Biomedical subjects

J Lewtas

Publications and source records attributed to J Lewtas.

At least 55 records · Page 3Linked to original sources

PubMed1993

Bioassay of complex mixtures of indoor air pollutants.

The development of short-term genetic bioassays has made it possible to conduct mutagenicity studies on complex mixtures of indoor air pollutants. Although most of the studies have used the standard S. typhimurium/microsome reversion assay developed by Ames et al. (1975), the development of microsuspension mutagenesis methods (see Methods 18 and 19, this volume) has facilitated wider use of bioassays in indoor air pollution studies. It is clear from the studies reviewed here that environmental tobacco smoke is the major source of mutagens indoors. Other sources which produce significant, but much smaller, increases in indoor air mutagenicity include outdoor air and other indoor combustion sources (e.g., cooking, kerosene heaters and open fireplaces burning wood). These studies have been based on measurement of the bacterial mutagenicity of extracts from particulate matter. A limited number of reports, however, indicate that volatile and semi-volatile constituents of indoor air may also be mutagenic and deserve further study.

Air Pollution, Indoor↗

PubMed1992

Detection of direct-acting mutagens in ambient air: a comparison of two highly sensitive mutagenicity assays.

Ambient air has been shown to contain numerous hazardous pollutants, many of which are known or suspected carcinogens and mutagens. Bioassays play a prominent role in the characterization of these genotoxic pollutants, and as new test methods are developed, it is incumbent upon researchers to evaluate assay performance and report relative merits. In this study, two Salmonella test methods (the spiral and preincubation assays) were assessed to determine their usefulness as screening methods for monitoring direct-acting mutagens in ambient air. The spiral assay automates the conventional plate-incorporation assay and has been shown to reduce the labor, materials, and sample mass required to perform mutagenicity testing. The preincubation assay has been shown to enhance test sensitivity for certain classes of compound, thereby reducing the amount of sample required for dose-response analysis. Both assays were used to test organic extracts of airborne particulate matter collected in Tokyo during the winters of 1988 and 1990. In addition to the conventional tester strains TA98 and TA100, two newly developed YG strains were evaluated. Strains YG1024 and YG1029-derived from TA98 and TA100, respectively-contain an acetyltransferase plasmid that confers upon the strains greater sensitivity towards nitroarenes. Results from this study indicated that both assays were able to detect direct-acting mutagens in the Tokyo air samples. The mutagenic activity associated with the samples was directly related to the particle mass present in a given volume of air. Mutagenic response was greater in the spiral assay relative to the preincubation assay, especially when YG tester strains were used. The YG strains were significantly more sensitive to mutation than the TA strains in both assays, which suggests that nitroaromatics are an important class of genotoxic contaminant present in Tokyo air.

Air Pollutants↗

PubMed1992

Desensitization to allopurinol in patients with gout and cutaneous reactions.

PURPOSE: To determine the efficacy and safety of slow oral desensitization in the management of allopurinol-related pruritic cutaneous eruptions. PATIENTS AND METHODS: Nine patients with renal insufficiency and chronic tophaceous gouty arthritis, who had to interrupt their allopurinol therapy because of an allergic-type pruritic maculopapular eruption, were enrolled in an allopurinol oral desensitization protocol using a schedule of gradually increasing doses. RESULTS: Cautious reinstitution of allopurinol was successfully accomplished in all nine patients, but four individuals required dose adjustment because of development of a mild, recurrent, macular rash early during the protocol at allopurinol doses of less than or equal to 5 mg/d. Transient, postdesensitization cutaneous reactions occurred in two patients, one of whom also had an early rash. CONCLUSION: Oral desensitization to the minor rashes induced by allopurinol is a feasible and acceptably safe approach to therapy, particularly for those with renal insufficiency in whom no substitute urate-lowering drug is available.

Administration, Oral↗

PubMed1992

Mutagenicity of organic emissions from unvented kerosene heaters in a chamber study.

