Anticlastogenic effect of Vitamin C on cisplatin induced chromosome aberrations in human lymphocyte cultures

Vitamin C (ascorbic acid) is an antioxidant that can scavenge free radicals and protect cellular macromolecules, including DNA, from oxidative damage induced by different agents. The protective effect of Vitamin C on cisplatin induced chromosome aberrations has been determined in the human peripheral lymphocyte chromosome aberration test in vitro. The results of treatments with Vitamin C indicated that it statistically significantly decreases the number of chromosome aberrations and number of metaphases with aberrations induced with cisplatin, but it can not completely protect cells from damage. The test concentrations of Vitamin C (10 and 100 μg/ml) had a limited antimutagen effect on cisplatin (0.5 μg/ml), which can cause genetic damage through free radical mechanisms. The antimutagen effect included the anticlastogenic effect of Vitamin C and its ability to decrease the number of aneuploid mitoses. Vitamin C showed the most efficient anticlastogenic effect during simultaneous treatment with cisplatin. Also, Vitamin C reduced cell toxicity of cisplatin during simultaneous treatment.     

Effects of caffeine on chromosome aberrations and sister-chromatid exchanges induced by mitomycin C in BrdU-labeled human chromosomes.

The BrdU-Hoechst staining technique has been used in analyzing the effect of caffeine (CAF) on chromosome aberrations and sister-chromatid exchanges (SCEs) induced by mitomycin C (MC). CAF increased the frequency of SCE in MC-treated chromosomes in all specimens. The combination of MC and CAF caused a remarkable increase in all types of chromosome aberrations, but the most startling effect was the appearance of many cells with multiple aberrations (shattered chromosomes). The BrdU-Hoechst technique showed that the shattered chromosomes did not appear in cells that had replicated only once, but did occur in cells which replicated twice in the presence of MC and CAF. The large majority of chromatid breaks observed did not involve areas common to SCE; and the SCE frequency significantly increased in spite of the existence of multiple breaks. This indicates that very few of the breaks are incomplete exchanges and that the mechanism for formation of SCE might be different from that of chromosome breaks. In another experiment, monofunctional-MC (M-MC) had a small effect on SCE rates, though it induced shattered chromosomes with CAF post-treatment. Possible differences in the mechanisms leading to SCE and chromosome breaks are discussed.     

Significance of structural chromosome aberrations in human sperm: analysis of induced aberrations

Summary A significant increase in the incidence of structural chromosome anomalies has been observed in the sperm of patients treated with radio and/or chemotherapy for different types of cancer when analyzed by the interspecific fertilization of hamster eggs. The analysis of these aberrations shows that while in controls only 9.4% of structural abnormalities are of the stable type, in treated patients this figure increases to 39.3%, thus indicating that the anomalies have not been produced during the fertilization of the hamster egg. However, it is possible that part, or even most, of the breaks appear as a result of a reduced repair capacity of sperm chromosomes in the cytoplasm of the hamster egg.     

Inhibition of condensation in the late-replicating X chromosome induced by 5-azadeoxycytidine in human lymphocyte cultures

Summary The cytidine analogue 5-azadeoxycytidine (5-aza-dC) induces a very distinct inhibition of condensation in the genetically inactive, late-replicating X chromosome (X_L) when applied to human lymphocyte cultures. One of the two X chromosomes in cytogenetically normal female cells becomes dramatically longer than its homologous partner. The highest rate of metaphases with an undercondensed X_L chromosome is achieved when 5-aza-dC is added at a final concentration of 10^-5 M 2 h before cell harvesting. The interactions between 5-aza-dC and chromosomal DNA as well as the factors involved in X chromosome inactivation are discussed.     

Induction of micronuclei and anaphase aberrations by cytochalasin B in human lymphocyte cultures

The frequency of micronucleated cells in isolated 72-h human lymphocyte cultures treated with cytochalasin B (Cyt-B; 1.5-6 micrograms/ml for the last 28 h) was 9-21 times higher (mean 14.6 times) among multinucleate than binucleate cells. At 3 micrograms/ml, the concentration of Cyt-B originally recommended for the human lymphocyte micronucleus assay, the frequency of micronucleated multinucleate cells was 8.5%, while 0.7% of the binucleate cells had a micronucleus. Although no dose-dependent induction of micronuclei could be observed for either of the cell types, increase in the concentration of Cyt-B was associated with a decrease in the ratio of multinucleate to binucleate cells. Treatment with Cyt-B (1.5-12 micrograms/ml) increased the frequency of anaphase cells with aberrations, especially lagging chromatids. This finding was explained by a dose-dependent increase in multipolar (greater than or equal to 3 poles) divisions which had a high frequency of anaphase aberrations (39-53%), irrespective of the concentration of Cyt-B. Bipolar anaphases did not show a significant increase in aberrant cells, although a suggestive dependence on the concentration of Cyt-B was observed. The findings indicate that the high frequency of micronuclei in multinucleate lymphocytes produced by Cyt-B is due to mitotic errors arising when bi- (and multi-) nuclear cells divide. To avoid possible artifactually high micronucleus frequencies due to inclusion of cells that have divided greater than or equal to 2 times in the presence of Cyt-B, it is recommended that, in the human lymphocyte micronucleus assay using the cytokinesis-block method, the cell culture time is reduced to minimize the frequency of such cells and that only good preparations and regularly shaped binucleates are included in the analysis.     

