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At least 145 records · Page 8Linked to original sources
PubMed1990

Enhanced G2 chromatid radiosensitivity in dyskeratosis congenita fibroblasts.

Dyskeratosis congenita (DC) is an inherited disorder characterized by reticular pigmentation of the skin, dystrophic nails, mucosal leukoplakia, and a predisposition to cancer in early adult life. In the majority of cases, DC is an X-linked recessive trait. However, one or more autosomal form(s) of DC may exist. Although excessive spontaneous chromatid breakage has been reported in DC, it is not a consistent cytological marker for this disorder. We examined the frequency and specificity of X-irradiation-induced G2 chromatid breakage in fibroblasts from three unrelated DC patients (two males and one female). Metaphase cells from DC patients had significantly more chromatid breaks (16-18-fold and 17-26-fold at 50 and 100 rad X-irradiation, respectively) and chromatid gaps (10-12-fold and 6-7-fold at 50 and 100 rad, respectively) than those from two different controls. Analysis of banded chromosomes revealed a nonrandom distribution of chromatid aberrations in DC but not in controls, a distribution corresponding to some of the known breakpoints for cancer-specific rearrangements, constitutive fragile sites, and/or loci for cellular proto-oncogenes. The significance of this finding for cancer predisposition in DC patients is uncertain, but the increased susceptibility of X-irradiation-induced chromatid breakage may serve as a cellular marker of diagnostic value.

Cell Line↗
PubMed1986

Differential inhibition of sister chromatid condensation induced by 5-azadeoxycytidine in human chromosomes.

The deoxycytidine analogue 5-azadeoxycytidine (5-aza-dC) induces differential inhibition of sister chromatid condensation when cells are treated with this substance for two replication cycles, as the subsequent staining of metaphase chromosomes with Giemsa shows. The bifilarly substituted chromatid is dramatically longer than the unifilar one. A percentage of the metaphases treated with 5-aza-dC even show a complete undercondensation of the bifilarly substituted chromatid. The optimum conditions for inducing sister chromatid differentiation were determined. No method has been developed as yet to permit enhancement of the differential staining in 5-aza-dC-treated preparations. The interactions between 5-aza-dC and chromosomal DNA as well as the factors involved in the differential staining of sister chromatids are discussed.

Azacitidine↗
PubMed1997

A change in sister chromatid behavior precedes nuclear division in Saccharomyces cerevisiae.

We used a genetic assay to monitor the behavior of sister chromatids during the cell cycle. We show that the ability to induce sister chromatid exchanges (SCE) with ionizing radiation is maximal in budded cells with undivided nuclei and then decreases prior to nuclear division. SCE can be induced in cells arrested in G2 using either nocodazole or cdc mutants. These data show that sister chromatids have two different states prior to nuclear division. We suggest that the sister chromatids of cir. III, a circular derivative of chromosome III, separate (anaphase A) prior to spindle elongation (anaphase B). Other interpretations are also discussed. SCE can be induced in cdc mutants that arrest in G2 and in nocodazole-treated cells, suggesting that mitotic checkpoints arrest cells prior to sister chromatid separation.

Anaphase↗
PubMed1995

Frontier questions about sister chromatid separation in anaphase.

Sister chromatid separation in anaphase is an important event in the cell's transmission of genetic information to a descendent. It has been investigated from different aspects: cell cycle regulation, spindle and chromosome dynamics within the three-dimensional cell architecture, transmission fidelity control and cellular signaling. Integrated studies directed toward unified understanding are possible using multidisciplinary methods with model organisms. Ubiquitin-dependent proteolysis, protein dephosphorylation, an unknown function by the TPR repeat proteins, chromosome transport by microtubule-based motors and DNA topological change by DNA topoisomerase II are all necessary for progression from metaphase to anaphase. Chromosome condensation, mitotic kinetochore function and spindle formation require a larger number of proteins, which are prerequisites for successful sister chromatid separation. Factors that help to retain sister chromatid connection after replication and prevent premature separation remain to be determined. Although sister chromatid separation occurs in anaphase, gene functions in other cell cycle stages also ensure the progression of correct chromatid separation.

