Genetic Hazards Research Articles

The influence of aluminum exposure on male reproduction and offspring in mice

The dominant lethal assay was utilized to assess the reproductive performance in male mice, possible genetic hazards, and persistent damage of aluminum (Al). Al chloride, AlCl 3, was administered subcutaneously to CD-1 adult male mice at dosages of 0, 7, or 13 mg Al/kg body weight/day for 2 weeks of pre-mating periods. Females were not dosed at any time during this study. At the end of the exposure period, each male was caged with three virgin females each day. The mean mating frequencies of the Al-treated groups reduced consistently from week 4 to 6, and a dramatic reduction in male fertility was also observed. However, the mating frequency restored to near normal control levels as the experiment terminated. Results showed significantly higher numbers of post-implantation losses, foetal mortality, and induced petechial haemorrhage; also significant decreases in body weights of viable foetus throughout weeks 3–8 in the Al-treated groups. The weights of the reproductive organs of the Al-dosed animals decreased significantly as Al accumulation increased in the testes. The spermatogenetic impairment within the seminiferous tubules was also apparent. Nevertheless, these disturbances disappeared at the end of the experiments. In summary, the results demonstrated that Al exerted substantial hazards on male reproductive function and produced genetic toxicity. However, these effects were found to be reversible.

Assessment of Genotoxicity of Lignin Substances as Risk Factors for Aquatic Ecosystems

The genotoxicity of lignin substances (LSs) isolated from biologically purified wastewaters of the pulp and paper industry was studied in experiments on Baikal endemic mollusks and corn. Both toxicity and mutagenic activity of LSs were revealed, with their levels depending on the physicochemical characteristics of LSs, the test organism, and experimental conditions. These results provide evidence that LSs discharged into water bodies create a genetic hazard to their ecosystems.

Monitoring of radiation-induced germline mutation in humans.

Estimating the genetic hazards of radiation and other mutagens in humans depends on extrapolation from experimental systems. Recent data have shown that minisatellite loci provide a useful and sensitive experimental approach for monitoring radiation-induced mutation in humans. This review describes the progress made in validating this approach and presents the results of recent publications on the analysis of minisatellite mutation rates in the irradiated families.

Monitoring of radiation-induced germline mutation in humans.

Estimating the genetic hazards of radiation and other mutagens in humans depends on extrapolation from experimental systems. Recent data have shown that minisatellite loci provide a useful and sensitive experimental approach for monitoring radiation-induced mutation in humans. This review describes the progress made in validating this approach and presents the results of recent publications on the analysis of minisatellite mutation rates in the irradiated families.

Future of radionuclide therapy

Despite much research over the last few decades, there still remains considerable uncertainty as to the genetic impact of ionizing radiation on human populations, particularly at low levels. The aim of the present study was to provide data on the genetic hazards due to occupational exposure of low doses of ionizing radiation in nuclear medicine departments. The assessment of primary DNA damage in peripheral blood leukocytes of medical staff was performed using the alkaline comet assay and the data obtained were compared with the results of conventional cytogenetic biodosimetry using the chromosome aberration (CA) test. Altogether 120 subjects (60 exposed and 60 controls) participated in the study. Statistically significant increases in primary DNA damage and increased frequencies of CAs compared to controls were observed. Within the exposed population, significant inter-individual differences in DNA damage were found, indicating differences in genome sensitivity. Age and gender were not confounding factors, while smoking enhanced the levels of primary DNA damage only in control subjects, as revealed by both biomarkers studied. The present study suggests that genotoxic damage results from exposure to chronic low doses of ionizing radiation in nuclear medicine departments. Therefore, the exposed medical personnel should carefully comply with the radiation protection procedures and should minimize radiation exposure where possible to avoid potential genotoxic effects. The results obtained in this study point to the significance of biological indicators providing information on the actual risk to the radiation exposed individuals. According to our results, the alkaline comet assay and CA test are sensitive biomarkers that can be used as additional complements to physical dosimetry for assessing exposure to radiation in nuclear medicine personnel.

