Baseline Sister Chromatid Exchange Frequencies Research Articles

Enhanced frequency of sister chromatid exchanges induced by diepoxybutane is specific characteristic of Fanconi anemia cellular phenotype

Fanconi anemia (FA) is a rare genetically heterogeneous disease characterized by developmental abnormalities, progressive bone marrow failure, and cancer susceptibility. We examined spontaneous, diepoxybutane (DEB)- induced and radiation-induced sister chromatid exchanges (SCEs) in wholeblood lymphocyte cultures of bone marrow failure (BMF) patients including Fanconi anemia, mothers of affected individuals, and healthy controls. The baseline frequency of SCE in FA cells was similar to that observed in controls. However, in response to DEB SCE frequencies in FA patients and their mothers were significantly increased compared to both non-FA BMF families and healthy controls. In response to ionizing radiation, cells displayed increased frequency of SCE, but no differences between FA patients and non-FA BMF patients were seen. Our data confirm and expand previous findings by showing that SCE induced by DEB can be used as an adjunct diagnostic test not only for FA patients, but also for female heterozygous carriers, at least for complementation groups FANCA and FANCD2.

RecA stimulates sister chromatid exchange and the fidelity of double-strand break repair, but not gene targeting, in plants transformed by Agrobacterium.

Expression of the bacterial RecA protein in plants stimulates homologous recombination in tobacco. Here we show that RecA plays a direct role in DNA strand exchange in vivo. The number of sister chromatid exchanges (SCEs) was increased 2.4-fold over wild type in transgenic tobacco plants expressing a nuclear-targeted RecA (nt-RecA) protein and could not be increased further by DNA damage, which caused a doubling of the baseline SCE frequency in wild-type plants. Although gene targeting requires homologous recombination, the number of targeted gene replacements was not increased markedly by the presence of nt-RecA by using Agrobacterium-mediated transformation. However, the number of double-strand breaks that were repaired at both sides by homologous recombination was increased 3.3-fold. Stimulation of SCE and fidelity of double-strand break repair by nt-RecA, but not by gene targeting, suggests that the stimulatory activity of RecA is linked to active DNA synthesis. Therefore, nascent replication-associated single strands may be a prerequisite for RecA action in plant cells.

RecA stimulates sister chromatid exchange and the fidelity of double-strand break repair, but not gene targeting, in plants transformed by Agrobacterium

Expression of the bacterial RecA protein in plants stimulates homologous recombination in tobacco. Here we show that RecA plays a direct role in DNA strand exchange in vivo. The number of sister chromatid exchanges (SCEs) was increased 2.4-fold over wild type in transgenic tobacco plants expressing a nuclear-targeted RecA (nt-RecA) protein and could not be increased further by DNA damage, which caused a doubling of the baseline SCE frequency in wild-type plants. Although gene targeting requires homologous recombination, the number of targeted gene replacements was not increased markedly by the presence of nt-RecA by using Agrobacterium-mediated transformation. However, the number of double-strand breaks that were repaired at both sides by homologous recombination was increased 3.3-fold. Stimulation of SCE and fidelity of double-strand break repair by nt-RecA, but not by gene targeting, suggests that the stimulatory activity of RecA is linked to active DNA synthesis. Therefore, nascent replication-associated single strands may be a prerequisite for RecA action in plant cells.

Meeting report

Meeting report

Monocytes affect cell‐cycle progression but not baseline frequency of sister chromatid exchanges of pig lymphocytes

The effect of pig monocytes (MNs) on the baseline frequency of sister chromatid exchanges (SCEs) and cell-cycle progression of pig lymphocytes was studied in plasma leukocyte (PLCs) and whole blood leukocyte cultures (WBCs). No variation in SCE frequency was observed between control WBC and PLC, nor did the addition of pig MNs to PLCs modify the baseline frequency of SCEs. Cell proliferation was slower in PLCs than in WBCs. Variations in cell-cycle progression of pig lymphocytes from PLC were observed both in the absence and presence of adherent cells in the culture. In MN-free cultures, lymphocytes proliferated faster than in parallel PLC cultures. However, when MNs were seeded into the cultures, cell-cycle progression gradually slowed as a function of the concentration of adherent cells present in the cultures. This finding shows that pig MNs modulate the in vitro cell-cycle progression of pig lymphocytes in a dose-dependent manner and that the low baseline SCE frequency of pig lymphocytes is independent of the presence or absence of MNs in the culture. Environ. Mol. Mutagen. 33:86–90, 1999 © 1999 Wiley-Liss, Inc.

