Analysis of radiation-induced micronuclei in two-cell human-hamster embryos using telomeric and centromeric FISH probes
ABSTRACT Simultaneous, fluorescent in situ hybridization using a centromeric human alpha satellite DNA probe and a telomeric DNA probe was used to analyze the chromosome content of micronuclei induced in two-cell human-hamster embryos by in vitro gamma-ray irradiation of human spermatozoa. In unirradiated samples, about 26% of micronuclei were centromere positive, indicating that both structural chromosome aberrations and numerical changes are involved in the spontaneous production of micronuclei. After exposure of spermatozoa to radiation, a significant increase in the number of micronuclei was found. About 77% of induced micronuclei contained only telomeric signals suggesting that they originated from acentric fragments. However, both centromere-positive and centromere-negative micronuclei increased with radiation dose. These results are consistent with the well known clastogenic effect of ionizing radiation and with its weak aneugenic effect.
- SourceAvailable from: Maitane Barasoain
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- "For FISH analysis, to identify centromeres, 50 MN for each individual and type of culture on coded slides in a blind study were analyzed for the presence of a fluorescent signal. MN were classified as centromere-negative (CN − ) or centromerepositive (CN + ) by considering the presence of a hybridization signal in a MN as a direct indication of the presence of a centromere . Among CN + MN, signals of different intensities were found due to variable amounts of alpha-satellite DNA on different human chromosomes . "
ABSTRACT: Angiotensin II receptor blockers (ARBs) are a new class of drugs for the treatment of hypertension. In this study, we studied the potential genotoxic effects of five ARBs in vivo and in vitro in human peripheral blood lymphocytes (PBLs) by means of the cytokinesis-block micronucleous (CBMN) assay in combination with fluorescence in situ hybridization (FISH) with a centromeric probe. The nuclear division index (NDI) was used as a measure of cytotoxicity. We also analyzed the association between sex, age, duration of treatment and MN formation. The in vivo study was carried out in 55 hypertensive patients. The in vitro study was performed in 10 control individuals by adding the drugs to the culture medium at a final concentration similar to the levels found in plasma in patients. Our results showed a significant increase in the frequencies of MN and binucleated cells with MN (BNMN) in vivo and especially in vitro. We observed variability in the mean frequency of MN and BNMN among the five drugs analyzed. In vivo, patients treated with Candesartan, Telmisartan and Valsartan showed a statistical significant increase in these parameters, while Olmesartan showed the highest effect in vitro. We also found that the drugs inhibit the NDI in vitro and that Eprosartan, Olmesartan and Telmisartan are the ARBs studied with the highest effect in decreasing the proliferation of the cells. FISH analysis revealed no significant difference between patients and controls in the frequency of centromeric signals. A slight variability, without statistical significance, in the frequency of micronuclei with a centromere signal (CN+MN) was found among the different ARBs analyzed, ruling out an aneugenic potential. When accounting for risk factors, we found that in patients there is a positive correlation between MN, BNMN and sex and a negative correlation with duration of treatment.Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis 04/2014; DOI:10.1016/j.mrgentox.2014.04.005 · 4.44 Impact Factor
- "Although the micronucleus test can be used to evaluate the residual lesions left after occupational or accidental exposure to radiation, this test cannot be used to estimate the dose received at the testicular level due to the different radiosensitivities of spermatogenic cells. Since micronuclei may contain acentric fragments  as well as centric fragments or whole chromosomes with damaged centromeres , and even groups of chromosomes that produce large micronuclei , we decided to carry out a FISH study with telomeric and centromeric probes for all human chromosomes to characterize the chromosomal content of micronuclei . The results showed that over 75% of micronuclei were centromere-negative, indicating that they originated from acentric fragments. "
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ABSTRACT: Ultrastructural and morphometric analyses of centromeric regions by scanning and transmission electron microscopy have been performed in chromosomes from male pronuclei obtained by heterologous fertilisation of hamster oocytes with human spermatozoa. In 1308 of 1323 chromosomes analysed, the primary constriction showed a defined biconcave constriction of variable length (0.56-1.34 microns) and constant width (0.64-0.7 micron). A positive correlation was observed between centromeric length and chromosome length. In some chromosomes, the primary constriction appears as decondensed regions of variable length (1.6-2.51 microns) composed of chromatin fibres with a minimum diameter of 30 nm.Zygote 03/2000; 8(1):79-85. DOI:10.1017/S096719940000085X · 1.32 Impact Factor