Spontaneous and induced aneuploidy,considerations which may influence chromosome malsegregation

Aneuploidy plays a major role in the production of human birth defects and is becoming increasingly recognised as a critical event in the etiology of a wide range of human cancers. Thus, the detection of aneuploidy and the characterisation of the mechanisms which lead to chromosome malsegregation is an important area of genotoxicological research. As an aid to aneuploidy research, methods have been developed to analyse the mechanisms of chromosome malsegregation and to investigate the role of aneuploidy in tumour progression. The presence of aneuploid cells is a common characteristic of many of tumour cell types as illustrated by the wide range of chromosome number changes detected in post-menopausal breast tumours. To investigate the time of occurrence of aneuploidy during tumour progression, we have studied the chromosome number status of Syrian hamster dermal (SHD) cells cultures progressing to morphological transformation. The production of both polyploid and aneuploid cells is a common feature of progressing cells in this model. The elevation of both progression to morphological transformation and aneuploid frequencies can be produced by exposure to a diverse range of carcinogens and tumour promoters. Analysis of the genotoxic activity of the hormone 17-beta oestradiol demonstrated its ability to induce both chromosome loss and non-disjunction in human lymphoblastoid cells implicating aneugenic activity in hormone related cancers. Mutations in the p53 tumour suppressor gene introduced into human fibroblasts produced modifications in chromosome separation at mitosis which may lead to the production of both aneuploidy and polyploid cells. Our studies indicate that the production of aneuploid cells can be influenced by both endogenous and exogenous factors and occur throughout the progression of normal cells to a malignant phenotype.     

In vitro and in vivo extrapolations of genotoxin exposures: consideration of factors which influence dose-response thresholds

The concept of a threshold of activity of a genotoxic agent is primarily based upon considerations of protective mechanisms and multiple cellular targets, which require inactivation before a toxic response is produced. In this paper, we have considered and evaluated the influences of compound metabolism, DNA lesion formation, mutation induction and sequence content, aneuploidy induction and the influence of repair enzymes upon genetic endpoints produced by both DNA reactive chemicals and by those chemicals which modify non-DNA cellular targets. Thresholds of activity have been evaluated by critical analysis of the published literature and original data analysing both the role of sequence context upon point mutation induction and DNA repair mechanisms upon the sensitivity of cultured cells to the induction of aneuploidy. In the case of DNA reactive chemicals, the presence of a threshold of chemical activity will be dependent upon cellular activities such as those of the Phase II enzymes reducing the activity of chemicals before lesion formation takes place and/or those of the DNA repair enzymes which reduce the proportion of DNA lesions which are processed into DNA sequence changes. Under such conditions, a given exposure of a DNA reactive chemical does not produce a linear or semi-linear increase in DNA lesions or in mutation frequency. However, even when these protective mechanisms are overwhelmed by the high exposures of genotoxic chemicals the biological effects of a genotoxin may be influenced by the sequence context of the gene under consideration. Here, we demonstrate that point mutations are detected at relatively higher frequencies in the non-coding introns compared with the coding exons. Many of the base changes detected in the exons do not produce amino acid changes in the proteins coded for by the genes being monitored for mutation induction. Both sequence context and the types of base changes induced may provide a "buffering" effect reducing the biological consequences of mutation induction. Spindle damaging chemicals, such as colcemid and vinblastine, induce aneuploidy by modifying the numbers of spindle fibres which regulate the segregation of chromosomes during mitosis and meiosis. The redundancy of spindle fibres in the dividing mammalian cell leads to the prediction that only chemical exposures which damage most, if not all, of the fibres will lead to the induction of polyploidy and/or aneuploidy. Such predicted thresholds of chemical activity can be observed when both chromosome loss and non-disjunction are measured in wild type cultures. However, we observed a substantial increase in sensitivity to aneugenic chemicals when measurements were made in primary cell cultures derived from xerodoma pigmentosum and trichothiodystrophy patients. Further studies are necessary to evaluate the consequences of the genetic background of tester strains upon the nature of the dose-response curve of aneugenic chemicals.     

