The Experts below are selected from a list of 56484 Experts worldwide ranked by ideXlab platform
János Szabad - One of the best experts on this subject based on the ideXlab platform.
-
Assaying benzene, a parquet varnish, and a synthetic thinner with respect to induction of in vivo Chromosome Loss in wing primordial cells of Drosophila
Mutation Research - Genetic Toxicology and Environmental Mutagenesis, 2014Co-Authors: Istvan Soos, János SzabadAbstract:An assay detecting the in vivo Loss of mwh+Y, a genetically engineered Y Chromosome, in cells of the Drosophila wing primordia was published recently. Loss of the mwh+Y Chromosome in any of the wing-disk cells - in a multiple wing hairs homozygous background - leads to the formation of an mwh mosaic spot (clone) in the emerging wing. The frequency and the size of the mwh clones allow detection and quantitative evaluation of environmental and/or genetic agents inducing Chromosome Loss. Using this novel technique, we analyzed the potential of vapors of benzene, a parquet varnish, and a synthetic thinner to induce Chromosome Loss. Exposure to 0.047μg/ml benzene vapor for one day or to 0.175μg/ml for four hours resulted in a significantly elevated mwh clone-frequency confirming the ability of benzene to induce Chromosome Loss. A one-day exposure to vapors of a parquet varnish or a 6-h exposure to vapors of a synthetic thinner slightly, yet significantly elevated the frequency of Chromosome Loss. Results of the present paper show the potential of vapors of the analyzed parquet varnish and synthetic thinner to induce Chromosome Loss, and illustrate the usefulness of the new technique. © 2014.
-
An Assay to Detect In Vivo Y Chromosome Loss in Drosophila Wing Disc Cells
G3 (Bethesda Md.), 2012Co-Authors: János Szabad, Hugo J. Bellen, Koen J. T. VenkenAbstract:Loss of the Y Chromosome in Drosophila has no impact on cell viability and therefore allows us to assay the impact of environmental agents and genetic alterations on chromosomal Loss. To detect in vivo Chromosome Loss in cells of the developing Drosophila wing primordia, we first engineered a Y Chromosome with an attP docking site. By making use of the ΦC31 integrase system, we site-specifically integrated a genomic transgene encompassing the multiple wing hair (mwh) locus into this attP site, leading to a mwh+Y Chromosome. This Chromosome fully rescues the mwh mutant phenotype, an excellent recessive wing cell marker mutation. Loss of this mwh+Y Chromosome in wing primordial cells then leads to manifestation of the mwh mutant phenotype in mwh-homozygous cells. The forming mwh clones permit us to quantify the effect of agents and genetic alterations by assaying frequency and size of the mwh mosaic spots. To illustrate the use of the mwh+Y Loss system, the effects of four known mutagens (X-rays, colchicine, ethyl methanesulfonate, and formaldehyde) and two genetic conditions (Loss- and gain-of-function lodestar mutant alleles) are documented. The procedure is simple, sensitive, and inexpensive.
Riccardo Crebelli - One of the best experts on this subject based on the ideXlab platform.
-
genetic effects of petroleum fuels ii analysis of Chromosome Loss and hyperploidy in peripheral lymphocytes of gasoline station attendants
Environmental and Molecular Mutagenesis, 1998Co-Authors: A Carere, Antonio Antoccia, Francesca Degrassi, Caterina Tanzarella, Daniela Cimini, Riccardo Crebelli, Paola Leopardi, Francesca Marcon, Antonella Sgura, Andrea ZijnoAbstract:Molecular cytogenetic methods were applied to investigate the effect of the occupational exposure to low concentrations of benzene and petroleum fuels on genomic stability. Twelve male gasoline station attendants (average benzene exposure of 0.32 mg/m3 as 8h TWA) and 12 age- and smoking-matched unexposed controls were selected for the study. The incidence of hyperploidy and polyploidy in peripheral lymphocytes was evaluated through in situ hybridization of interphase cells, harvested 24 hr after stimulation, with centromeric probes of Chromosomes 7, 11, 18, and X. For half of the subjects, metaphases harvested 24 hr later were analyzed. The incidence of Chromosome Loss in vitro was determined in cytokinesis-blocked cells, harvested at 66 hr, through the hybridization of micronuclei with a pancentromeric probe. Ten thousand Chromosomes (more than 200 metaphases equivalent) and 2,000 binucleated cells/person were scored for hyperploidy and micronucleus analysis, respectively. The results obtained did not show any exposure-related excess of hyperploidy or micronucleus formation. Conversely, the age of the subjects was significantly correlated with several markers of genomic instability, such as the incidence of Chromosome X and Chromosome 18 hyperploidy, total hyperploidy and polyploidy, and close to statistical significance with Chromosome Loss. Smoking habits did not appear to contribute significantly to the effects measured. The parallel analysis of hyperploidy and polyploidy in interphase nuclei in 24-hr cultures and in metaphase cells harvested 24 hr later showed basically similar incidences of aneuploid cells, indicating that no significant selection against hyperploid and polyploid types occurred during the first cell cycle in vitro.
