Incidence Of Structural Chromosome Aberrations Research Articles

An investigation of the potential effect of vacuoles in human sperm on DNA damage using a chromosome assay and the TUNEL assay

The aims of this study were to establish whether individual differences exist in the frequency and size of vacuoles found in human sperm and to ascertain whether such vacuoles are involved in causing DNA damage. Morphologically normal sperm were obtained from 15 IVF and 2 ICSI patients and 3 fertile donors. (i) Sperm heads were analyzed for the presence of vacuoles under a 1000× differential interference contrast microscope. (ii) In three patients and two donor samples, structural chromosomal damage was evaluated in normal sperm containing large vacuoles and selected at 1000× magnification for injection into mouse oocytes. (iii) In 10 patients and two donor samples, confocal laser microscopy detected DNA damage in sperm-exhibiting large vacuoles and stained with an in situ cell death detection kit. (i) Vacuoles were observed in almost all normal sperm from patient and donor ejaculates and were mainly located at the tip or middle area of the sperm heads. However, average incidence of normal sperm exhibiting large vacuoles was 4.6 and 4.2% in the patient and donor groups, respectively. (ii) Sperm chromosome assays did not reveal any differences in the incidence of structural chromosome aberrations between sperm exhibiting large vacuoles and those without them (9.1 versus 4.1%). (iii) No significant difference in frequency of TUNEL-positive cells was found between normal sperm with large vacuoles and those without them in the samples examined. Among 227 sperm exhibiting large vacuoles, only 7 cells were TUNEL positive. The results showed that large vacuoles were not responsible for DNA damage, suggesting that intra-cytoplasmic injection of morphologically selected sperm may not be required for patients who produce high-quality semen.

Structural chromosomal aberrations, aneuploidy, and mosaicism in early cleavage mouse embryos derived from spermatozoa exposed to γ-rays

Purpose: To quantitatively and qualitatively investigate the changes in chromosomal aberrations during early cleavage in mouse embryos derived from γ-irradiated spermatozoa.Materials and methods: Mature males were exposed to 2 Gy or 4 Gy of 137Cs γ-rays, and their spermatozoa were used to produce embryos via in vitro fertilisation (IVF). The metaphase chromosomes were prepared from one-cell, two-cell, and four-cell embryos. In the chromosome preparations from two-cell and four-cell embryos, the separation of the sister blastomeres was precluded by treatment of the embryos with concanavalin A. The incidence of embryos with structural chromosomal aberrations, aneuploidy, or mosaicism was estimated. The fates of the different types of γ-ray-induced structural chromosomal aberrations were also investigated in those embryos.Results: The exposure of spermatozoa to 2 Gy or 4 Gy γ-rays caused structural chromosomal aberrations in 25.9% and 35.7% of the resultant one-cell embryos, respectively. At two-cell embryonic stage, the incidence of structural chromosomal aberrations was 17.4% in the 2 Gy group and 27.1% in the 4 Gy group. At the four-cell embryonic stage, although the incidence of control embryos with structural chromosomal aberrations was considerably high, the net incidence of embryos with radiation-induced structural chromosomal aberrations was similar to that at the one-cell stage. The incidence of aneuploidy was high in two-cell and four-cell embryos after both doses of γ-rays. The incidence of mosaicism increased significantly in dose- and embryonic-stage-dependent manners. Anaphase lag, and the degeneration and non-disjunction of the aberrant chromosomes were frequently observed in aneuploid and mosaic embryos.Conclusions: Mouse sperm DNA is highly vulnerable to γ-rays. The structural chromosomal aberrations of sperm origin are unstable in their behaviour and structure during cleavage, and therefore cause secondary aneuploidy and mosaicism in the early cleavage embryos.