When 2 + 2 = 5: The origins and fates of aneuploid and tetraploid cells

Aneuploid cells are frequently observed in human tumors, suggesting that aneuploidy may play an important role in the development of cancer. In this review, I discuss the processes that may give rise to aneuploid cells in normal tissue and in tumors. Aneuploid cells may arise directly from diploid cells through errors in chromosome segregation, as a consequence of incorrect microtubule-kinetochore attachments, or through failure of the spindle checkpoint. A second route to formation of aneuploid cells is through a tetraploid intermediate, where division of tetraploid cells can yield very high rates of chromosome missegregation as a consequence of multipolar spindle formation. Diploid cells may become tetraploid through a variety of mechanisms, including endoreduplication, cell fusion, and cytokinesis failure. Although aneuploid cells may arise from either diploid or tetraploid cells, the fate of the resulting aneuploid cells may be distinct. It is therefore important to understand the different pathways that can give rise to aneuploid cells, and how the varied origins of these cells affect their subsequent ability to survive or proliferate.     

When 2+2=5: the origins and fates of aneuploid and tetraploid cells

Aneuploid cells are frequently observed in human tumors, suggesting that aneuploidy may play an important role in the development of cancer. In this review, I discuss the processes that may give rise to aneuploid cells in normal tissue and in tumors. Aneuploid cells may arise directly from diploid cells through errors in chromosome segregation, as a consequence of incorrect microtubule-kinetochore attachments, or through failure of the spindle checkpoint. A second route to formation of aneuploid cells is through a tetraploid intermediate, where division of tetraploid cells can yield very high rates of chromosome missegregation as a consequence of multipolar spindle formation. Diploid cells may become tetraploid through a variety of mechanisms, including endoreduplication, cell fusion, and cytokinesis failure. Although aneuploid cells may arise from either diploid or tetraploid cells, the fate of the resulting aneuploid cells may be distinct. It is therefore important to understand the different pathways that can give rise to aneuploid cells, and how the varied origins of these cells affect their subsequent ability to survive or proliferate.     

The use of the in vitro micronucleus assay to detect and assess the aneugenic activity of chemicals

The successful validation of the in vitro micronucleus assay by the SFTG now provides the opportunity for this highly cost effective assay to be used to screen chemicals for their ability to induce both structural (clastogenic) and numerical (aneugenic) chromosome changes using interphase cells. The use of interphase cells and a relatively simple experimental protocol provides the opportunity to greatly increase the statistical power of cytogenetic studies on chemical interactions. The application of molecular probes capable of detecting kinetochores and centromeres provides the opportunity to classify mechanisms of micronucleus induction into those which are primarily due to chromosome loss or breakage. When a predominant mechanism of micronucleus induction has been shown to be based upon chromosome loss then further investigation can involve the determination of the role of non-disjunction in the induction of aneuploidy. The binucleate cell modification of the in vitro micronucleus assay can be combined with the use of chromosome specific centromere probes to determine the segregation of individual chromosomes into daughter nuclei. The combination of these methods provides us with powerful tools for the investigation of mechanisms of genotoxicity particularly in the low dose regions.     

Aneuploidy induction in human fibroblasts: comparison with results in Syrian hamster fibroblasts

