Variations of SCE frequencies in peripheral lymphocytes of ex-smokers

We measured SCE frequencies over a period of 8 months in 14 smokers who stopped smoking at the start of the study. In a first group of 10 subjects, who did not resume smoking during the period of cytogenetic follow-up, a lowering of SCE frequencies was already evident after 18 days and this became statistically significant after 78 days. SCE decrease was related to the logarithm of the period (in days) for which smoking was interrupted (r = 0.98; p less than 0.001). In a second group of 4 subjects, who at various times resumed smoking, the decrease of SCE followed the same pattern as in the first group during the period of nonsmoking, but SCE frequencies rose even higher once smoking was resumed. Our study indicates that the decrease of SCE in ex-smokers is rather rapid during the first 78 days after stopping smoking, and much slower from the 78th to the 233rd day.     

SCE frequencies in lymphocytes of systemic lupus erythematosus patients

The frequency of sister-chromatid exchanges (SCEs) was investigated in peripheral lymphocytes of lupus erythematosus patients and compared with values obtained for healthy controls. Irrespective of the kind of medical treatment, an increased level of spontaneously occurring SCEs could be demonstrated in lupus patients. In addition to spontaneously occurring SCEs, mitomycin C (MMC)-induced SCEs were evaluated. No difference between patients and controls was found with respect to MMC-induced SCEs.     

Bimodal distribution of sensitivity to 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 SCEs, it should be investigated as a source of bias and/or a potential marker of sensitivity to environmental mutagens in future cytogenetic studies.     

Measurement of SCE frequencies in plants: a simple Feulgen-staining procedure for vicia faba

A relatively straightforward approach is described to obtain differential contrast in sister chromatids for SCE detection in Vicia faba after BrdUrd incorporation. The hydrolysis time of the well-known Feulgen reaction was extended to differentially degrade the DNA, the BrdUrd-substituted strands being more resistant. The procedure may easily be adapted for other plant species with large chromosomes.     

Comparison of baseline sister-chromatid exchanges (SCE), cyclophosphamide-, ethylnitrosourea (ENU)-induced SCE, ENU-induced cell-cycle delay and chromosome aberrations between Peru and laboratory mice

The baseline sister-chromatid exchange (SCE) frequency and sensitivity to the effects of the mutagens cyclophosphamide (CPP) and ethylnitrosourea (ENU) in bone-marrow cells of descendants of wild mice trapped from Rimac valley in Peru (Peru mice) were studied and compared to the same effects in laboratory mice. Baseline SCE of the Peru mice were significantly higher than those of the C57BL/6J and DBA/2 mice. The average SCE/cell of 4 Peru mice was 5.4 (range 3.8-7.6), while the average of SCE/cell of either 4 C57BL or 5 DBA mice was 3.2 (range 3.0-3.4). The variation of SCE/cell among Peru mice studied was statistically significant whereas among C57BL or DBA mice it was not. SCE frequencies of primary cultures derived from the ear tissue of 10 Peru (mean SCE/cell = 8.5) were also significantly higher than those of 6 C57BL mice (mean SCE/cell = 7.4). CPP treatment resulted in a dose-dependent increase of SCE frequencies in bone-marrow cells of all the mice. However, some of Peru mice treated with CPP had significantly higher SCE than the other Peru mice and than all of the C57BL and DBA mice treated with equivalent dose. ENU induced increased SCE frequencies in Peru and C57BL mice. Again some of Peru mice either had significantly higher SCE, greater extent induced cell-cycle delay or chromosome aberrations (CA) than other Peru mice and than of all the C57BL mice treated with equivalent dose.(ABSTRACT TRUNCATED AT 250 WORDS)     

Spontaneous and busulphan-induced sister-chromatid exchange (SCE) frequencies in haematologically normal human bone marrow and lymphocytes

In a study of 14 patients who were not treated with either chemotherapy or irradiation, 13 patients had lower siste-chromatid exchange (SCE) frequencies in their bone-marrow cells than in their lymphocytes. For both bone marrow and lymphocytes, there was significant inter-patient variability in SCE frequencies, but there was no correlation between the bone-marrow and lymphocyte values.

The effect of exposing bone-marrow cells to busulphan (BUS) in vitro was investigated using doses up to 5.0 μg/ml. The dose-response relationships between BUS and SCEs in vitro were found to be similar for bone marrow and lymphocytes.     

