Pentachlorophenol

Pentachlorophenol (PCP) is a substance whose widespread use, mainly in wood protection and pulp and paper mills, has led to a substantial environmental contamination. This in turn accounts for a significant exposure of the general human population, with rather high exposure levels being attained in occupational settings.Investigations on the genotoxic activity of PCP have given rise to divergent results which would seem to make an evaluation difficult. By grouping them into 3 categories a somewhat clearer picture, allowing finally an (admittedly tentative) assessment, can be obtained.PCP does seem to be at most a weak inducer of DNA damage: it produces neither DNA-strand breaks nor clear differential toxicity to bacteria in rec-assays in the absence of metabolic activation. Also in SCE induction no increase can be observed in vivo, while PCP is found marginally active in a single in vitro experiment. Metabolic activation, however, leads to prophage induction and to DNA strand breaks in. human lymphocytes, presumably through the formation of oxygen radicals. A possible further exception in this area might be the positive results in the yeast recombination tests, although their inadequate reporting makes a full evaluation difficult.PCP does not seem to induce gene (point) mutations, as most bacterial assays, the Drosophila sex-linked recessive lethal test and in vitro assays with mammalian cells did not demonstrate any effects. Marginally positive results were obtained in the mammalian spot test in vivo and in one bacterial test; the positive result in the yeast assay for cycloheximide resistance is fraught somewhat with its questionable genetic basis.PCP does, however, induce chromosomal aberrations in mammalian cells in vitro and in lymphocytes of exposed persons in vivo. Those in vivo results that were unable to provide evidence of chromosomal damage are hampered either by methodological inadequacies or by too low exposure levels.The (rodent) metabolite tetrachlorohydroquinone might be a real genotoxic agent, capable of binding to DNA and producing DNA strand breaks; this activity is probably due to semiquinone radical formation and partly mediated through active oxygen species. Since this compound has not been tested in the common bacterial and mammalian mutagenicity assays, the few ancillary results on this substance cannot be used in a meaningful human risk assessment of PCP. Furthermore, this metabolite has only been produced by human liver microsomes in vitro, but has not been detected in exposed humans in vivo. Its formation in mutagenicity test systems and its activity involving radicals might, however, help to explain some of the divergencies in the genotoxicity results.The review concludes that PCP is a weak human clastogen which may lack other genotoxic properties, although it may add somewhat to the normal oxidative damage.     

Vanadate induces DNA strand breaks in cultured human fibroblasts at doses relevant to occupational exposure

To study possible genotoxic effects of occupational exposure to vanadium pentoxide, we determined DNA strand breaks (with alkaline comet assay), 8-hydroxy-2'deoxyguanosine (8-OHdG) and the frequency of sister chromatid exchange (SCE) in whole blood leukocytes or lymphocytes of 49 male workers employed in a vanadium factory in comparison to 12 non-exposed controls. In addition, vanadate has been tested in vitro to induce DNA strand breaks in whole blood cells, isolated lymphocytes and cultured human fibroblasts of healthy donors at concentrations comparable to the observed levels of vanadium in vivo. To investigate the impact of vanadate on the repair of damaged DNA, co-exposure to UV or bleomycin was used in fibroblasts, and DNA migration in the alkaline and neutral comet assay was determined. Although, exposed workers showed a significant vanadium uptake (serum: median 5.38microg/l, range 2.18-46.35microg/l) no increase in cytogenetic effects or oxidative DNA damage in leukocytes could be demonstrated. This was consistent with the observation that in vitro exposure of whole blood leukocytes and lymphocytes to vanadate caused no significant changes in DNA strand breaks below concentrations of 1microM (50microg/l). In contrast, vanadate clearly induced DNA fragmentation in cultured fibroblasts at relevant concentrations. Combined exposure of fibroblasts to vanadate/UV or vanadate/bleomycin resulted in non-repairable DNA double strand breaks (DSBs) as seen in the neutral comet assay. We conclude that exposure of human fibroblasts to vanadate effectively causes DNA strand breaks, and co-exposure of cells to other genotoxic agents may result in persistent DNA damage.     