Monocytes affect cell‐cycle progression but not baseline frequency of sister chromatid exchanges of pig lymphocytes

The effect of pig monocytes (MNs) on the baseline frequency of sister chromatid exchanges (SCEs) and cell-cycle progression of pig lymphocytes was studied in plasma leukocyte (PLCs) and whole blood leukocyte cultures (WBCs). No variation in SCE frequency was observed between control WBC and PLC, nor did the addition of pig MNs to PLCs modify the baseline frequency of SCEs. Cell proliferation was slower in PLCs than in WBCs. Variations in cell-cycle progression of pig lymphocytes from PLC were observed both in the absence and presence of adherent cells in the culture. In MN-free cultures, lymphocytes proliferated foster than in parallel PLC cultures. However, when MNs were seeded into the cultures, cell-cycle progression gradually slowed as a function of the concentration of adherent cells present in the cultures. This finding shows that pig MNs modulate the in vitro cell-cycle progression of pig lymphocytes in a dose-dependent manner and that the low baseline SCE frequency of pig lymphocytes is independent of the presence or absence of MNs in the culture.

Chromosome Breaks and Sister Chromatid Exchange as Predictors of Second Cancers in Hodgkin's Disease

Hodgkin's disease (HD) survivors face an increased risk of developing second cancers. We evaluated baseline cytogenetic biomarkers, sister chromatid exchange (SCE) and chromosome breaks [spontaneous (SCB) and bleomycin-induced (BIB)], as predictors of second cancer risk in a cohort of 105 adult HD patients. During follow-up, seven second cancers occurred. SCBs and BIBs showed no association with risk of second primaries. Multivariate Cox regression revealed that high levels of SCEs (relative risk (RR) = 11.3, p = 0.02) and age (RR = 1.08, p = 0.02) predicted second cancer risk. Histology, stage, and treatment were not associated with elevated risk. In conclusion, baseline SCE frequencies may be a useful biomarker for identifying HD patients at increased risk of developing second cancers. These results need to be verified in a larger cohort with a longer follow-up time.

Baseline and mitomycin C(MMC)-induced sister-chromatid exchanges in XX and XY cells of Callithrix jacchus chimeric twins

Lymphocytes from chimetric individuals of the species Callithrix jacchus (Primates) were examined to evaluate differences in the frequency of sister chromatid exchanges (SCE) between XX and XY cells. The aim was to discover whether SCE differ according to genetic sex and whether XX and XY cells show a different sensitivity to SCE inducing agents. This experimental model has enabled us to eliminate the possible differences caused by environmental factors. The results obtained do not reveal significant differences between male and female cells, in either the baseline SCE frequency or that induced by mitomycin C at concentrations of 0.01 and 0.03 μg/ml. No signficant differences were observed in the distribution of high SCE frequency cells (HFC), even if it is possible to observe a higher level of exchanges in XX cells in each trial. With regard to the phenotypic sex, there appears to be a trend towards slightly higher SCE rates in females, even if results are not statistically significant.

Increased frequencies of sister chromatid exchange in soldiers deployed to Kuwait.

Frequencies of sister chromatid exchange (SCE), a measure of genotoxic exposure, were assessed in military troops deployed to Kuwait in 1991. Soldiers completed health questionnaires and had blood collected prior to, during and following deployment to Kuwait. Frequency of spontaneous SCE was determined on blood samples as a measure of mutagenic exposure. Compared to pre-deployment baseline SCE frequency means, levels obtained 2 months into the Kuwaiti deployment were significantly increased (P < 0.001) and persisted for at least 1 month after return to Germany. Outcome was unaffected by known personal SCE effect modifiers including smoking, age and diet. Potential sources of the apparent mutagenic exposure are discussed.

