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F Degrassi

Publications and source records attributed to F Degrassi.

At least 37 records · Page 2Linked to original sources
PubMed1997

Topoisomerase II inhibition in mitosis produces numerical and structural chromosomal aberrations in human fibroblasts.

We investigated the effects of treatment of mitotic human fibroblasts with the topoisomerase II inhibitor etoposide (VP-16) on chromosome segregation at anaphase and the genetic consequence to daughter cells of topoisomerase inhibition during mitosis. The most striking effect of VP-16 treatment during mitosis was the production of anaphase cells with several entangled chromosomes (catenated anaphase cells). To analyze the effects of sister chromatid catenation at anaphase on the daughter cells, several interphase methodologies were applied to binucleated human fibroblasts that were blocked during cytokinesis. Post-treatment of mitotic cells with the cytokinesis inhibitor cytochalasin-B maintains the reciprocal products of a mitotic division in the same cytoplasm, allowing the distribution of whole chromosomes or chromosome fragments in daughter nuclei or micronuclei to be followed. The presence of micronuclei containing kinetochores, as detected by antikinetochore staining, suggested that VP-16 treatment during mitosis induces chromosome loss in binucleated fibroblasts. Induction of aneuploid cells for chromosomes 7 and 11 was observed by double in situ hybridization using chromosome-specific alphoid probes in binucleated fibroblasts. In addition, double in situ hybridization with adjacent alphoid and classical satellite DNA probes to chromosome 1 demonstrated that both numerical and structural aberrations contribute to the genetic effects of topoisomerase II inhibition in mitosis.

Cell Line↗
PubMed1997

Deregulated expression of the RanBP1 gene alters cell cycle progression in murine fibroblasts.

RanBP1 is a molecular partner of the Ran GTPase, which is implicated in the control of several processes, including DNA replication, mitotic entry and exit, cell cycle progression, nuclear structure, protein import and RNA export. While most genes encoding Ran-interacting partners are constitutively active, transcription of the RanBP1 mRNA is repressed in non proliferating cells, is activated at the G1/S transition in cycling cells and peaks during S phase. We report here that forced expression of the RanBP1 gene disrupts the orderly execution of the cell division cycle at several stages, causing inhibition of DNA replication, defective mitotic exit and failure of chromatin decondensation during the telophase-to-interphase transition in cells that achieve nuclear duplication and chromosome segregation. These results suggest that deregulated RanBP1 activity interferes with the Ran GTPase cycle and prevents the functioning of the Ran signalling system during the cell cycle.

3T3 Cells↗
PubMed1997

Age-related increase of baseline frequencies of sister chromatid exchanges, chromosome aberrations, and micronuclei in human lymphocytes.

Intra- and interindividual variations of baseline frequencies of cytogenetic end points in lymphocytes of human populations have been reported by various authors. Personal characteristics seem to account for a significant proportion of this variability. Several studies investigating the role of age as a confounding factor in cytogenetic biomonitoring found an age-related increase of micronucleus (MN) frequency, whereas contradictory results were reported for chromosomal aberrations (CAs) and sister chromatid exchanges (SCEs). We have quantitatively evaluated the effect of age on SCE, CA, and MN through the analysis of a population sample that included data from several biomonitoring studies performed over the last few decades in 12 Italian laboratories. The large size of the data set, i.e., more than 2000 tests for each end point, allowed us to estimate the independent effect of age, taking into account other covariates, such as sex, smoking habits, occupational exposure, and inter- and intralaboratory variability. A greater frequency of the mean standardized values by increasing of age was observed for all of the end points. A leveling off was evident in the last age classes in the trend of MN frequencies. Frequency ratios (FRs), which express the increase of the cytogenetic damage with respect to the first age classes, i.e., 1-19 years, were estimated using Poisson regression analysis after adjustment for the potential confounding factors and confirmed the increasing trend by age class for all three end points. The most dramatic increase was observed for MN, with a FR that approaches the value of 2 at the age class 50-59 (FR, 1.97; 95% confidence interval, 1.43-2.71) and remains substantially unchanged thereafter. The trend of FRs for CA is more homogeneous, with a constant rise even in the older classes, whereas the frequency of SCE increases with age to a lesser extent, reaching a plateau in the age class 40-49 and the maximum value of FR in the age class over 70 (FR, 1.14; 95% confidence interval, 1.07-1.23). In conclusion, our results point to an age-related increase of the chromosome damage in lymphocytes and emphasize the need to take into account the potential confounding effect of this variable in the design of biomonitoring studies based on chromosome damage.