Studies on the in vivo and in vitro mutagenicity and the lipid peroxidation of chlorinated surface (drinking) water in rats and metabolically competent human cells

In the present study, DNA damaging and mutagenic effects of chlorinated drinking water (CDW) extracts obtained from polluted raw water resources were examined in metabolically competent human Hep G2 hepatoma cells using the in vitro micronucleus assay and the single cell gel electrophoresis (SCGE, comet assay). Additionally, the in vivo induction of micronuclei (MN) was studied in polychromatic erythrocytes (PCEs) derived from bone marrow of CDW-treated Wistar rats. Furthermore, we examined the influence of CDW on the lipid peroxidation (LpO) in blood, liver, kidney and testicle of rats. The results demonstrated significant increases of micronucleated PCEs in the bone marrow of rats fed with relatively low CDW doses (33.3 ml/kg body weight per day). Similar effects, i.e. increases of MN frequencies, were found in Hep G2 hepatoma cells after CDW treatment (41 MN/1000 binucleated cells (BNCs) for 167 ml CDW) in comparison to the vehicle control (24 MN/1000 BNC). Additionally, DNA damages caused by CDW were observed in the comet assay. As a product of LpO, the levels of malondialdehyde (MDA) were significantly enhanced almost in all animals and organs tested after CDW treatment. In livers and serum of rats dose-dependent increases of MDA were observed. The data indicated that extracts from CDW obtained from polluted raw water were able to cause oxidative damages and to induce various biological effects in mammalian cells in vivo and in vitro, i.e. clastogenicity and/or aneugenicity, DNA strand breaks and/or alkali-labile damages. The consistency of the results among the various biological systems and endpoints led to the conclusion that the consumption of chlorinated drinking water obtained from polluted raw water may enhance the body burden with mutagenic and/or carcinogenic substances and therefore, means a potential genetic hazard for human health.

Chromosomal aberration and single cell gel electrophoresis (Comet) assay in the longitudinal risk assessment of occupational exposure to pesticides.

In recent years the use of pesticides in agriculture has been increasing steadily. At present there are more than 1000 chemicals classified as pesticides. Therefore, the widespread use of pesticides and their potential genetic hazard suggests that evaluation of their genotoxicity should be extended using the newer assays now available. In the present study chromosomal aberration analysis and the alkaline single cell gel electrophoresis (Comet) assay were used to evaluate the extent of DNA damage and DNA repair in peripheral blood lymphocytes of subjects employed in pesticide production. In order to determine possible primary genotoxic effects in workers blood samples were taken after an 8 month long period of exposure to a complex mixture of pesticides. To detect the possible occurrence of DNA repair in lymphocytes of the same subjects the second blood sample was taken after an 8 month long period of absence from the pesticide exposure zone. Regardless of the period of sampling, in the exposed group statistically significantly increased numbers of aberrant cells, chromatid and chromosome breaks, acentric fragments and dicentric chromosomes compared with the controls were found. After the workers had spent 8 months out of the pesticide exposure zone the number of aberrant cells and all types of chromatid and chromosome aberrations decreased significantly compared with sampling after the high exposure period, but it still remained significantly higher in comparison with the control group. After the period of high exposure to a mixture of pesticides statistically significantly increased levels of DNA damage in the Comet assay in terms of tail length and tail moment were found. After the workers were removed from production for 8 months both Comet assay end-points decreased significantly compared with the first sampling point, but they remained increased compared with the control.

Untargeted mutation of the maternally derived mouse hypervariable minisatellite allele in F1 mice born to irradiated spermatozoa.

Length change mutation at the Ms6hm hypervariable mouse minisatellite locus was analyzed in C57BL/6N x C3H/HeN F(1) mice and the F(1) of the reciprocal cross born to irradiated male parents. Spontaneous mutant frequencies were 8.4% and 9.8% for the paternally derived and maternally derived C3H/HeN alleles, respectively. The mutant frequencies for the paternally derived allele increased to 22% and 19% when the male parents were irradiated with 6 Gy at the postmeiotic spermatozoa stage and the spermatogonia stage, respectively. These increases in the mutant frequency were at least 10 to 100 times higher than those expected from the frequency of hits to the 3- to 4-kb allele, suggesting that the length change mutation at this minisatellite locus was not a targeted event due directly to DNA damage in the region. Further analysis demonstrated that the mutant frequency increased also at the maternally derived C3H/HeN allele to 20% when the male parents were irradiated at the spermatozoa stage. This increase in the maternal allele mutation was not observed in F(1) born to irradiated spermatogonia. The present study suggests that introduction of DNA damage by irradiated sperm triggers genomic instability in zygotes and in embryos of subsequent developmental stages, and this genomic instability induces untargeted mutation in cis at the paternally derived minisatellite allele and in trans at the maternally derived unirradiated allele. Untargeted mutation revealed in the present study defines a previously unnoticed genetic hazard to the maternally derived genome by the paternally introduced DNA damage.

