Significant Increase In Sister Chromatid Exchanges Research Articles

The effect of valproic acid on SCE and chromosome aberrations in epileptic children

Sister-chromatid exchange (SCE) and chromosome aberrations have been studied in peripheral lymphocytes of 20 epileptic children treated in monotherapy with valproic acid (VPA) for 6–52 months and in 2 matched control groups. The frequencies of SCE in the VPA-treated epileptic children were significantly higher than in the 2 control groups ( p<0.01); rates of chromosome aberrations were slightly higher but not significantly different from the 2 control groups. We also examined SCE in 10 epileptic children before and after they took sodium valproate for 6–7 months; there was a statistically significant change in SCE following VPA. 9 normal children whose lymphocytes were exposed in vitro to sodium valproate (5–20 μg/ml) showed a significant increase in SCE.

Unequal SCE is a rare event in homologous recombination between duplicated neo gene fragments in CHO cells

The frequency of sister-chromatid exchange (SCE) was studied in Chinese hamster ovary (CHO) cell lines with stable insertions of the vector pIII-14gpt which contains 2 truncated neomycin resistance (neo) gene fragments. Recombination between regions of homology in the 2 fragments can restore a functional neo gene and make the cell resistant to the antibiotic G418, a neomycin analogue. Unequal SCE would be one of several possible mechanisms for this event. The observed spontaneous rate of formation of G418-resistant subclones was approximately 6.4 × 10 −6 per cell per generation, as compared to the estimated spontaneous frequency of 3 SCE per cell per generation. Given this SCE frequency, the probability of an SCE occurring in a target site of about 1600 bp (the distance separating the homologous regions in the neo fragments) would be about 8 × 10 −7 per cell per generation, or approximately one tenth of the estimated rate of recombination. Treatment of the cells with methyl methanesulfonate (MMS, 50 × 10 −6 M) induced about 80–90 SCE per cell, corresponding to a probability of 2 × 10 −5 SCE per 1600-bp target per cell. In the same cell culture, MMS treatment induced 4–8 × 10 −4 recombination events per cell giving rise to G418 resistance. Cells treated with HN 2 (up to 4 × 10 −6 M) showed a significant increase in SCEs, but no change in the frequency of G418-resistant revertants. These results suggest that the 2 pathways leading to SCE and recombination respectively are uncoupled, and only a small fraction of the recombination events, if any, are due to unequal SCE in this system.

Sister-chromatid exchange studies on direct- and indirect-acting clastogens in mouse primary cell cultures

An in vitro sister-chromatid exchange (SCE) assay using mouse primary bone marrow and spleen cells was conducted with both direct- and indirect-acting genotoxic agents. 2,4,7-Trinitrofluorenone, a direct-acting genotoxic agent, induced a significant dose-related increase in SCEs. In both bone marrow and spleen cells, 2.0 μg/ml caused an approx. 3-fold increase in SCE level over control values. Cyclophosphamide, an indirect-acting genotoxicant which requires metabolic activation for its clastogenicity, induced a significant increase in SCEs in the presence of S9 from liver of rats pretreated with Aroclor-1254. A dose of 2 μg/ml resulted in a 2-fold increase in bone marrow and a greater than 5-fold increase in spleen cells. Benzo[ a]pyrene, another indirect-acting genotoxicant, also induced significant dose-related SCE responses in both cell types. It seems that primary bone marrow and spleen cell culture systems can detect both direct- and indirect-acting genotoxicants and may be useful for routine and/or comparative cytogenetic studies.

The effect of agent dose and treatment time on the intercellular distribution of sister-chromatid exchanges induced by genotoxic agents in mouse bone marrow cells in vivo