The susceptibility of human fibroblast cells in culture to neoplastic transformation by chemical carcinogens is appreciably lower than that of rodent fibroblasts. We have proposed that a key step in the neoplastic progression of Syrian hamster embryo fibroblasts is the induction of aneuploidy by carcinogens. It is possible that the different sensitivity to neoplastic transformation of Syrian hamster versus human cells is due to a difference in genetic stability following treatment with chemicals inducing aneuploidy. Therefore, we measured the induction of numerical chromosome changes in normal human fibroblasts and Syrian hamster fibroblasts by 4 specific aneuploidogens. Dose- and time-dependent studies were performed. Nondisjunction, resulting in aneuploid cells with a near-diploid chromosome number, in up to 14-28% of the hamster cells was induced by colcemid (0.1 microgram/ml), vincristine (30 ng/ml), diethylstilbestrol (DES) (1 microgram/ml) or 17 beta-estradiol (10 micrograms/ml). In contrast, human cells displayed far fewer aneuploid (near-diploid) cells, i.e., 8% following treatment with colcemid (0.02 micrograms/ml) or vincristine (10 ng/ml) and only 3% following treatment with DES (6 micrograms/ml) or 17 beta-estradiol (20 micrograms/ml). The doses at which the maximum effect was observed are given. Treatment of human cells induced a higher incidence of cells with a near-tetraploid chromosome number, which was similar to the level observed in treated hamster cells except at the highest doses. These results indicate that human cells respond differently from hamster cells to agents that induce aneuploidy. In particular, nondisjunction yielding aneuploid human fibroblasts with a near-diploid chromosome number was less frequent. The magnitude of the observed species differences varied with different chemicals. The difference in aneuploidy induction may contribute, in part, to species differences in susceptibility of fibroblasts to neoplastic transformation.     

Aneuploidy: a matter of bad connections

Proper chromosome segregation is required to maintain the appropriate number of chromosomes from one cell generation to the next and to prevent aneuploidy, the condition in which a cell has gained or lost one or several chromosomes during cell division. Aneuploidy is a hallmark associated with birth defects and cancer, and is observed at relatively high frequencies in human somatic cells. Recent studies in mammalian tissue culture cells suggest that the persistence of kinetochore-microtubule misattachments through mitosis is a major cause of chromosome mis-segregation and aneuploidy. Furthermore, studies in mice and humans suggest that small changes in the expression, rather than complete inactivation, of genes encoding specific proteins might be associated with aneuploidy in living organisms. In this article (which is part of the Chromosome Segregation and Aneuploidy series), we survey the outcome of these studies, focusing on the importance of kinetochore misattachments in producing aneuploid cells.     

Mitotic Spindle Defects and Chromosome Mis-Segregation Induced by LDL/Cholesterol—Implications for Niemann-Pick C1, Alzheimer’s Disease,and Atherosclerosis

Elevated low-density lipoprotein (LDL)-cholesterol is a risk factor for both Alzheimer’s disease (AD) and Atherosclerosis (CVD), suggesting a common lipid-sensitive step in their pathogenesis. Previous results show that AD and CVD also share a cell cycle defect: chromosome instability and up to 30% aneuploidy–in neurons and other cells in AD and in smooth muscle cells in atherosclerotic plaques in CVD. Indeed, specific degeneration of aneuploid neurons accounts for 90% of neuronal loss in AD brain, indicating that aneuploidy underlies AD neurodegeneration. Cell/mouse models of AD develop similar aneuploidy through amyloid-beta (Aß) inhibition of specific microtubule motors and consequent disruption of mitotic spindles. Here we tested the hypothesis that, like upregulated Aß, elevated LDL/cholesterol and altered intracellular cholesterol homeostasis also causes chromosomal instability. Specifically we found that: 1) high dietary cholesterol induces aneuploidy in mice, satisfying the hypothesis’ first prediction, 2) Niemann-Pick C1 patients accumulate aneuploid fibroblasts, neurons, and glia, demonstrating a similar aneugenic effect of intracellular cholesterol accumulation in humans 3) oxidized LDL, LDL, and cholesterol, but not high-density lipoprotein (HDL), induce chromosome mis-segregation and aneuploidy in cultured cells, including neuronal precursors, indicating that LDL/cholesterol directly affects the cell cycle, 4) LDL-induced aneuploidy requires the LDL receptor, but not Aß, showing that LDL works differently than Aß, with the same end result, 5) cholesterol treatment disrupts the structure of the mitotic spindle, providing a cell biological mechanism for its aneugenic activity, and 6) ethanol or calcium chelation attenuates lipoprotein-induced chromosome mis-segregation, providing molecular insights into cholesterol’s aneugenic mechanism, specifically through its rigidifying effect on the cell membrane, and potentially explaining why ethanol consumption reduces the risk of developing atherosclerosis or AD. These results suggest a novel, cell cycle mechanism by which aberrant cholesterol homeostasis promotes neurodegeneration and atherosclerosis by disrupting chromosome segregation and potentially other aspects of microtubule physiology.     