Differences in the response to mutagens between two V79 sublines

Two V79 Chinese hamster sublines (V79-UL and V79-MZ) which differed markedly with respect to their spontaneous pattern of mutations at the HPRT locus were comparatively investigated in genotoxicity tests with ethyl methanesulphonate (EMS). EMS-induced frequencies of HPRT mutations and sister chromatid exchanges (SCEs) were much higher in V79-MZ than in V79-UL. V79-MZ were not hypersensitive against EMS and had normal frequencies of spontaneous gene mutations and chromosome aberrations. Baseline SCE frequencies at various BrdUrd concentrations were slightly increased compared to V79-UL. EMS induced a similar amount of chromosome aberrations in both cell lines but exchange figures occurred with lower frequency in V79-MZ. The results indicate that specific and significant differences in the response to mutagens may exist between ‘normal’ Chinese hamster cell lines which might be relevant for genotoxicity testing.

Pig plasma modulates cell cycle kinetics but not the baseline frequency of sister-chromatid exhanges in human lymphocytes

The effect of human and pig plasma on the baseline frequency of sister-chromatid exchanges (SECs) in human and porcine plasma leukocyte cultures was studied. Human plasma leukocyte cultures, but not porcine plasma leukocyte cultures, showed at least a twofold increase in SCE frequency over whole blood culture values. Addition of pig plasma to human plasma leukocyte cultures and human plasma to pig plasma leukocyte cultures did not modify the baseline SCE frequencies. In both human and porcine cultures, cell proliferation was slower in plasma leukocyte cultures than in whole blood cultures. The addition of pig plasma to human plasma leukocyte cultures, but not the incorporation of human plasma in pig plasma leukocyte cultures, accelerated the cell cycle progression of lymphocytes. With 10% pig plasma in the plasma leukocyte culture medium, lymphocyte proliferation was similar to that in whole blood cultures. Smaller concentrations of pig plasma rendered cell cycle progression intermediate between the basal plasma leukocyte culture and whole blood culture values. Exchanging fetal calf serum for human AB serum in human plasma leukocyte cultures did not affect the cell cycle kinetics of lymphocytes but it did decrease their baseline SCE frequency.

Baseline frequency of sister-chromatid exchanges in 142 persons of the general Korean population

Baseline frequencies of sister-chromatid exchange (SCE) were measured in lymphocytes of 142 healthy Koreans ranging in age from newborn infants to the fifties. The overall mean frequency of SCE was 8.78±0.24/cell. However, highly significant differences were found between individuals. The mean SCE values of the newborn babies and small children less than 10 years old were significantly lower than those of other age groups. No age effect was, however, observed in adolescent and adult subjects. Females had statistically higher SCE levels than males. The mean SCE frequencies of smokers, measured in male subjects more than 10 years old, were slightly, but statistically significantly, higher than those of non-smokers.

Bimodal distribution of sensitivity of SCE induction by diepoxybutane in human lymphocytes. II. Relationship to baseline SCE frequency

Environmental and genetic factors have been implicated as important sources of individual variation in baseline sister-chromatid exchange (SCE) frequency in humans. The current study was designed to test whether the frequency of baseline SCEs in 58 normal blood donors is associated with previously observed variations in SCE frequencies induced by diepoxybutane (DEB). Because 12 subjects were current cigarette smokers and smoking is known to be an in vivo inducer of baseline SCE frequencies, we specifically tested whether higher baseline SCE frequencies in smokers would be associated with in vitro sensitivity to SCE induction by DEB. Analysis of variance showed that DEB-induced SCE frequencies were significantly associated with baseline SCE frequencies; those who were sensitive to SCE induction by DEB were more likely to have higher baseline SCE frequencies. This effect, however, was independent of in vivo induction of SCE by smoking. Chromosomal sensitivity to the induction of SCE by DEB explained approx. 15–20% of the variation in baseline SCE. This was similar in magnitude to the effect of cigarette smoking. Because increased sensitivity to DEB-induced SCEs is common in normal blood donors (approx. 24%) and is associated with an increase in baseline SCE s, it should be investigated as a source of bias and/or a potential marker of sensitivity to environmental mutagens in future cytogenetic studies.