Using two methods of bromodeoxyuridine (BrdUrd) administration and three genotoxic chemicals, the effects of dose and treatment time on the intercellular distribution of sister-chromatid exchanges (SCE) in the bone marrow of male B6C3F1 mice were evaluated. The dispersion of SCE among solvent control mice infused intravenously with BrdUrd or implanted subcutaneously with a BrdUrd tablet partially coated with paraffin was largely consistent with a Poisson model. Intraperitoneal treatment with cyclophosphamide (CP; solvent = phosphate-buffered saline), 7,12-dimethylbenzanthracene (DMBA; solvent = corn oil) and, in mice infused with BrdUrd, mitomycin C (MMC; solvent = phosphate-buffered saline) induced a significant increase in SCE, the distribution of which was not distributed as a Poisson. For CP and MMC, the increase in dispersion was dose-dependent and independent of treatment time (−1, +1 or +8 h in relation to the start of the BrdUrd treatment). The lack of a treatment time effect suggests that there were no significant differences among treatment times in the distribution of the reactive forms of these two chemicals, no variation in cell-stage sensitivity, and no cellular toxicity to modulate the response. For DMBA, the increased dispersion of induced SCE depended on treatment time and was not simply related to dose. The increased in dispersion was agent-specific; at equal levels of SCE induction, the distribution of SCE in mice treated with DMBA exhibited greater dispersion than SCE in mice treated with either CP or MMC. These differences between DMBA and CP/MMC are probably due to DMBA's slower absorption/distribution kinetics, its requirement for metabolic activation to genotoxic metabolites and its extended half-life. These data suggest that analyzing the distribution of SCE, in addition to mean frequency, is a useful method for evaluating agent specific patterns in SCE induction.

Lack of sister-chromatid exchange induction by polymethyl methacrylate bone cement in human lymphocytes cultured in vitro

Human lymphocytes cultured in vitro were used to assess the ability of polymethyl methacrylate (PMMA), currently used in orthopedic surgery as bone cement, to induce sister-chromatid exchanges (SCE). Under the conditions used in this study, PMMA bone cement did not produce any significant increase in SCEs in phytohemagglutini-stimulated human lymphocytes. However, a significant decline ( p<0.05) in the proliferation rate index was observed, which might be an indicator of the cytotoxic effect of PMMA.

Analysis of sister-chromatid exchanges, cell kinetics and mitotic index in lymphocytes of smoking pesticide sprayers

Whole blood of 50 smokers who were exposed to pesticides was set up in RPMI 1640 medium, and observed for sister-chromatid exchanges (SCEs), cell kinetics (CK) and mitotic index (MI). As controls, blood samples were collected from 20 non-smokers (control I) and 27 smokers (control II) who were not exposed to pesticides. A significant increase in SCEs was observed as the duration of exposure increased. The frequency of M 1 metaphases increased significantly whereas M 2 and M 3+ metaphases decreased in the exposed group. The mitotic index increased in control II and in the exposed population while it showed a decrease at 11–25 years' exposure.

Comparative study of sister-chromatid exchanges and chromosomal aberrations induced by airborne particulates from an urban and a highly industrialized location in human lymphocyte cultures

Cytogenetic effects induced by extracts of airborne particulates in human lymphocyte cultures were studied with regard to local and seasonal variations. Samples of airborne particulates were collected from an urban and a highly industrialized area in March and October, respectively. All extracts of particulates induced a significant increase of sister-chromatid exchanges (SCE) in a dose-dependent manner. Referring to the induction of sister-chromatid exchanges, local and seasonal differences were observed. Samples from the industrialized area revealed the highest activities. In addition, SCE rates found for the March samples were always higher than those for October for both locations. Furthermore, a remarkable, significant induction of chromosomal aberrations occurred with all samples from both locations and sampling periods. Aspects of health risk evaluation for exposed human populations are discussed with respect to the observed cytogenetic effects of airborne particulates in human lymphocyte cultures.

Induction of sister chromatid exchanges and cell division delays in human lymphocytes by the anti-tumour agent homo-aza-steroidal ester of p-bis(2-chloroethyl)aminophenoxy acetic acid.

3 beta-Hydroxy-13 alpha-amino-13,17-seco-5 alpha-androstan-17-oic-13,17-lactam-p-bis(2-chloroethyl) aminophenoxyacetate (NSC 294859) is a new modified steroidal alkylating agent. This compound was given by i.p. administration to mice bearing different types of tumour. It was found to exhibit good activity in L1210 and P388 leukaemias with maintenance of activity against advanced tumours. The treatment of colon 26 tumour and B16 melanoma resulted in positive antineoplastic activity. The drug was not shown to be active in a melphalan-resistant P388 line. In this study, NSC 294859 was found to be effective in causing statistically significant increases in sister-chromatid exchange (SCE) rates and cell division delays. The alkylating agent component, p-bis-(2-chloroethyl)aminophenoxy acetic acid, was shown to be less effective than the parent compound, while the modified steroid component, 3 beta-hydroxy-13 alpha-amino-13,17-seco-5 alpha-androstan-17-oic-13,17-lactam, showed no effect. There were no statistically significant differences among donors regarding the induction of SCEs and replication indices (RIs) for the compounds tested.