Growth inhibition and mutagenesis induced in Escherichia coli by dihydropyrazines with DNA strand-cleaving activity

Dihydropyrazine (DHP) causes DNA strand breaks in vitro. We evaluated the cytotoxic and genotoxic potential of DHP in Escherichia coli. DHP exposure dose-dependently caused inhibition of cell growth in the wild-type strain, death in recA and uvrB, and an increase in mutation frequency in uvrB. These findings indicate that DHP causes DNA strand breaks in vivo.     

Comparative in vitro and in vivo assessment of genotoxic effects of etoposide and chlorothalonil by the comet assay.

The alkaline single cell gel electrophoresis (comet) assay was used to assess in vitro and in vivo genotoxicity of etoposide, a topoisomerase II inhibitor known to induce DNA strand breaks, and chlorothalonil, a fungicide widely used in agriculture. For in vivo studies, rats were sacrificed at various times after treatment and the induction of DNA strand breaks was assessed in whole blood, bone marrow, thymus, liver, kidney cortex and in the distal part of the intestine. One hour after injection, etoposide induced DNA damage in all organs studied except kidney, especially in bone marrow, thymus (presence of HDC) and whole blood. As observed during in vitro comet assay on Chinese hamster ovary (CHO) cells, dose- and time-dependent DNA effects occurred in vivo with a complete disappearance of damage 24 h after administration. Even though apoptotic cells were detected in vitro 48 h after cell exposure to etoposide, such a result was not found in vivo. After chlorothalonil treatment, no DNA strand breaks were observed in rat organs whereas a clear dose-related DNA damage was observed in vitro. The discrepancy between in vivo and in vitro models could be explained by metabolic and mechanistic reasons. Our results show that the in vivo comet assay is able to detect the target organs of etoposide and suggest that chlorothalonil is devoid of appreciable in vivo genotoxic activity under the protocol used.     

Review of the genotoxicity of styrene in humans

Styrene (CAS No. 100-42-5) is an important industrial chemical for which positive results have been reported in in vitro and in vivo genotoxicity assays. Styrene-exposed workers have been studied extensively over two decades for the induction of various types of genotoxic effects. The outcomes of these studies have been conflicting, and where positive responses have been reported, it has proved difficult to demonstrate clear relationships between levels of damage reported and exposure levels. In this review, we have assessed studies addressing mutagenicity (chromosome aberrations, micronuclei and gene mutations) and other endpoints (sister chromatid exchanges, DNA breaks and DNA adducts) using criteria derived from the IPCS guidelines for the conduct of human biomonitoring studies. Based on the re-evaluated outcomes, the data are not convincing that styrene induces gene mutations. The evidence for induction of clastogenicity in occupationally exposed workers is less clear, with a predominant lack of induction of micronuclei in different studies, but conflicting responses in chromosome aberration assays. The results of numerous studies on sister chromatid exchanges do not provide evidence of a clear positive response, despite these being induced in animals exposed to styrene at high concentrations. However, there is evidence that both DNA adducts and DNA single strand breaks are induced in styrene workers. These types of damage are considered indicative of exposure of the target cells and interaction with cellular DNA but do not necessarily result in heritable changes. There is evidence that the metabolism of styrene in humans is affected by genetic polymorphisms of metabolizing genes and that these polymorphisms affect the outcome of in vitro mutagenicity studies on styrene. Therefore, studies that have addressed the potential of this factor to affect in vivo responses were considered. To date, there are no consistent relationships between genetic polymorphisms and induction of genotoxicity by styrene in humans, but further work is warranted on larger samples. The analyses of individual studies, together with a consideration of dose-response relationships and the lack of a common profile of positive responses for the various endpoints in different studies, provide no clear evidence that styrene exposure in workers results in detectable levels of mutagenic damage. However, evidence of exposure to genotoxic metabolites is demonstrated by the formation of DNA adducts and strand breaks.     