Untargeted mutation of the maternally derived mouse hypervariable minisatellite allele in F1 mice born to irradiated spermatozoa

Length change mutation at the Ms6hm hypervariable mouse minisatellite locus was analyzed in C57BL/6N x C3H/HeN F(1) mice and the F(1) of the reciprocal cross born to irradiated male parents. Spontaneous mutant frequencies were 8.4% and 9.8% for the paternally derived and maternally derived C3H/HeN alleles, respectively. The mutant frequencies for the paternally derived allele increased to 22% and 19% when the male parents were irradiated with 6 Gy at the postmeiotic spermatozoa stage and the spermatogonia stage, respectively. These increases in the mutant frequency were at least 10 to 100 times higher than those expected from the frequency of hits to the 3- to 4-kb allele, suggesting that the length change mutation at this minisatellite locus was not a targeted event due directly to DNA damage in the region. Further analysis demonstrated that the mutant frequency increased also at the maternally derived C3H/HeN allele to 20% when the male parents were irradiated at the spermatozoa stage. This increase in the maternal allele mutation was not observed in F(1) born to irradiated spermatogonia. The present study suggests that introduction of DNA damage by irradiated sperm triggers genomic instability in zygotes and in embryos of subsequent developmental stages, and this genomic instability induces untargeted mutation in cis at the paternally derived minisatellite allele and in trans at the maternally derived unirradiated allele. Untargeted mutation revealed in the present study defines a previously unnoticed genetic hazard to the maternally derived genome by the paternally introduced DNA damage.

Radiation-induced chromosome aberrations in guinea-pig growing oocytes, and their relation to follicular atresia

The female guinea-pig has been shown to represent a good model to investigate the genetic hazard of ionizing radiation in humans. The sensitivity of the guinea-pig oocytes to radiation-induced chromosome aberrations was, therefore, studied at different stages of oocyte and follicular growth. The sensitivity of oocytes enclosed in small follicles (15 weeks before ovulation) was found to be low and comparable to that of immature oocytes present at birth. The sensitivity of growing oocytes remained low and almost constant until 3 weeks before ovulation, from which time it began to increase. The most dramatic increase of sensitivity occurred during the last week preceding ovulation: about 90% of oocytes X-irradiated with 4 Gy, 2 days before ovulation showed one or more chromatid interchanges, as compared to 20% for those irradiated with the same dose 1 week earlier. A comparison of our results with those found by others in the mouse shows that considerable differences of sensitivity exist between oocytes of these two species irradiated at similar stages of development. The possible reasons for these differences are discussed.

Follow-up study of the genetic damage in lymphocytes of pharmacists and nurses handling antineoplastic drugs evaluated by cytokinesis-block micronuclei analysis and single cell gel electrophoresis assay

A follow-up study was carried out 4 years after an initial evaluation of the micronucleus frequency in 10 healthy individuals who had been occupationally exposed to antineoplastic drugs in a Brazilian hospital. Upon the first evaluation, these 10 exposed individuals were compared with 10 non-exposed individuals matched for age, sex and smoking habits; the results revealed that the frequency of micronucleated lymphocytes in individuals exposed to antineoplastic drugs was significantly higher ( P=0.038) than in controls. The frequency of dicentric bridges was also increased, although not significantly ( P=0.0545). After the first analysis, the workers handling antineoplastic drugs were advised to modify their work schedule to limit exposure, and the number of workers in the group was increased from 10 to 12 individuals. In the follow-up study, 12 individuals from the same work area were assessed. In addition to micronucleus frequency, alkaline single cell gel electrophoresis was also used to monitor genetic hazard. This exposed group was compared to 12 non-exposed workers from the same hospital, matched for age, sex and smoking habits. In the follow-up study, no statistical difference was found between exposed workers and controls in terms of micronucleus and dicentric bridge frequency with the Mann–Whitney U-test ( P=0.129 and 0.373, respectively). However, the mean value of SCGE analysis was significantly higher in the exposed group than in the controls ( P=0.0006). Although the micronucleus analysis seems to be less sensitive to assess DNA damage, it detects chromosome aberrations and not just repairable DNA breakage and alkali-labile sites. Combination of the alkaline single cell gel electrophoresis and cytokinesis blocked micronucleus assay appears to be commendable to monitor populations chronically exposed to genotoxic agents.