-
Sex Chromosome Loss and non-disjunction in women: analysis of chromosomal segregation in binucleated lymphocytes.
Chromosoma, 1996Co-Authors: Andrea Zijno, Francesca Marcon, Paola Leopardi, Riccardo CrebelliAbstract:Chromosomal lagging and non-disjunction are the main mechanisms of chromosomal malsegregation at mitosis. To date, the relative importance of these two events in the genesis of spontaneous or induced aneuploidy has not been fully elucidated. A methodology based on in situ hybridization with centromeric probes in binucleated lymphocytes was previously developed to provide some insight into this matter. With this method, both chromosomal Loss and non-disjunction can be simultaneously detected by following the distribution of specific Chromosomes in the nuclei and micronuclei of binucleated cells. In this study, this approach was used for studying the role of chromosomal Loss and non-disjunction in the age-related malsegregation of sex Chromosomes in females. For this purpose, cultures of cytokinesis-blocked lymphocytes were established from 12 healthy women ranging in age from 25 to 56. The occurrence of malsegregation of X Chromosomes in vitro was estimated in binucleated cells that contained four signals, which orginates from the division of normal disomic cells. In this cell population, the frequencies of X Chromosome Loss and non-disjunction ranged from 0% to 1.69% (mean 0.75%), and from 0.20% to 1.33% (mean 0.57%), respectively. This indicates that both events contribute to malsegregation of X Chromosomes in vitro. Moreover, a small but not negligible fraction of binucleated cells with two or six copies of the X Chromosome was noticed in all donors. These cells, which are thought to arise from parental monosomic and trisomic types, may indicate the malsegregation of X Chromosomes in vivo. The frequency of X Chromosome aneuploidy both in vivo and in vitro significantly correlated with the age of donors. Analysis of chromosomal distribution in unbalanced cells demonstrated that both X homologues were frequently involved. The frequency of such multiple events (0.17%) was far greater than that expected by mere chance, indicating a tendency to multiple malsegregation events in the cell population investigated. Finally, parallel analysis of the segregation of Chromosomex X and 1 in five of the donors confirmed the greater (about tenfold) susceptibility of X Chromosomes to malsegregate compared with autosomes.
-
simultaneous detection of x Chromosome Loss and non disjunction in cytokinesis blocked human lymphocytes by in situ hybridization with a centromeric dna probe implications for the human lymphocyte in vitro micronucleus assay using cytochalasin b
Mutagenesis, 1994Co-Authors: Andrea Zgno, Francesca Marcon, Paola Leopardi, Riccardo CrebelliAbstract:: A methodology for the simultaneous detection of Chromosome Loss and gain in mammalian cells has been developed which is based upon the analysis of Chromosome distribution in daughter nuclei of binucleated human lymphocytes. X-Chromosome distribution was followed by in situ hybridization, using a commercial biotinylated DNA probe specific for the centromeric alphoid sequences of human X-Chromosome. In order to optimize the experimental protocol for the use of cytokinesis-blocked lymphocytes in aneuploidy assays, the effect of harvest time and cytochalasin B (Cyt B) dosage upon Chromosome distribution was investigated. To this end, lymphocyte cultures were treated 44 h after mitogen stimulation with different dosages of Cyt B and collected at 60, 66 and 72 h. High rates of binucleated cells with unbalanced Chromosome distribution (two spots in one nucleus and none in the other in male cells; three spots in one nucleus and one in the other in female cells) and abnormal spot number (more than or less than two per male cell or four per female cell) were observed at 66 and 72 h in cultures treated with the lowest Cyt B dose (3 micrograms/ml). In contrast, low frequencies of unbalanced or abnormal binucleated cells were observed at 60 h with both 3 and 6 micrograms/ml Cyt B. These results indicate that binucleated lymphocytes with low background frequencies of malsegregation (required for the analysis of induced aneuploidy), can be obtained by harvesting lymphocyte cultures 60 h after stimulation (16 h after Cyt B block).(ABSTRACT TRUNCATED AT 250 WORDS)
Robert Benezra - One of the best experts on this subject based on the ideXlab platform.