A mutagenicity assessment of acetaldehyde

Acetaldehyde has been shown in studies by several different laboratories to be a clastogen (chromosome-breaking) and inducer of sister-chromatid exchanges in cultured mammalian cells (Chinese hamster cells and human lymphocytes). Although there have been very few studies in intact mammals, the available evidence suggests that acetaldehyde produces similar cytogenetic effects in vivo. The production of cytogenetic abnormalities may be related to the ability of acetaldehyde to form DNA-DNA and/or DNA-protein cross-links. Acetaldehyde apparently has not been evaluated for its ability to cause gene mutations in cultured mammalian cells, but it has been shown to produce sex-linked recessive lethals in Drosophila. In general, bacteria tests have been negative. Although acetaldehyde is a genotoxic cross-linking agent, it does not appear to cause DNA strand breaks. There were no studies available regarding the potential of acetaldehyde to produce genetic damage in mammalian germ cells in vivo. Most mutagenicity testing on acetaldehyde has been motivated by attempts to define the proximate mutagen in ethanol metabolism.     

Camptothecin induces protein-linked DNA breaks via mammalian DNA topoisomerase I

Camptothecin, a cytotoxic drug, is a strong inhibitor of nucleic acid synthesis in mammalian cells and a potent inducer of strand breaks in chromosomal DNA. Neither the equilibrium dialysis nor the unwinding measurement indicates any interaction between camptothecin and purified DNA. However, camptothecin induces extensive single strand DNA breaks in reactions containing purified mammalian DNA topoisomerase I. DNA breakage in vitro is immediate and reversible. Analyses of camptothecin-induced DNA breaks show that topoisomerase I is covalently linked to the 3' end of the broken DNA. In addition, camptothecin inhibits the catalytic activity of mammalian DNA topoisomerase I. We propose that camptothecin blocks the rejoining step of the breakage-reunion reaction of mammalian DNA topoisomerase I. This blockage results in the accumulation of a cleavable complex which resembles the transient intermediate proposed for eukaryotic DNA topoisomerase I. The inhibition of nucleic acid synthesis and the induction of DNA strand breaks observed in vivo may be related to the formation of this drug-induced cleavable complex.     

Inflammatory and genotoxic effects of diesel particles in vitro and in vivo

Recent studies have identified an indirect genotoxicity pathway involving inflammation as one of the mechanisms underlying the carcinogenic effects of air pollution/diesel exhaust particles (DEP). We investigated the short-term effects of DEP on markers of inflammation and genotoxicity in vitro and in vivo. DEP induced an increase in the mRNA level of pro-inflammatory cytokines and a higher level of DNA strand breaks in the human lung epithelial cell line A549 in vitro. For the in vivo study, mice were exposed by inhalation to 20 or 80 mg/m3 DEP either as a single 90-min exposure or as four repeated 90-min exposures (5 or 20 mg/m3) and the effects in broncho-alveolar lavage (BAL) cells and/or lung tissue were characterized. Inhalation of DEP induced a dose-dependent inflammatory response with infiltration of macrophages and neutrophils and elevated gene expression of IL-6 in the lungs of mice. The inflammatory response was accompanied by DNA strand breaks in BAL cells and oxidative DNA damage and increased levels of bulky DNA adducts in lung tissue, the latter indicative of direct genotoxicity. The effect of a large single dose of DEP was more pronounced and sustained on IL-6 expression and oxidative DNA damage in the lung tissue than the effect of the same dose administered over four days, whereas the reverse pattern was seen in BAL cells. Our results suggest that the effects of DEP depend on the rate of delivery of the particle dose. The mutation frequency (MF), after DEP exposure, was determined using the transgenic Muta Mouse and a similar exposure regimen. No increase was observed in MF in lung tissue 28-days after exposure. In conclusion, short-term exposure to DEP resulted in DNA strand breaks in BAL cells, oxidative DNA damage and DNA adducts in lungs; and suggested that DNA damage in part is a consequence of inflammatory processes. The response was not associated with increased MF, indicating that the host defence mechanisms were sufficient to counteract the adverse effects of inflammation. Thus, there may be thresholds for the inflammation-associated genotoxic effects of DEP inhalation.     