Induction of sister chromatid exchanges by tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone in human and hamster cells.

4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), a nicotine derived N-nitrosamine, is a carcinogen that induces tumors in mice, rats and hamsters. To assess the ability of NNK to interact with the cellular DNA as an essential step in carcinogenesis, the induction of sister chromatid exchange (SCE) was examined in cultured normal human lymphocytes (HL) and Chinese hamster V79 cells. SCE formation is a sensitive indicator of carcinogen-DNA interaction that correlates with the induction of mutation and neoplastic cell transformation. HL and V79 cells were treated for 2 h with 20, 50, 100 and 200 micrograms/ml of NNK with and without application of metabolic activation S-9 mixture, and subsequently incubated for two rounds of replication in the presence of 5-bromodeoxyuridine required for SCE visualization. In V79 cells NNK produced a dose-dependent increase in SCE only with metabolic activation. In HL NNK induced a small but statistically significant increase in SCE with or without metabolic activation. These data provide the first evidence that NNK and/or its metabolic derivatives are able to induce DNA damage leading to SCE formation both in hamster and human cells. The differences in response between the two cell types suggests the existence of a difference in susceptibility associated with NNK metabolism and its interaction with cellular DNA.

Variable sister-chromatid exchange response in human lymphocytes exposed in vitro to gossypol acetic acid

Gossypol has potential for widespread use as a male oral antifertility agent in humans since it appears to be highly efficacious, with reversible spermatostatic effects and minimal side effects. Furthermore, it is both inexpensive and readily available. Therefore, a thorough understanding of gossypol's genotoxic potential is critical. Although genotoxicity studies have produced conflicting reports, increased sister-chromatid exchange (SCE) and DNA-strand breaks have been reported in human cells exposed to gossypol in vitro. In the present study, SCE was examined in purified human lymphocytes and whole blood cultures exposed to gossypol acetic acid at various concentrations in serum-free medium. A small but statistically significant increase in SCE was observed in pooled analysis of 7 donors in whole blood cultures exposed to 0.70 μM gossypol acetic acid ( p < 0.02). Individual analyses revealed only one donor with a significant SCE response ( p < 0.001). In subsequent experiments, exposure at higher doses had no effect on SCE frequencies. A small but significant increase in SCE was observed in ficoll/hypaque purified lymphocytes exposed to 0.07 and 0.70 μM gossypol acetic acid. Interpretation of SCE data with variable response is discussed. This work was supported by Public Health Service toxicology training grant ES-07046.

SCE induction in Chinese hamster ovary cells (CHO) exposed to G agents

Cultured Chinese hamster ovary (CHO) cells were exposed to two neurotoxic organophosphates, either satin (GBI, GBII) at 1.4 x 10 −3 M or soman (GD) at 1.1 and 2.2 x 10 t-3 M for 1 h, grown and their metaphase chromosomes scored for sister-chromatid exchanges (SCE). No cytotoxicity was seen with either agent at any dose level tested. Since histograms of SCE per cell showed that they were non-symmetrically arrayed around the mean, the number of SCEs were analyzed by using the nonparametric tests, Mann-Whitney and Kruskall-Wallis. Agents GBI and GBII did not show any significant increase in SCE over baseline. On the other hand, GD demonstrated a statistically significant increase in SCE with and without metabolic activation. Ethyl methanesulfonate (EMS) alone at 5 x 10 −3 M and cyclophosphamide (CP) at 10 −4 M in the presence of rat microsomes (S9) induced a 3- and 8-fold increase in SCE per cell, respectively.

Induction of sister chromatid exchanges (SCE) in G0 lymphocytes by plutonium-238 alpha-particles.