Chromosome analysis of mouse zygotes produced by intracytoplasmic injection of spermatozoa exposed to acrosome reaction inducing agents methyl-β-cyclodextrin and calcium ionophore A23187

This study was performed to investigate whether removal of cholesterol from the plasma membrane and collapse of the acrosome can prevent structural chromosome aberrations of paternal origin in mouse zygotes produced by intracytoplasmic sperm injection (ICSI). Mouse spermatozoa were treated with methyl-beta-cyclodextrin (M beta CD) to remove cholesterol from the plasma membrane and with calcium ionophore A23187 to collapse the acrosome. Chromosomes of zygotes derived from M beta CD- and ionophore-treated spermatozoa were analyzed at the first mitotic metaphase. Both chemical agents effectively induced the acrosome reaction. Incidence of structural chromosome aberrations in ICSI zygotes derived from M beta CD-treated spermatozoa was similar to that in zygotes produced by in vitro fertilization (IVF) with the same spermatozoa, but significantly lower compared to ICSI zygotes derived from acrosome-intact spermatozoa. Chromosome aberration rates in ICSI zygotes derived from ionophore-treated spermatozoa were evidently high compared to IVF zygotes. Induction of the acrosome reaction through cholesterol efflux by M beta CD can prevent chromosome aberrations of paternal origin, while use of ionophore to induce the acrosome reaction exerts detrimental effect on paternal chromosomes in ICSI zygotes.

Protection of rat chromosomes by melatonin against gamma radiation-induced damage

The aim of this study was to investigate the protective effect of melatonin (2.5 mg/kg/day, given to rats five times by intra-peritoneal injection) against damage in bone-marrow chromosomes induced by a single dose of gamma radiation (4.0 Gy whole-body irradiation; WBI). Ninety-six male albino rats were divided into four equal groups of 24 rats each. They were designated as I—non-irradiated, non-treated control rats, II—non-irradiated rats treated with melatonin for five successive days, III—whole-body gamma-irradiated rats and IV—rats injected with melatonin daily for five successive days, then subjected to whole-body gamma irradiation 2 h after the final melatonin injection. Six rats from each group were sacrificed at days 1, 3, 7 and 10 following treatment and/or irradiation and their bone marrows were flushed out for micronuclei scoring and chromosomal analysis. WBI resulted in significant elevations in bone-marrow polychromatic erythrocytes containing micronuclei in their cytoplasm, and caused a significant decrease in the mitotic index of bone-marrow cells; in addition, there was a significant increase in the frequency of aberrant bone-marrow cells and in the different types of structural chromosomal aberration. Melatonin injection prior to WBI significantly reduced the mean frequencies of micro-nucleated polychromatic erythrocytes and of aberrant cells, as well as the incidence of structural chromosomal aberrations in bone-marrow cells; it also caused a highly significant elevation in the value of the mitotic index of bone-marrow cells. This investigation clarifies the protective and/or ameliorative role played by melatonin against deleterious effects of gamma radiation.

Possible causal factors of structural chromosome aberrations in intracytoplasmic sperm injection of the mouse.

Incidence of structural chromosome aberrations in mouse one-cell embryos produced by intracytoplasmic sperm injection (ICSI) with mature epididymal spermatozoa were influenced by sperm incubation medium and time. When spermatozoa were incubated in bicarbonate-buffered TYH for ≤0.5 h, the embryo aberration rates were significantly higher than in vitro fertilization (IVF) embryos. However, after the incubation of spermatozoa in the same medium for ≥2 h, the aberration rates were close to the IVF embryo level. When spermatozoa were incubated in bicarbonate-buffered mCZB, hepes-buffered H-TYH and H-mCZB, and phosphate-buffered PB1, the increased incidences of aberrations were observed at any incubation time. In the case of sperm incubation in H-TYH, H-mCZB and PB1, the aberration rates increased in a time-dependent manner. Chromosome aberrations generated by ICSI were transmissible to offspring. On the other hand, the aberration rate in embryos derived from testicular spermatozoa was independent of the medium type and incubation time. Thus, the incubation media appears to have no effect on sperm chromatin. TYH can effectively induce capacitation and acrosome reaction, while H-TYH, H-mCZB and PB1 never induce these spermatozoal events. It is probable that the cholesterol-rich plasma membrane and intact acrosome injected into the ooplasm affect sperm chromatin remodeling, thus resulting in the generation of chromosome damage in ICSI embryos.