Cytogenetic effects of pulsing electromagnetic field of human lymphocytes in vitro: chromosome aberrations, sister-chromatid exchanges and cell kinetics

Exposure of human lymphocyte cultures to a pulsing electromagnetic field (PEMF; 50 Hz, 1.05 mT) for various duration (24, 48 and 72 h) resulted in a statistically significant suppression of mitotic activity and a higher incidence of chromosomal aberrations. Furthermore, the shorter exposure times (24 and 48 h) did not cause a significant delay in cell turnover (cell proliferation index) or an increase in the baseline frequency of sister-chromatid exchanges (SCE). However, cultures continuously exposed to PEMF for 72 h exhibited significant reduction of the cell proliferation index (CPI) and an elevation of SCE rate. The results suggest that exposure to PEMF may induce a type of DNA lesions that lead to chromosomal aberrations and cell death but not to SCE, except probably at longer exposure times.

Erythrocytes modulate the baseline frequency of sister-chromatid exchanges and the kinetics of lymphocyte division in culture

The baseline sister-chromatid exchange (SCE) frequencies of human plasma lymphocyte cultures (PLC), but not pig PLC, were nearly twice as high as those of whole-blood cultures (WBC). Addition of human red blood cells (RBCs) to human PLC decreased the SCE frequency in proportion to the RBC-leukocyte co-incubation interval. When the period of RBC-leukocyte co-incubation was equivalent to the total length of the culture period (72 h), the SCE frequency was similar to that observed in WBC. Shorter co-incubation periods yielded SCE frequencies intermediate between those of PLC and WBC. Regardless of the species, cell proliferation was slower in PLC than in WBC. Experiments where RBCs were added to PLC showed that the time sequence of RBC incorporation also affects the cell-cycle progression of human and pig lymphocytes. When either human or pig RBCs were added immediately after PLC stimulation, the cell-cycle kinetics was similar to that of WBC. Shorter co-incubation periods made cell-cycle progression intermediate between PLC and WBC values. Thus, PBCs modulate the baseline frequency of SCEs in human PLC and the cell-cycle progression of both human and pig lymphocytes in a time-dependent manner. Two possible hypotheses for the heightened frequency of SCEs of human lymphocytes in RBC-free cultures were assessed. The loss of RBC-to-lymphocyte cellular contact in PLC did not influence the SCE frequencies of lymphocytes. Finally, the increase of SCEs in human PLC could not be related to differences in the generation time of lymphocytes in culture.

Baseline and phosphoramide mustard-induced sister-chromatid exchanges in cancer patients treated with cyclophosphamide

Determinations of baseline sister-chromatid exchanges (SCE) have been used extensively as indicators of previous mutagen exposure in both animals and humans. Hypersensitivity to mutagen-induced SCE levels has also been studied in a variation on the basic technique as an indication of previous mutagen exposure in a stressed or provocative test system. The genotoxicity of the alkylating anti-cancer drugs including cyclophosphamide (CP) has been examined previously by determining baseline SCEs in peripheral blood lymphocytes from treated cancer patients. This study examined the in-vivo genotoxic effects of CP therapy by comparing baseline and phosphoramide mustard (PM)-induced SCEs in therapeutically (in-vivo) treated cancer patients with SCE levels in newly diagnosed, but not treated patients. Therapeutically treated patients showed statistically higher baseline SCE frequencies than untreated control patients with a mean SCE/cell of 6.95 vs. 5.25, p < 0.016. When net SCE values (induced minus baseline) were determined in PM-exposed cells in-vitro both at low dose (0.1 μg/ml PM) and high dose (0.25 μg/ml PM) however, the difference was not significant between therapeutically treated and untreated control patients. The return to control SCE levels as a function of time since last therapeutic treatment was also evaluated and no difference was found between the rate of decline of PM-induced SCEs and baseline SCE levels over time.

Effects of antiepileptic drugs and organic brain damage on SCE frequency in cerebral palsies.