Trichlorfon effects on mouse oocytes following in vivo exposure

Trichlorfon (TCF) is a widely used pesticide, which according to some epidemiological and experimental data, is suspected of being aneugenic in human and mouse cells. In particular, in vitro studies in mouse oocytes showed the induction of aneuploidy and polyploidy at the first meiotic division and of severe morphological alterations of the second meiotic spindle. We have tested the hypothesis that an acute treatment of mice with TCF might similarly affect chromosome segregation in maturing oocytes. Superovulated MF-1 mice were intraperitoneally injected with 400mg/kg TCF or orally administered with 600mg/kg TCF either at the time of or 4h after human chorionic gonadotrophin (HCG) injection. Oocytes were harvested 17h after HCG and metaphase II chromosomes were cytogenetically analyzed. No significant increase of aneuploid or polyploid cells was detected at any treatment condition. A significant (p<0.001) decrease of metaphases showing premature chromatid separation or premature anaphase II in all TCF-treated groups with respect to controls suggested that TCF treatment may have delayed the first meiotic division. To evaluate possible effects of the pesticide upon the second meiotic division, a group of females orally treated with 600mg/kg TCF at resumption of meiosis was mated with untreated males and zygotes were collected for cytogenetic analysis. No evidence of aneuploidy induction was obtained, but the frequency of polyploid zygotes was increased fivefold over the control level (p<0.01). Such polyploid embryos might have arisen from fertilization of oocytes that were either meiotically delayed and still in metaphase I at fertilization or progressed through anaphase II without cytokinesis. These findings show that in vivo studies on aneuploidy induction in oocytes may yield results different from those obtained by in vitro experiments and that both kinds of data may be necessary for risk assessment of environmentally relevant exposures.     

Dynamic karyotype,dynamic proteome: buffering the effects of aneuploidy

Despite its ubiquity in cancer, link with other pathologies, and role in promoting adaptive evolution, the effects of aneuploidy or imbalanced chromosomal content on cellular physiology have remained incompletely characterized. Significantly, it appears that the detrimental as well as beneficial effects of aneuploidy are due to the altered gene expression elicited by the aneuploid state. In this review we examine the correlation between chromosome copy number changes and gene expression in aneuploid cells. We discuss the primary effects of aneuploidy on gene expression and describe the cellular response to altered mRNA and protein levels. Moreover, we consider compensatory mechanisms that may ameliorate imbalanced gene expression and restore protein homeostasis in aneuploid cells. Finally, we propose a novel hypothesis to explain the hitherto enigmatic abundance compensation of proteins encoded on supernumerary chromosomes.     

Alzheimer Aβ Peptide Induces Chromosome Mis-Segregation and Aneuploidy,Including Trisomy 21: Requirement for Tau and APP

Both sporadic and familial Alzheimer''s disease (AD) patients exhibit increased chromosome aneuploidy, particularly trisomy 21, in neurons and other cells. Significantly, trisomy 21/Down syndrome patients develop early onset AD pathology. We investigated the mechanism underlying mosaic chromosome aneuploidy in AD and report that FAD mutations in the Alzheimer Amyloid Precursor Protein gene, APP, induce chromosome mis-segregation and aneuploidy in transgenic mice and in transfected cells. Furthermore, adding synthetic Aβ peptide, the pathogenic product of APP, to cultured cells causes rapid and robust chromosome mis-segregation leading to aneuploid, including trisomy 21, daughters, which is prevented by LiCl addition or Ca²⁺ chelation and is replicated in tau KO cells, implicating GSK-3β, calpain, and Tau-dependent microtubule transport in the aneugenic activity of Aβ. Furthermore, APP KO cells are resistant to the aneugenic activity of Aβ, as they have been shown previously to be resistant to Aβ-induced tau phosphorylation and cell toxicity. These results indicate that Aβ-induced microtubule dysfunction leads to aneuploid neurons and may thereby contribute to the pathogenesis of AD.     