Studies on endoreduplication. III. Endoreduplication induced by mitomycin C in PHA-stimulated tonsillar lymphocyte cultures

The activity of mitomycin C (MMC) to induce endoreduplication was examined in PHA-stimulated tonsillar lymphocyte cultures. Cellular kinetics of both diploid and endoreduplicated cells was studied by the combined techniques of thymidine and BrdU incorporation. The kinetic data showed that the increase of endoreduplicated cells induced by MMC correlated with the increase of the second-generation diploid cells. Also, it showed that the majority of endoreduplicated cells found at the time when the maximal number of endoreduplicated cells was obtained had incorporated BrdU only once into their diplochromosomes. This suggests that MMC-induced endoreduplicated cells originate from MMC-induced G2-arrested cells.     

The effect of oxygen concentration on the X-ray induction of chromosome aberrations in human lymphocytes.

In vitro dose--response curves, for unstable chromosome aberration induction in human lymphocytes under conditions of full oxygenation or of anoxia, have been obtained using 250 kVp X-rays. Dicentric yields have been fitted to the quadratic function Y = alpha D + beta D2. An Oxygen Enhancement Ratio (OER) ranging from 7.2 to 2.7 was calculated from the coefficients of these curves possibly indicating that the data do not fit to the dose-modifying model of the oxygen effect although the differences are not statistically significant. A similar analysis of total aberration data yields an OER of 3.6 to 2.7 fitting much more satisfactorily to the dose-modifying model. Variations in dicentric yield induced by 3.0 Gy and 0.75 Gy of X-rays with increasing oxygen concentration were plotted and for each dose a constant dicentric yield was observed at oxygen levels of 2 and 250 ppm. Above 250 ppm yields increased steeply up to about 1% oxygen and then more gradually to a maximum at 100% oxygen.     

The fate of X-ray-induced chromosome aberrations in blood lymphocyte culture

The BrdU-Giemsa method which facilitates an unequivocal identification of metaphases at different cycles has been utilized to investigate the fate of X-ray-induced chromosome aberrations in the blood lymphocyte culture system of the Indian muntjac which has the lowest diploid number (2n = 6 female/7 male) and easily distinguishable large-sized chromosomes. The results demonstrate that about 50% of dicentrics and only 12% of rings were transmitted from the first cycle to the second. There were as high as 73% abnormal cells in the second cycle as against 94% in the first cycle following 4.0 Gy. However, the frequencies of dicentrics, rings and of abnormal cells were greatly reduced in the third+ cycles. The frequencies of acentric fragments per post-irradiated first, second and third+ division cell were 2.21, 0.64 and 0.24, respectively. In sharp contrast to all earlier reports, about 75% of them were retained as a single acentric fragment in the second cycle. Analysis of fragment segregation during anaphase separation supports this finding. The survival probability of dicentrics and rings was found to be more than 60% in the second and only 18% in the third+ cycle.     

Dose-response relationship for chromosome aberrations induced by fecapentaene-12 in human lymphocytes

Chromosome analyses were carried out in human lymphocytes exposed to a synthetic racemic all-trans fecapentaene-12 at 2-24 microM. A dose-dependent increase of the incidences of chromatid-type changes with distinct saturation at higher doses could be observed. The results reveal for the first time that fec-12 is a potent direct-acting mutagen in human lymphocytes.     

Comparative frequency of chromosome aberrations in a human lymphocyte culture depending on the mode of neutron irradiation

The study of the dose dependence of the incidence of chromosome aberrations in a human lymphocyte culture exposed at the G0 stage to pulsed neutrons of 0.7 MeV revealed no significant distinctions in the cytogenetic effect at the pulse frequency of 1 and 5 Hz, but at 100 Hz it was somewhat increased. As to the biological effectiveness, pulsed neutrons of 0.7 MeV occupied an intermediate position between 0.35 and 0.85 MeV static neutrons.     

Analysis of chromosome aberrations in human peripheral lymphocytes induced by 5.4 keV x-rays

Irradiation of human lymphocytes by x-rays has been seen, in past studies, to produce increasing frequencies of chromosome aberrations at lower x-ray energies. However, in one earlier irradiation experiment with chromium x-rays, the relative biological effectiveness (RBE) did not appear to be larger than that of hard x-rays, especially at higher doses. A possible reason for this unexpected result may have been the irradiation and culture conditions. We have, therefore, in the present study used a technique that has been developed in our laboratory to ensure uniformity of irradiation within lymphocytes and to avoid artefacts due to the cell cycle kinetics. Monolayers of 3-h-stimulated lymphocytes were exposed to 5.4 keV x-rays. A linear-quadratic dose-response was found for dicentrics. The comparison to an earlier finding with 220 kV x-rays shows the expected result of the RBE of the 5.4 keV x-rays to be above that of 220 kV x-rays. The intercellular distribution of dicentrics did not differ significantly from a Poisson distribution. Received: 17 January 1997 / Accepted in revised form: 17 July 1997     

Anticlastogenic effects of a polyvitamin product, 'Pharmavit', on gamma-ray induction of somatic and germ cell chromosome aberrations in the mouse.

The polyvitamin product 'Pharmavit' (Pv), comprising vitamins A, D2, B1, B2, B6, C, E, nicotinamide, and calcium pantothene, was tested for anticlastogenic properties against gamma-rays in mice. Pretreatment with Pv consisted of daily administration by gavage for 30 days at dose levels corresponding to clinical recommendations for an adult human, as recalculated in terms of mg/kg. Findings indicated a reduction of chromosome aberrations in bone marrow cells from mice exposed to 3.0 Gy 137Cs gamma-rays; the reduction concerned predominantly fragments of the chromatid type. Furthermore, a reduction factor of 1.6 was obtained for the frequency of reciprocal translocations induced by spermatogonial irradiation in mice exposed to 4.0 Gy gamma-rays. Pretreatment with vitamin C alone, at the dose present in Pv, proved nearly ineffective in protecting from chromosome aberrations in bone marrow cells. Pharmavit is believed to be a promising agent for application to human populations exposed to the carcinogenic and genetic hazards of ionizing radiation.