Anaphase↗
PubMed1993

Chromatid contamination can impair the purity of flow-sorted metaphase chromosomes.

We describe a phenomenon where a second flow karyotype is superimposed on the normal metaphase flow karyotype of human lymphoblastoid chromosomes. The events of this second flow karyotype contain half the DNA content of corresponding events in the normal flow karyotype and, when sorted, show the morphology of single chromatids. DNA amplified by degenerate oligonucleotide-primed PCR of 300 chromatids sorted from a single peak and hybridized onto normal metaphase spreads painted the entire length of the corresponding chromosome type. In retrospect, we could observe this phenomenon at a low level in 37% of randomly selected, previously analyzed flow karyotypes and in several published flow karyotypes from other laboratories. Experimentally, chromatid frequency was shown to vary with the colcemid blocking conditions as well as with the cell line used. At the extreme, we obtained a 4:10 ratio of chromatids to metaphase chromosomes, whereas this could be reduced tenfold by altering the colcemid blocking conditions in the same cell line. The presence of chromatids in chromosome preparations has important implications for the purity of isolated fractions used for generating libraries and in PCR analysis.

Cell Line↗
PubMed1993

Chromatin bridges between sister chromatids induced in late G2 mitosis in CHO cells by trimethylpsoralen + UVA.

A hypothesis is proposed that sister chromatid pairing is due, at least in part, to the pairing between DNA strands belonging to each of the two sister chromatids (conservative pairing). To test this hypothesis interstrand DNA cross-links were induced in late G2-mitosis in CHO cells in order to bind covalently paired DNA strands eventually coming from both sister chromatids and detect the consequent chromatin bridges between sister chromatids (SCCBs). Therefore cells were treated with trimethylpsoralen (TMP) + UVA (365 or 405 nm). Chromatin bridges in ana-telophase were induced by an UVA irradiation at 365 nm, which gives rise to both monoadducts and cross-links, but not by a 405-nm irradiation, which gives rise only to monoadducts. An analysis of colchicine-induced c-anaphases demonstrated that such chromatin bridges were really SCCBs and that terminal regions of chromosomes were particularly involved. The evolution of SCCBs was studied to rule out that they were masked isochromatid exchanges. So TMP + UVA-treated cells were induced to polyploidize with colchicine and labeled with 5-bromodeoxyuridine. Cells treated with TMP + UVA in G2-mitosis appeared as M1 tetraploid c-metaphases; in such cell populations there was not an increase in isodicentric chromosomes, which are derived from isochromatid exchanges. The present data, as a whole, support the hypothesis that a "conservative pairing" between DNA strands of sister chromatids can be present in mitosis.

Animals↗
PubMed1980

Chromosome aberration formation and sister chromatid exchange in relation to DNA repair in human cells.

Apparent association between the ability to induce chromosome aberrations and sister chromatid exchanges and mutagenic-carcinogenic potential found in a variety of physical and chemical agents has led us to speculate that these cytogenetic changes might be reflection of DNA damage and repair and might provide induces of mutagenic changes. However, the mechanisms of their formation and their relation to DNA repair as well as the mechanism of their linking to mutation are by no means well understood. Studies in some human genetic mutant cells defective in their ability to repair DNA damage indicate, as a testable proposition, that sister chromatid exchanges and chromosome aberrations are cytological manifestations of replication-mediated dual-step repair pathways that are in operation to tolerate DNA damage when damage-bearing DNA enters and passes through semiconservative replication. The observations are also in line with idea that the majority of sister chromatid exchanges can arise when damage DNA attempts replication possibly by a process relating with the replicative bypass repair mechanisms such as those proposed by Fujiwara and Tatsumi [34] and Higgins et al. [54], while chromosome aberration formation and some fraction of sister chromatid exchanges are related with the post-replication repair processes which attempt to rescue damaged template post-replicationally by de novo synthesis or recombination type repair systems. The former sister chromatid exchange-relating process seems to link mutation to less extent, if any, than the latter process, which is caffeine sensitive and likely to be error-prone.