Irradiation of human G0 lymphocytes with plutonium-238 alpha-particles and X-rays was performed to investigate the production of sister chromatid exchanges (SCE). Alpha-particles produce a significant increase in SCE and this elevation is more significant when separated lymphocytes are irradiated. X-ray irradiation did not induce any significant increase in SCE. Therefore the relative biological effectiveness (RBE) for the induction of SCE by alpha-particles in this system is undefined and effectively infinite.

Induction of sister chromatid exchange and dominant lethal mutation by ‘Katha’ (catechu) in male mice

Induction of sister chromatid exchange (SCE) and dominant lethal mutation by ‘Katha’ were studied on mice following acute and prolonged oral treatment. A significant increase in SCEs was observed in all the treated series except 1.5 mg/kg body weight treated series when compared to control. Post-implantation loss was also increased by both acute and chronic treatments with ‘Katha’. Thus the above results indicate that ‘Katha’ has a strong clastogenic and mutagenic effect.

In vivo induction of sister chromatid exchanges in mice by nitrosated coal dust extract

The genotoxicity of coal dust extract nitrosated with sodium nitrite (NaNO2) was investigated in mice with the in vivo sister chromatid exchange (SCE) assay system. The SCEs in bone marrow cells of mice were examined following single and double oral dosings of coal dust extract, NaNO2, and nitrosated coal dust extract. Coal dust extract and NaNO2 separately did not cause significant increases of SCEs either in single or in double dosings. Nitrosated coal dust extract in single doses did not increase SCEs but in two doses significant increases in SCEs were observed (P less than 0.02). The mutagenicity of the same extracts was tested in Salmonella typhimurium with the Ames tester strain TA98. Coal dust extract was either non- or weakly mutagenic and NaNO2 was nonmutagenic. The nitrosated coal dust extract caused pronouced increases in his+ revertants both with and without rat liver S9 activation. These findings provide additional evidence that nitrosation of ingested coal dust which may occur in the stomach environment could be one of the factors leading to the higher incidence of gastric cancer in coal miners.

Sister-chromatid exchanges in human lymphocytes exposed in vitro to therapeutic ultrasound

Human lymphocytes were exposed in vitro to therapeutic levels of ultrasound (1 W/cm 2, CW, 0.87 MHz, durations of 80 and 160 sec). There were no significant differences in sister-chromatid exchange frequencies between controls and ultrasound-exposed cells. Exposure of lymphocytes to the positive control (mitomycin C) resulted in a significant increase in sister-chromatid exchanges. The data do not verify a report by Stella et al. (Mutation Res., 138 (1984) 75–85) that such exposures result in increased frequencies of SCEs.

Increased sister-chromatid exchange frequency in preimplantation mouse embryos after maternal cyclophosphamide treatment before implantation

The effect of a toxic agent in vivo on sister-chromatid exchange (SCE) frequency of preimplantation mouse embryos and bone marrow cells was determined using combined in vivo treatment and in vitro culture in the presence of 5-bromo-2-desoxyuridine (BrdU) for differential staining of the chromatids. In mice exposed to cyclophosphamide (CPA) on day 2 of pregnancy SCE frequency was increased dose-dependently both in embryos and bone marrow cells 1h after treatment. It returned to control values in bone marrow cells obtained 24 h after exposure but was still significantly increased in the embryos. A closer time-related evaluation of SCE on day 2 of gestation showed a significant increase in SCE in bone marrow cells and in embryos obtained 20–60 min after CPA treatment. Furthermore, SCE frequency was the most sensitive toxicological endpoint to detect embryotoxic effects of CPA treatment before implantation, since it was significantly increased in embryos exposed to 5 mg/kg CPA on day 2 of pregnancy while embryolethality at term and both cytogenetical (structural chromosomal aberrations, micronuclei) and developmental parameters (cell number, differentiation in culture) before implantation did not indicate any toxic effect.

Study of the genotoxicity of the total aqueous extract of betel nut and its tannin.