Retinyl palmitate is non-genotoxic in Chinese hamster ovary cells in the dark or after pre-irradiation or simultaneous irradiation with UV light

Retinyl palmitate (RP), an ingredient of cosmetic and medical skin-care preparations, has been reported to be photo-genotoxic/photo-clastogenic in mouse lymphoma cells ( Tk locus) as well as in human Jurkat T-cells, as measured by use of the comet assay. Given that these results were obtained under exploratory conditions, we re-investigated the photo-genotoxicity of RP following a protocol consistent with current recommendations for photo-genotoxicity testing of drugs and chemicals. We tested RP in Chinese hamster ovary (CHO) cells in the dark (standard chromosome aberration test), under pre-irradiation (UVA irradiation of cells and subsequent treatment with RP) or simultaneous irradiation (irradiation of cells and RP together, standard photo-genotoxicity protocol) conditions. UVA irradiation was applied at 350 and 700 mJ/cm 2 with the high UV dose targeted to produce a small increase in the incidence of structural chromosome aberrations (CA) in cells not treated with RP. RP was tested up to and above its limit of solubility in the culture medium (20–40 μg/mL). There was no overt cytotoxicity under dark or different irradiation conditions. Treatment of cells with RP in the dark, as well as treatment under pre- or simultaneous irradiation conditions failed to produce biologically significant increases in the incidence of CA, whereas the positive control substances 4-nitroquinolone and 8-methoxypsoralene produced significantly positive effects in the dark or under simultaneous irradiation, respectively. Overall, our results failed to confirm the reported positive photo-genotoxic effects, and suggest that they may have been due to the test conditions, i.e. high irradiation doses, high cytotoxicity or re-irradiation of photo-products. In conclusion, our data suggest that, under standard conditions for testing photo-genotoxicity, RP had no in vitro genotoxic or photo-genotoxic potential and is therefore unlikely to pose a local or systemic genotoxic or photo-genotoxic risk.

THE ROLE OF CD40 LIGAND IN EARLY DETECTION OF CHROMIUM-INDUCED IMMUNOTOXIC EFFECTS AMONG EXPOSED WORKERS

Objective: the immunological effect of chromium exposure among workers in tanning industry was assessed by studying the incidence of structural chromosomal aberrations in lymphocytes. Early monitoring of immunotoxicity and possibility ofcarcinogenecity using the CD40 ligand member of the tumor necrosis superfamily of trans-membrane proteins was evaluated. Methods: levels of chromium in blood (S-Cr) and urine (U-Cr) were measured using the graphite furnace atomic absorption spectrophotometer. The CD40 Ligand (CD40L) was determined using ELISA. Whole blood was treated and chosen metaphases were recorded and scored for morphological aberrations and rearrangements.Results: the study was carried out on 30 workers exposed to chromium in chrome-tanning for 17.13 ± 8.09 years, and a matched group of 28 administrative workers. The mean serum chromium levels (S-Cr) were 4.35 ± 1.46 and 1.89 ± 1.31 μg/dl among exposed and control groups, respectively, and the difference was statistically significant, P<0.00±. The CD40L assessment revealed decreased levels among the exposed population, difference was statistically non-significant. The genotoxic changes in the form of chromosomal and chromatid gaps and breaks as well as dicen-actitricchanges were significantly higher among exposed workers, correlating significantlywith the levels of chromium in both serum and urine after correction for creatinine.A significant negative correlation was reported for the deletions with the level of CD40 ligand. The effect of age, smoking and cumulative exposure indices were dealt with.Conclusion: the use of cytogenetic monitoring of human populations exposed to chemicals has proved to be a useful tool for detecting the chemical's mutagenic effects.CD40L, which is a member of the tumor necrosis factor, may be used as an additional marker to assess the presence of chronic inflammation, an early warning indication of potentially adverse genetic and somatic effects.