Quantitation of chemically induced DNA strand breaks in human cells via an alkaline unwinding assay

DNA strand breaks induced in human CCRF-CEM cells by electrophilic chemicals (carcinogens/mutagens) can be readily quantitated via a facile alkaline unwinding assay. This procedure estimates the number of chemically induced DNA strand breaks on the basis of the percentage DNA converted from double-stranded to single-stranded form during an exposure to the alkaline unwinding conditions. The assay is based on the assumption that each strand break serves as a strand unwinding point during the alkaline denaturation. The extent of strand separation can be standardized with respect to the initial level of induced strand breaks by the use of X-rays, which produce known levels of DNA strand breaks per rad in mammalian cells. Subsequent to the alkaline exposure, the single- and double-stranded DNA were separated by use of thermostated hydroxylapatite columns (60 degrees C), and the DNA was quantitated via a fluorescence assay (Hoechst 33258 compound). A correlation was shown between mammalian DNA strand-breaking potential (as measured in this procedure) and the propensity of these chemicals to revert Salmonella typhimurium TA100.     

Ionizing radiation sensitivity of DNA polymerase lambda-deficient cells

Ionizing radiation induces a diverse spectrum of DNA lesions, including strand breaks and oxidized bases. In mammalian cells, ionizing radiation-induced lesions are targets of non-homologous end joining, homologous recombination, and base excision repair. In vitro assays show a potential involvement of DNA polymerase lambda in non-homologous end joining and base excision repair. In this study, we investigated whether DNA polymerase lambda played a significant role in determining ionizing radiation sensitivity. Despite increased sensitivity to hydrogen peroxide, lambda-deficient mouse embryonic fibroblasts displayed equal survival after exposure to ionizing radiation compared to their wild-type counterparts. In addition, we found increased sensitivity to the topoisomerase inhibitors camptothecin and etoposide in the absence of polymerase lambda. These results do not reveal a major role for DNA polymerase lambda in determining radiosensitivity in vivo.     

Detection of DNA strand breaks in Escherichia coli treated with platinum(IV) antitumor compounds

DNA strand breaks were observed in bacteria treated with Pt(IV) but not Pt(II) antitumor compounds by two methods. First, compounds which cause DNA strand breaks produced an SOS induction signal which was detected by a rapid bacterial assay. In addition, the capacity of these compounds to cut DNA in vivo was directly measured by agarose gel electrophoresis of pBR322 DNA extracted from bacteria treated with these drugs. cis-Diamminetetrachloroplatinum(IV) (cis-DTP) and cis-dichloro-trans-dihydroxo-cis-bis(isopropylamine)-platinum(IV) (iproplatin) produced strand breaks in both assays while cis-diamminedichloroplatinum(II) (cisplatin) did not. These results indicate that Pt(IV) antitumor complexes may cause DNA damage in vivo which is not produced by Pt(II) compounds.     

Genotoxicity of environmental tobacco smoke: a review

Environmental tobacco smoke (ETS), or second-hand smoke, is a widespread contaminant of indoor air in environments where smoking is not prohibited. It is a significant source of exposure to a large number of substances known to be hazardous to human health. Numerous expert panels have concluded that there is sufficient evidence to classify involuntary smoking (or passive smoking) as carcinogenic to humans. According to the recent evaluation by the International Agency for Research on Cancer, involuntary smoking causes lung cancer in never-smokers with an excess risk in the order of 20% for women and 30% for men. The present paper reviews studies on genotoxicity and related endpoints carried out on ETS since the mid-1980s. The evidence from in vitro studies demonstrates induction of DNA strand breaks, formation of DNA adducts, mutagenicity in bacterial assays and cytogenetic effects. In vivo experiments in rodents have shown that exposure to tobacco smoke, whole-body exposure to mainstream smoke (MS), sidestream smoke (SS), or their mixture, causes DNA single strand breaks, aromatic adducts and oxidative damage to DNA, chromosome aberrations and micronuclei. Genotoxicity of transplacental exposure to ETS has also been reported. Review of human biomarker studies conducted among non-smokers with involuntary exposure to tobacco smoke indicates presence of DNA adducts, urinary metabolites of carcinogens, urinary mutagenicity, SCEs and hypoxanthine-guanine phosphoribosyltransferase (HPRT) gene mutations (in newborns exposed through involuntary smoking of the mother). Studies on human lung cancer from smokers and never-smokers involuntarily exposed to tobacco smoke suggest occurrence of similar kinds of genetic alterations in both groups. In conclusion, these overwhelming data are compatible with the current knowledge on the mechanisms of carcinogenesis of tobacco-related cancers, occurring not only in smokers but with a high biological plausibility also in involuntary smokers.     

