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E Huberman

Publications and source records attributed to E Huberman.

At least 145 records · Page 8Linked to original sources
PubMed1976

Mutability of different genetic loci in mammalian cells by metabolically activated carcinogenic polycyclic hydrocarbons.

The relationship between carcinogenesis and mutagenesis in mammalian cells has been determined with 10 polycyclic hydrocarbons with different degrees of carcinogenicity. Mutagenesis was determined in Chinese hamster cells with genetic markers that affect the surface membrane, nucleic-acid synthesis, and protein synthesis. The mutations were characterized by resistance to ouabain, 8-azaguanine, and temperature. Mutagenesis by the carcinogens required metabolic activation and this was provided by the presence of lethally irradiated metabolizing cells. The degree of carcinogenicity was related to the degree of mutagenicity for all three genetic markers. The most potent carcinogen, 7,12-dimethylbenz[a]anthracene, gave the highest mutagenicity and mutagenicity was obtained with 0.01 mug/ml. Treatment of the cells with aminophylline, which increases polycyclic hydrocarbon metabolism, increased mutagenesis by the carcinogens. It is suggested that such an experimental system with these and other mammalian cells should be useful as a sensitive assay for hazardous environmental chemicals.

Aminophylline↗
PubMed1976

Identification of mutagenic metabolites of benzo(a)pyrene in mammalian cells.

The mutagenicity of benzo[a]pyrene and 15 of its derivatives, which included phenols, the benzo[a]yrene-4,5-epoxide (the K-region epoxide), dihydrodiols, two isomeric 7,8-diol-9,10-epoxides, a 6-methyl derivative, and a 6-hydroxymethyl derivative, were tested with Chinese hamster V79 cells in order to identify the mutagenic metabolites of benzo[a]pyrene. Mutations were characterized by resistance to ouabain or 8-azaguanine. Since V79 cells do not metabolize polycyclic hydrocarbons, mutagenesis was tested both in the presence and absence of benzo[a]pyrene-metabolizing normal golden hamster cells. All the tested phenols, 4,5-diols, trans-9,10-diol, 6-methyl, and 6-hydroxymethyl derivatives of benzo[a]pyrene showed little or no mutagenicity for both genetic markers. The (+/-)7alpha,8beta-dihydroxy-9alpha,10alpha-epoxy-7,8;9,10-tetrahydrobenzo[a]pyrene and K-region 4,5-epoxide exhibited similar and moderate mutagenicity in the absence of benzo[a]pyrene-metabolizing cells, but the (+/-)7alpha,8beta-dihydroxy-9beta,10beta-epoxy-7,8,9,10-tetrahydrobenzo[a]-pyrene showed a 2000- and 270-fold higher mutation frequency for ouabain and 8-azaguanine resistance, respectively, than did the K-region 4,5-epoxide. The trans-7,8-diol which was not mutagenic in the absence of benzo[a]pyrene-metabolizing cells was more mutagenic than benzo[a]pyrene after metabolism and mutagenesis by trans-7,8-diol in these cells was inhibited by 7,8-benzoflavone, an inhibitor of mixed-function oxidases. Metabolically formed trans-7,8-diol was isolated and incubated with rat liver microsomes in the presence of co-factors. High-pressure liquid chromatography analysis indicated that the major metabolite of trans-7,8-diol is 7alpha,8beta-dihydroxy-9beta,10beta-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene. The results indicate that the latter compound is metabolically formed and the major mutagenic intermediate of benzo[a]yrene metabolism.

Benzopyrenes↗
PubMed1975

Mutation induction in Chinese hamster V79 cells by two vinyl chloride metabolites, chloroethylene oxide and 2-chloroacetaldehyde.

Chloroethylene oxide and 2-chloroacetaldehyde, two possibly carcinogenic metabolities of vinyl chloride in mammals, caused a dose-dependent induction of 8-azaguanine- and ouabain-resistant mutants in Chinese hamster V79 cells in vitro. Up to one-hundred-fold higher concentrations of 2-chloroethanol or monochloroacetic acid, a urinary vinyl chloride metabolite in rats and man, were inactive.