In order to assess the genotoxic effects of antiepileptic drugs (AEDs), analysis of sister chromatid exchanges (SCEs) was performed in lymphocytes of three groups of males; epileptics with AED therapy, non-epileptics without AED therapy and healthy controls. The epileptics and non-epileptics were cases of severe cerebral palsies due to perinatal asphyxia. Possible confounding factors of SCE frequencies, such as age, smoking habit, drug usage other than AED, recent history of viral infection, etc. were controlled. The frequency of SCE per cell was 4.63 +/- 0.71 (mean +/- S.D.) in epileptics, 4.70 +/- 0.89 in non-epileptics and 3.84 +/- 0.56 in healthy controls. SCEs in both epileptics and non-epileptics were significantly higher (p less than 0.05) than those in controls. There was no significant difference of SCE frequency between epileptics and non-epileptics. These results suggested that no mutagenic effect of AED could be demonstrated as revealed by SCEs, and organic brain damage per se might influence the baseline SCE frequency. The possible explanations for such observations are discussed.

Cytogenetic observations of a human ovarian carcinoma clinically resistant to therapy

Resistance of a cancer cell to therapy represents an important problem in tumor biology. Hypotheses concerning the mechanism of resistance have included genetic instability, gene amplification, aneuploidy, and altered cell growth kinetics. Cytogenetic assays allow for the analysis of each of these parameters and provide important information concerning tumor heterogeneity. In using cytogenetic analysis, we have analyzed a human ovarian carcinoma clinically resistant to therapy. The ovarian carcinoma had a complex but stable karyotype with a mean chromosome number of 61.7 chromosomes/cell. Approximately 50% of the metaphase preparations examined had double-minute chromosomes, which have sometimes been associated with gene amplification. The frequency of double minutes varied from one pair to hundreds of pairs per cell. Cell cycle kinetic analysis revealed an average generation time of 38 hours. The cells were relatively resistant to doxorubicin in vitro, and the baseline sister chromatid exchange frequency, a measure of genetic instability, was elevated. These results suggest that cytogenetic assays have potential as predictive assays of tumor chemoresistance and may provide information regarding the biologic aggressiveness encountered clinically.

Sister chromatid exchanges and dementia of the Alzheimer type

Sister chromatid exchange (SCE) analysis was carried out on ten patients with the clinical diagnosis of dementia of the Alzheimer type (DAT) and 11 cognitively-intact controls of a comparable age. There was no significant difference between patient and control groups in either the mean baseline SCE frequency or the SCE frequency following in vitro exposure to mitomycin-C. The present results, based on the largest sample (n = 21) published to date, fail to confirm the single positive report in the literature of an increased baseline SCE frequency in DAT patients compared to controls.

Cytogenetic analysis in workers occupationally exposed to nickel carbonyl

Chromosomal aberration and sister-chromatid exchange (SCE) base-line frequencies and SCE frequencies induced by 10 ng/ml mitomycin C (MMC) were analysed in cultured peripheral lymphocytes of 65 workers occupationally exposed to nickel carbonyl Ni(CO) 4. The subjects were divided into 4 groups: (1) control; (2) exposed to nickel carbonyl ( = exposed); (3) cigarette smokers; (4) smoking-exposed. The results show that there are no significant differences in chromosomal aberration frequencies, breaks or gaps, between the various groups. However, the SCE base-line frequency of the smoking-exposed group, with an average of 7.7/cell, was significantly higher than that of the control group, with an average of 6.5/cell ( P < 0.01), and also than that of the exposed group with an average of 5.9/cell ( P < 0.01). Similarly, the SCE frequency induced by 10 ng/ml MMC in the smoking-exposed group which averaged 15.5/cell was significantly higher than that of the control group (average of 13.2/cell ( P < 0.05), and also than that of the exposed group with an average of 12.3/cell ( P < 0.01). Under our experimental conditions, it may be that the level of exposure was not high enough to elicit an increase in chromosomal aberrations and SCE frequencies in the non-smoker exposed group. The fact that an increase in SCE frequencies was only found in the smoking-exposed group implies that the two factors, smoking and exposure to nickel carbonyl, are jointly responsible for the result.