Frequent structural chromosome aberrations in immotile human sperm exposed to culture media.

The influence of culture media or centrifugation on chromosomes of immotile human sperm was examined using ICSI into mouse oocytes. In experiment 1, immotile and motile human sperm retrieved directly from ejaculates were injected into mouse oocytes. In experiment 2, immotile human sperm were exposed to seminal plasma or one of four kinds of culture media (HEPES-BWW, modified-BWW, modified-human tubal fluid (HTF) and Dulbecco's phosphate-buffered saline) for 1.5-2.5 h at 18 degrees C in air before microinjection. In experiment 3, immotile human sperm were centrifuged along with HEPES-BWW before microinjection. In experiment 4, frozen-thawed immotile human sperm washed with seminal plasma or HEPES-BWW were injected into mouse oocytes. The hybrid oocytes were prepared for chromosome slides at first cleavage metaphase and were then examined cytogenetically. In experiment 1, there was no significant difference in the incidences of structural chromosome aberrations between motile and immotile sperm (4.3% versus 5.8%). In experiment 2, culture media caused more frequent structural chromosome aberrations (14.3-32.6%) in immotile sperm than did seminal plasma (5.4%). In experiment 3, structural chromosome aberrations were found in 48.1% of the centrifuged immotile sperm, and a live/dead sperm viability test intimated that the aberrant sperm were probably dead. In experiment 4, the incidence of structural chromosome aberrations in frozen-thawed immotile sperm was significantly higher in HEPES-BWW (62.2%) than in seminal plasma (17.2%). The results indicate that immotile sperm do not have significantly more DNA lesions than motile sperm, although DNA of immotile sperm appears to be vulnerable to damage caused by different culture media.

Chromosome aberrations study of pupils in high radon level elementary school.

The ICRP Publication 65 recommends 200-600 Bq x m(-3) as the indoor radon action level for the general public. In Slovenia, a value of 400 Bq x m(-3) has been proposed but not yet approved. In a nation-wide radon project financed by the Health Inspectorate of Slovenia, it was discovered that the elementary school named "S3" belongs to a group of schools with elevated winter indoor radon concentrations up to 7,000 Bq x m(-3). Opening windows and doors during classes substantially decreased radon concentrations, but very seldom below 1,000 Bq x m(-3). Yearly effective doses for pupils, estimated according to ICRP 65, ranged from 7 to 11 mSv. Because the pupils have been subjected to the elevated radon concentrations, special preventive health checks have been performed. The examination protocol included mutagenetic tests, one for structural chromosomal aberrations and the other a micronucleus test. Altogether 85 pupils (37 girls and 48 boys) from the first four grades between the ages of 9 and 12 y were examined. An increase in cytogenetic damage was found for these pupils, compared to the control group, composed of pupils of the same age from another area with indoor radon concentrations in their school of below 400 Bq x m(-3). The incidence of structural chromosomal aberrations reached 2.0% (0.5-4) and micronucleus test was 6.52 per 500 cells with a maximum of 15 in some cases. In the control group structural chromosomal aberrations varied from 0.5 to 2.5%, while the maximum incidence of micronucleus was 9 micronucleus per 500 CB cells. The results obtained are preliminary and suggest a need to expand the study. A long-term radon survey, at least over a year, of the homes and wider residential environment of the pupils would be necessary to assess the correlation between radon exposure and both structural chromosomal aberrations and micronucleus findings.

Effect of injected spermatozoa morphology on the outcome of intracytoplasmic sperm injection in humans

Effect of injected spermatozoa morphology on the outcome of intracytoplasmic sperm injection in humans

Sonication per se is not as deleterious to sperm chromosomes as previously inferred.