SCE frequencies in rabbit lymphocytes as a function of time after an acute dose of cyclophosphamide

Following acute and chronic exposures to various chemicals in vivo, the average SCE frequency in human and rabbit lymphocytes has generally been shown to decrease with time posttreatment. The rate of this decline varies, however, and little data have been published pertaining to the decrease in SCEs soon exposure. To gain more information about the immediate decline in SCEs with time, we injected rabbits with a single dose of 35 mg/kg cyclophosphamide (CP) and determined SCE levels in circulating lymphocytes at various times 5 h to 2 weeks after treatment. We observed a rapid decline in SCE frequencies within 5 days, and by 10 days post-exposure the SCE levels were back to control values. The distributions of SCEs among cells and the number of circulating lymphocytes were also analyzed at each time. Within 2–3 days posttreatment we observed a rapid loss of cells with high SCE levels concomitantly with a rapid decline in circulating lymphocytes and a decrease in the average SCE frequency. When the number of lymphocytes began to increase, the number of cells with normal SCE values also increased. By 10–11 days after CP, the lymphocyte count had recovered, the SCE frequency had returned to control levels, and the distribution of SCEs among cells was almost identical to the control distribution. These data, in addition to published information on rabbit lymphocyte lifespan, suggest that the decline in SCE levels with time posttreatment is a function of lymphocyte turnover.     

Correlations between sister chromatid exchange frequencies and replicon sizes: A model for the mechanism of SCE production

Baseline sister chromatid exchanges (SCEs) increase as a function of average replicon size in a variety of human and Chinese hamster cell lines. This observation is the basis for a model in which SCEs are generated by errors in the unravelling of daughter double helices by topo isomerases. These errors cause exchanges behind the replication fork, not at the fork as several current models assume. This model also provides an explanation for SCEs induced when residual DNA damage that has been replicated interferes with the normal processes of unravelling the daughter strands.     

Sister chromatid exchange (SCE) frequencies differ between directly prepared cytotrophoblasts and cultured mesenchymal core cells

Summary Analysis of sister chromatid exchange (SCE) in chorionic villus cells may become useful in measuring the response of fetal tissues to clastogens or mutagens or for prenatal diagnosis of chromosome breakage syndromes such as Bloom syndrome. Previous studies have failed to analyze cytotrophoblastic cells and mesenchymal core cells, or have found no difference between SCE frequencies in directly prepared and cultured cells. Our data indicate significant differences in SCE frequencies between the two cell types: SCE frequency in directly prepared cytotrophoblasts was 6.73 SCE/cell ± 1.6, whereas SCE frequency in cultured mesenchymal core cells was 10.31 SCE/cell ± 0.49 (P < 0.001). SCE analyses involving chorionic villi must take into account cell type.Presented at the 41st Meeting of the American Society of Human Genetics, Cincinnati, Ohio     

Paradoxical changes in SCE frequencies persistently elevated in vivo, on exposure to a mutagen in vitro

Lymphocyte cultures from 4 individuals with persistently significantly elevated frequencies of sister-chromatid exchange (SCE) were examined with no treatment, and with 2 concentrations of mitomycin C. In each of the 4 cases, the mean level of SCEs in the untreated lymphocytes exhibited a paradoxical reduction in SCE frequency when exposed to the lower (0.005 microgram/ml) of the two doses of mitomycin C. At the second higher dose of mitomycin C (0.025 microgram/ml) the mean level of SCE/cell exceeded the untreated mean. When the distributions of SCE/cell were examined it appeared that the untreated cultures had two or more populations of cells; one was in the normal SCE frequency range, while the second population was in an elevated SCE frequency range. The paradoxical reduction in SCE frequency was apparently due to elimination of, or mitotic inhibition of cells in the highest range of SCE frequency, while a small elevation in SCEs was initiated in the cells with a normal SCE frequency. Thus, mean levels of SCE/cell can be misleading. This data suggests that new exposure to the same or a different genotoxic agent might possibly result in a misleading lowering of the mean SCE frequency.     

