RESUMO
Current risk assessments of 1,3-butadiene (BD*) are complicated by limited evidence of its carcinogenicity in humans. Hence, there is a critical need to identify early events and factors that account for the heightened sensitivity of mice to BD-induced carcinogenesis and to deter-mine which animal model, mouse or rat, is the more useful surrogate of potency for predicting health effects in BD-exposed humans. HEI sponsored an earlier investigation of mutagenic responses in mice and rats exposed to BD, or to the racemic mixture of 1,2-epoxy-3-butene (BDO) or of 1,2,3,4-diepoxybutane (BDO2; Walker and Meng 2000). In that study, our research team demonstrated (1) that the frequency of mutations in the hypoxanthine-guanine phosphoribosyl transferase (Hprt) gene of splenic T cells from BD-exposed mice and rats could be correlated with the species-related differences in cancer susceptibility; (2) that mutagenic-potency and mutagenic-specificity data from mice and rats exposed to BD or its individual epoxy intermediates could provide useful information about the BD metabolites responsible for mutations in each species; and (3) that our novel approach to measuring the mutagenic potency of a given chemical exposure as the change in Hprt mutant frequencies (Mfs) over time was valuable for estimating species-specific differences in mutagenic responses to BD exposure and for predicting the effect of BD metabolites in each species. To gain additional mode-of-action information that can be used to inform studies of human responses to BD exposure, experiments in the current investigation tested a new set of five hypotheses about species-specific patterns in the mutagenic effects in rodents of exposure to BD and BD metabolites: 1. Repeated BD exposures at low levels that approach the occupational exposure limit for BD workers (set by the U.S. Occupational Safety and Health Administration) are mutagenic in female mice. 2. The differences in mutagenic responses of the Hprt gene to BD in similarly exposed rodents of a given species (reported in various earlier studies) are primarily associated with age-related thymus activity and trafficking of T cells and with sex-related differences in BD metabolism. 3. The mutagenic potency of the stereochemical forms of BD's epoxy intermediates plays a significant role in the species-related mutagenicity of BD. 4. The hydrolysis-detoxification pathway of BD through 1,2-dihydroxy-3-butene (BD-diol) is a major contributor to mutagenicity at high-level BD exposures in mice and rats. 5. Significant and informative species-specific differences in mutation spectra can be identified by examining both large- and small-scale genetic alterations in the Hprt gene of BD-exposed mice and rats. The first four hypotheses were tested by exposing mice and rats to BD, meso-BDO2, or BD-diol and measuring Hprt Mfs as the primary biomarker. For this, we used the T-cell-cloning assay of lymphocytes isolated from the spleens of exposed and control (sham-exposed) mice and rats. The first hypothesis was tested by exposing female B6C3F1 mice (4 to 5 weeks of age) by inhalation for 2 weeks (6 hours/day, 5 days/week) to 0 or 3 ppm BD. Hprt Mfs were measured at the time of peak mutagenic response after exposure for this age of mice. We then compared the resulting data to those from mutagenicity studies with mice of the same age that had been exposed in a similar protocol to higher levels of BD (Walker and Meng 2000). In mice exposed to 3 ppm BD (n = 27), there was a significant 1.6-fold increase over the mean background Hprt Mf in control animals (n = 24, P = 0.004). Calculating the efficiency of Hprt mutant induction, by dividing induced Hprt Mfs by the respective BD exposure levels, demonstrated that the mutagenic potency of 3 ppm BD was twice that of 20 ppm BD and almost 20 times that of 625 or 1250 ppm BD in exposed female mice. Sample-size calculations based on the Hprt Mf data from this experiment demonstrated the feasibility of conducting a future experiment to find out whether induced