Spermatogenesis and mutagenicity of environmental hazards: extrapolation of genetic risk from mouse to man

To perform germ cell mutagenicity studies it is mandatory to know the duration of the different stages of spermatogenesis. The timing of male germ cell development determines the test protocols. Chemical mutagens are characterized by their differential spermatogenic responses, e.g. different chemicals induce mutations in different germ cell stages. Knowledge of the sensitive germ cell stages for a test agent is essential for the evaluation of the genetic hazard, i.e. stem cell effects present permanent genetic hazards and post-stem cell effects present transient hazards. A variety of assays are available to determine germ cell mutagenicity in treated animals or in the progeny of treated animals. Germ cell cytogenetics in differentiating spermatogonia and the dominant lethal assay are used for genetic hazard identification. Their results allow categorization of chemicals as germ cell mutagens (Maximale Arbeitsplatz Konzentration categories for germ cell mutagens). Gene mutations or reciprocal chromosome translocations induced in germ cells are assessed by observation of mutant offspring of treated males. These results are applicable to the quantification of genetic hazards for chemical exposures which cannot be avoided, i.e. for occupational exposures to chemicals such as butadiene.

Genetic engineering of crops as potential source of genetic hazard in the human diet

The benefits of genetic engineering of crop plants to improve the reliability and quality of the world food supply have been contrasted with public concerns raised about the food safety of the resulting products. Debates have concentrated on the possible unforeseen risks associated with the accumulation of new metabolites in crop plants that may contribute to toxins, allergens and genetic hazards in the human diet. This review examines the various molecular and biochemical mechanisms by which new hazards may appear in foods as a direct consequence of genetic engineering in crop plants. Such hazards may arise from the expression products of the inserted genes, secondary or pleiotropic effects of transgene expression, and random insertional mutagenic effects resulting from transgene integration into plant genomes. However, when traditional plant breeding is evaluated in the same context, these mechanisms are no different from those that have been widely accepted from the past use of new cultivars in agriculture. The risks associated with the introduction of new genes via genetic engineering must be considered alongside the common breeding practice of introgressing large fragments of chromatin from related wild species into crop cultivars. The large proportion of such introgressed DNA involves genes of unknown function linked to the trait of interest such as pest or disease resistance. In this context, the potential risks of introducing new food hazards from the applications of genetic engineering are no different from the risks that might be anticipated from genetic manipulation of crops via traditional breeding. In many respects, the precise manner in which genetic engineering can control the nature and expression of the transferred DNA offers greater confidence for producing the desired outcome compared with traditional breeding.

Induction of micronuclei in vitro by organochlorine compounds in beluga whale skin fibroblasts

Beluga whales ( Delphinapterus leucas) inhabiting the St. Lawrence estuary are highly contaminated with environmental pollutants and have a high incidence of cancer. Environmental contaminants may be partly responsible for the high cancer incidence observed in this population. DNA damage plays an important role in the development of cancer. The micronuclei (MN) assay was used to test the genotoxic potential of organochlorine (OC) pesticides with and without external metabolic factor in skin fibroblasts of an Arctic beluga whale. Toxaphene, chlordane and p, p′-DDT induced significant ( p<0.05) concentration-response increases of micronucleated cells (MNCs). Statistically significant increases in MNCs, ranging from 1.7- to 5-folds when compared to control cultures, were observed for 0.05, 0.5, 5 and 10 μg/ml toxaphene, 2, 5 and 10 μg/ml chlordane and 10 and 15 μg/ml p, p′-DDT. Presence of exogeneous metabolic factor (S9) completely abolished the MN induction potency of chlordane and p, p′-DDT, and toxaphene induced MN formation at higher concentrations (0.5 μg/ml) than without S9 mix. The ecotoxicological significance of MN induction by low concentrations of toxaphene is unknown and do not imply that toxaphene is involved in the etiology of cancer in St. Lawrence beluga whales. However, because of the known genotoxicity of toxaphene and the long lifespan of beluga whales, it cannot be excluded that toxaphene may pose a long-term genetic hazard to the more contaminated whales of this population.

12 Nondisjunction in the Human Male

This chapter describes the current understanding of human male nondisjunction—considering studies aimed at determining the incidence and stage of origin of nondisjunction and describing recent information on etiological factors associated with the male nondisjunctional process. Aneuploidy is the most common class of chromosome abnormality in humans and, according to some, the most important genetic hazard facing man. Most aneuploidy derives from errors in maternal meiosis, and accordingly, most nondisjunction research is directed at understanding abnormalities in oogenesis. In addition to its clinical importance, male meiosis has several features that make studies of male nondisjunction an attractive alternative to studies of female nondisjunction. These include the ability to collect products of meiosis without resorting to invasive procedures, the availability of a virtually limitless supply of gametic products, and the ability to study nondisjunction free from the confounding effects of increasing maternal age. For these reasons, several investigators have suggested that analysis of male nondisjunction may serve as a useful model for several aspects of human nondisjunction.