The induction of aneuploidy by nitrogen mustard in a human lymphoblastoid cell line

We report that the presence of an extra Y chromosome can be used as a marker for the induction of aneuploidy (mitotic non-disjunction) in a human lymphoblastoid cell line. This endpoint is easily visualized in metaphase chromosome preparations after staining with quinacrine mustard. The induction of cells with two Y chromosomes by nitrogen mustard (NM) was examined. Exposure to 150 ng/ml nitrogen mustard induced a 6-fold increase in aneuploid frequency relative to untreated control levels; maximal induction of aneuploidy was observed 2 days after treatment. Lower concentrations of nitrogen mustard (36 and 75 ng/ml) induced smaller increases in aneuploid frequency, with maximal induction observed 1 day after treatment. This system has the potential to be used as an assay for the induction of aneuploidy in cultured human cells.     

Aneugenic Effects of Epirubicin in Somatic and Germinal Cells of Male Mice

The ability of the antineoplastic agent epirubicin to induce aneuploidy and meiotic delay in the somatic and germinal cells of male mice was investigated by fluorescence in situ hybridization assay using labeled DNA probes and BrdU-incorporation assay. Mitomycin C and colchicine were used as positive controls for clastogen and aneugen, respectively, and these compounds produced the expected responses. The fluorescence in situ hybridization assay with a centromeric DNA probe for erythrocyte micronuclei showed that epirubicin is not only clastogenic but also aneugenic in somatic cells in vivo. By using the BrdU-incorporation assay, it could be shown that the meiotic delay caused by epirubicin in germ cells was approximately 48 h. Disomic and diploid sperm were shown in epididymal sperm hybridized with DNA probes specific for chromosomes 8, X and Y after epirubicin treatment. The observation that XX- and YY-sperm significantly prevailed over XY-sperm indicates missegregation during the second meiotic division. The results also suggest that earlier prophase stages contribute less to epirubicin-induced aneuploidy. Both the clastogenic and aneugenic potential of epirubicin can give rise to the development of secondary tumors and abnormal reproductive outcomes in cured cancer patients and medical personnel exposed to epirubicin.     

Whole chromosome aneuploidy: Big mutations drive adaptation by phenotypic leap

Despite its widespread existence, the adaptive role of aneuploidy (the abnormal state of having an unequal number of different chromosomes) has been a subject of debate. Cellular aneuploidy has been associated with enhanced resistance to stress, whereas on the organismal level it is detrimental to multicellular species. Certain aneuploid karyotypes are deleterious for specific environments, but karyotype diversity in a population potentiates adaptive evolution. To reconcile these paradoxical observations, this review distinguishes the role of aneuploidy in cellular versus organismal evolution. Further, it proposes a population genetics perspective to examine the behavior of aneuploidy on a populational versus individual level. By altering the copy number of a significant portion of the genome, aneuploidy introduces large phenotypic leaps that enable small cell populations to explore a wide phenotypic landscape, from which adaptive traits can be selected. The production of chromosome number variation can be further increased by stress- or mutation-induced chromosomal instability, fueling rapid cellular adaptation.     