Ataxia Telangiectasia↗
PubMed1986

Induction of sister chromatid exchanges by inhibitors of topoisomerases.

To investigate the role of topoisomerases in the production of sister chromatid exchanges, the effects of inhibitors of type I and II topoisomerases on baseline and mutagen-induced sister chromatid exchanges were compared. V79 cells were treated with VM-26 and m-AMSA, known inhibitors of type II topoisomerase, or with camptothecin, the only known inhibitor of type I topoisomerase. We observed that inhibitors of both type I and II topoisomerases induced high levels of sister chromatid exchanges at 10(-6) M, and that the dose-response curves of these drugs were very similar. A clear heterogeneity in the distribution patterns of exchanges induced by inhibitors of topoisomerases was observed. We believe that this heterogeneity in response to these compounds is due to variation in sensitivity within the cell cycle. We also studied interactions of these agents with mitomycin-C and with PUVA (8-methoxypsoralen + UVA), both cross-linking agents and potent sister chromatid exchange inducers, and with x-rays, an agent that induces high levels of DNA strand breaks. No significant change in exchange levels was observed in interactions between topoisomerase inhibition and the levels induced by the agents studied. We conclude that double-strand break prevalence, known to be increased through inhibition of type II topoisomerase, is not the primary mechanism for induction of sister chromatid exchanges. We further conclude that acute inhibition of type I and type II topoisomerases does not influence substantially the induction of exchanges by other agents.

Amsacrine↗
PubMed1981

Topographic examination of sister chromatid differential staining by Nomarski interference microscopy and scanning electron microscopy.

BrdU-substituted Chinese hamster chromosomes were treated with a hog Na2HPO4 solution and stained with Giemsa to produce sister chromatid differential staining (SCD). The process of SCD was examined with the Nomarski differential interference microscope and the scanning electron microscope. After the Na2HPO4 treatment alone, unifilarly BrdU-substituted (TB) chromatids appeared somewhat more severely collapsed than the bifilarly substituted (BB) chromatids. Subsequent Giemsa staining, however, brought about pronounced piling up of the Giemsa dye on the TB-chromatids but not on the BB-ones, causing highly distinct differential Giemsa staining as well as a marked differentiation in surface topography between the sister chromatids. Removal of the Giemsa dye from the differentially Giemsa stained chromosomes resulted in a disappearance of such a pronounced topographic differentiation.

Animals↗
PubMed1979

Chromosome breakage and rejoining of sister chromatids in Bloom's syndrome.

The occurrence of chromosome breaks and reunion of sister chromatids in lymphocytes of two patients with Bloom's syndrome has been compared with those found in X-rayed and control cells. The distribution of breaks in BS is non-random both between and within chromosomes, the centric regions of certain chromosomes being preferentially involved. The following working hypotheses are put forward: When chromosome breaks in human lymphocytes occur in G0-G1, practically no sister chromatid reunion (SCR) takes place, whereas ends created by an S-G2 break show a considerable tendency to SCR. We propose further that chromosome aberrations in BS mainly result from breaks in S-G2, including possible U-type rejoining of sister chromatid exchanges. Fragments extra to an intact chromosome complement result from a chromatid break or an asymmetrical chromatid translocation in a previous mitosis.

Cells, Cultured↗
PubMed1981

Selective differential staining of sister chromatids of the facultative heterochromatic X chromosome in the female mouse.

Selective differential staining of sister chromatids for the facultative heterochromatic X chromosome in the female mouse has been achieved by the combination of two differential staining techniques; one for the heterochromatic X chromosome and the other for sister chromatids. Thermal hypotonic treatment moderately destroyed the chromosome structure except for the heterochromatic X in BrdU labelled metaphase cells, resulting in the selective sister chromatid differentiation of this X with Giemsa stain. This technique enables us to know the exact frequency of th spontaneous sister chromatid exchanges in the heterochromatic X without using 3H-TdR labelling for detecting the late DNA replication. The results indicate that the sister chromatid exchange frequency of the heterochromatic X chromosome is not affected by its late DNA replication during S phase, or by the genetic inactivation and the resulting heterochromatinization.