-
whole Chromosome Loss and associated breakage fusion bridge cycles transform mouse tetraploid cells
The EMBO Journal, 2018Co-Authors: Rozario Thomas, Daniel Henry Marks, Yvette Chin, Robert BenezraAbstract:Abstract Whole Chromosome gains or Losses (aneuploidy) are a hallmark of ~70% of human tumors. Modeling the consequences of aneuploidy has relied on perturbing spindle assembly checkpoint (SAC) components, but interpretations of these experiments are clouded by the multiple functions of these proteins. Here, we used a Cre recombinase‐mediated Chromosome Loss strategy to individually delete mouse Chromosomes 9, 10, 12, or 14 in tetraploid immortalized murine embryonic fibroblasts. This methodology also involves the generation of a dicentric Chromosome intermediate, which subsequently undergoes a series of breakage–fusion–bridge (BFB) cycles. While the aneuploid cells generally display a growth disadvantage in vitro , they grow significantly better in low adherence sphere‐forming conditions and three of the four lines are transformed in vivo , forming large and invasive tumors in immunocompromised mice. The aneuploid cells display increased chromosomal instability and DNA damage, a mutator phenotype associated with tumorigenesis in vivo . Thus, these studies demonstrate a causative role for whole Chromosome Loss and the associated BFB‐mediated instability in tumorigenesis and may shed light on the early consequences of aneuploidy in mammalian cells.
-
Whole Chromosome Loss and associated breakage–fusion–bridge cycles transform mouse tetraploid cells
The EMBO journal, 2017Co-Authors: Rozario Thomas, Daniel Henry Marks, Yvette Chin, Robert BenezraAbstract:Abstract Whole Chromosome gains or Losses (aneuploidy) are a hallmark of ~70% of human tumors. Modeling the consequences of aneuploidy has relied on perturbing spindle assembly checkpoint (SAC) components, but interpretations of these experiments are clouded by the multiple functions of these proteins. Here, we used a Cre recombinase‐mediated Chromosome Loss strategy to individually delete mouse Chromosomes 9, 10, 12, or 14 in tetraploid immortalized murine embryonic fibroblasts. This methodology also involves the generation of a dicentric Chromosome intermediate, which subsequently undergoes a series of breakage–fusion–bridge (BFB) cycles. While the aneuploid cells generally display a growth disadvantage in vitro , they grow significantly better in low adherence sphere‐forming conditions and three of the four lines are transformed in vivo , forming large and invasive tumors in immunocompromised mice. The aneuploid cells display increased chromosomal instability and DNA damage, a mutator phenotype associated with tumorigenesis in vivo . Thus, these studies demonstrate a causative role for whole Chromosome Loss and the associated BFB‐mediated instability in tumorigenesis and may shed light on the early consequences of aneuploidy in mammalian cells.
-
Whole Chromosome Loss in tetraploid cells confers tumorigenic potential in a mouse allograft model
2017Co-Authors: Rozario Thomas, Daniel Henry Marks, Yvette Chin, Robert BenezraAbstract:Whole Chromosome gains or Losses (aneuploidy) are a hallmark of ~70% of human tumors. Modeling the consequences of aneuploidy has relied on perturbing spindle assembly checkpoint (SAC) components but interpretations of these experiments are clouded by the multiple functions of these proteins. Here we used a Cre recombinase-mediated Chromosome Loss strategy to individually delete mouse Chromosomes 9, 10, 12 or 14 in tetraploid immortalized murine embryonic fibroblasts. While the aneuploid cells generally display a growth disadvantage in vitro, they grow significantly better in low adherence sphere-forming conditions and 3 of the 4 lines are transformed in vivo, forming large and invasive tumors in immunocompromised mice. The aneuploid cells display increased chromosomal instability and DNA damage, a mutator phenotype associated with tumorigenesis in vivo. Thus, these studies demonstrate a causative role for whole Chromosome Loss in tumorigenesis and may shed light on the early consequences of aneuploidy in mammalian cells.
Mika Yamamoto - One of the best experts on this subject based on the ideXlab platform.
-
Chromosome Loss caused by DNA fragmentation induced in main nuclei and micronuclei of human lymphoblastoid cells treated with colcemid.