Total aneuploidy and age-related sex chromosome aneuploidy in cultured lymphocytes of normal men and women

Summary In PHA-cultured lymphocytes, about 8% of metaphases from 32 women were aneuploid compared to 4% of metaphases from 35 men. A significant part of this aneuploidy was characterized by sex chromosome involvement: in women, the loss or gain of X chromosomes; in men, the gain of X chromosomes and the loss or gain of Y chromosomes. The incidence of this aneuploidy was positively age-related for both sexes. Premature division of the X-chromosome centromere was closely associated with X-chromosome aneuploidy in women and men, and appeared to be the mechanism of nondisjunction causing this aneuploidy. Premature centromere division (PCD) indicated a dysfunction of the X-chromosome centromere with aging, and this dysfunction was the basic cause of age-related aneuploidy. A similar mechanism of nondisjunction may operate for the Y chromosome of men, but could not be clearly demonstrated because of the low incidence of Y-chromosome aneuploidy.The balance of the aneuploidy was characterized by chromosome loss and the involvement of all chromosome groups. It was consistent with chromosome loss from metaphase cells damaged during preparation for cytogenetic examination.     

Mechanistic investigations of low dose exposures to the genotoxic compounds bisphenol-A and rotenone

A complete hazard and risk assessment of any known genotoxin requires the evaluation of the mutagenic, clastogenic and aneugenic potential of the compound. In the case of aneugenic chemicals, mechanism of action (MOA) and quantitative responses may be investigated by studying their effects upon the fidelity of functioning of components of the cell cycle. These present studies have demonstrated that the plastics component bisphenol-A (BPA) and the natural pesticide rotenone induce micronuclei and modify the functioning of the microtubule organising centres (MTOCs) of the mitotic spindles of cultured mammalian cells in a dose-dependent manner. BPA and rotenone were used as model compounds in an investigation of dose response relationships for the hazard/risk assessment of aneugens. Thresholds of action for the induction of aneuploidy have been predicted for spindle poisons on the basis of the multiple targets, which may need disabling before a quantitative response can be detected. The cytokinesis blocked micronucleus assay (CBMA) methodology was utilised in the human lymphoblastoid cell lines AHH-1, MCL-5 and Chinese hamster V79 cell lines. A no observable effect level (NOEL) at 10.8 microg/ml BPA was observed for MN induction. Rotenone showed a small increase in MN induction with the first significant effect at 0.25 ng/ml in V79 cells but there was no significant effect in the metabolically competent cell line, MCL-5. For a mechanistic evaluation of the aneugenic effects of BPA and rotenone, fluorescently labelled antibodies were used to visualise microtubules (alpha-tubulin) and MTOCs (gamma-tubulin). The NOELs for tripolar mitotic spindle induction in V79 cells were 7 microg/ml for BPA and 80 pg/ml for rotenone (concentrations which produced similar changes to mitotic index (M.I.)). Interestingly there was close proximity to the NOEL of 10.8 microg/ml BPA for micronucleus (MN) induction in the human lymphoblastoid AHH-1 cell. Multiple MTOCs can therefore be predicted as a possible mechanism for MN induction. The similarity in concentration inducing tripolar mitosis, M.I. and MN changes suggests immunofluorescence analysis to be a useful dose setting assay with emphasis on the mechanism.     

5-Azacytidine- and Hoechst-induced aneuploidy in Indian muntjac

Hoechst 33258 (bis-benzimidazole) and 5-azacytidine (5-AC) cause decondensation of the pericentric heterochromatin in mouse and aberrations in the sequence of centromere separation apparently by different mechanisms. We treated the male Indian muntjac cells (2n=7), which do not undergo decondensation of the pericentric heterochromatin, to study if these chemicals would result in induction of aneuploidy limited to the Y(2) chromosome. This paper reports that both agents result in aneuploidy primarily limited to one chromosome, the Y(2). It is likely that other chromosomes are not tolerated in aneuploid condition because every chromosome carries some household genes including those essential for mitotic progression. The loss/gain of the Y(2) chromosome is tolerated because it is the smallest chromosome and is almost entirely composed of constitutive heterochromatin. Since Indian muntjac has only three pairs of large chromosomes comprising its basic genome, which can be clearly viewed under high dry objective, these cells are very suitable for the preliminary analysis of aneuploidy-inducing ability of various chemicals.     