Animals↗
PubMed1985

Reverse differential staining of sister chromatids induced by Hoechst plus black light and endonuclease.

A differential Giemsa staining between sister chromatids was obtained by treating chromosomes replicated twice in medium containing 5-bromodeoxyuridine (BrdU) with Hoechst 33258 plus black light at 55 degrees C (HB pretreatment) and deoxyribonuclease (DNase) I, II, or micrococcal nuclease. In this staining pattern the BrdU bifilarly substituted chromatids were darkly and the unifilarly substituted chromatids lightly stained. This staining pattern was obtained only by staining the HB-DNase I-treated chromosomes with Giemsa and methylene blue, not by several other dyes tested. Relatively more DNA labelling was removed from the non-BrdU-substituted than the BrdU-substituted chromosomes, when the HB-pretreated chromosomes were digested with DNase I. But the protein labelling was not removed appreciably in the same treatment. The differential DNase I sensitivity between the non-BrdU-substituted and BrdU-substituted chromosomes disappeared when the HB-pretreated chromosomes were incubated with proteinase K before The DNase I digestion. Moreover, no differential DNase I sensitivity was found between the HB-pretreated isolated DNA containing and not containing BrdU. We propose that during the HB pretreatment, more DNA-protein cross-linkings are induced in BrdU bifilarly substituted than the unifilarly substituted chromatids. This structure protects the chromosomal DNA against the DNase I digestion. Thus, a reverse differential Giemsa staining between sister chromatids is obtained by the HB-DNase I treatment.

Animals↗
PubMed1975

X-ray and UV-induced chromatid aberrations: evidence for polynemic chromosomes?

Ikushima and Wolff have recently interpreted both their observation of chromatid aberrations in second and third mitoses following X-irradiation and the production of chromatid type chromosomal aberrations by UV light administered during the G1 phase of the cell cycle in terms of a polyneme model of eukaryote chromosome structure. They were led to do so, however, largely because of their X-ray data, which the interpreted as evidence for the induction of sub-chromosomal lesions (by G1 irradiation; sub-chromatid for G2 irradiation) which appear as chromatid type aberrations only in later divisions. We here report data from similar X-ray experiments in which synchronized Chinese hamster tissue culture cells were irradiated in either G1 or G2 and then scored for chromatid aberrations in their first, second and third post-irradiation mitoses. Our results do not show the effect reported by Ikushima and Wolff. We conclude that all of the data available of aberration production is compatible with a simple mononeme model of eukaryote chromosome structure.

Animals↗
PubMed1988

Influence of dietary carrot on cytostatic drug activity of cyclophosphamide and its main directly acting metabolite: induction of sister-chromatid exchanges in normal human lymphocytes, Chinese hamster ovary cells, and their DNA repair-deficient cell lines.

We have utilized an in vivo drug metabolism technique (i.e. injecting the chemical into rat and isolating plasma with metabolites from blood) for detecting the genotoxicity of indirectly acting cyclophosphamide and its directly acting metabolite phosphoramide mustard in cultures of human peripheral blood lymphocytes of normal individuals, Fanconi's anaemia (FA) and aplastic anaemia (AA) patients, wild-type Chinese hamster ovary cells (CHO) and its DNA repair-deficient mutant 43-3B cells. In addition, the influence of dietary carrot on the clastogenic activity of these 2 chemicals in all the different cell types was studied. The genotoxicity was assessed by the ability of the metabolites of these agents to induce sister-chromatid exchanges in the treated cells. A dose-dependent increase in the frequencies of sister-chromatid exchanges was observed in all cell strains following treatment with activated metabolites of cyclophosphamide or phosphoramide mustard. The sensitivity of lymphocytes from normal donors, FA and AA patients to these 2 chemicals was similar. In CHO cell lines the induced frequency of sister-chromatid exchanges was slightly higher after treatment with the metabolites of cyclophosphamide than with phosphoramide mustard. The mutant 43-3B cells responded with higher frequencies of SCEs when compared to the wild-type CHO cells, about 1.5-2-fold, at low doses. Pretreating of rats with fresh carrot juice effectively inhibited the increase in the frequencies of sister-chromatid exchanges induced by cyclophosphamide in wild-type and mutant CHO cells (P less than 0.01), and to a lesser extent in human lymphocytes (p less than 0.05). In contrast, no inhibitory effect was observed in any of these cell types in combination of dietary carrot for direct acting phosphoramide mustard on the frequency of induced sister-chromatid exchanges. The possibility that dietary carrot exerts its antimutagenic effect by affecting the processes of enzymatic activation of cyclophosphamide is discussed.

