Biomedical subjects

J Lewtas

Publications and source records attributed to J Lewtas.

102 records · Page 6Linked to original sources

PubMed1981

Development of a toxicity test system using primary rat liver cells.

A model in vitro rat liver parenchymal cellular toxicity system employing cells obtained by the in situ collagenase perfusion technique has been developed to detect potential liver toxicants. The initial evaluation of this test system was accomplished using cadmium chloride, chromium chloride, cobalt chloride, mercuric chloride, nickelous chloride, sodium arsenite, sodium selenite, and ammonium vanadate. Linear regression analysis of the dose response curves was used to determine the effective concentration at which the viability was reduced to 50% (EC50). The relative toxicity of the compounds was as follows: Cd greater than V = As greater than Se greater than Hg greater than Cr = Co greater than Ni. Since several of the compounds with very similar EC50s had significantly different dose response slopes, an additional parameter, lowest effective concentration tested (LECT) was employed to assess the relative toxicity. The LECT was determined using the Williams test and the relative toxicity of the compounds was found to be Cd = Se greater than V greater than As = Hg greater than Co greater than Cr = Ni. The primary objective in developing this rat liver cellular toxicity test system was to employ an in vitro test system utilizing metabolically active primary cells from a potential target organ. This study demonstrates the utility of this test system in determining the relative liver cell toxicity of a series of inorganic agents of differing toxicity.

Animals↗

PubMedDate not supplied

Design and implementation of a collaborative study of the mutagenicity of complex mixtures in Salmonella typhimurium.

In 1987, the International Programme on Chemical Safety (IPCS) in collaboration with the U.S. Environmental Protection Agency (U.S. EPA) and the U.S. National Institute of Standards and Technology (U.S. NIST) initiated an international collaborative study of the mutagenicity of complex environmental mixtures in the Ames Salmonella typhimurium mutation assay. The objectives of this study were: (1) to estimate the inter- and intra-laboratory variability associated with the extraction of mixtures for bioassay, (2) to estimate the inter- and intra-laboratory variability associated with the Salmonella typhimurium bioassay when applied to complex mixtures, and (3) to determine whether standard reference complex mixtures would be useful in mutagenicity studies and to evaluate whether reference or certified mutagenicity values determined from this collaborative study should be reported. The complex mixtures used in this study were selected from standard reference materials (SRMs) which had previously been issued by the U.S. NIST as SRM 1597 (coal tar), SRM 1649 (diesel particulate matter) and SRM 1650 (urban air particulate matter) with certified values for polycyclic aromatic hydrocarbons. These SRM complex mixtures are available to scientists as reference standards for analytical chemistry research and are under consideration as SRMs for mutagenicity studies of complex environmental mixtures. This paper briefly describes the final study design, protocol, selection of the complex mixtures, and implementation of this international study.

Laboratories↗

PubMedDate not supplied

Standard reference materials for chemical and biological studies of complex environmental samples.

Standard Reference Materials (SRMs) from the National Institute of Standards and Technology (NIST) are often used in methods development and interlaboratory comparison studies since they are homogeneous and readily available to the scientific community. SRM 1649 (urban dust/organics), SRM 1650 (diesel particulate matter), and SRM 1597 (complex mixture of polycyclic aromatic hydrocarbons from coal tar) are three environmental samples which have been used by the scientific community for these purposes. These SRMs were originally developed to assist laboratories in validating analytical procedures for the determination of polycyclic organic compounds in complex mixtures. In addition, these SRMs have been valuable for the comparison of methodologies for bacterial bioassays and the development of bioassay-directed fractionation and bioassay-directed chemical analysis techniques. Most recently these SRMs were chosen for use as test samples in a collaborative study coordinated by the World Health Organization--International Programme on Chemical Safety. This paper provides a summary of much of the work to date (published and unpublished) on the chemical and biological characterization of these three SRMs. Information regarding the availability of other NIST SRMs that might be useful for these types of studies are provided also.

Air Pollutants↗

PubMedDate not supplied

Overview, conclusions, and recommendations of the IPCS collaborative study on complex mixtures.

