In Vitro Test Systems for Identifying Potential Chemical Carcinogens

The development of in vitro test systems for evaluating the toxic, mutagenic, and carcinogenic potential of substances has led to a reduction of animal experiments besides of being better suited for many differentiated mechanistic investigations. In a pre-screening approach, substances which exhibit the aforementioned characteristics in vitro may be excluded from further studies in vivo. On the other hand, negative results obtained with in vitro test systems cannot be extrapolated to the animal or to the human situation with sufficient certainty. Thus, if a substance has proven to be nontoxic and non-mutagenic in vitro there is still the need to set up an animal experiment. In this review we report on suborganismic systems which have proven to be useful tools for evaluating the mutagenic and carcinogenic potential of chemicals which are relevant to human health

Genotoxicity testing of nanomaterials

Nanomaterials have outstanding and unprecedented advantageous material properties but may also cause adverse effects in humans upon exposure. Testing nanomaterials for genotoxic properties is challenging because traditional testing methods were designed for small, soluble molecules and may not be easily applicable without modifications. This review critically examines available genotoxicity tests for use with nanomaterials, including DNA damage tests such as the comet assay, gene mutation tests such as the mouse lymphoma and hprt assay, and chromosome mutation tests such as the micronucleus test and the chromosome aberration test. It presents arguments for the relative usefulness of various tests, such as preferring the micronucleus test over the chromosome aberration test for scoring chromosome mutations and preferring mammalian cell gene mutation tests because the Ames test has limited utility. Finally, it points out the open questions and further needs in adapting genotoxicity tests for nanomaterials, such as validation, reference nanomaterials, and the selection of top test concentrations, as well as the relevance and applicability of test systems and the need to define testing strategies. This article is categorized under: Toxicology and Regulatory Issues in Nanomedicine > Toxicology of Nanomaterials Toxicology and Regulatory Issues in Nanomedicine > Regulatory and Policy Issues in Nanomedicin

Monooxygenase, epoxide hydrolase, and glutathione-S-transferase activities in human lung. Variation between groups of bronchogenic carcinoma and non-cancer patients and interindividual differences

Activities of microsomal monooxygenases (MO) and epoxide hydrolase (EH) and cytoplasmic glutathione-S-transferases (GST) will contribute to controlling the pool of reactive intermediates, enzymatically derived from polynuclear aromatic hydrocarbons (PAH) within the cells of target organs such as the human lung. Therefore, we studied what interindividual differences exist in these enzyme activities and whether there is a correlation between the activities of these epoxide forming and metabolizing enzymes in preparations from peripheral lung samples and the occurrence of bronchogenic carcinomas in smokers and non-smokers. 57 samples obtained from surgery were studied. Among them were 12 samples from non-smoking patients without cancer as a control group. It is not known whether this control group behaves, with respect to the investigated parameters, identically to fully healthy people, since in all cases indications existed which justified the removal of lung biopsies. Using very sensitive standard assays with benzo[a]pyrene, biphenyl, 7-ethoxyresorufin and 7-ethoxycoumarin as substrates, MO activity could only be determined as O-deethylation of 7-ethoxycoumarin and only after modification of the assay method. Evidence was obtained for the presence of a diffusible, but not dialyslble, MO Inhibitor in human lung microsomes. The MO activity (substrate: 7-ethoxycoumarin) in this fraction was extremely low in human (100-fold lower than in rat lung preparations), whereas EH (substrate: benzo[a]pyrene 4, 5-oxide) was slightly (about 2-fold) higher in human and GST (substrate: 2, 4-dinitrochlorobenzene) had similar activities in both species. Interindividual variations of enzyme activities in human lung were considerable: MO, 40-fold: EH, 5-fold; GST 10-fold. Compared to the control group (non-smokers without cancer) MO activities were slightly but significantly higher in lungs from bronchogenic carcinoma patients whether they were smokers (170% of controls, p 0.1) between the various groups studied. The substrate specificity of human lung EH, which was studied using five K-region epoxides of various PAH as substrates, corresponded to that in human and rat liver and in human, mouse and rat skin and to the pure enzyme isolated from rat liver. In contrast to rat liver hepatoma preparations, where EH had been shown to be increased in the tumor tissue and had been identified as a preneoplastic antigen, EH activity in lung microsomal preparations from samples of peripheral squamous cell carcinomas of two subjects had in the tumor tissue only one third of the activity of non-diseased areas of the same lun

Effects of sodium butyrate on DNA content, glutathione S-transferase activities, cell morphology and growth characteristics of rat liver nonparenchymal epithelial cells in vitro

The effects of sodium butyrate, which has been shown to act as a differentiation promoting agent in several different tumor cell lines, were studied in a rat liver nonparenchymal epithelial cell line. Exposure of these cells to 3.75 mM butyrate resulted in an inhibition of cell proliferation and, at the same time, an increase in cell diameter (2- to 6-fold) and size of the nuclei (∼2-fold) after 3 days in culture. Binucleated cells arose, comprising ∼12% of the cells investigated, and the number of cells with an abnormal set of chromosomes was increased. Intercellular communication, measured by dye transfer of Lucifer Yellow, was unchanged. From the various xenobiotic metabolizing enzyme activities measured, only those of glutathione S-transferases were significantly altered (increases of 4- to 9-fold) by butyrate treatment. These increases were mainly due to the predominant rise in the π class isoenzyme which is a well-known tumour marker in rat hepatocarcinogenesis. Thus, our results cannot be interpreted as being either due to promotion of differentiation or due to transformation. The state and type of cell under study has to be considered and investigations of further differentiation parameters are needed to obtain a deeper insight into the biological activity and the underlying mechanisms of cell state modifying agents like butyrat