Anemia, Aplastic↗
PubMed1996

Effect of melatonin on mitotic and proliferation indices, and sister chromatid exchange in human blood lymphocytes.

Cells from human peripheral blood were cultured in vitro in the presence of 0.05 to 1.00 mM melatonin, 10(-7) M mitomycin C (positive control) and 0.5% ethanol (solvent control) for 72 h at 37 +/- 1 degree C. Lymphocytes were examined for mitotic and proliferation indices, and for the incidence of sister chromatid exchange. The results indicate that the lymphocytes which were cultured in the presence of > or = 0.20 mM concentrations of melatonin exhibited a significant and concentration-dependent decrease in mitotic index and alteration in proliferation kinetics. This was demonstrated by an increase in the frequency of lymphocytes in their first division, with a concomitant decrease in the second and third or later division cells. The incidence of sister chromatid exchange was similar in the lymphocytes exposed to 0.05 to 1.00 mM melatonin and untreated controls. Exposure of the cells to ethanol, the solvent used, did not alter either the mitotic or proliferation indices, or the frequency of sister chromatid exchange. The lymphocytes treated with mitomycin C showed the expected decrease in mitotic and proliferation indices, and an increased incidence of sister chromatid exchange. These observations indicate that melatonin, when continuously present in the cultures for 72 h at the concentrations tested, while not genotoxic as indicated by the sister chromatid exchange assay, inhibits the proliferation of mitogen stimulated (and proliferating) human blood lymphocytes at supraphysiological concentrations.

Adult↗
PubMed1984

Sister-chromatid exchanges and chromosome aberrations in lymphocytes from monkeys exposed to ethylene oxide and propylene oxide by inhalation.

The ability of long-term exposures to inhaled ethylene oxide (EO) and propylene oxide (PO) to induce sister-chromatid exchanges (SCEs) and chromosome aberrations in peripheral lymphocytes of monkeys was investigated. Five groups of adult male cynomolgus monkeys were exposed at 0 (shared control), 50, or 100 ppm EO, and at 100 or 300 ppm PO (7 hr/day, 5 days/week) for 2 years. EO exposures at 50 and 100 ppm resulted in statistically significant increases in sister-chromatid exchange rates and in the incidence of chromosome aberrations in monkey lymphocytes. Both EO-exposed groups had increased numbers of SCEs/metaphase compared to controls, with the SCEs/metaphase of the EO 100 ppm group also significantly elevated versus the EO 50 ppm group. Variability of SCEs/metaphase within each monkey increased even more than the increase in total SCEs/metaphase group with increasing EO exposure. Chromatid-type aberrations were also significantly increased for both EO 50 and EO 100 ppm groups compared to controls. Statistically significant increases in the number of chromosome-type aberrations (excluding gaps) were found only in the EO 100 ppm group. Combined chromatid- and chromosome-type aberrations were increased in both EO 50 and EO 100 ppm groups. No group differences in the number of gaps were found. In lymphocytes from monkeys exposed at 100 and 300 ppm PO, there were no group differences compared to controls for any variable-chromatid or chromosome-type aberrations, gaps, or SCEs/metaphase. These results indicate that EO is a more potent clastogen than PO and demonstrate, for the first time, statistically significant effects of EO on both SCEs and chromosome aberrations in lymphocytes of nonhuman primates.