Mfs at even lower exposure levels (between 0.1 and 1.0 ppm BD) fit the supralinear exposure-response curve found with exposures between 3.0 and 62.5 ppm BD, or whether they deviate from the curve as Mf values approach the background levels found in control animals. The second hypothesis was tested by estimating mutagenic potency for female mice exposed by inhalation for 2 weeks to 0 or 1250 ppm BD at 8 weeks of age and comparing this estimate to that reported for female mice exposed to BD in a similar protocol at 4 to 5 weeks of age (Walker and Meng 2000). For these two age groups, the shapes of the mutant splenic T-cell manifestation curves were different, but the mutagenic burden was statistically the same. These results support our contention that the disparity in responses reported in earlier Hprt-mutation studies of BD-exposed rodents is related more to age-related T-cell kinetics than to age-specific differences in the metabolism of BD. The third hypothesis was tested by estimating mutagenic potency for female mice and rats (4 to 5 weeks of age) exposed by inhalation to 2 or 4 ppm meso-BDO2 and comparing these estimates to those previously obtained for female mice and rats of the same age and exposed in a similar protocol to (+/-)-BDO2 (Meng et al. 1999b; Walker and Meng 2000). These exposures to stereospecific forms of BDO2 caused equivalent mutagenic effects in each species. This suggests that the small differences in the mutagenic potency of the individual stereoisomers of BDO2 appear to be of less consequence in characterizing the sources of BD-induced mutagenicity than the much larger differences between the mutagenic potencies of BDO2 and the other two BD epoxides (BDO and 1,2-dihydroxy-3,4-epoxybutane [BDO-diol]). The fourth hypothesis was tested in several experiments. First, female and male mice and rats (4 to 5 weeks of age) were exposed by nose only for 6 hours to 0, 62.5, 200, 625, or 1250 ppm BD or to 0, 6, 18, 24, or 36 ppm BD-diol primarily to establish BD and BD-diol exposure levels that would yield similar plasma concentrations of BD-diol. Second, animals were exposed in inhalation chambers for 4 weeks to 0, 6, 18, or 36 ppm BD-diol to determine the mutagenic potency estimates for these exposure levels and to compare these estimates with those reported for BD-exposed female mice and rats (Walker and Meng 2000) in which similar blood levels of BD-diol had been achieved. Measurements of plasma concentrations of BD-diol (via a gas chromatography and mass spectrometry [GC/MS] method developed for this purpose) showed these results: First, BD-diol accumulated in a sublinear manner during a single 6-hour exposure to more than 200 ppm BD. Second, BD-diol accumulated in a linear manner during single (6-hour) or repeated (4-week) exposure to 6 or 18 ppm BD and in a sublinear manner with increasing levels of BD-diol exposure. Third, exposure of female mice and rats to 18 ppm BD-diol produced plasma concentrations equivalent to those produced by exposure to 200 ppm BD (exposure to 36 ppm BD-diol produced plasma concentrations of about 25% of those produced by exposure to 625 ppm BD). In general, 4-week exposure to 18 or 36 ppm BD-diol was significantly mutagenic in female and male mice and rats. The differences in mutagenic responses between the species and sexes were not remarkable, except that the mutagenic effects were greatest in female mice. The substantial differences in the exposure-related accumulation of BD-diol in plasma after rodents were exposed to more than 200 ppm BD compared with the relatively small differences in the mutagenic responses to direct exposures to 6, 18, or 36 ppm BD-diol in female mice provided evidence that the contribution of BD-diol-derived metabolites to the overall mutagenicity of BD has a narrow range of effect that is confined to relatively high-level BD exposures in mice and rats. This conclusion was supported by the results of parallel analyses of adducts in mice and rats concurrently exposed to BD-diol (Powley et al. 2005b), which showed