Detoxification Strategy of Epoxide Hydrolase

The human microsomal epoxide hydrolase, a single enzyme, has to detoxify a broad range of structurally diverse, potentially genotoxic epoxides that are formed in the course of xenobiotic metabolism. The enzyme has developed a unique strategy to combine a broad substrate specificity with a high detoxification efficacy, by immediately trapping the reactive compounds as covalent intermediates and by being expressed at high levels for high trapping capacity. Computer simulation and experimental data as well as existing epidemiologic studies reveal this detoxification strategy as the mechanistic basis for a threshold in the tumorigenesis of mutagenic carcinogens

Dietary topoisomerase II-poisons

DNA topoisomerases are nuclear enzymes inducing transient breaks in the DNA allowing DNA strands or double helices to pass through each other. The clinically used DNA topoisomerase II-poison etoposide is known to induce DNA double strand breaks leading to chromosomal aberrations and leukemias. Recently, some alarming studies have been published, suggesting that maternal exposure to low doses of dietary topoisomerase II-poisons, including bioflavonoids such as genistein or quercetin, may contribute to the development of infant leukemia: approximately 80% of infants with acute myelogenous leukemia (AML) and acute lymphoblastic leukemia (ALL) have chromosome translocations involving the MLL (mixed lineage leukemia) gene. It has been shown that antineoplastic chemotherapy with the leukemogenic topoisomerase II-poison etoposide induced identical chromosomal aberrations involving the MLL gene compared to children with infant leukemia. Interestingly, the MLL cleavage sites induced by etoposide colocalized with the cleavage sites observed in infant leukemia. In addition, an almost 10-fold higher risk of infant AML has been reported for mothers consuming relatively high levels of topoisomerase II-poison containing foods. These observations are relevant, since many foods contain topoisomerase II-poisons, predominantly soy and soy products, but also coffee, wine, tea, cocao, as well as some fruits and vegetables. Further studies on the role of dietary topoisomerase II-poisons are urgently required. If the causal relationship between dietary exposure to topoisomerase II-poisons and infant leukemia will be confirmed, care should be taken to reduce exposure to critical foods during pregnancy

Genotoxicity characteristics of reverse diol-epoxides of chrysene

Trans-3,4-dihydroxy-3,4-dihydrochrysene (chrysene-3,4-diol), a major metabolite of chrysene, is further metabolized by rat liver enzymes to products which effectively revert the his− Salmonella typhimurium strain TA98 to histidine prototrophy, but are only weakly mutagenic in strain TA100 and in Chinese hamster V79 cells (acquisition of resistance to 6-thioguanine). The liver enzyme mediated mutagenicity of chrysene-3,4-diol is substantially enhanced in the presence of 1,1,1-trichloropropene 2,3-oxide, an inhibitor of microsomal epoxide hydrolase. The predominant metabolites of chrysene-3,4-diol, namely the anti- and syn-isomers of its 1,2-oxide (termed reverse diol-epoxides), proved to be extraordinarily effective mutagens in S.typhimurium strain TA98, but were only moderately active in strains TA100 and TA104, and in the SOS induction in Escherichia coli PQ37. These genotoxicity spectra in bacteria are completely different from those observed with the bay-region diol-epoxides of chrysene and 3-hydroxychrysene. In V79 cells, the reverse diol-epoxides formed low levels of DNA adducts and were very weak inducers of gene mutations. In M2 mouse prostate cells, however, high numbers of transformed foci were induced by chrysene-3,4-diol and its diastereomeric 1,2-oxides. Chrysene-3,4-diol was somewhat more potent than chrysene-1,2-diol. The potency of both reverse diol-epoxides was similar to that of the syn-diastereomers of the bay-region diol-epoxides of chrysene and 3-hydroxychrysene, but lower than that of their anti-diastereomers. The reverse diol-epoxides of chrysene, unlike the bay-region diol-epoxides, were inactivated by purified microsomal epoxide hydrolase. Noteworthy findings were also made with regard to the chemical stability of the diol-epoxides in buffer, determined from the decline in mutagenicity after preincubation in the absence of the target cells. Despite its lower ΔEdeloc/β value for the formation of the benzylic carbocation, anti-chrysene-3,4-diol 1,2-oxide was shorter-lived (t½ = 46 min) than anti-chrysene-l,2-diol 3,4-oxide (t½ = 74 min). Unlike other investigated diastereomeric pairs of diol-epoxides, it was also shorter-lived than its syn-diastereomer (t½12 = 340 min

The evolution of the cosmic molecular gas density

One of the last missing pieces in the puzzle of galaxy formation and evolution through cosmic history is a detailed picture of the role of the cold gas supply in the star-formation process. Cold gas is the fuel for star formation, and thus regulates the buildup of stellar mass, both through the amount of material present through a galaxy's gas mass fraction, and through the efficiency at which it is converted to stars. Over the last decade, important progress has been made in understanding the relative importance of these two factors along with the role of feedback, and the first measurements of the volume density of cold gas out to redshift 4, (the "cold gas history of the Universe") has been obtained. To match the precision of measurements of the star formation and black-hole accretion histories over the coming decades, a two orders of magnitude improvement in molecular line survey speeds is required compared to what is possible with current facilities. Possible pathways towards such large gains include significant upgrades to current facilities like ALMA by 2030 (and beyond), and eventually the construction of a new generation of radio-to-millimeter wavelength facilities, such as the next generation Very Large Array (ngVLA) concept.Comment: 7 pages, 2 figures, Science White paper submitted to Astro2020 Decadal Surve