A comparison of calcium phosphate coprecipitation and electroporation

Plasmid-based transfection assays provide a rapid means to measure homologous and nonhomologous recombination in mammalian cells. Often it is of interest to examine the stimulation of recombination by DNA damage induced by radiation, genotoxic chemicals, or nucleases. Transfection is frequently performed by using calcium phosphate coprecipitation (CPP), because this method is well suited for handling large sample sets, and it does not require expensive reagents or equipment. Alternative transfection methods include lipofection, microinjection, and electroporation. Since DNA strand breaks are known to stimulate both homologous and nonhomologous recombination, the induction of nonspecific damage during transfection would increase background recombination levels and thereby reduce the sensitivity of assays designed to detect the stimulation of recombination by experimentally induced DNA damage. In this article, we compare the stimulatory effects of nuclease-induced double-strand breaks (DSBs) on homologous and nonhomologous recombination for molecules transfected by CPP and by electroporation. Although electroporation yielded fewer transfectants, both nonhomologous and homologous recombination were stimulated by nuclease-induced DSBs to a greater degree than with CPP. Ionizing radiation is an effective agent for inducing DNA strand breaks, but previous studies using CPP generally showed little or no stimulation of homologous recombination among plasmids damaged with ionizing radiation. By contrast, we found clear dose-dependent enhancement of recombination with irradiated plasmids transfected using electroporation. Thus, electroporation provides a higher signal-to-noise ratio for transfection-based studies of damage-induced recombination, possibly reflecting less nonspecific damage to plasmid DNA during transfection of mammalian cells.     

Evaluation of the genotoxicity induced by the fungicide fenarimol in mammalian and plant cells by use of the single-cell gel electrophoresis assay

Fenarimol, a systemic pyrimidine carbinol fungicide, is considered to be not genotoxic or weakly genotoxic, although the available toxicological data are controversial and incomplete. Our results obtained in vitro with leukocytes of two different rodent species (rat and mouse) show that fenarimol affects DNA, as detected by the single-cell gel electrophoresis (SCGE, Comet) assay. This fungicide is able to induce DNA damage in a dose-related manner, with significant effectiveness at 36 nM, but without significant interspecies differences. Simultaneous exposure of rat leukocytes to fenarimol (36-290 nM) and a model genotoxic compound (50 microg/ml bleomycin) produced a supra-additive cytotoxic and genotoxic effect. This supports previous findings suggesting possible co-toxic, co-mutagenic, cancer-promoting and co-carcinogenic potential of fenarimol, and modification of the effects of other xenobiotics found to be influenced by this agrotoxic chemical, with consequent different toxicological events. The potential for DNA strand breaks to act as a biomarker of genetic toxicity in plants in vivo was also considered, in view of the fact that higher plants represent reliable sensors in an ecosystem. Significant DNA breakage was observed in the nuclei of Impatiens balsamina leaves after in vivo treatment with fenarimol (145 nM, 1h). More than 50% of the cells showed such DNA damage.     

Inhibition of replicon initiation in human cells following stabilization of topoisomerase-DNA cleavable complexes.