Acetaldehyde↗
PubMed1975

Regulation of aryl hydrocarbon (benzo-(A)-pyrene) hydroxylase activity in mammalian cells. Induction of hydroxylase activity by N6,O2'-dibutyryl8 adenosine 3':5'-monophosphate and aminophylline.

Treatment of hamster BHK cells with N6,O2'-dibutyryl adenosine 3':5'-monophosphate (Bt2cAMP), aminophylline, theophylline, or papaverine increased the level of aryl hydrocarbon (benzo(a)pyrene) hydrolxylase activity. The highese increase, 100-fold, was obtained with Bt2cAMP plus aminophylline or theophylline. N2,O2-Dibutyryl guanosine 3':5'-monophosphate gave a lower induction than Bt2cAMP. The level of hydroxylase activity started to decrease 6 hours after treatment with the inducer and was reduced to almost the uninduced level after 24 hours. Repeated addition of Bt2cAMP and aminophylline did not prevent this decrease. The hydroxylase can also be induced by treating cells with benz(a)anthracene, and the level of this induced activity was maintained for 24 hours. Aminophylline gave a 2- to 8-fold stimulation of the induction by benz(a)anthracene. The enzyme activity induced by Bt2cAMP, aminophylline, and benz(a)anthracene converted benzo(a)pyrene to similar alkali-extractable metabolities with a fluorescence spectra similar to that of 3-hydroxybenzo(a)pyrene. These induced enzyme activities also showed a similar heat stability. Induction by Bt2cAMP and aminophylline, like induction by benz(a)anthracene, required continued protein synthesis and only an initial period of RNA synthesis. Compared to the benz(a)anthracene-induced hydroxylase with a Km of 4.3 muM, the hydroxylase induced by Bt2cAMP and aminophylline showed a Km of 0.14 muM, and was 100-fold more sensitive to inhibition by 7,8-benzoflavone. Increasing the serum concentration in the culture medium stimulated the induction by aminophylline but did not stimulate induction by benz(a)anthracene. The results indicate that aryl hydrocaarbon (benzo(a)pyrene) hydroxylase can be induced by compounds that increase the level of adenosine 3':5'-monophosphate and that this induction and induced enzyme activity differs from that caused by benz(a)anthracene.

Aminophylline↗
PubMed1975

Activation of carcinogenic polycyclic hydrocarbons in polyoma-virus-transformed cells as a prerequisite for polyoma virus induction.

Polyoma-virus (PV)-transformed cell clones, which are inducible for virus synthesis by various physical and chemical agents, metabolize the chemically non-reactive carcinogen benzo(a)pyrene (BP) into water soluble products. In cultures of such clones, which metabolize BP to a level of 30-6-% of that of normal cells, up to 10.4% of the cells were induced for PV synthesis by BP, 20-methylcholanthrene (MCA) and 7,12-dimethylbenz(a)anthracene (CMBA). No PV induction was observed with the non-carcinogenic polycyclic hydrocarbons pyrene chrysene and benz(a)-anthracene. A proportion of subclones, isolated from a PV- inducible clone, which metabolized 0.1 mug or less BP per 10-6 cells were all inducible for PV synthesis by these carcinogens. Subclones isolated from an inducible clone pretreated with BP were shown to metabolize less than 0.1 mu BP per 10-6 cells and were resistant to virus induction by the carcinogenic polycyclic hydrocarbons. Benzoflavone, which inhibited the metabolism of BP in clones metabolizing high levels of this carcinogen, also prevented the induction of PV antigen and infectious virus synthesis in these clones. The data indicate a relationship between the carcinogenicity of polycyclic hydrocarbons and their ability to induce virus in the PV-transformed cells and suggest that virus induction depends on metabolic conversion of these hydrocarbons into similar reactive compounds that are responsible for malignant transformation and mutagenesis.

Animals↗
PubMed1975

Mammalian cell transformation and cell-mediated mutagenesis by carcinogenic polycyclic hydrocarbons.