A study was conducted to assess the mutagenicity of semivolatile organics and particle-bound organics emitted from unvented kerosene space heaters. The units tested included a well-tuned radiant heater and a maltuned convective heater. The tests were conducted in a 27-m3 chamber with a prescribed on/off heater usage pattern. The organic emissions were collected on Teflon-coated glass filters backed by XAD-2 resin. The dichloromethane-extractable organics from both the filters and the XAD were analyzed for nitropolycyclic hydrocarbons using gas chromatography/mass spectrometry, and were bioassayed for mutagenicity in microsuspension assays using Salmonella typhimurium strains TA98 with and without S9 and TA98NR (a nitroreductase-deficient strain) without S9. The results showed that both the semivolatile and particle-bound organics emitted from the kerosene heaters were mutagenic, and the presence of nitropolycyclic hydrocarbons in these organic emissions substantiated these findings.

Air Pollutants↗

PubMed1992

Sources of genotoxicity and cancer risk in ambient air.

Products of incomplete combustion are identified as a major source of carcinogenic risk in urban areas, especially those from small non-industrial sources. The major ubiquitous emission sources outdoors in populated areas are residential home heating and motor vehicles. Indoors the major combustion source is environmental tobacco smoke. Polycyclic organic matter adsorbed onto the particles emitted from incomplete combustion are estimated to make the largest contribution to human genotoxic and cancer risk. Mutagenic emission factors combined with dispersion modelling indicated that automobiles and heating sources were major sources of mutagens. Ambient air studies to apportion the sources of mutagens in non-industrial areas confirmed this prediction. To apportion and estimate the cancer risk of ambient organic matter from particles in vivo animal tumour data, receptor modelling and human exposure data were combined. Tumourigenicity studies of the source apportioned ambient organic matter provided the relative tumour potencies of two ambient samples of different source composition. The human cancer unit risks were developed based on the comparative potency method using tumour data from these ambient samples. Residential wood combustion accounted for 75% of the exposure to particle associated organics, but only 20% of the estimated cancer risk. The remaining 80% of the risk appears to be associated with the mobile source component and atmospheric transformation products from these source emissions.

Air Pollutants↗

PubMed1992

Development of source testing, analytical, and mutagenicity bioassay procedures for evaluating emissions from municipal and hospital waste combustors.

Incineration is currently being used for disposal of about 10% of the solid waste generated in the United States, and this percentage will likely increase as land disposal declines. Siting new incinerators, however, is often controversial because of concerns related to the possibility of adverse health effects and environmental contamination from long-term exposure to stack emissions. Specific concerns relate to the adequacies of a) stack emission testing protocols, b) existing regulations, and c) compliance monitoring and enforcement of regulations. U.S. Environmental Protection Agency laboratories are cooperatively conducting research aimed at developing new testing equipment and procedures that will allow a more comprehensive assessment of the complex mixture of organics that is present in stack emissions. These efforts are directed specifically toward developing source testing equipment and procedures, analytical procedures, and bioassay procedures. The objectives of this study were to field test two types of high-volume source dilution samplers, collect stack samples for use in developing analytical and mutagenicity bioassay procedures, and determine mutagenicity of organics associated with emission particles from two municipal waste combustors and a hospital waste combustor. Data are presented for particle concentrations and emission rates, extractable organic concentrations and emission rates, and Salmonella (Ames) mutagenic potency and emission rates. The mutagenic emission rates and emission factors are compared to other incinerators and combustion sources.

Air Pollutants↗

PubMed1991

Relative sensitivity of 32P-postlabelling of DNA and the autoradiographic UDS assay in the liver of rats exposed to 2-acetylaminofluorene (2AAF).