Aneuploidy in Drosophila, IV. Inhalation studies on the induction of aneuploidy by nitriles

The Drosophila ZESTE system was used to monitor the induction of sex chromosome aneuploidy following inhalation exposure of adult females to four nitriles: acetonitrile, propionitrile, acrylonitrile and fumaronitrile. Acetonitrile and propionitrile were highly effective aneuploidogens, inducing both chromosome loss and chromosome gain following brief exposures to low concentrations of these chemicals, and these nitriles also induced rapid paralysis. Acrylonitrile-induced chromosome loss only but did not induce paralysis. Fumaronitrile, in contrast with the results reported in yeast, was ineffective in inducing chromosome loss or gain. Virtually all exceptional offspring induced by acetonitrile and propionitrile were recovered in the first sampled eggs, corresponding to treated mature oocytes. Additionally, the time interval between treatment and sampling was shown to be important, suggesting rapid loss or detoxification of the nitriles. Genetic analysis demonstrated that most aneuploids resulted from induced segregation errors during the first division of meiosis. Cold treatments were found to be ineffective in enhancing the effects of acetonitrile, suggesting important differences between the Drosophila and yeast aneuploidy detection systems. Possible mechanisms by which nitriles may disrupt chromosome segregation in Drosophila oocytes are considered.     

The detection and assessment of the aneugenic potential of selected oestrogens, progestins and androgens using the in vitro cytokinesis blocked micronucleus assay

The use of 17-beta-oestradiol, testosterone, progesterone, zearanol, trenbolone acetate and melengesterol acetate in animal feed as growth promoters has been banned in the European Union since 1989. However, the data available on their genotoxicity is limited. To bridge this gap the present study was carried out with the aim of evaluating these hormones for their ability to induce aneuploidy. Aneuploidy has been recently considered sufficiently important to be included in the routine testing of chemicals and radiation. These types of numerical chromosomal aberrations may arise by at least two mechanisms, chromosome loss and non-disjunction. Over the past few years, the cytokinesis blocked micronucleus (CBMN) technique has evolved into a robust assay for the detection of aneuploidy induction. At the present time, it is the only assay which can reliably detect both chromosome loss and non-disjunction when the basic methodology is coupled with appropriate molecular probing techniques such as immunoflourescent labelling of kinetochores and Fluorescence in situ Hybridisation. In this present study, aneuploidy induction by three groups of hormones was studied using CBMN assay coupled with Fluorescence in situ Hybridisation. The results from the present study demonstrate that 17-beta-oestradiol, diethylstilboestrol, progesterone and testosterone are genotoxic and induce aneuploidy by non-disjunctional mechanism, whereas trenbolone is also genotoxic by a clastogenic mechanism. However, melengesterol acetate and zearanol proved to be non-genotoxic in vitro.     

Karyotypic determinants of chromosome instability in aneuploid budding yeast

Recent studies in cancer cells and budding yeast demonstrated that aneuploidy, the state of having abnormal chromosome numbers, correlates with elevated chromosome instability (CIN), i.e. the propensity of gaining and losing chromosomes at a high frequency. Here we have investigated ploidy- and chromosome-specific determinants underlying aneuploidy-induced CIN by observing karyotype dynamics in fully isogenic aneuploid yeast strains with ploidies between 1N and 2N obtained through a random meiotic process. The aneuploid strains exhibited various levels of whole-chromosome instability (i.e. chromosome gains and losses). CIN correlates with cellular ploidy in an unexpected way: cells with a chromosomal content close to the haploid state are significantly more stable than cells displaying an apparent ploidy between 1.5 and 2N. We propose that the capacity for accurate chromosome segregation by the mitotic system does not scale continuously with an increasing number of chromosomes, but may occur via discrete steps each time a full set of chromosomes is added to the genome. On top of such general ploidy-related effect, CIN is also associated with the presence of specific aneuploid chromosomes as well as dosage imbalance between specific chromosome pairs. Our findings potentially help reconcile the divide between gene-centric versus genome-centric theories in cancer evolution.