Although sonication is a simple way to immobilize ("kill") spermatozoa prior to injection into oocytes, this has been thought to be destructive to sperm chromosomes. Mouse and human spermatozoa were immobilized by sonication and kept in various media for up to 2 h, then their nuclei were individually injected into mouse oocytes for the analysis of chromosomes at the first cleavage metaphase. In both the mouse and human, incidence of structural chromosome aberrations was much higher in the spermatozoa sonicated and stored in Biggers-Whitten-Whittingham medium for 2 h at 37.5 degrees C than in those stored for 5 min in the same medium. We concluded, therefore, that it is not sonication per se but a prolonged exposure of sperm nuclei to extracellular milieu that is detrimental to sperm chromosomes. The incidence of structural chromosome aberrations of mouse and human spermatozoa was significantly reduced when the spermatozoa were sonicated and stored in K(+)-rich nucleus isolation medium containing EDTA. This suggests that sperm chromosome degradation following sperm immobilization by sonication is partly due to detrimental effects of a Na(+)-rich medium and of DNase on sperm chromatin. Ideally, it should be possible to prepare artificial media that maintain the integrity of sperm chromosomes for many hours after immobilization.

Dithiothreitol induces sperm nuclear decondensation and protects against chromosome damage during male pronuclear formation in hybrid zygotes between Chinese hamster spermatozoa and Syrian hamster oocytes.

The present study was undertaken to investigate whether a time lag in sperm nuclear decondensation and male pronuclear formation in the course of development of eggs is associated with any occurrence of structural chromosome aberrations in male genomes of hybrid zygotes between Chinese hamster spermatozoa and zona-free Syrian hamster oocytes. Shortly after insemination, hybrid zygotes were treated with dithiothreitol (DTT) at different concentrations (0.1-10.0 mM) for 30 min to reduce protamine disulphide (S-S) bonds and thereby accelerate sperm nuclear decondensation and male pronuclear formation. The incidence of sperm nuclear decondensation and male pronuclear formation increased with increasing DTT concentrations, indicating that a reduction in S-S bonds effectively induces these cytological events. Chromosomes of male genomes in hybrid zygotes generated by treatment with 1.0 mM, 2.5 mM and 10.0 mM DTT were analysed at the first cleavage metaphase. Incidence of structural chromosome aberrations in each treatment was 34.5%, 27.1% and 24.7%, respectively. There was a significant difference between the incidences with 1.0 mM and 10.0 mM DTT treatment. As the time lag in nuclear decondensation and male pronuclear formation was greatest in the 1.0 mM treatment condition, followed in order by 2.5 mM and 10.0 mM, it is suggested that the lag in sperm nuclear development behind egg development is responsible for structural chromosome aberrations in male genomes of hybrid zygotes.

Improvement of male pronuclear formation during cross-fertilization between Chinese hamster spermatozoa and Syrian hamster oocytes by nocodazole, and chromosome analysis of hybrid zygotes

During cross-fertilization between Chinese hamster spermatozoa and Syrian hamster oocytes, incorporated sperm heads frequently fail to develop into male pronuclei, whereas the group of oocyte chromosomes develop into female pronuclei. The present study applies this cross-fertilization system to the cytogenetic investigation of mammalian hybrid embryos. Immediately after insemination, oocytes were exposed to 0.1 μg/ml nocodazole for 1 hr (1 hr group) or 2 hr (2 hr group), then further cultured. Although the rates of sperm penetration in the 1 hr (48.0%) and 2 hr (75.8%) groups were significantly lower than that in the control group (89.8%), the ratios of male pronuclear formation were higher in both exposed groups (79.4% and 74.2%, respectively) than in the control group (10.6%). These results were apparently due to sperm head decondensation induced during the meiotic arrest of oocytes at metaphase II by nocodazole. Chromosomes of hybrid zygotes obtained after nocodazole exposure were analyzed at the first cleavage metaphase. The incidence of structural chromosome aberrations in the Chinese hamster genome of hybrid zygotes was high in the control (42.1%) and 1 hr (48.8%) groups. This incidence was reduced to 14.4% in the 2 hr group. Because the lag of sperm head decondensation behind the second meiotic division of oocytes was greater in the control and 1 hr groups than in the 2 hr group, untimely sperm head decondensation may be implicated in occurrence of structural chromosome aberrations in the male genomes of hybrid zygotes.