Adolescent↗
PubMed1996

The detection and evaluation of aneugenic chemicals.

Although aneuploidy makes a significant contribution to both somatic and inherited disease the mechanisms by which environmental chemicals may induce numerical chromosome aberrations are only poorly defined. The European Union Project was aimed to further our understanding of those chemical interactions with the components of the mitotic and meiotic cell division cycle which may lead to aneuploidy and to characterise the parameters such as cellular metabolism which may influence the activity of aneugenic chemicals. C-mitosis can be induced by the highly lipophilic polychlorinated biphenyl and the completion of mitosis and cleavage can be modified by agents which deplete cellular levels of reduced glutathione. Modifications of the fidelity of chromosome segregation were produced by inhibiting the functioning of topoisomerase II during chromatid separation. In contrast, the modification of centromere integrity resulted in chromosome breakage as opposed to disturbance of segregation. Modifiers of tubulin assembly and centriolar functioning in somatic cells such as acrylamide, vinblastine and diazepam reproduced their activity in rodent bone marrow and male germ cells. The analysis of chromosome malsegregation in Aspergillus nidulans by a structurally related series of halogenated hydrocarbons was used to develop a QSAR model which had high predictive value for the results of fungal tests for previously untested related chemicals. Metabolic studies of potential aneugens in genetically engineered human lymphoblastoid cells demonstrated the detoxification of the aneugenic activity of chloral hydrate and the activation of 2,3-dichlorobutane, 1,1,2-trichloroethane and trichloroethylene by Phase I biotransforming enzymes. Cell transformation studies in Syrian hamster dermal cultures using a panel of 22 reference and or potential aneugens indicated that 15 of the 22 produced positive results following single exposures. Five of the aneugens which were negative following single exposures produced positive results where cultures were continuously exposed for up to 6 weeks to low concentrations following a single non-transforming exposure to the mutagen dimethyl sulphate. The transformation studies indicate that a significant proportion of chemical aneugens are potential complete carcinogens and/or co-carcinogens. To optimise the enumeration of chromosomes following exposure to potential chemical aneugens whole chromosome paints and centromere specific probes suitable for use in fluorescence in situ hybridisation (FISH) were developed for the rat, mouse and Chinese hamster and selected human probes evaluated for their suitability for routine use. Molecular chromosome probes were used to develop protocols for enumerating chromosomes in metaphase cells and centromeres and micronuclei in interphase cells. The analysis of segregation of specific centromeres in binucleate cells following cytochalasin B treatment was shown to be a potentially valuable system for characterising non-disjunction following chemical exposure. Whole chromosome paints and centromere specific probes were used to demonstrate the presence of dose-response thresholds following treatment with a reference panel of spindle inhibiting chemicals. These data indicate that the FISH technology is suitable for evaluating the relative hazards of low-dose exposures to aneugenic chemicals.

Aneuploidy↗
PubMed1996

Topoisomerase I activity and cellular response to radiation in Chinese hamster cells.

The aim of the present study was to investigate the synergistic potential of the combination of camptothecin, a specific inhibitor of topoisomerase I, and radiation in the the induction of chromosome aberrations and cell cycle delay in actively proliferating mammalian cells. Synergistic effects of the combined treatments were obtained for induced frequencies of aberrations in exponentially growing Chinese hamster ovary cells. The potentiating effects were more pronounced for aberrations of the exchange type, suggesting that interaction of unrepaired radiation- and camptothecin-induced lesions during replication may be involved in the observed drug-radiation synergism. Cytofluorimetric analysis of cell cycle progression in cells receiving the combined treatments displayed enhanced responses of CHO cells to S- and G2 phase delay induced by the single treatments. To investigate the determinants of the synergistic response, the influence of radiation exposure on the catalytic activity of topoisomerase I was assayed. A decreased plasmid supercoiled DNA relaxation capacity of crude extracts derived from irradiated CHO cells was found which suggests a decrease in the topoisomerase I catalytic activity following irradiation. In addition, a lower sensitivity of the enzyme from irradiated cells to inhibition of topoisomerase I activity by camptothecin was also observed using the same DNA relaxation test.

Animals↗
PubMed1996

Effects of 5-azacytidine on the centromeric region of human fibroblasts studied by CREST staining and in situ hybridization on cytokinesis-blocked cells.