that the exposure-response curves for the formation of N-(2,3,4-trihydroxybutyl)valine (THB-Val) in hemoglobin, formation of N7-(2,3,4-trihydroxybutyl)guanine (THB-Gua) in DNA, and induction of Hprt mutations in exposed rodents were remarkably similar in shape (i.e., supralinear). Combined, these data suggest that trihydroxybutyl (THB) adducts are good quantitative indicators of BD-induced mutagenicity and that BD-diol-derived BDO-diol (the major source of the adducts) might be largely responsible for mutagenicity in rodents exposed to BD-diol or to hight levels of BD. The mutagenic-potency studies of meso-BDO2 and BD-diol reported here, combined with our earlier studies of BD, (+/-) BDO, and(+/-)-BDO2 (Walker and Meng 2000), revealed important trends in species-specific mutagenic responses that distinguish the relative degree to which the epoxy intermediates contribute to mutation induction in rodents at selected levels of BD exposures. These data as a whole suggest that , in mice, BDO2 largely causes mutations at exposures less than 62.5 ppm BD and that BD-diol-derived metabolites add to these mutagenic effects at higher BD exposures. In rats, it appears that the BD-diol pathway might account for nearly all the mutagenicity at the hight-level BD exposures where significant increases in Hprt Mfs are found and cancers are induced. Additional exposure-response studies of hemoglobin and DNA adducts specifics to BDO2, BDO-diol, and other reactive intermediates are needed to determine more definitively the relative contribution of each metabolite to the DNA alkylation and mutation patterns induced by BD exposure in mice and rats. For the fifth hypothesis, a multiplex polymerase chain reaction (PCR) procedure for the analysis of genomic DNA mutations in the Hprt gene of mice was developed. (ABSTRACT TRUNCATED)
Assuntos
Butadienos/toxicidade , Exposição Ambiental/efeitos adversos , Compostos de Epóxi/toxicidade , Alquilantes , Animais , Butadienos/sangue , Butadienos/metabolismo , Testes de Carcinogenicidade , Análise Mutacional de DNA , Compostos de Epóxi/sangue , Compostos de Epóxi/metabolismo , Feminino , Humanos , Neoplasias Pulmonares/induzido quimicamente , Neoplasias Pulmonares/epidemiologia , Neoplasias Pulmonares/etiologia , Neoplasias Pulmonares/genética , Masculino , Camundongos , Mutagênese , Testes de Mutagenicidade , Ratos , Ratos Endogâmicos F344 , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Medição de Risco , Fatores de Risco , Estados Unidos/epidemiologiaRESUMO
The disposition of [14C]methyltetrahydrofuran (14C-MTHF) in rats and mice was determined by following changes in the radioactivity in tissue and excreta with time after dosing. MTHF administered orally (1, 10, or 100 mg/kg) or intravenously (1 mg/kg) to either rats or mice was rapidly metabolized and excreted with <8% (mice) or 8-22% (rats) of the dose remaining in the body after 24 h (1 and 10 mg/kg doses) or 72 h (100 mg/kg dose). Based on recovery of radioactivity in excreta (other than feces) and tissues (other than the gastrointestinal [GI] tract), absorption of orally administered MTHF was essentially complete (93-100%). There were no overt signs of toxicity observed at any dose studied. The major route of excretion in mice was in urine followed by exhaled CO2. In rats the major route of excretion was exhaled CO2 followed by urinary excretion. The excretion of exhaled volatile organic compounds (VOC) was dose-dependent in both species; at lower doses exhaled VOC represented 1-5% of dose, but at the highest dose (100 mg/kg) this proportion rose to 14% (mice) and 27% (rats). Analysis of the VOCs exhaled at the high dose indicated that the increase was due to exhalation of the parent compound, 14C-MTHF. Analysis of urine showed three highly polar peaks in the mouse urine and two polar peaks in the rat urine. Because the 14C label in MTHF was in the methyl group, the polar metabolites were considered likely due to the one-carbon unit getting into the metabolic pool and labeling intermediate dietary metabolites.