Animals↗
PubMed1990

Enhanced assays detect increased chromosome damage and sister-chromatid exchanges in heroin addicts.

To refine previous studies of chromosome damage (CD) and sister-chromatid exchanges (SCE) in heroin addicts, we applied new methods developed in our laboratory to enhance detection of the cytogenetic effects of low-level radiation exposure in hospital workers. For CD analysis, we applied our thymidine-fluorodeoxyuridine-caffeine (TFC) enhancement procedure in which cells at setup receive 1 x 10(-7) M fluorodeoxyuridine to inhibit thymidylate synthetase and 4 X 10(-5) M thymidine to satisfy the induced requirement, and then in G2 receive 2.2 mM caffeine to modulate DNA repair. For SCE enhancement, caffeine treatment was initiated in G1 at 19 h before harvest. Using both standard and enhanced procedures for CD and SCE analysis, blood samples were evaluated from 20 street heroin addicts and 22 controls. Standard 2-day CD and 3-day SCE assays showed small, insignificant genotoxic increases in addicts while the enhanced CD and SCE assays showed highly significant increases. Most CD events were in the form of chromatid and chromosome breaks. There were no rings and only a few dicentrics were observed in the TFC-enhanced cultures. Although quadriradials are rare, 10 were found in addict TFC-cultures and 3 in control TFC-cultures. With the standard CD assay, the mean number of chromosome breaks per 100 cells was 0.727 for controls and 1.056 for addicts (not significant). With the TFC-enhanced assay, the same measure showed 1.483 chromosome breaks for controls and 5.143 for addicts (highly significant, ANOVA: p less than 0.0001). A highly significant difference was also observed for chromatid-type damage with the TFC-enhanced assay (chromatid breaks per 100 cells: 16.793 for controls; 48.191 for addicts). The SCE data also showed significant differences with the enhanced assay. Scoring 25 cells/condition, standard SCE cultures showed 10.892 SCE/cell for controls and 11.732 SCE/cell for addicts (not significant). With CAF enhancement there were 13.08 SCE/cell for controls and 17.05 SCE/cell for addicts (ANOVA: p less than 0.008). These findings indicate that detection of CD and SCE effects can be significantly enhanced by the use of these new procedures. The finding of greatly increased chromatid damage in the addicts with the TFC procedure suggests that at least part of the CD detected occurred in vitro and is not a product of prior in vivo damage. Therefore exposure to this drug and perhaps other environmental agents may not only leave a residue of DNA or chromosome damage but may also induce a sensitivity to further genotoxic damage that is revealed by using the enhanced procedures.

Analysis of Variance↗
PubMed1986

Studies on sister chromatid exchanges in patients with Hodgkin's disease.

Sister chromatid exchange frequencies were analyzed in bone marrow cells and in stimulated peripheral blood lymphocytes from 20 patients with Hodgkin's disease. Sister chromatid exchange studies were also carried out in the bone marrow of eight and peripheral blood of 11 normal subjects separately. Among the patients, ten were newly diagnosed and ten patients were survivors who had received therapy 30-64 months prior to studies. The mean sister chromatid exchange frequencies in bone marrow and peripheral blood of untreated patients were 3.93 +/- 0.7/cell and 4.09 +/- 0.91/cell, respectively, which were not significantly different from those of normal subjects (3.22 +/- 0.7/cell and 5.16 +/- 1.3/cell in bone marrow and peripheral blood, respectively). The mean sister chromatid exchange frequencies in bone marrow and peripheral blood in five of the untreated patients 3-4 months after initial therapy and in a treated (30-64 months after therapy) group were close to the sister chromatid exchange values of the untreated group. Analysis of the distribution of exchanges per chromosome or chromosome groups showed a nonrandom distribution of exchanges in chromosomes #1, #2, and #3, and in E-, F-, and G-group chromosomes in the normal controls and in Hodgkin's disease patients.

Adolescent↗