Diploid human fibroblast strains were treated for 10 min with inhibitors of type I and type II DNA topoisomerases, and after removal of the inhibitors, the rate of initiation of DNA synthesis at replicon origins was determined. By alkaline elution chromatography, 4'-(9-acridinylamino)methanesulfon-m-anisidide (amsacrine), an inhibitor of DNA topoisomerase II, was shown to produce DNA strand breaks. These strand breaks are thought to reflect drug-induced stabilization of topoisomerase-DNA cleavable complexes. Removal of the drug led to a rapid resealing of the strand breaks by dissociation of the complexes. Velocity sedimentation analysis was used to quantify the effects of amsacrine treatment on DNA replication. It was demonstrated that transient exposure to low concentrations of amsacrine inhibited replicon initiation but did not substantially affect DNA chainelongation within operating replicons. Maximal inhibition of replicon initiation occurred 20 to 30 min after drug treatment, and the initiation rate recovered 30 to 90 min later. Ataxia telangiectasia cells displayed normal levels of amsacrine-induced DNA strand breaks during stabilization of cleavable complexes but failed to downregulate replicon initiation after exposure to the topoisomerase inhibitor. Thus, inhibition of replicon initiation in response to DNA damage appears to be an active process which requires a gene product which is defective or missing in ataxia telangiectasia cells. In normal human fibroblasts, the inhibition of DNA topoisomerase I by camptothecin produced reversible DNA strand breaks. Transient exposure to this drug also inhibited replicon initiation. These results suggest that the cellular response pathway which downregulates replicon initiation following genotoxic damage may respond to perturbations of chromatin structure which accompany stabilization of topoisomerase-DNA cleavable complexes.     

Illumination of human keratinocytes in the presence of the sunscreen ingredient Padimate-O and through an SPF-15 sunscreen reduces direct photodamage to DNA but increases strand breaks.

On illumination with simulated sunlight, the UVB-absorbing sunscreen chemical 2-ethylhexyl-4-dimethylaminobenzoate (Padimate-O) generates excited species which inflict non-ligatable strand breaks on DNA in vitro and it also becomes mutagenic to yeast in vivo. Padimate-O is known to penetrate human skin but its effects on human cells are not clear. Here, we first simulate the sunlight which penetrates human skin and use it to illuminate human keratinocytes. The DNA damage observed in terms of UV-endonuclease-sensitive sites (ESS) and direct strand breaks per kilobase (kb) of DNA per joule per square metre agrees well with that predicted from action spectra based on monochromatic light. Using plasmid DNA in vitro, we find a very similar pattern of results. Next, we simulate the spectrum that results when the incident light is first attenuated by a film of sunscreen (SPF-15; 2 mg/cm(2)) containing benzophenone-3 (a UVA absorber), octyl methoxycinnamate (a UVB absorber), and Padimate-O. If the sunscreen is not in contact with keratinocytes it reduces direct DNA damage from sunlight (ESS). However, any Padimate-O in contact with the cells substantially increases indirect damage (strand breaks) even though the film of sunscreen reduces direct photodamage. We estimate that applying an SPF-15 sunscreen which contains Padimate-O to human skin followed by exposure to only 5 minimum erythemal doses (MED) of sunlight could, while suppressing the formation of ESS, increase strand breaks in cells under the epidermis by at least 75-fold compared to exposure to 1 MED in the absence of sunscreen.     

Genetic toxicity of dopamine

The genetic toxicity of dopamine was studied in a battery of test systems including DNA single-strand break analysis in cultured human skin fibroblasts, the Salmonella/mammalian-microsome mutagenicity test, sister-chromatid exchange analysis in human lymphocytes, the mouse-lymphoma forward mutation assay, the sex-linked recessive lethal test in Drosophila melanogaster and the micronucleus test in mouse and rat. Dopamine at concentrations of 50-300 micrograms/ml induced DNA strand breaks in human fibroblasts. It also gave a positive response in the mouse-lymphoma forward mutation assay, where a dose-dependent increase in the frequency of mutant cells was observed in the presence of dopamine, 94-750 micrograms/ml. All other tests showed no response to dopamine. The dopamine-induced DNA strand breaks in human fibroblasts were inhibited by superoxide dismutase or dithiothreitol. Furthermore, dopamine caused nicking of circular Col El DNA and bound to calf thymus DNA in vitro. It is suggested that this genetic activity of dopamine in vitro relates to oxidation of dopamine and the generation of reactive oxygen radicals, semiquinones and quinones. It is unlikely that similar reactions would occur and cause genotoxic activity of dopamine in vivo.     