Assuntos
Poluentes Ambientais/farmacocinética , Furanos/farmacocinética , Administração Oral , Animais , Radioisótopos de Carbono , Cromatografia Líquida de Alta Pressão , Injeções Intravenosas , Masculino , Camundongos , Camundongos Endogâmicos , Ratos , Ratos Endogâmicos F344RESUMO
Studies were performed to determine if the detoxification pathway of 1,3-butadiene (BD) through 3-butene-1,2-diol (BD-diol) is a major contributor to mutagenicity in BD-exposed mice and rats. First, female and male mice and rats (4-5 weeks old) were exposed by nose-only for 6h to 0, 62.5, 200, 625, or 1250 ppm BD or to 0, 6, 18, 24, or 36 ppm BD-diol primarily to establish BD and BD-diol exposure concentrations that yielded similar plasma levels of BD-diol, and then animals were exposed in inhalation chambers for 4 weeks to BD-diol to determine the mutagenic potency estimates for the same exposure levels and to compare these estimates to those reported for BD-exposed female mice and rats where comparable blood levels of BD-diol were achieved. Measurements of plasma levels of BD-diol (via GC/MS methodology) showed that (i) BD-diol accumulated in a sub-linear fashion during single 6-h exposures to >200 ppm BD; (ii) BD-diol accumulated in a linear fashion during single or repeated exposures to 6-18 ppm BD and then in a sub-linear fashion with increasing levels of BD-diol exposure; and (iii) exposures of mice and rats to 18 ppm BD-diol were equivalent to those produced by 200 ppm BD exposures (with exposures to 36 ppm BD-diol yielding plasma levels approximately 25% of those produced by 625 ppm BD exposures). Measurements of Hprt mutant frequencies (via the T cell cloning assay) showed that repeated exposures to 18 and 36 ppm BD-diol were significantly mutagenic in mice and rats. The resulting data indicated that BD-diol derived metabolites (especially, 1,2-dihydroxy-3,4-epoxybutane) have a narrow range of mutagenic effects confined to high-level BD (>or=200 ppm) exposures, and are responsible for nearly all of the mutagenic response in the rat and for a substantial portion of the mutagenic response in the mouse following high-level BD exposures.
Assuntos
Compostos de Epóxi/sangue , Compostos de Epóxi/urina , Glicóis/sangue , Glicóis/toxicidade , Glicóis/urina , Hipoxantina Fosforribosiltransferase/genética , Exposição por Inalação , Mutação/genética , Animais , Butadienos/toxicidade , Relação Dose-Resposta a Droga , Feminino , Hemoglobinas/metabolismo , Masculino , Camundongos , Ratos , Ratos Endogâmicos F344 , Reprodutibilidade dos Testes , Baço/citologia , Baço/efeitos dos fármacos , Baço/enzimologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/enzimologia , Fatores de TempoRESUMO
Inhalation of subclinical doses of sarin suppresses the antibody-forming cell (AFC) response, T-cell mitogenesis, and serum corticosterone (CORT) levels, and high doses of sarin cause lung inflammation. However, the duration of these changes is not known. In these studies, rats were exposed to a subclinical dose of sarin (0.4 mg/m3/h/day) for 1 or 5 days, and immune and inflammatory parameters were assayed up to 8 weeks before sarin exposure. Our results showed that the effects of a 5-day sarin exposure on the AFC response and T-cell receptor (TCR)-mediated Ca2+ response disappeared within 2-4 weeks after sarin exposure, whereas the CORT and adrenocorticotropin hormone (ACTH) levels remained significantly decreased. Pretreatment of rats with chlorisondamine attenuated the effects of sarin on the AFC and the TCR-mediated Ca2+ response, implicating the autonomic nervous system (ANS) in the sarin-induced changes in T-cell function. Moreover, exposure to a single or five repeated subclinical doses of sarin upregulated the mRNA expression of proinflammatory cytokines in the lung, which is associated with the activation of NFkappaB in bronchoalveolar lavage cells. These effects were lost within 2 weeks of sarin inhalation. Our results suggest that while sarin-induced changes in T cells and cytokine gene expression were short lived, suppression of CORT and ACTH levels were relatively long lived and might represent biomarkers of sarin exposure. Moreover, while the effects of sarin on T-cell function were regulated by the ANS, the decreased CORT levels by sarin might result from its effects on the hypothalamus-pituitary-adrenal axis.