Groups of male Alderley Park rats were dosed concomitantly with 2-acetylaminofluorene (2AAF) by gavage at doses between 0.01 mg/kg and 40 mg/kg, and livers sampled 2-72 h later. The liver of one group of animals was perfused to yield hepatocytes which were assayed in vitro for unscheduled DNA synthesis (UDS) via incorporation of tritiated thymidine and autoradiography. DNA was extracted from the livers of the other group and DNA adduct levels determined using the 32P-postlabelling technique. The major C-8 2-aminofluorene/guanosine adduct and 3 minor adducts were quantitated, enabling the relative sensitivity of the 2 techniques to be compared. A dose- and time-related UDS response was observed, which, at the most sensitive time-point (12 h) enabled DNA repair to be discerned at a dose level of 0.1-1 mg/kg of 2AAF, a response classified as formally positive at 5 mg/kg 2AAF. Only the C-8 adduct, as determined by 32P-postlabelling, was discernible at 0.01 mg/kg of 2AAF, although other adducts were visible on autoradiograms at higher dose levels. It is concluded that as part of a well-defined dose response, UDS can be discerned with confidence for doses of 2AAF between approximately 0.1 and 5 mg/kg, and DNA adducts for doses of 2AAF between approximately 0.01 and 1 mg/kg. Discernible UDS for 2AAF in the rat liver is apparent at approximately 13 DNA (total) adducts/10(8) nucleotides, or approximately 8 DNA (C-8) adducts/10(8) nucleotides. The presumed C-8 2-acetylaminofluorene/guanosine adduct, prepared by reaction of 2-acetoxy-2-acetylaminofluorene (2AAAF) with DNA, was a significant but unreliable marker of 2AAF/DNA adducts in the rat liver in vivo. DNA repair did not appear to remove DNA adducts selectively, and adducts remained in DNA when discernible DNA repair had ceased.

2-Acetylaminofluorene↗

PubMed1991

Relative sensitivity of 32P-postlabelling of DNA and the autoradiographic UDS assay in the liver of mice exposed to 2-acetylaminofluorene (2AAF).

In contrast to earlier studies conducted at lower dose levels, 2AAF is shown to induce a positive UDS response in the liver of mice dosed orally at dose levels between 500 and 1000 mg/kg. Similarly exposed mice had low levels of 2AAF-related hepatic DNA adducts at dose levels in the range 10-1000 mg/kg 2AAF, as determined by 32P-postlabelling analysis. It is concluded that the attenuated UDS response observed in the mouse liver, as compared to the rat liver, is due primarily to metabolic differences between these two species, coupled to a reduced capacity for UDS in the mouse liver for a given level of total 2AAF-related adducts per unit of DNA. These observations are compared and contrasted with identical studies conducted in the rat and reported in the preceding paper (Gallagher et al., 1991).

2-Acetylaminofluorene↗

PubMed1991

Formation of mutagenic urinary metabolites from 1-nitropyrene in germ-free and conventional rats: role of the gut flora.

1-Nitropyrene (NP), an environmental pollutant, a potent mutagen and an animal carcinogen, undergoes reduction, acetylation, ring-hydroxylation and conjugation in the rat in vivo to form mutagenic metabolites which are excreted in the urine. In order to investigate the role of the gut flora in the generation of these metabolites, germ-free rats of the AGUS strain, and conventional AGUS rats matched for sex and age, were injected i.p. with NP labelled with 14C. The germ-free rats excreted significantly less of the dose in urine than did the conventional rats. When urines were examined for mutagenicity with the Ames plate incorporation assay, the highest mutagenic activity was seen in the presence of S9 in 8-24 h urine from conventional rats. The conventional urines exceeded the germ-free urines by 10-fold in their content of 6-hydroxy-1-acetamidopyrene (NAAP-6-OH), previously identified as the predominant contributor to the mutagenicity of the urines of rats dosed with NP and excreted mainly as its beta-glucuronide conjugate. Conventional Charles River CD rats treated orally with D-glucaro-1,4-lactone, an inhibitor of beta-glucuronidase activity, excreted somewhat less NP-derived 14C in their urines over 48 h than did matched untreated rats, and their 8-24 h urines contained less than half as much of the mutagenic NAAP-6-OH as was found in the urines of the control rats. These results indicate that the gut flora are necessarily involved in the formation of NAAP-6-OH, and that both nitroreduction and the hydrolysis of glucuronides released for enterohepatic recirculation are essential in generating mutagenic metabolites from NP.