Frequencies of sister-chromatid exchanges (SCEs) were measured to study the genotoxicity of the total aqueous extract of betal nut and its tannin on mouse bone-marrow cells in vivo. Betal nut aqueous extract and tannin were injected i.p. into mice at doses of 12.5, 25 or 50 micrograms/g body weight and 50, 100 or 200 micrograms/g body weight respectively for 5, 10 or 15 days. Betal nut extract induced a dose-related increase in the frequency of SCEs after five daily doses of 12.5, 25 or 50 micrograms. Significant increases in SCEs were also observed in animals dosed for 10 days with 50 micrograms/g and for 15 days with 25 or 50 micrograms/g extract. After five daily doses of 50, 100 or 200 micrograms/g of betel nut tannin there was no significant induction of SCE. Mice dosed with 200 micrograms/g tannin for 10 days, or 100 or 200 micrograms/g for 15 days, showed significant increases in the frequency of SCEs.

Sister-chromatid exchange and cell replication kinetics in fetal and maternal cells after treatment with chemical teratogens

Effects of selected chemical teratogens on sister-chromatid exchange (SCE) frequencies and cell replication kinetics (CRK) in pregnant mice and their fetuses were investigated. Maternal and fetal cells were analyzed for SCE and classified as to whether they had gone through 1 (M 1), 2 (M 2), or 3 or more (M 3+) cell cycles for quantifying cell replication kinetics and estimating average generation time (AGT). The teratogens tested in this system were mitomycin C (MMC), cyclophosphamide (CP), ethylnitrosourea (ENU), dimethylnitrosamine (DMN), lead acetate (LA), benzene (BEN), diethylstilbestrol (DES), diphenyldantoin (DPH), N-methyl- N′-nitro- N-nitrosoguanidine (MNNG), and dimethyl sulfoxide (DMSO). MMC, CP, ENU, DMN, LA and BEN induced significant increases in SCE in both maternal and fetal cells compared with control values. DES and DPH induced increases in SCE in fetal cells only, whereas MNNG and DMSO did not induce any increase in SCE in either maternal or fetal cells. Chi-square analysis of the relative numbers of M 1, M 2 and M 2+ cells revealed significant heterogeneity among test chemical doses in both maternal and fetal cells treated with all of the chemicals except DES. DES had an effect on fetal cells but not on maternal cells. A significant linear relationship between AGT and test chemical dose was noted for ENU and DMSO in maternal cells and DPH and MNNG in fetal cells. Although statistically significant, the effects of DPH, MNNG, and DMSO on AGT were small, with increases in AGT of only 1 h or less at the highest doses tested. ENU, however, resulted in more than a 2-h increase in the AGT of maternal cells. Average generation time was consistently lower in fetal cells as compared with maternal cells (overall mean AGT ± S.D. from solvent controls was 8.6 ± 0.3 h for fetal cells and 11.6 ± 0.7 h for maternal cells). The results indicate that SCE induction and cell replication inhibition can occur independently, and that the assessment of SCE and CRK in maternal and fetal cells may be a promising approach to the identification of teratogenic agents.

In vivo cytogenetic studies on mice exposed to ethylene dibromide

The pesticide, ethylene dibromide (EDB), was evaluated with in vivo cytogenetic assays to determine its genotoxicity. CD 1 male mice were exposed to EDB through intraperitoneal injections. Bone marrow cells isolated from femora were analyzed for sister-chromatid exchange (SCE), chromosome aberration and micronucleus formation. The results showed that only certain concentrations of EDB tested caused a slight but significant increase in SCEs and chromosome aberrations. However, these increases were not dose-related. No increase in the polychromatic erythrocytes with micronuclei was observed following EDB exposure. Aslo, EDB did not cause cell-cycle delay in comparison with controls. Thus, it appears that EDB is not an effective genotoxic agent in vivo in mice.

Increased rates of sister chromatid exchanges induced by the herbicide 2,4-D.

The potential for genetic damage from widely used hormonic herbicides, such as 2,4-dichlorophenoxyacetic acid (2,4-D), continues to be of serious concern. The mutagenic effect as reflected by the rates of sister chromatid exchanges (SCE) was determined in cultured human lymphocytes. Data were based on the analysis of 50 cells for the control and each of the three treatments. A 50 micrograms/ml dosage caused a highly significant increase in SCE. Dosages of 100 and 250 micrograms/ml elevated the rate of SCE, but not significantly. Since 2,4-D biodegrades rapidly in soil and water, its continued use is not in serious question until safer compounds are available. However, the results of this study suggest that the danger of genetic damage from direct exposure to commercial samples of 2,4-D should not be ignored.