Several interphase methodologies have been applied in this study to investigate whether chemically induced undercondensation of the pericentromeric region of human fibroblast chromosomes promotes structural or numerical aberrations at the cell cycles following treatment. To achieve this aim, the effects of the hypomethylating agent 5-azacytidine (5- azaC) were studied on cytokinesis-blocked binucleated fibroblasts. This approach allowed the distribution of whole chromosomes or chromosome fragments in the daughter nuclei and micronuclei of treated cells to be followed, since the daughter nuclei of a single mitosis are maintained in one cytoplasm by treatment with the actin inhibitor cytochalasin B. Antikinetochore staining and in situ hybridization with an alpha-satellite probe capable of detecting all human centromeres on 5-azaC-induced micronuclei in binucleated fibroblasts indicated that the predominant effect of the chemical is to induce micronuclei lacking centromeres, suggestive of induced chromosome aberrations. Double in situ hybridization with alphoid and classical satellite DNA probes specific for chromosome 1 on binucleated fibroblasts was used to discriminate induced aneuploidy from breakage effects in the target area of hybridization. The results showed that undercondensation of the pericentromeric heterochromatin of human fibroblast chromosomes by treatment with 5-azaC produces structural chromosome aberrations involving the classical satellite DNA, whereas there was no evidence that the chemical induced chromosomal aneuploidy.

Azacitidine↗
PubMed1995

Genetic effects of petroleum fuels: cytogenetic monitoring of gasoline station attendants.

Workers in the petroleum distribution trades experience relatively high-level exposures to fuel vapours whose consequences have not been fully elucidated. In this study, the possible relationship between occupational exposure to petroleum fuels and cytogenetic damages in peripheral lymphocytes was investigated. Twenty-three male, non-smoking workers from the area of Rome were enrolled in the study, together with age-paired controls with no occupational exposure to fuels. Peripheral lymphocyte cultures were set up for the analysis of structural chromosome aberrations (CAs), sister chromatid exchanges (SCEs) and micronuclei (MN) in cytokinesis-blocked lymphocytes. Frequencies of CAs, SCEs and MN were compared between exposed and control groups, and evaluated in relation to blood lead level (as an indicator of engine exhausts exposure) for the whole group under study, and to yearly averaged exposure to benzene (8-h time weighted averages, as determined by repeated personal sampling) for fillingstation attendants only. Both CAs and SCEs were slightly increased in station attendants: 1.97 versus 1.46 aberrations per 100 cells, and 4.73 +/- 0.15 versus 4.48 +/- 0.11 SCEs/cell in exposed and control individuals, respectively. The difference between cumulative CA rates in the exposed and control populations was of borderline statistical significance (p = 0.066). However, when the exposed population was dichotomized for benzene exposure, a significant (p = 0.018) correlation of CAs with benzene exposure was found. The analysis of SCE data highlighted a significant increase of cells with more than 6 exchanges (HFCs), corresponding to the 75 degrees percentile of the overall distribution, in fillingstation attendants (relative risk (RR) = 1.3, 95% CI = 1.1-1.5) in comparison with controls. In the pooled population, the frequency of HFCs showed a statistically significant upward trend at increasing blood lead levels (chi 2 for trend = 27.8, p < 0.0001). A complex relationship between SCEs and benzene exposure was observed, with an increased frequency of HFCs in the medium exposure intensity class (RR = 1.5, 95% CI = 1.2-1.7), and no difference for exposure to higher benzene levels (RR = 1.0, 95% CI = 0.9-1.2), compared to reference subjects. Finally, the analysis of MN in both phytohemagglutinin- and pokeweed-stimulated cell cultures did not show significant excess of MN in binucleated lymphocytes of exposed workers with respect to the age-paired controls.

Adult↗
PubMed1995

Genotoxic activity of nitrilotriacetic acid in Chinese hamster cells.

Nitrilotriacetic acid (NTA), a chelating agent, was tested for its ability to induce chromosomal damage in Chinese hamster cells. The chemical was shown to exert a weak genotoxic activity increasing the frequency of micronuclei after prolonged treatments. The analysis of kinetochore containing-micronuclei showed that NTA prevailingly induces chromosomal aberrations as compared to chromosome loss in hamster cells. Furthermore, immunostaining with an alpha-tubulin antibody showed clear alterations in the interphase microtubule network of cells treated for 24 h with 3 mM NTA. The microtubule effects of the chemical may be partly responsible for its cytotoxic effects.

Animals↗
PubMed1995

Are chromosome aberrations in circulating lymphocytes predictive of future cancer onset in humans? Preliminary results of an Italian cohort study.