SCE variability in lymphocytes and fibroblasts

Summary To determine whether the sister chromatid exchange (SCE) distributions obtained in lymphocytes and fibroblasts from different individuals are comparable, a controlled study was set up. Peripheral blood and skin biopsies were taken on the same day from five individuals living for years under the same environmental conditions. All samples were treated in the same fashion, and the SCEs were scored in 50 metaphases of peripheral blood lymphocytes and of skin fibroblasts in an early and in a late passage. A repeat blood sample was taken from the same five indivuduals 1 year later. Based on the results obtained in this first part of the study, five randomly chosen healthy blood donors were sampled at different times and studied in the same fashion. Each chromosome was identified, and the SCE scores were tabulated per chromosome over 50 metaphases. The statistical analysis consisted of fitting log linear models to these scores and examining the best fit by determining the exceedance probabilities (observed significance level). For lymphocytes, the results indicated that the SCE distributions depended only on the chromosome examined, and not on BrdU-exposure time, individuals, or time of sampling. Treatment with ethyl methane sulfonate (EMS) increased the number of SCEs proportionally on all chromosomes. Analysis of the SCE scores on lymphocytes and fibroblasts of the five individuals and on their low and high passage fibroblast cultures revealed the necessity of including higher order interactions in order to fit a suitable model to the data. Therefore comparison of the SCE scores of lymphocytes with those of fibroblasts or comparison of scores on fibroblasts from different individuals could not be done. In practice, to compare samples or individuals, it suffices to score the SCE on a limited number of chromosomes (e.G., the A group) of 50 metaphases.     

SCE induction and cell-cycle delay by toxaphene

Toxaphene is genotoxic in mammalian cell systems and also inhibits cell replication. It was therefore used to investigate possible masking of SCE induction due to cell-cycle delay. In this study, toxaphene-treated Chinese hamster lung (Don) cells exhibited a dose-dependent decrease in cell-cycle progression compared with untreated cells. At high, nontoxic toxaphene levels (15 micrograms/ml), cell cycling also slowed as the toxaphene treatment time was increased. Toxaphene induced significantly higher numbers of SCEs in treated cells, demonstrating a dose- and treatment time-relationship. Slopes of dose-response curves were 0.29, 0.43 and 0.77 SCE/micrograms toxaphene for 20.5 h, 24.5 h and 28.5 h incubation, respectively. There were no changes in SCE values in control cultures even when slower dividing cells were sampled e.g. at longer incubation times. Thus, higher SCE values in Chinese hamster cells were not associated per se with slower or more delayed cells. The results demonstrate that longer toxaphene treatment times were not necessary for obtaining sufficient harlequin-stained cells for SCE analysis, but that higher numbers of SCEs occurred in slower dividing cells, following prolonged incubation of cultures treated with toxaphene.     

Studies of mammalian chromosome replication. I. BrdU-induced differential staining patterns in interphase and metaphase chromosomes

The patterns of differential staining based on the effects of BrdU-substitution in chromosomal DNA have been examined in both metaphase chromosomes and prematurely condensed chromosomes (PCC) of interphase Chinese hamster cells. Results indicate that differential staining may be obtained in chromosomes from all stages of the cell cycle and correspond to the semi-conservation mode of DNA replication. Such fidelity of differential staining in both interphase and metaphase chromosomes suggests that components essential for induction of differential staining are present throughout the cell cycle and chromosomes may contain similar structures and organization throughout the cycle.     

A pulse BrdU method for SCE

Using ‘reverse’ harlequin staining (bromouracil-substituted chromatin staining dark), it is possible to detect at metaphase a pulse of bromodeoxyuridine (BrdU) incorporated during S-phase. Provided that this pulse is of reasonable duration, fairly uniform staining along the chromatids of some S-cells is achieved, and no difficulty is encountered in observing and scoring SCE in such cells at second division.Thus, it is possible to define within an asynchronous population a narrow cohort of target cells and recover these for SCE scoring at second division irrespective of treatment perturbation. This serves to reduce the heterogeneity found in the usual terminal BrdU SCE protocols for such populations and should lead to more reliable and repeatable quantitative results.The method is illustrated for mitomycin C given to dividing human blood lymphocytes using both simultaneous and delayed pulse modes.     

Comparison of in vivo and in vitro SCE induction

In vivo and in vitrod sister-chromatid exchange (SCE) induction and cell replication kinetics were compared in P388 cells exposed to 4 mutagens. While concordance was observed between SCE induction and inhibition of cell replication kinetics, certain mutagens were more potent in vivo while others were more potent in vitro. These results indicate that caution should be applied before equating in vivo and in vitro mutagen exposures.     

Lymphocytes proliferation kinetics and SCE variation after rubella vaccination

The effect of anti-rubella vaccination on lymphocyte proliferation kinetics in vitro and on SCE frequency was studied in three young women. The studies were carried out by taking blood samples before vaccination (day 0) and subsequently on days 7, 14, 28 and 42. The mitotic index (MI) shows a decrease at day 14 and 28 followed at day 42 by an increase above day 0 levels. The average cell cycle (ACC) shows a decrease at day 14 followed by an increase at day 28. Complex variations were also found in the percentage distribution of cells in the various division classes. The SCE frequency showed variations inverse to the MI. The whole picture seems to indicate the existence of changes in the lymphocyte population which can be correlated with immune response.