The introduction of a polycyclic hydrocarbon such as benzo(alpha)pyrene (BP) into normal golden hamster embryo cell cultures results, in addition to cytotoxicity, in malignant cell transformation. Studies on the effect of different doses of BP on the normal cells showed that the frequency of transformed colonies was directly related to the dose of the carcinogen. Analysis of this dose-response curve suggests a one-event ("one-hit") response for transformation by this carcinogen. The one-event response for transformation by carcinogenic polycyclic hydrocarbons and the fact that these carcinogens bind to DNA in susceptible cells suggests that transformation can involve a single alteration in the genetic constitution of the treated cells. Carcinogens may, therefore, produce somatic mutations, some of which may involve the genes that control malignancy. Recently, considerable progress has been made in developing models for the study of chemical mutagenesis in mammalian cells. Using resistance to 8-azaguanine as a marker, positive correlations between mutagenicity and transformation were obtained with chemically reactive carcinogens such as N-acetoxy-N-2-fluorenyl-acetamide, N-methyl-N'-nitro-N-nitrosoguanidine and K-region epoxides of polycyclic hydrocarbons. However, no such correlations were obtained with the carcinogenic polycyclic hydrocarbons themselves, since the cell lines used in chemical mutagenesis do not metabolize these carcinogens. In order to obtain better correlations, we have developed a cell-mediated mutagenic assay with carcinogenic hydrocarbons in which Chinese hamster cells, which are susceptible for mutagenesis, were co-cultivated with lethally irradiated rodent cells that can metabolize these compounds. Using this cell mediated assay, we obtained mutagenesis with the carcinogenic hydrocarbons 7,12-dimethylbenz(alpha)anthracene (DMBA), BP, 3-methylcholanthrene and 7-methylbenz(alpha)anthracene; the most potent carcinogen, DMBA, gave the highest frequency of mutations. The polycyclic hydrocarbons, pyrene and benz(alpha)anthracene, which are not carcinogenic were also not mutagenic. We have therefore demonstrated a relationship between the carcinogenecity of polycyclic hydrocarbons and their mutagenicity in mammalian cells, without having to isolate their reative metabolic intermediates. It should be possible to use in this system human cells from different organs and individuals to screen for environmental chemicals hazardous to humans which have to be metabolically activated.

Acetoxyacetylaminofluorene↗
PubMed1974

Genetic control of the regulation of cell susceptibility to carcinogenic polycyclic hydrocarbons by cyclic AMP.

The metabolism of benzo (a) pyrene (BP) in normal golden hamster and BHK cells in culture was increased by treating the cells with dibutyryl cyclic AMP (dcAMP), prostaglandin E1, theophylline or aminophylline. The largest increase, 6-fold for the normal cells and 20-fold for the BHK cells, was obtained by treatment with both dcAMP and aminophylline. Treatment with aminophylline also stimulated the metabolism of 20-methylcholanthrene and 7,12-dimethylbenz (a) anthracene. The increased metabolism of these three carcinogeneic polycyclic hydrocarbons was associated with an increased cytotoxicity. Treatment with aminophylline increased the cytotoxicity of five other potent and weak carcinogenic polycyclic hydrocarbons, but not of two non-carcinogenic polycyclic hydrocarbons. The amount of BP metabolism in 27 different cell types from various mammals, including humans, ranged from less than 0.1 mug to 2.3 mug metabolized BP per 10-6 cells. Treatment of these different cell types with aminophylline gave either an increase in BP metabolism, an induction of metabolism in cells that did not metabolize without aminophylline, or no induction after treatment with aminophylline and dcAMP. The existence of responding and non-responding cell lines indicates that the regulation of the level of polycyclic hydrocarbon metabolism by dcAMP is genetically controlled. The induction of metabolism in cells that did not metabolize without aminophylline resulted in the conversion of cell resistance to cell susceptibility to the cytotoxic effect of BP. Treatment with dcAMP and aminophylline can therefore be used to increase the sensitivity of screening tests for chemical carcinogens.

Aminophylline↗