Assuntos
Sistema Nervoso Autônomo/efeitos dos fármacos , Substâncias para a Guerra Química/toxicidade , Inibidores da Colinesterase/toxicidade , Inflamação/induzido quimicamente , Exposição por Inalação , Neuroimunomodulação/efeitos dos fármacos , Sistemas Neurossecretores/efeitos dos fármacos , Sarina/toxicidade , Hormônio Adrenocorticotrópico/sangue , Animais , Sistema Nervoso Autônomo/metabolismo , Biomarcadores/sangue , Cálcio/metabolismo , Clorisondamina/farmacologia , Inibidores da Colinesterase/administração & dosagem , Corticosterona/sangue , Citocinas/genética , Citocinas/metabolismo , Bloqueadores Ganglionares/farmacologia , Sistema Hipotálamo-Hipofisário/efeitos dos fármacos , Sistema Hipotálamo-Hipofisário/metabolismo , Inflamação/metabolismo , Inflamação/patologia , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Pulmão/patologia , Masculino , NF-kappa B/metabolismo , Sistemas Neurossecretores/metabolismo , Sistema Hipófise-Suprarrenal/efeitos dos fármacos , Sistema Hipófise-Suprarrenal/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Endogâmicos F344 , Receptores de Antígenos de Linfócitos T/efeitos dos fármacos , Receptores de Antígenos de Linfócitos T/metabolismo , Sarina/administração & dosagem , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo , Fatores de Tempo , Transcrição Gênica/efeitos dos fármacosRESUMO
Experiments were performed: (i) to investigate potential age- and gender-dependent differences in mutagenic responses in T cells following exposures of B6C3F1 mice and F344 rats by inhalation for 2 weeks to 0 or 1250 ppm butadiene (BD), and (ii) to determine if exposures for 2 weeks to 62.5 ppm BD produce a mutagenic effect in female rats. To evaluate the effect of age on mutagenic response, mutant manifestation curves for splenic T cells of female mice exposed at 8-9 weeks of age were defined by measuring Hprt mutant frequencies (MFs) at multiple time points after BD exposure using a T cell cloning assay and comparing the resulting mutagenic potency estimate (calculated as the difference of areas under the mutant manifestation curves of treated versus control animals) to that reported for female mice exposed to BD in the same fashion beginning at 4-5 weeks of age. The shapes of the mutant T cell manifestation curves for spleens were different [e.g., the maximum BD-induced MFs in older mice (8.0+/-1.0 [S.D.]x10(-6)) and younger mice (17.8+/-6.1 x 10(-6)) were observed at 8 and 5 weeks post-exposure, respectively], but the mutagenic burden was the same for both age groups. To assess the effect of gender on mutagenic response, female and male rodents were exposed to BD at 4-5 weeks of age and Hprt MFs were measured when maximum MFs are expected to occur post-exposure. The resulting data demonstrated that the pattern for mutagenic susceptibility from high-level BD exposure is female mice>male mice>female rats>male rats. Exposures of female rats to 62.5 ppm BD caused a minor but significant mutagenic response compared with controls (n=16/group; P=0.03). These results help explain part of the differing outcomes/interpretations of data in earlier Hprt mutation studies in BD-exposed rodents.
Assuntos
Envelhecimento/genética , Butadienos/administração & dosagem , Butadienos/toxicidade , Exposição por Inalação , Mutagênese/efeitos dos fármacos , Caracteres Sexuais , Linfócitos T/efeitos dos fármacos , Animais , Células Clonais , Intervalos de Confiança , Feminino , Hipoxantina Fosforribosiltransferase/genética , Masculino , Camundongos , Testes de Mutagenicidade , Mutagênicos/administração & dosagem , Mutagênicos/toxicidade , Proteínas Mutantes/genética , Mutação/genética , Ratos , Ratos Endogâmicos F344 , Especificidade da Espécie , Baço/citologia , Baço/efeitos dos fármacos , Baço/enzimologia , Linfócitos T/enzimologia , