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Although aneuploidy makes a significant contribution to both somatic and inherited disease the mechanisms by which environmental chemicals may induce numerical chromosome aberrations are only poorly defined. The European Union Project was aimed to further our understanding of those chemical interactions with the components of the mitotic and meiotic cell division cycle which may lead to aneuploidy and to characterise the parameters such as cellular metabolism which may influence the activity of aneugenic chemicals. C-mitosis can be induced by the highly lipophilic polychlorinated biphenyl and the completion of mitosis and cleavage can be modified by agents which deplete cellular levels of reduced glutathione. Modifications of the fidelity of chromosome segregation were produced by inhibiting the functioning of topoisomerase II during chromatid separation. In contrast, the modification of centromere integrity resulted in chromosome breakage as opposed to disturbance of segregation. Modifiers of tubulin assembly and centriolar functioning in somatic cells such as acrylamide, vinblastine and diazepam reproduced their activity in rodent bone marrow and male germ cells. The analysis of chromosome malsegregation in Aspergillus nidulans by a structurally related series of halogenated hydrocarbons was used to develop a QSAR model which had high predictive value for the results of fungal tests for previously untested related chemicals. Metabolic studies of potential aneugens in genetically engineered human lymphoblastoid cells demonstrated the detoxification of the aneugenic activity of chloral hydrate and the activation of 2,3-dichlorobutane, 1,1,2-trichloroethane and trichloroethylene by Phase I biotransforming enzymes. Cell transformation studies in Syrian hamster dermal cultures using a panel of 22 reference and or potential aneugens indicated that 15 of the 22 produced positive results following single exposures. Five of the aneugens which were negative following single exposures produced positive results where cultures were continuously exposed for up to 6 weeks to low concentrations following a single non-transforming exposure to the mutagen dimethyl sulphate. The transformation studies indicate that a significant proportion of chemical aneugens are potential complete carcinogens and/or co-carcinogens. To optimise the enumeration of chromosomes following exposure to potential chemical aneugens whole chromosome paints and centromere specific probes suitable for use in fluorescence in situ hybridisation (FISH) were developed for the rat, mouse and Chinese hamster and selected human probes evaluated for their suitability for routine use. Molecular chromosome probes were used to develop protocols for enumerating chromosomes in metaphase cells and centromeres and micronuclei in interphase cells. The analysis of segregation of specific centromeres in binucleate cells following cytochalasin B treatment was shown to be a potentially valuable system for characterising non-disjunction following chemical exposure. Whole chromosome paints and centromere specific probes were used to demonstrate the presence of dose-response thresholds following treatment with a reference panel of spindle inhibiting chemicals. These data indicate that the FISH technology is suitable for evaluating the relative hazards of low-dose exposures to aneugenic chemicals.

In order to investigate the chemical nature and diversity of the mutagens in ambient airborne particles (AAP) and coal fly ash (CFA), extracts of these particles were subjected to bioassay-directed fractionation. Open-column as well as high-pressure liquid chromatography were used as fractionation techniques after purification by means of liquid-liquid partition. Extracts and fractions were tested with the Salmonella /microsome test using various tester strains, among them TA98NR and analysed for polycyclic aromatic hydrocarbons (PAH) and some nitrated PAH. Seven distinct groups of mutagens could be discerned in AAP. Two of them were identified as PAH and mono-nitro PAH; the first accounted for about 5% of the total effect in TA98 with S9 and about 20% of the total effect in TA100 with S9. For the mono-nitro PAH, these percentages were about 12, 24, 14 and 13 for TA98 with and without S9 and TA100 with and without S9, respectively. A significant contribution of dinitropyrenes (possibly together with other multiply nitrated PAH) could be ruled out. Five more polar groups of mutagens could be isolated in AAP, which showed clearly nitroreductase-dependent direct mutagenicity. Besides direct-acting nitrocompounds, these groups also comprised indirect acting compounds (S9 depenent); possibly, but not necessarily, these are also nitro-compounds. The distribution of the mutagenicity over the various active chromatographical fractions was clearly strain and S9 dependent. The use of strains TA100 and TA97 pointed to the importance of indirect mutagens, also for the more polar fractions. PAH did not significantly contribute to the mutagenicity of CFA. These particles contained two groups of mutagens that were not found in AAP: a group, the effect of which is most probably caused by dinitropyrenes and comparable compounds and a polar group containing nitro-reductase dependent direct mutagens.

The Health Council of The Netherlands advises the Dutch government on scientific issues concerning health and environmental protection. In 1978 a committee of the Council established mechanistic criteria for quantitative risk assessment of carcinogenic substances. For this purpose it adopted the multistage model of carcinogenesis. Based on this model two categories of carcinogens were distinguished. The first category includes complete carcinogens and tumor initiators: these agents induce irreversible modification of DNA and act by a stochastic mechanism. The second category includes promotors and other substances acting as cocarcinogens; these agents act by nonstochastic mechanisms. Stochastic action implies that there is no threshold for the contribution to the carcinogenic effect, whereas nonstochastic action implies a threshold dose. Recently, reevaluation of the method for carcinogenic risk assessment was considered due because of remarkable scientific progress since 1978, such as the identification of cellular oncogenes and tumor suppressor genes, and the partial clarification of mechanisms at the cellular and/or molecular level by which tumor promotors stimulate tumor development. The present paper discusses the Dutch method for carcinogenic risk assessment in the light of the recent scientific developments. It concludes that this method for risk assessment is still a valid and appropriate one.

The ability of aflatoxins B1 and G1 to induce back mutations to arg+ in Escherichia coli K-12/343/113 was compared with the induction of mitotic gene conversion to ade+ in the diploid yeast strain Saccharomyces cerevisiae D4, ade-2. In analogy to previous results with other microorganisms, the compounds were not genetically active per se, indicating that under the experimental conditions employed none of the tester strains were able to activate the compounds to mutagenic products. In experiments using liver homogenates (S-9 fraction) of male Golden Syrian hamsters previously treated with phenobarbital, aflatoxin B1 exhibited strong genetic activity both in E. coli and in S. cerevisiae, whereas the mutagenic activity of aflatoxin G1 was markedly lower and could be detected only in the E. coli tester strain. These results correlate the findings that aflatoxin G1 is a less potent carcinogen and mutagen than aflatoxin B1.

The activation of nitrofurans to mutagenic intermediates by testicular tissue was investigated. AF-2 and nitrofurazone were tested in a microsomal suspension assay with strain E. coli K-12 343/113 as indicator and subcellular fractions from rabbit testes. Different mutation patterns were observed in the presence or absence of testicular homogenate, indicating the presence of different mutagenic intermediates. The frequency of arg + reversion increased proportionally to the homogenate concentration suggesting that the nitrofurans were activated by testicular components to intermediates that induced base-pair substitutions. Other experiments showed that a component of low molecular weight, present in the soluble fraction of homogenates of testes from rabbits, rats and monkeys, was responsible for the increased mutation frequency. It is concluded that this “co-mutagen-like” factor either alters the metabolism of nitrofurans in E. coli and/or promotes the formation of base-pair substitution-type mutations. This direct interaction between a non-enzymic component of mammalian testes and the mutation induction/expression process in E. coli suggests the role of co-mutagen-like factors in the sensitivity of testes to nitrofurans.