Human bronchus-mediated mutagenesis of mammalian cells by carcinogenic polynuclear aromatic hydrocarbons.

Hsu, Ih‐Chang; Stoner, Gary D.; Autrup, Herman; Trump, Benjamin F.; Selkirk, James K.; Harris, Curtis C.

doi:10.1073/pnas.75.4.2003

Cited by 37 publications

(9 citation statements)

References 16 publications

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“…In such assays, PAH metabolites produced by cultured human epithelial cells or tissues induce mutations in cocultured fibroblasts, which grow better in simple culture conditions and for which mutagenesis at several loci has been quantitated and characterized (10,11). Human mammary epithelial cells (12), epidermal keratinocytes (13,14), bronchial tissue explants (15), and a human kidney carcinoma line (16) have been shown to produce PAH metabolites that can induce mutation in fibroblasts. A fundamental limitation of this approach, resulting from the instability of many mutagenic PAH intermediates and the differences in DNA repair capacity among various cell types, is that the genomes of the target fibroblasts may not sustain the same type and degree of damage as the genomes of the epithelial cells that produce the metabolites.…”

mentioning

confidence: 99%

Polycyclic aromatic hydrocarbon mutagenesis of human epidermal keratinocytes in culture.

Allen-Hoffmann¹,

Rheinwald²

1984

Proc. Natl. Acad. Sci. U.S.A.

182

103

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We have developed a culture system for detecting and isolating rare hypoxanthine phosphoribosyltransferase-deficient mutants of human epidermal keratinocytes. A thioguanine-resistant variant, 3T3M1, of the Swiss mouse fibroblast line 3T3 was used as a feeder layer to support clonal growth of mutant keratinocytes. A near-diploid, epidermal squamous cell carcinoma line, SCC-13Y, was used as a prototype to determine mutagen treatment conditions, plating density, and phenotypic expression time for maximum mutant recovery. To extend this system to normal keratinocytes, we improved the culture conditions by adding insulin, adenine, and Ham's nutrient mixture F-12, which increased colony-forming efficiencies to 30% in early passage and made feasible the detection of rare mutants in normal epidermal keratinocyte populations. We have quantitated mutation in SCC-13Y and three strains of normal human epidermal keratinocytes after exposure to polycyclic aromatic hydrocarbons, which are activated to their mutagenic forms by cellular mixed-function oxidases.7,12-Dimethylbenz[a]anthracene and benzo[a]pyrene caused almost no cytotoxicity, but induced thioguanine-resistant mutants at frequencies as much as 50-fold higher than the spontaneous frequency of 10-6. The mutants were aminopterin-sensitive and possessed no measurable hypoxanthine phosphoribosyltransferase activity; their behavior was indistinguishable from that of keratinocytes cultured from individuals with Lesch-Nyhan syndrome. This mutagenesis assay system should also be applicable to other feeder layer-dependent human epithelial cell types, such as urothelial, mammary, and tracheal epithelial cells.The epidermis and many other epithelial tissues possess a family of mixed-function oxidases (flavoprotein-linked monoxygenase, EC 1.14.14.1) and related enzymes, which metabolize complex organic chemicals to excretable and generally less toxic intermediates (1-4). Although the mixed-function oxidase system probably evolved as a protective mechanism, some classes of chemicals, such as the polycyclic aromatic hydrocarbons (PAHs), are converted by these enzymes to intermediates that are more cytotoxic, mutagenic, and carcinogenic than the parent compounds (5-7). More than half of all human cancers arise in surface epithelia that are in direct contact with the environment (i.e., the epithelia of the epidermis, esophagus, trachea, bronchus, cervix, and intestine) (8). Exposure of these tissues to PAHs and other environmental agents is thought to contribute significantly to the initiation and/or progression of human carcinomas (for review, see ref. 9).A criterion such as frequency of induced mutation at a defined genetic locus or acquisition of a phenotype associated with malignant transformation in cultures of chemically treated human epithelial cells could be useful in identifying both potentially dangerous chemicals and individuals particularly susceptible to chemical carcinogenesis. The proliferative capacity and clonal growth ability of normal human epithelial cell...

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mentioning

confidence: 99%

Polycyclic aromatic hydrocarbon mutagenesis of human epidermal keratinocytes in culture.

Allen-Hoffmann¹,

Rheinwald²

1984

Proc. Natl. Acad. Sci. U.S.A.

182

103

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“…In addition to the theoretical utility of ratios of transformation to mutation frequencies obtained in the same cells, comparisons of mutagenesis and transformation are also potentially helpful to standardize the use of the mutagenic potential of agents as a predictor of their possible oncogenic potential (10)(11)(12)(13)(14)(21)(22)(23)). …”

mentioning

confidence: 99%

Chemical carcinogens produce mutations to ouabain resistance in transformable C3H/10T1/2 Cl 8 mouse fibroblasts.