The effect of a mammalian repair endonuclease on x-irradiated DNA.

DNA was extracted from rat liver of non-irradiated animals, and was irradiated in vitro, and from animals which received whole body doses of X-radiation. Sedimentation on neutral and alkaline sucrose gradients as well as measurements of 32P release after sequential treatment with endonuclease and alkaline phosphatase and determination of triphosphate incorporation after the sequential treatment with endonuclease, alkaline phosphatase and DNA polymerase indicated that DNA irradiated in vivo and in vitro were effective substrates for the mammalian repair endonuclease. The experiments suggest that in addition to strand breaks, X-radiation causes base damage and they have provided a plausible explanation for the formation of double strand breaks in DNA irradiated in vivo.     

In vitro mammalian mutagenesis as a model for genetic lesions in human cancer

Recently in vitro assays of mutagenesis have been criticized as being poorly predictive of long-term in vivo rodent assays of carcinogenicity. Questions have also been raised concerning the relevance of rodent assays to human risk. In vitro assays using mammalian cells can detect most types of genetic lesions thought to be important in human malignant disease. Molecular and cytogenetic analyses of mutations induced by a variety of genotoxic compounds at the heterozygous thymidine kinase locus in mouse lymphoma cells indicate that this in vitro assay does indeed register the range of genetic lesions recently found in a wide variety of human tumors. The types and complexity of the induced lesions are reflected in mutant colony phenotype in a compound-specific fashion. These studies point to the use of appropriate in vitro mammalian mutagenesis assays as new model systems for dissecting the genetic lesions important in human carcinogenesis, and as a means of determining the potential for compounds to induce such lesions.     

Quantitative comparative mutagenesis in bacteria, mammalian cells, and animal-mediated assays A convenient way of estimating genotoxic activity in vivo?

The accumulation of environmental compounds which exhibit genotoxic properties in short-term assays and the increasing lag of time for obtaining confirmation or not in long-term animal mutagenicity and carcinogenicity tests, makes it necessary to develop alternative, rapid methodologies for estimating genotoxic activity in vivo. In the experimental approach used here, it was assumed that the genotoxic activity of foreign compounds in animals, and ultimately humans, is determined among others by exposure level, organ distribution of (DNA) dose, and genotoxic potency per unit of dose, and that knowledge about these 3 parameters may allow to rapidly determine the expected degree of genotoxicity in various organs of exposed animals. In view of the high degree of qualitative correlation between mutagenic activity of chemicals in bacteria and in cultured mammalian cells, and their mutagenic and carcinogenic properties in animals, and in order to be able to distinguish whether mutagenic potency differences were due to differences in (DNA) dose rather than other physiological factors, the results of mutagenicity tests obtained in the present experiments using bacteria and mammalian cells were compared on the basis of DNA dose rather than exposure concentrations, with the following questions in mind: Is there an absolute or a relative correlation between the mutagenic potencies of various ethylating agents in bacteria (E. coli K12) and in mammalian cells (V79 Chinese hamster) after treatment in standardized experiments, and can specific DNA adducts be made responsible for mutagenicity? Is the order of mutagenic potency of various ethylating agents observed in bacteria in vitro representative of the ranking of mutagenic potency found in vivo? Since the answer to this last question was negative, a further question addressed to was whether short-term in vivo assays could be developed for a rapid determination of the presence (and persistence) of genotoxic factors in various organs of mice treated with chemicals. In quantitative comparative mutagenesis experiments using E. coli K12 and Chinese hamster cells treated under standardized conditions in vitro with 5 ethylating agents, there was no indication of an absolute correlation between the number of induced mutants per unit of dose in the bacteria and the mammalian cells. The ranking of mutagenic potency was, however, identical in bacteria and mammalian cells, namely, ENNG greater than ENU greater than or equal to DES greater than DEN congruent to EMS, the mutagenic activity of DEN being dependent on the presence of mammalian liver preparations.(ABSTRACT TRUNCATED AT 400 WORDS)