counterstained with DAPI. The metaphase spreads were relocated and 1000 cells from each sample were scored under a fluorescence microscope equipped with a dual band excitation filter set for orange and green fluorescence and filters for single dyes including DAPI and Cy5. Images of aberrant or potentially aberrant cells were captured and classified by two independent scorers using the image analysis software ISIS (MetaSystems) according to the Protocol for Aberration Identification and Nomenclature.29 Three or more cells with the same rearrangement were considered clones and counted only once. Translocations (1-way and 2-way translocations), dicentric chromosomes (dic), acentric fragments (ace), complex rearrangements (complex) (3 or more breaks in 2 or more chromosomes) and aberrant cells (ab.c.) (carrying the above aberrations) were evaluated. To calculate the whole genome translocation frequency, each complete reciprocal translocation (2-way translocation) and incomplete translocation (1-way translocation) were considered as one event. Apparently stable complex exchanges (metaphases containing 46 centromeres and neither dicentric chromosome nor acentric fragment) were broken down into simple translocations and included in the total translocation count.30,31 The genomic frequency of apparently stable chromosomal translocations (FG) was calculated according to a standard formula proposed by Lucas and Sachs32: is the genomic frequency of translocations, is the frequency of translocations detected by FISH, is the fraction of the genome painted red (chromosome 1), and is the fraction of the genome painted green (chromosomes 2 a 4).