Mutation Research, 2014Co-Authors: Mika Yamamoto, Akihiro Wakata, Yoshinobu Aoki, Yoichi Miyamae, Seiji KodamaAbstract:Aneuploidy, a change in the number of Chromosomes, plays an essential role in tumorigenesis. Our previous study demonstrated that a Loss of a whole Chromosome is induced in human lymphocytes by colcemid, a well-known aneugen. Here, to clarify the mechanism for colcemid-induced Chromosome Loss, we investigated the relationship between Chromosome Loss and DNA fragmentation in human lymphoblastoid cells treated with colcemid (an aneugen) compared with methyl methanesulfonate (MMS; a clastogen). We analyzed the number of fluorescence in situ hybridization (FISH) signals targeted for a whole Chromosome 2 in cytokinesis-blocked binucleated TK6 cells and WTK-1 cells treated with colcemid and MMS, and concurrently detected DNA fragmentation by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. Results revealed that DNA fragmentation occurred in 60% of all binucleated TK6 cells harboring colcemid-induced Chromosome Loss (30% of micronuclei and 30% of main nuclei). DNA fragmentation was observed in colcemid-induced micronuclei containing a whole Chromosome but not in MMS-induced micronuclei containing Chromosome fragments. In contrast, colcemid-induced nondisjunction had no effect on induction of DNA fragmentation, suggesting that DNA fragmentation was triggered by micronuclei containing a whole Chromosome but not by micronuclei containing Chromosome fragments or nondisjunction. In addition, the frequency of binucleated cells harboring Chromosome Loss with DNA fragmentation in micronuclei or main nuclei was higher in wild-type p53 TK6 cells than in mutated-p53 WTK-1 cells treated with colcemid. Taken together, these present and previous results suggest that colcemid-induced Chromosome Loss is caused by DNA fragmentation, which is triggered by a micronucleus with a whole Chromosome and controlled by the p53-dependent pathway.
-
induction of a whole Chromosome Loss by colcemid in human cells elucidated by discrimination between fish signal overlap and Chromosome Loss
Mutation Research, 2013Co-Authors: Mika Yamamoto, Akihiro Wakata, Yoshinobu Aoki, Yoichi Miyamae, Seiji KodamaAbstract:Aneuploidy is a change in the number of Chromosomes and an essential component in tumorigenesis. Therefore, accurate and sensitive detection of aneuploidy is important in screening for carcinogens. In vitro micronucleus (MN) assay has been adopted in the recently revised International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH) S2 guideline and can be employed to predict both clastogenic and aneugenic chromosomal aberrations in interphase cells. However, distinguishing clastogens and aneugens is not possible using this assay. The Organization for Economic Co-operation and Development (OECD) guideline TG487 therefore recommends the use of centromere/kinetochore staining in micronuclei to differentiate clastogens from aneugens. Here, we analyzed numerical changes of a specific Chromosome in cytokinesis-blocked binucleated cells by fluorescence in situ hybridization (FISH) using the specific centromere probe in human lymphoblastoid TK6 cells treated with aneugens (colcemid and vincristine) or clastogens (methyl methanesulfonate [MMS] and 4-nitroquinoline-1-oxide [4-NQO]). Colcemid and vincristine significantly increased the frequencies of nondisjunction and Loss of FISH signals, while MMS and 4-NQO slightly increased only the frequency of Loss of FISH signals. The Loss of FISH signals of a specific Chromosome from two to one per nucleus implies either a Loss of a whole Chromosome or an overlap of two signals. To distinguish a Chromosome Loss from signal overlap, we investigated the number of FISH signals and the fluorescent intensity of each signal per nucleus using a probe specific for whole Chromosome 2 in binucleated TK6 cells and primary human lymphocytes treated with colcemid and MMS. By discriminating between Chromosome Loss and FISH signal overlap, we revealed that colcemid, but not MMS, induced a Loss of a whole Chromosome in primary lymphocytes and TK6 cells.
Istvan Soos - One of the best experts on this subject based on the ideXlab platform.
-
Assaying benzene, a parquet varnish, and a synthetic thinner with respect to induction of in vivo Chromosome Loss in wing primordial cells of Drosophila
Mutation Research - Genetic Toxicology and Environmental Mutagenesis, 2014Co-Authors: Istvan Soos, János SzabadAbstract:An assay detecting the in vivo Loss of mwh+Y, a genetically engineered Y Chromosome, in cells of the Drosophila wing primordia was published recently. Loss of the mwh+Y Chromosome in any of the wing-disk cells - in a multiple wing hairs homozygous background - leads to the formation of an mwh mosaic spot (clone) in the emerging wing. The frequency and the size of the mwh clones allow detection and quantitative evaluation of environmental and/or genetic agents inducing Chromosome Loss. Using this novel technique, we analyzed the potential of vapors of benzene, a parquet varnish, and a synthetic thinner to induce Chromosome Loss. Exposure to 0.047μg/ml benzene vapor for one day or to 0.175μg/ml for four hours resulted in a significantly elevated mwh clone-frequency confirming the ability of benzene to induce Chromosome Loss. A one-day exposure to vapors of a parquet varnish or a 6-h exposure to vapors of a synthetic thinner slightly, yet significantly elevated the frequency of Chromosome Loss. Results of the present paper show the potential of vapors of the analyzed parquet varnish and synthetic thinner to induce Chromosome Loss, and illustrate the usefulness of the new technique. © 2014.