The International Programme on Chemical Safety (IPCS) sponsored an international collaborative study to examine the variability associated with the extraction and bioassay of standard reference materials (SRMs) that are complex environmental mixtures provided by the U.S. National Institute of Standards and Technology (NIST). The study was also intended to evaluate the feasibility of establishing bioassay reference values and ranges for the SRMs. Twenty laboratories from North America, Europe, and Japan participated in the study. As part of the mandatory core protocol, each laboratory extracted the organic material from two particulate samples and bioassayed these extracts. A coal tar polycyclic aromatic hydrocarbon (PAH) solution and two mutagenic control compounds were also subjected to bioassay without prior extraction by the participating laboratories. The bioassay used was the Salmonella/microsomal plate incorporation assay. For the optional portion of the study, a laboratory was free to use the SRMs for any type of exploratory research. The primary purpose of the required portion of the study was to estimate the intra- and inter-laboratory variability in mutagenic potencies of the test materials and to determine whether or not the NIST mixtures could be used as reference materials by others performing the Salmonella assay. Repeatability (intra-laboratory variance) of the bioassay results ranged from 16% to 88% depending on the SRM and the bioassay conditions (tester strain and metabolic activation), whereas reproducibility (inter-laboratory variance) ranged from 33% to 152%. Between-laboratory variability was the main source of variation accounting for approximately 55-95% of the total variation for the three environmental samples. Variation in the mutagenic potency of the control compounds was comparable, with the exception of 1-nitropyrene for which the reproducibility ranged from 127% to 132%. In summary, NIST SRMs provided useful materials for an international inter-laboratory study of complex mixtures. By establishing both intra- and inter-laboratory variance for the mutagenicity results for these materials, the usefulness of these SRMs as reference materials for the Salmonella bioassay was established, critical procedures within the bioassay protocol were identified, and recommendations for future efforts were delineated.

International Cooperation↗

PubMedDate not supplied

Mutagenicity of derivatives and metabolites of 1-nitropyrene: activation by rat liver S9 and bacterial enzymes.

The mutagenicity and activation requirements of purified synthetic derivatives and potential metabolites of 1-nitropyrene have been characterized in the Ames plate incorporation assay with the Salmonella tester strains TA98, TA98NR and TA98/1,8-DNP6, in the presence or absence of exogenous metabolic activation provided by Aroclor-induced rat liver S9. All the compounds tested (1-aminopyrene, N-acetyl-1-aminopyrene, N-hydroxy-N-acetyl-1-aminopyrene, 3-hydroxy-1-nitropyrene, 6-hydroxy-1-nitropyrene, and 8-hydroxy-1-nitropyrene) exhibited mutagenic activity under one or more assay conditions. 1-Nitropyrene was metabolized to 3-hydroxy-1-nitropyrene, 6- or 8-hydroxy-1-nitropyrene, 1-aminopyrene, N-acetyl-1-aminopyrene and other unidentified products (including some bound to protein) by an S9 preparation analogous to that used for exogenous metabolic activation in the Ames assay. 1-Nitropyrene and 3-hydroxy-1-nitropyrene were activated primarily by the 'classical' nitroreductase, while the other compounds, particularly in the presence of S9 metabolic activation, were dependent on transesterification for expression of their mutagenicity.

Animals↗

PubMedDate not supplied

In vitro metabolism and DNA adduct formation from the mutagenic environmental contaminant 2-nitrofluoranthene.

The metabolism and DNA adduct formation by the mutagenic environmental contaminant 2-nitrofluoranthene (2-NFA) were studied. Incubation under aerobic conditions with liver microsomes of rats pretreated with 3-methylcholanthrene yielded trans-7,8-dihydroxy-7,8-dihydro-2-nitrofluoranthene, trans-9,10-dihydroxy-9,10-dihydro-2-nitrofluoranthene, and 7-, 8-, and 9-phenolic metabolites. When the epoxide hydrolase inhibitor 3,3,3-trichloropropylene was present in the incubation, only phenolic metabolites were detected. Under hypoxic conditions, 2-aminofluoranthene was obtained, together with a trace of the ring-oxidized metabolites. The activated metabolite, N-hydroxy-2-aminofluoranthene, was prepared in situ and reacted with calf thymus DNA. Upon enzymatic hydrolysis of the DNA and purification by HPLC, a C8-substituted deoxyguanosine adduct, N-(deoxyguanosin-8-yl)-2-aminofluoranthene, was identified by mass and proton NMR spectral analysis. This adduct was also formed at a level of 10 pmol/mg of DNA when 2-NFA was metabolized by xanthine oxidase, 6 pmol/mg of DNA from incubation with liver microsomes of rats pretreated with 3-methylcholanthrene, and 3-pmol/mg of DNA from metabolism by liver microsomes of rats pretreated with phenobarbital.

Animals↗

PubMedDate not supplied

Separation of 32P-postlabeled DNA adducts of polycyclic aromatic hydrocarbons and nitrated polycyclic aromatic hydrocarbons by HPLC.