Animals↗

PubMed1991

Improvement in the diagnostic potential of 32P-postlabeling analysis demonstrated by the selective formation and comparative analysis of nitrated-PAH-derived adducts arising from diesel particle extracts.

Two versions of the 32P-postlabeling assay (nuclease P1 and butanol extraction) enhance the detection limit of polycyclic aromatic hydrocarbon (PAH)-modified DNA. Previously published studies suggest that DNA adducts derived from N-substituted aryl compounds are poorly recovered in the nuclease P1 version. In this study, both versions were employed to ascertain whether the apparent differences in sensitivity could be used to select diagnostically for nitroaromatic-DNA adducts derived by treating calf thymus DNA with organic extracts isolated from four diesel and one gasoline vehicle emission particles. We enhanced the formation of nitrated-PAH-derived adducts through xanthine oxidase (XO)-catalyzed nitroreduction of nitrated-PAHs, constituents previously detected in the diesel emissions. Chromatographic mobilities of the XO-derived DNA adducts were compared to adducts detected in calf thymus DNA resulting from rat liver S9-mediated metabolism. All four diesel organic extracts treated with XO resulted in the formation of one major DNA adduct, chromatographically distinct from the multiple DNA adducts detected in the rat liver S9-treated incubations. This adduct was detectable with the butanol extraction but not the nuclease P1 version of the 32P-postlabeling assay and was chromatographically similar to DNA adducts formed following XO nitroreduction of 1-nitropyrene or ascorbic acid treatment of 1-nitro-8-nitroso-pyrene and 1-nitro-6-nitrosopyrene. In contrast, when S9 activation was used, multiple DNA adducts were detected along a diagonal zone of radioactivity and were radioactively labeled with equivalent efficiency irrespective of the assay version employed. The in vitro calf thymus DNA model described in this study enhances the diagnostic potential of the 32P-postlabeling assay through the selective formation of nuclease P1-sensitive N-substituted aryl-derived DNA adducts.

Air Pollutants↗

PubMed1990

Metabolism of 1-nitropyrene by human, rat, and mouse intestinal flora: mutagenicity of isolated metabolites by direct analysis of HPLC fractions with a microsuspension reverse mutation assay.

The metabolism of [14C]-1-nitropyrene by human, rat and mouse intestinal microflora and a bioassay-directed chemical analysis of the isolated metabolites by assaying HPLC fractions with a microsuspension reverse mutation assay were examined. [14C]-1-Nitropyrene was metabolized by human, rat, and mouse intestinal microflora to 1-aminopyrene, N-acetyl-1-aminopyrene, N-formyl-1-aminopyrene, and two unknown metabolites identified as A and B. The predominant metabolite produced by human, rat, or mouse intestinal microflora following a 12-h incubation with [14C]-1-nitropyrene was 1-aminopyrene, which accounted for 93, 79, and 88% of the total 14C, respectively. Only minor amounts of N-formyl-1-aminopyrene (1.4, 1.2, and 1.0%), N-acetyl-1-aminopyrene (4.4, 3.0, and 3.4%), unknown A (1.0, 1.2, and 1.0%), and unknown B (3.3, 5.0, and 1.2%) were detected. These data suggest that a similar mechanism exists in the biotransformation of 1-nitropyrene by intestinal microflora from all three sources. Direct mutagenicity analysis of the HPLC fractions produced by intestinal microflora with the microsuspension reverse mutation assay indicated that mutagenic fractions can be resolved using this methodology.

Animals↗

PubMed1990

Dose-related differences in DNA adduct levels in rodent tissues following skin application of complex mixtures from air pollution sources.

Dose-related differences in the binding of DNA reactive intermediates for three environmentally important complex mixture particulate extracts and a well-studied carcinogen, benzo[a]pyrene (BaP), were examined in female C-57 mice following multiple topical treatments ranging from 1 to 120 mg/mouse. Particulate extracts from coke oven, coal soot and diesel exhaust were selected as model complex mixtures based on short-term mutagenicity assays, animal bioassays for carcinogenicity or epidemiological studies, where increased incidences of lung cancer in exposed populations were detected. Positive and negative control animals were treated with 1.2 mg BaP or acetone respectively. DNA was isolated from skin, lung and liver 24 h following the last application and analyzed for DNA adducts using the nuclease P1 version of the 32P-postlabeling assay. Each of the particulate extracts produced distinct patterns of DNA adducts. A diagonal zone of radioactivity, presumably representing multiple putative DNA adducts, was observed for coke-oven, coal-soot- and diesel-modified DNA samples. One adduct, common to all three complex-mixture-modified DNA samples, co-migrated with the major BaP adduct observed following treatment with BaP alone. Based on the BaP concentration for each of the extracts it seems unlikely that this adduct is derived from BaP alone. It is possible that an adduct is formed with chromatographic properties similar to the major BaP-derived adduct detected in mice treated with BaP alone. This adduct was detected in all tissues examined and represented approximately 12-34% of the total number of adducts detected within the diagonal radioactive zone for all coke-oven- and coal-soot-exposed tissues (skin, lung and liver). In contrast, this adduct represented 49-67% of the total radioactivity recovered from the diagonal zone of DNA isolated from lungs of animals exposed to diesel extract. The highest total number of adducts resulted from the metabolism of coke oven extract followed by coal soot and diesel treatments respectively. A dose-dependent increase in adduct formation was observed for all tissues in the diesel- and coal-soot-treatment mice. Liver and lung, but not skin, DNA adduct levels increased in a dose-dependent manner in the coke-oven-treated mice. The percentage of dose administered, detected as DNA adducts increased in all tissues as the dose decreased for all three complex mixtures. These data have important implications for risk assessment of these complex mixtures.

Adenosine Triphosphate↗

PubMed1990

Bioassay-directed fractionation of 1-nitropyrene metabolites: generation of mutagrams by coupling reverse-phase HPLC with microsuspension mutagenicity assays.

We have performed bioassay-directed fractionation of a model complex mixture (rabbit lung S9-generated metabolites of 14C-radiolabeled 1-nitropyrene) by assaying reverse-phase HPLC fractions using two microsuspension mutagenicity assays. A forward-mutation assay measuring mutation at the gpt locus (8-azaguanine resistance) in Salmonella typhimurium TM677 was performed in a total volume of 100 microliters, and a reverse-mutation assay measuring mutation at the hisD3052 allele in S. typhimurium TA98 was performed in a total volume of 200 microliters. HPLC fractions were collected every 30 s for 45 min, resulting in 90 fractions per run. The HPLC chromatogram (absorbance at 280 nm) and the 14C profile were compared to the mutagenicity profiles (mutagrams) and to the mutagenic potencies of pure metabolites studied separately. The results indicate that a fine dissection of the mutagenic fractions can be obtained by coupling HPLC to microsuspension mutagenicity assays. Differences observed between the mutagrams generated by the two bacterial strains were most likely due to metabolic (nitroreductase) differences between the two strains. This method should be generally applicable to the bioassay-directed chemical analysis of complex mixtures.

Animals↗

PubMed1990

Cotinine analytical workshop report: consideration of analytical methods for determining cotinine in human body fluids as a measure of passive exposure to tobacco smoke.

A two-day technical workshop was convened November 10-11, 1986, to discuss analytical approaches for determining trace amounts of cotinine in human body fluids resulting from passive exposure to environmental tobacco smoke (ETS). The workshop, jointly sponsored by the U.S. Environmental Protection Agency and Centers for Disease Control, was attended by scientists with expertise in cotinine analytical methodology and/or conduct of human monitoring studies related to ETS. The workshop format included technical presentations, separate panel discussions on chromatography and immunoassay analytical approaches, and group discussions related to the quality assurance/quality control aspects of future monitoring programs. This report presents a consensus of opinion on general issues before the workshop panel participants and also a detailed comparison of several analytical approaches being used by the various represented laboratories. The salient features of the chromatography and immunoassay analytical methods are discussed separately.

Animals↗

PubMed1990

Complex mixtures of urban air pollutants: identification and comparative assessment of mutagenic and tumorigenic chemicals and emission sources.

Current research on the cancer risks of complex mixtures of urban air pollutants addresses the identification and comparative assessment of mutagenic and carcinogenic chemicals and emission sources. To identify genotoxic compounds in urban air, micromutagenesis methods are coupled to analytical fractionation and new chemical characterization methods. Among polar mutagenic compounds, many appeared to be nitrated and or oxygenated aromatic compounds. Source apportionment of the mutagenic activity observed in urban air shows that vehicles and residential heating are major contributors to the ambient mutagenicity of the aerosol fraction. Comparative tumorigenesis and DNA adduct dosimetry studies are also being conducted with these same combustion emission sources and urban aerosols. The potential hazard and sources of the gaseous emissions have not been determined, but appear to be significantly increased by atmospheric transformation by ozone and nitrogen oxides.

Air Pollutants↗

PubMed1990

Sources of mutagenic activity in urban fine particles.

Samples were collected during the winter of 1984-1985 in the cities of Albuquerque, NM and Raleigh NC as part of a US Environmental Protection Agency study to evaluate methods to determine the emission sources contributing to the mutagenic properties of extractable organic matter (EOM) present in fine particles. Data derived from the analysis of the composition of these fine particles served as input to a multi-linear regression (MLR) model used to calculate the relative contribution of wood burning and motor vehicle sources to mutagenic activity observed in the extractable organic matter. At both sites the mutagenic potency of EOM was found to be greater (3-5 times) for mobile sources when compared to wood smoke extractable organics. Carbon-14 measurements which give a direct determination of the amount of EOM that originated from wood burning were in close agreement with the source apportionment results derived from the MLR model.

Air Pollutants↗

PubMed1989

Cytotoxicity and effect on mutagenicity of buffers in a microsuspension assay.

We have examined the effects of three buffers--Vogel-Bonner minimal E (VBM) and 0.15 and 0.015 M sodium phosphate, pH 7.4--and two concentrations of overnight cells (5x and 10x) on the mutagenicity of two pure compounds and two complex mixtures in strain TA98 using a modification of a microsuspension assay developed by Kado et al. (Mutation Research 121:25-32, 1983). The assay was performed by adding 50 microliters of cell concentrate of an overnight culture of TA98 resuspended in the appropriate buffer; 50 microliters of the same buffer or S9 mix; and 2 microliters of mutagen or dimethyl sulfoxide to a 1-dram vial or 13 x 150-mm test tube. The suspension was incubated for 90 min at 37 degrees C, top agar was added, and the contents poured onto a bottom agar of minimal medium. Cell concentration (5x vs. 10x) had little effect on the mutagenic potencies of the agents tested. The mutagenic potencies of the direct-acting agents (1-nitropyrene and diesel exhaust) were consistently lower with 0.15 M buffer compared to the mutagenic potencies of these agents with the other two buffers. The three buffers gave similar results for the indirect-acting agents (2-aminoanthracene and environmental tobacco smoke). The 0.15 M buffer was considerably cytotoxic in the absence of S9, which may explain why the direct-acting agents were less potent with this buffer compared to the other buffers. VBM was also somewhat cytotoxic, but this did not appear to affect the mutagenic potencies of the agents studied.(ABSTRACT TRUNCATED AT 250 WORDS)

Anthracenes↗

PubMed1989

Utility of short-term tests for genetic toxicity.

By definition, short-term tests (STTs) for genetic toxicity detect genotoxic agents, not carcinogens specifically. However, there is sufficient evidence, based on mechanistic considerations alone, to say that genotoxic agents are potential carcinogens. STTs have high statistical power, are almost always replicated, can be performed rather easily under various sets of experimental conditions, are relatively inexpensive, and detect a variety of endpoints relevant to carcinogenesis. In addition, several STTs have shown considerable utility in evaluating the genotoxic effects of real-world, environmental complex mixtures as well as the antimutagenic effects of various pure compounds and complex mixtures. STTs are likely to continue to be refined, resulting in STTs that are increasingly more relevant to human mutation and disease. Their utility should not be judged solely against the questionable standard of a rodent carcinogenicity assay.

Animals↗