To investigate the existence of an association between the frequency of chromosome aberrations (CA) in non-target tissues and cancer risk, a historical cohort study was carried out in a group of 1455 subjects screened for CA over the last 20 years in Italy. Statistically significant increases in standardized mortality ratio (SMR) for all cancers were found in subjects with medium and high levels of CA in peripheral blood lymphocytes (SMR = 178.5 and SMR = 182.0, respectively) and in subjects with high levels of CA for respiratory tract cancers (SMR = 250.8) and lymphatic and hematopoietic tissue neoplasms (SMR = 548.8). Significant trends in the SMRs were observed for these latter causes of death.

Chromosome Aberrations↗
PubMed1994

Mitotic indirect non-disjunction in phytohemagglutinin stimulated human lymphocytes.

In a previous publication we demonstrated that in cells of Vicia faba micronuclei derived from whole lagging chromosomes or chromatids may perform DNA synthesis and mitotic condensation in synchrony with main nuclei and be regained by main nuclei at the next mitosis, giving rise to trisomic cells together with diploids. This process was called 'mitotic indirect non-disjunction' (MIND). In the present work the occurrence of MIND was studied in human lymphocytes cultivated in vitro. Human lymphocytes were treated with low colcemid concentrations until fixation; BrUdR was supplied together with colcemid to distinguish the number of mitoses performed by the cells (M1, M2 and M3 cells). The frequencies of M1 ana-telophases with single lagging chromosomes/chromatids and of M2+ prophases with single micronuclei in synchronous motitic condensation with main nuclei were evaluated. On this basis the expected frequencies of both monosomic and trisomic M2 cells were calculated, according to the hypothesis of MIND. Their observed frequencies were very close to those expected. These results support the hypothesis of the occurrence of MIND in human lymphocytes.

Aneuploidy↗
PubMed1994

The effect of cytochalasin-B concentration on the frequency of micronuclei induced by four standard mutagens. Results from two laboratories.

In a previous collaborative work, we have recently shown that the cytochalasin-B (Cyt-B) concentration used in the human lymphocytes cytokinesis-block micronucleus (CBMN) assay is an important variable in the baseline micronuclei (MN) frequency as well as in the percentage of binucleated cells obtained. Now we have investigated how Cyt-B concentration modulates the MN frequency induced in whole blood human lymphocyte cultures by two clastogens (ethyl methanesulphonate and mitomycin-C) and two aneugens (colchicine and vincristine sulphate). The experimental design includes six donors, two concentrations of Cyt-B (3 and 6 micrograms/ml), two concentrations of the four chemicals tested and the exchange of slides between laboratories. The statistical analysis of the results shows: (i) non-significant differences in the MN frequencies and in the toxicity results between scorers from each laboratory, except for 0.06 microM colchicine at 3 micrograms/ml Cyt-B; (ii) an induction of MN by all genotoxic agents tested, the frequencies being lower with 6 than with 3 micrograms/ml Cyt-B, in control and aneugen-treated cultures; and (iii) significant differences between Cyt-B concentrations in several treatments, obtaining lower MN frequencies and higher values for nuclear division index and % binucleated cells when 6 micrograms/ml Cyt-B was used. Bearing in mind these results as well as the toxicity data showing that 6 micrograms/ml Cyt-B is much more effective in blocking cytokinesis, we can conclude that the use of 3 micrograms/ml Cyt-B may overestimate the induced frequency of MN.

Adult↗
PubMed1993

The production of chromosomal alterations by beta-lapachone, an activator of topoisomerase I.

The frequencies of chromosomal aberrations and sister-chromatid exchanges (SCE) after exposure to beta-lapachone, an activator of mammalian topoisomerase I, were studied in Chinese hamster cells. A dose-dependent increase in the frequencies of SCE was observed in continuous treatments with beta-lapachone. Chromatid-type aberrations were obtained in cells exposed to beta-lapachone for one cell cycle but also in cells exposed during the G2 phase of the cell cycle, with a marked induction of exchange-type aberrations for both treatment schedules. We therefore propose that activation of topoisomerase I by beta-lapachone results in the production of chromosomal alterations. The cell cycle dependence of beta-lapachone clastogenic effects strongly suggests a mechanism for the formation of chromosomal aberrations after this drug closely resembling the one observed for the topoisomerase I inhibitor, camptothecin.

Animals↗
PubMed1993

Cytokinesis-block micronucleus assay with kinetochore detection in colchicine-treated human fibroblasts.