Genotoxic effects in workers exposed to benzene: with special reference to exposure biomarkers and confounding factors.

The aim of the study was to establish a correlation between biomarkers of exposure and cytogenetic test results in workers occupationally exposed to benzene and toluene, with special reference to confounding factors influencing the outcome of the cytogenetic test. The incidence of structural chromosome aberrations and sister chromatid exchanges was studied in the peripheral blood lymphocytes cell genome of 49 female shoe-makers, mean age 38 years, mean length of occupational exposure 17 years and in a group of 27 well-matched controls. Workers were exposed to concentrations of benzene up to 15 ppm and of toluene up to 50 ppm. The presence of benzene and toluene in the workers' blood samples, and the presence of phenol in pre- and post-shift urine were considered proof of occupational exposure. Chromosomal aberration analysis revealed a significant increase in dicentric incidence in the exposed group compared to the controls (P = 0.004). However, significant correlation between cytogenetic test results and the exposure biomarkers was not established. On the contrary, correlation between the cytogenetic test results and data on confounding factors (e.g. age and alcohol consumption), was marked. The major point raised by this study is the influence of confounding factors on the cytogenetic test outcome. This imposes the need for caution in the interpretation of cytogenetic test results, and ultimately in the estimation of individual genotoxicity risk related to low level benzene exposure.

Analysis of chromosome constitution of human spermatozoa with normal and aberrant head morphologies after injection into mouse oocytes.

The chromosome constitution of human spermatozoa was determined after injecting individual spermatozoa into mouse oocytes. Of a total 279 eggs arrested at first cleavage metaphase, 200 (71.7%) were suitable for the analysis of sperm chromosomes. Incidences of spermatozoa with numerical and structural chromosome aberrations were 1.3 and 6.9% respectively in spermatozoa with normal head morphology, showing values comparable with those found in previous studies using the hamster oocyte-human sperm fusion system. The ratio of X- to Y-bearing spermatozoa did not differ significantly from the expected 1:1 ratio. The incidence of structural chromosome aberrations was about four times higher in spermatozoa with amorphous, round and elongated heads (26.1%) than in those with morphologically normal heads, whereas the incidence of aneuploidy was not significantly different between the two groups. No increase in chromosome aberrations was found in spermatozoa with large heads. The same was true for spermatozoa with small heads. Although the sample size used in this study is rather small, the results nevertheless indicate that some morphological abnormalities in the sperm heads are associated with their chromosome defects.

Potential genotoxic risk related to simultaneous exposure to radionuclides and cytostatics

The aim of the study was to evaluate the genotoxic risk to medical personnel concurrently exposed to ionizing radiation and antineoplastics, using changes in their lymphocyte cell genome as a bioindicator. The study comprised 12 female nurses employed in the nuclear medicine hospital department and an equal number of matched controls. For each examinee, both conventional structural chromosomal aberration analysis and sister chromatid exchange test (SCE) were carried out. According to Student's t-test, neither the incidence of structural chromosomal aberrations (p > 0.6) nor the mean SCE-frequency rate (p > 0.3) were significantly increased among the exposed subjects. Nevertheless, in those exposed, irreparable chromosomal damages and wide SCE-ranges were observed. Such findings suggest the possibility of genotoxic implications of concurrent occupational exposure to ionizing radiation and antineoplastic drugs.

Karyotype analyses of 20 human glioma cell lines.