Landolph¹,

Heidelberger²

1979

Proc. Natl. Acad. Sci. U.S.A.

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Chemical carcinogens induce mutations to ouabain resistance in the transformable mouse fibroblast cell line C3H/1OT1/2 Cl 8. The mutant phenotype is stable and heritable in the absence of selective agent, and dose-response curves for mutant frequency were obtained with N-methyl-N'-nitro-N-nitrosoguanidine, 3-methylcholanthrene, benzo[apyrene, N-acetoxy-N-2-acetylaminofluorene, and the anti-7,8-diol-9,10-epoxide of benzo [a]pyrene. The ratio of the malignant transformation frequency to the mutation frequency was 12 for benzo[a pyrene and 21 for-N-acetoxy-N-2-acetylaminofluorene. The development of the mutational assay reported here allows the use ofthis permanent cell line for comparison of mutation and transformation frequencies and as a screening system for xenobiotics that pose mutagenic or carcinogenic hazards to mammalian cells.It is well known that chemical carcinogens can induce malignant transformation in mammalian cell systems (1-9). However, it is not known with certainty if chemically induced transformation is a mutational event, and the precise molecular and cellular alterations that result in transformation have not been identified.The carcinogen-induced transformation frequencies reported (1-9) are generally of the order of 102 to 103 times greater than carcinogen-induced mutation frequencies measured in different mammalian cells (10)(11)(12)(13)(14), and these comparisons have recently been reviewed (15). When mutation and transformation were induced in the same cells by the same agent, ratios of transformation to mutation frequencies of 20 (16), 20-500 (17, 18), and 10 (19, 20) have been reported. However, some of these ratios of transformation to mutation frequencies could be artificially high due to the nontumorigenicity of some of the morphologic transformants in some systems. These ratios could be useful because they might help to define the number of genes that are targets for transformation.In addition to the theoretical utility of ratios of transformation to mutation frequencies obtained in the same cells, comparisons of mutagenesis and transformation are also potentially helpful to standardize the use of the mutagenic potential of agents as a predictor of their possible oncogenic potential (10-14, 21-23).Consequently we have established and characterized a mutational assay for ouabain-resistant mutants in the C3H/LOT1/2 Cl 8 mouse fibroblast cell line (24), in which we have established a correspondence between morphologic and malignant transformation (7).Ouabain resistance (Ouar), which has been shown to be induced in many mammalian cell types by known mutagens (25-29), results from an altered sodium-potassium adenosinetriphosphatase (Na+,K+-ATPase) that no longer binds ouabain (Oua) or binds it less tightly than the wild-type enzyme does (27). During the preparation of this manuscript, Chan and Little demonstrated mutagenesis to Ouar in these cells and showed that ultraviolet light produced a transformation/ouabain resistance ratio of 10 (20). MATERIALS AND METHODSCell Culture ...

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“…The or mutation frequency was dependent on the number of cocultivated PAM and on the concentration of either (+) 7,X, 8a-dihydroxy-7,8-dihydrobenzo[a]pyrene (7,8-diol) or B(a)P. Chinese hamster V79 cells do not effectively metabolize either B(a)P or 7,8-diol into ultimate mutagens (10). In the present report, these studies have been extended to evaluate the ability of PAM from 20 individuals to mediate mutations in V79 cells.…”

Section: Abstract Pulmonary Macrophages (Pam)mentioning

confidence: 99%

Induction of Ouabain-resistant Mutation and Sister Chromatid Exchanges in Chinese Hamster Cells with Chemical Carcinogens Mediated by Human Pulmonary Macrophages

Hsu¹,

Harris²,

Yamaguchi³

et al. 1979

J. Clin. Invest.

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Human bronchus-mediated mutagenesis of mammalian cells by carcinogenic polynuclear aromatic hydrocarbons.

Abstract: Cultured human bronchial explants activated benzo[alpyrene (BzaP) We have modified these cell-mediated mutagenesis systems

Cited by 37 publications

References 16 publications

Polycyclic aromatic hydrocarbon mutagenesis of human epidermal keratinocytes in culture.

Polycyclic aromatic hydrocarbon mutagenesis of human epidermal keratinocytes in culture.

Chemical carcinogens produce mutations to ouabain resistance in transformable C3H/10T1/2 Cl 8 mouse fibroblasts.

Induction of Ouabain-resistant Mutation and Sister Chromatid Exchanges in Chinese Hamster Cells with Chemical Carcinogens Mediated by Human Pulmonary Macrophages

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