Evaluation of the genetic toxicity of synthetic chemicals (II), a pyrethroid insecticide, fenpropathrin

The detection of many synthetic chemicals used in industry that may pose a genetic hazard in our environment is subject of great concern at present. In this respect, the genetic toxicity of fenpropathrin ((RS)-α-cyano-3-phenoxybenzyl-2,2,3,3-tetramethyl cyclopropane carboxylate, CAS No.: 39515-41-8), a pyrethroid insecticide, was evaluated in bacterial gene mutation system, chromosome aberration in mammalian cell system andin vivo micronucleus assay with rodents. In bacterial gene mutation assay, no mutagenicity of fenpropathrin (62–5000 μg/plate) was observed inSalmonella typhimurium TA 98, 100, 1535 and 1537 both in the absence and in the presence of S-9 metabolic activation system. In mammalian cell system using chinese hamster lung fibroblast, no clastogenicity of fenpropathrin was also observed both in the absence andin the presence of metabolic activation system in the concentration range of 7–28 μg/ml. And also,in vivo micronucleus assay using mouse bone marrow cells, fenpropathrin also revealed no mutagenic potential in the dose range of 27–105 mg/kg body weight of fenpropathrin (i.p.). Consequently, no mutagenic potential of fenpropathrin was observedin vitro bacterial, mammalian mutagenicity systems andin vivo micronucleus assay in the dose ranges used in this experiment.

The Comet assay for the evaluation of the genetic hazard of pollutants in cetaceans: Preliminary results on the genotoxic effects of methyl-mercury on the bottle-nosed dolphin ( Tursiops truncatus) lymphocytes in vitro

In this study the single cell microgel electrophoresis (Comet assay) was used to evaluate the genetic effects of methyl-mercury in dolphin ( Tursiops truncatus) lymphocytes in the dose range 1–8 μg ml −1. The results of in vitro exposure showed that methyl-Hg induces DNA single-strand breaks and cytotoxicity in a dose-dependent manner. Dolphin lymphocytes have greater resistance to the genotoxic and cytotoxic effects of methyl-Hg than human and rat cells. The ecological significance of these results are discussed.

Assessment of the potential in vivo genotoxicity of fluoranthene.

Fluoranthene is a ubiquitous environmental pollutant. Although fluoranthene is mutagenic in bacterial and mammalian in vitro cell systems following metabolic activation by rat liver fraction, information on in vivo mutagenicity is lacking and studies on tumour initiating activity in mice are equivocal. In the present study, the potential genetic hazard to man was assessed using the mouse bone marrow micronucleus and rat liver unscheduled DNA synthesis test systems. Fluoranthene did not show any evidence of genotoxicity in either of the in vivo assays following acute oral administration at levels of up to 2000 mg/kg b.w.

A Framework for Assessing Genetic Vulnerability

Abstract Genetic risk assessment is a tool for describing and communicating genetic risks and hazards of natural resource management objectively, on a case-by-case basis, to reduce conflicts and ambiguities that arise from generalized perceptions of such risks. Here, we describe a general framework and some technical approaches to quantifying genetic risks and hazards. We develop a model of genetic vulnerability of managed populations that links sources of potential technological hazards, protective mechanisms and responses, and potential losses using artificial propagation of Pacific salmon in the Columbia River as an example. Unlike other models of ecological risk assessment that emphasize containment of hazards, risk assessment must consider both the protective measures against genetic hazards (reliability) and the potential rate of recovery from failure of protective measures (resilience). We illustrate the essential components of these safeguards, and we illustrate how genetic vulnerability can be de...

An introduction and study design

Under the sponsorship of the International Programme on Chemical Safety (IPCS), 17 laboratories from diverse regions of the world participated in evaluating the utility of four plant bioassays for detecting genetic hazards of environmental chemicals. The bioassays included in this collaborative study were: Arabidopsis thaliana embryo and chlorophyll assay and Tradescantia stamen hair assay, Tradescantia paludosa micronucleus assay and Vicia Faba root tip assay. Four to six laboratories participated in the performance of each of the bioassays. All laboratories participating in a particular bioassay were supplied with uniform plant material as well as standardized protocol. Five direct acting water soluble test chemicals, i.e. maleic hydrazide, methyl nitrosourea, ethyl methanesulfonate, sodium azide and azidoglycerol, were selected for this study. The study was designed to be completed in three phases. Ethyl methanesulfonate was used as a positive control and has already been reported earlier (Sandhu et al., 1991). The data from the remaining four chemicals used for the evaluation of four plant test systems in the first phase of the collaborative study are reported in this issue.