Human malignant gliomas are frequently associated with loss of gonosomes and chromosomes 13, 17, and 22. Their progression from anaplastic glioma to glioblastoma is marked by additional loss of chromosome 10. In addition, structural and numerical aberrations of chromosome 7 are frequently found. We report on the karyotypes of a series of 20 human gliomas of which 11 were analysed as established cell lines; 9 cases were investigated in early culture, 5 of which later also became established lines. In addition to the frequently reported overrepresentation of chromosome 7, four cell lines with polysomy for chromosome 22 were seen. A high incidence of structural chromosomal aberrations was present in early cultures as well as in cell lines after various in vitro passages. We found that the general characteristics of karyotypic aberrations found in early cultures or direct preparations of dispersed tumour material were reflected in the pattern of aberrations present in cell lines at much later time points. Thus it appears as if no systematic changes can be attributed to long-term cultures. Suspicious losses of chromosomes 14, 18, and 19 or gain of chromosome 22 indicate that individual cases may have originated due to other mechanisms than the ones already hypothesized, i.e., different suppressor genes or amplification of genes other than the EGF-R-gene. None of the established cell lines had a genomic rearrangement of c-erbB 1, c-erbB 2 or of the p 53 gene.

Mutagenicity studies of prednisolone farnesylate (PNF)

Prednisolone farnesylate (PNF) was tested for mutagenicity by Ames test using Salmonella typhimurium (TA100, TA1535, TA98, TA1537) and Escherichia coli (WP2 uvrA), for clastogenic activity in vitro by the chromosomal aberration test in a Chinese hamster fibroblast cell line (CHL), and for induction of micronuclei by the micronucleus test in male ICR mice. 1) In Ames test, PNF with and without metabolic activation showed no mutagenicity in any strains at any dose levels (312-5,000 micrograms/plate). 2) In the chromosomal aberration test, PNF with metabolic activation produced a slight increase in the incidence of structural chromosomal aberrations in CHL cells at 1,500 micrograms/ml. 3) In the micronucleus test, a single administration of PNF caused no significant increase of micronucleated polychromatic erythrocytes at any doses (250-2,000 mg/kg).

Human sperm chromosomes: Long-term effect of cancer treatment

The long-term cytogenetic effect of radio- or chemotherapy or both on male germ cells was evaluated by study of the chromosomal abnormalities in spermatozoa of four men treated for cancer 5–18 years earlier. The cytogenetic analysis of 422 sperm metaphases showed no differences in the aneuploidy rate. The incidence of structural chromosome aberrations was 14.0%, however, which is much higher than in controls. Thus, the high incidence of structurally aberrant spermatozoa observed in our long-term study indicates that antitumoral treatments affect stem-cell spermatogonia and that aberrant cells can survive germinal selection and produce abnormal spermatozoa.

Co-administration of ascorbic acid with cyclophosphamide (CPA) to pregnant mice inhibits the clastogenic activity of CPA in preimplantation murine blastocysts.

Cyclophosphamide (CPA) administered at a dose of 15 mg/kg body weight to pregnant inbred CBA/CaH mice 60 h after copulation significantly elevated the incidence of structural chromosomal aberrations and sister-chromatid exchanges in 96 h blastocysts. When ascorbic acid (800 mg/kg body weight) was co-administered with CPA (15 mg/kg) using different injection sites, the incidence of structural chromosomal aberrations and the number of aberrant metaphases were significantly lower as compared to the CPA-exposed embryos, but still significantly higher than untreated controls. Sister-chromatid exchanges were not significantly different in embryos exposed to CPA only when compared to those exposed to CPA and ascorbic acid. In terms of the developmental potential of embryos, pregnant mice injected with both CPA and ascorbic acid had significantly fewer resorptions per mouse, and a significantly higher number of viable fetuses retrieved on the 18th day of gestation when compared to mice injected with CPA alone. The data in this study demonstrate that the co-administration of ascorbic acid with CPA to pregnant mice ameliorates the CPA-induced clastogenicity and improves the developmental potential of these embryos. This study suggests that some of the CPA-induced genotoxicity is due to the generation of reactive oxygen molecules, but further research is necessary before this can be firmly concluded.