The 32P-postlabeling assay, thin-layer chromatography, and reverse-phase high-pressure liquid chromatography (HPLC) were used to separate DNA adducts formed from 10 polycyclic aromatic hydrocarbons (PAHs) and 6 nitrated polycyclic aromatic hydrocarbons (NO2-PAHs). The PAHs included benzo[j]fluoranthene, benzo[k]fluoranthene, indeno[1,2,3-cd]pyrene, benzo[a]pyrene, chrysene, 6-methylchrysene, 5-methylchrysene, and benz[a]anthracene. The NO2-PAHs included 1-nitropyrene, 2-nitrofluoranthene, 3-nitrofluoranthene, 1,6-dinitropyrene, 1,3-dinitropyrene, and 1,8-dinitropyrene. Separation of seven of the major PAH-DNA adducts was achieved by an initial PAH HPLC gradient system. The major NO2-PAH-DNA adducts were not all separated from each other using the initial PAH HPLC gradient but were clearly separated from the PAH-DNA adducts. A second NO2-PAH HPLC gradient system was developed to separate NO2-PAH-DNA adducts following one-dimensional TLC and HPLC analysis. HPLC profiles of NO2-PAH-DNA adducts were compared using both adduct enhancement versions of the 32P-postlabeling assay to evaluate the use of this technique on HPLC to screen for the presence of NO2-PAH-DNA adducts. To demonstrate the application of these separation methods to a complex mixture of DNA adducts, the chromatographic mobilities of the 32P-postlabeled DNA adduct standards (PAHs and NO2-PAHs) were compared with those produced by a complex mixture of polycyclic organic matter (POM) extracted from diesel emission particles. The diesel-derived adducts did not elute with the identical retention time of any of the PAH or NO2-PAH standards used in this study. HPLC analyses of the NO2-PAH-derived adducts (butanol extracted) revealed the presence of multiple DNA adducts.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMedDate not supplied

Evaluation of a personal air sampler for twenty-four hour collection of fine particles and semivolatile organics.

The U.S. EPA has conducted an evaluation of a commercially available lightweight fine particle personal sampler for use in human exposure and biomarker studies involving 24-h collections of particulate matter, particle-bound organics such as polycyclic aromatic hydrocarbons (PAHs), and semivolatile organics (PAHs). Independent laboratory evaluation of the prototype design, intended to produce a 2.5-micron aerodynamic diameter cut-point at a flow between 1.5 and 1.7 lpm (liters per minute), indicated that at a challenge flow rate of 1.5 lpm, the sampler provided an aerodynamic cut-point (dp50) of only 1.7 microns. The variance between expected size selection resulted from the prototype's jet diameter dimension being inadvertently based upon a flow rate of 2.0 lpm rather than an intended 1.5-1.7 lpm. Other aerodynamic factors not presently accounted for may also have played an influence. Extrapolated cut-points for flow rates at 1 and 3 lpm for the prototype were determined to be 2.1 and 1.2 microns, respectively. Total losses attributed to unwanted particle retention within the sampler ranged from 10% for 0.91 micron size particles to essentially zero approaching diameters greater than 2.0 microns. The ambient concentration of particles (< 1.7 microns) needed for acceptable gravimetric filter measurements involving 24-h sample collection was determined to be 10 micrograms/m3. Investigation of the sampler to retain and recover PAHs using XAD-2 resin, often of importance in human exposure biomarker studies, indicated that corrected recoveries between 94% and 108% could be obtained for 16 priority pollutant PAH species. Overall evaluation of the personal monitor indicates that it has research utility due to its modular features and size but reconfiguration should be performed that would permit true PM2.5 size selection. The current configuration collects particles less than 2.5 microns at approximately 95% collection efficiency.

Aged↗

PubMedDate not supplied

Mutagenicity and cytotoxicity of coal fly ash from fluidized-bed and conventional combustion.

In summary, fly-ash samples from a pressurized fluidized-bed combustion miniplant were found to consist of submicron, irregular particles that were cytotoxic and contained bioavailable mutagens. The fly-ash emission sample from a conventional coal-fired power plant was found to consist of spherical particles that were also cytotoxic but less mutagenic. The FBC fly ash investigated here was collected from an experimental miniplant and should not be considered representative of fly ash that may be obtained in the future from larger commercial-scale FBC plants. Further health and environmental assessment studies of coal fly-ash samples collected at multiple sites, including commercial-scale fluidized-bed and other conventional combustors, are needed to evaluate the potential health effects of coal fly ash from both types of combustion technology.

Air Pollutants↗

PubMedDate not supplied

Role of endotoxin in tumor necrosis factor alpha expression from alveolar macrophages treated with urban air particles.

The effects of urban air and diesel particles on inflammatory cytokine gene expression, tumor necrosis factor alpha (TNF-alpha) in particular, were studied in rat alveolar macrophages. TNF-alpha, interleukin (IL)-1, IL-6, cytokine-induced neutrophil chemoattractant (CINC), and macrophage inflammatory protein (MIP)-2 gene expression and TNF-alpha secretion were increased in cells treated with 50 to 200 micrograms/mL of urban air particles in a concentration-related manner. There was no cytokine induction by diesel particles at any of the concentrations tested. Cytokine expression was not related to reactive oxygen species since antioxidants, such as catalase, TMTU, or DMSO, had no effect on TNF-alpha secretion. However, cytokine induction by urban air particles was completely prevented by polymyxin B, an antibiotic capable of neutralizing bacterial lipopolysaccharide (LPS) activities. Furthermore, LPS was detected on the urban air particles, but not on diesel particle. These results suggest that activation of cytokine gene expression and secretion in rat alveolar macrophages by urban air particles is due to the presence of endotoxin on the particles.

Air Pollutants↗

PubMedDate not supplied

32P-postlabeling analysis of DNA adducts in human sperm cells from smokers and nonsmokers.

To determine the feasibility of using human sperm cells for DNA 32P-postlabeling analyses, and to evaluate the baseline level and the possible presence of smoking-related DNA adducts in these cells, sperm DNA was isolated from specimens obtained from 12 heavy smokers, 12 light smokers, and 12 nonsmokers. Background levels of radioactivity were minimized by using magnet transfer of 32P-labeled mononucleotides to new polyethyleneimine cellulose plates. Compared with placental tissues, few adducts were observed. Diffuse radioactivity observed in some of the autoradiograms was minimally above background but the level of radioactivity expressed as putative adducts/nucleotide was not related to smoking status. It was not clear, in some cases, whether this radioactivity was associated with chemically bound adducts or was from nonspecifically bound chemicals, radiolabeled enzymes, or other proteins. One major discrete DNA adduct of unknown chemical structure was detected in three of the 36 samples analyzed (one nonsmoker and two smokers). Based on the level of radioactivity associated with various dilutions of a benzo(a)pyrene-derived adduct, our limit of sensitivity was at least 1.2 adducts/10(9) nucleotides. Our study emphasizes the need to more clearly define the significance of background radioactivity associated with DNA adduct maps where the measured adduct levels approximate detection limits defined by visual observance of adduct spots. This point is particularly relevant given that the 32P-postlabeling procedures rely, in part, on visual verification of the presence of DNA adducts.

Adult↗

PubMedDate not supplied

Airborne exposures to PAH and PM2.5 particles for road paving workers applying conventional asphalt and crumb rubber modified asphalt.

Personal exposure monitoring was conducted for road paving workers in three states. A research objective was to characterize and compare occupational exposures to fine respirable particles (< 2.5 microns) and particle-bound polycyclic aromatic hydrocarbons (PAHs) for road paving workers applying conventional (petroleum derived) asphalt and asphalt containing crumb rubber from shredded tires. Workers not exposed to asphalt fume were also included for comparison (to support the biomarker component of this study). The rubber content of the crumb rubber modified (CRM) asphalt at the three study sites was 12, 15, and 20%. A comparison of some specific job categories from two sites indicates greater potential carcinogenic PAH exposures during CRM asphalt work, however, the site with the greatest overall exposures did not indicate any differences for specific jobs. A statistical analysis of means for fine particle, pyrene and total carcinogenic PAH personal exposure shows, with two exceptions, there were no differences in exposures for these three measurement variables. One site shows significantly elevated pyrene exposure for CRM asphalt workers and another site similarly shows greater carcinogenic PAH exposure for CRM asphalt workers. Conventional and CRM asphalt worker airborne exposures to the PAH carcinogen marker, BaP, were very low with concentrations comparable to ambient air in many cities. However, this study demonstrates that asphalt road paving workers are exposed to elevated airborne concentrations of a group of unknown compounds that likely consist of the carcinogenic PAHs benz(a)anthracene, chrysene and methylated derivatives of both. The research described in this article has been reviewed in accordance with U.S. Environmental Protection Agency policy and approved for publication. Mention of trade names or commercial products does not constitute endorsement or recommendation for use.

Air Pollution↗