A modified micronucleus assay using antikinetochore antibody has been developed in cytokinesis-blocked human fibroblasts as a simple method to identify aneuploidy-inducing agents. Different protocols for inducing binucleated cells by cytochalasin B in colchicine-treated human fibroblasts were investigated. A dose-related increase in kinetochore-positive micronuclei was obtained when cytochalasin B was given subsequent to colchicine treatment. No induction of micronuclei was observed in combined treatments of the two substances. These results indicate that the detection of kinetochores in micronucleated cytokinesis-blocked human fibroblasts can be effectively applied to the identification of environmental agents with aneuploidy-inducing potential. However, in testing such compounds particular attention should be paid to the protocol used for inducing cytokinesis-blocked cells.

Cell Division↗
PubMed1993

Hypersensitivity of lymphoblastoid lines derived from ataxia telangiectasia patients to the induction of chromosomal aberrations by etoposide (VP-16).

Mammalian DNA topoisomerase II represents the cellular target of many antitumor drugs, such as epipodophyllotoxin VP-16 (etoposide). The mechanism by which VP-16 exerts its cytotoxic and antineoplastic actions has not yet been firmly established, although the unique correlation between sensitivity to ionizing radiation and to topoisomerase II inhibitors suggest the involvement of DNA double-strand breaks. In the present study we analyzed the chromosomal sensitivity of lymphoblastoid cell lines derived from ataxia telangiectasia (AT) patients to low concentrations of the drug. Our results indicate that AT derived cells are hypersensitive to the clastogenic activity of VP-16 either when the drug is present for the whole duration of the cell cycle or specifically in the G2 phase, confirming that the induction of DNA double strand breaks, to which AT cells seem typically sensitive, could have an important role in the biological activity of VP-16.

Ataxia Telangiectasia↗
PubMed1993

Sensitivity of lymphocytes from vulcanizers to the in vitro induction of sister chromatid exchanges.

Spontaneous frequencies of sister chromatid exchanges (SCEs) and SCEs induced in vitro by chemicals with different mechanisms of action such as mitomycin C, 4-nitroquinoline oxide, and 3-aminobenzamide were examined in phytohemagglutinin-stimulated peripheral blood lymphocytes from a group of workers in a rubber plant and a control group, both of which had been analyzed for levels of spontaneous SCEs 2 years earlier. An interindividual variability in the induction of SCEs was found after in vitro treatments with the different mutagens, which did not correlate with occupational exposure. This variability in the sensitivity to the induction of SCEs might be correlated to genetic differences among individuals, which have to be taken into account in environmental monitoring programs.

Adult↗
PubMed1992

A collaborative study on the improvement of the micronucleus test in cultured human lymphocytes.

A collaborative study between two laboratories has been carried out in order to investigate the reproducibility of the cytokinesis-block micronucleus assay in human lymphocytes. A modified protocol to obtain good quality slides from whole blood cultures has been developed. The spontaneous frequencies of micronuclei in binucleated lymphocytes from five male subjects were evaluated in each laboratory at two cytochalasin-B concentrations (3 and 6 micrograms/ml). Our data show that 6 micrograms/ml cytochalasin-B was more effective than 3 micrograms/ml to arrest cytokinesis in all the donors analysed. The mean frequency of micronuclei in binucleated cells was significantly lower at 6 micrograms/ml compared to 3 micrograms/ml cytochalasin-B in both laboratories. The results obtained indicate a very high concordance between both sets of data supporting the validity of the proposed protocol.

Adult↗
PubMed1991

In vitro micronucleus test with kinetochore staining: evaluation of test performance.

In the framework of the coordinated programme 'Genomic Mutations' sponsored by the Commission of European Communities, eight known or suspected spindle poisons (cadmium chloride, chloral hydrate, colchicine, diazepam, econazole, hydroquinone, pyrimethamine, thiabendazole, thimerosal and vinblastine) were tested in a modified in vitro micronucleus test in Chinese hamster cells. Micronuclei (MN) with or without kinetochore were analyzed by staining of the kinetochore with an antikinetochore antibody (CREST staining). Mitotic index and ana-telophase:mitosis ratio were also recorded to evaluate cytotoxicity and c-mitotic effects of tested chemicals. CREST-positive MN were induced by diazepam, thiabendazole and vinblastine. Hydroquinone, pyrimethamine, econazole and cadmium chloride induced MN that contained both entire chromosomes and acentric fragments. Negative results were obtained with thimerosal. The results obtained indicate that the detection of CREST-positive MN is a powerful assay to identify spindle poisons. Furthermore, this method provides a useful tool to ascertain the origin(s) of induced MN.

Aneuploidy↗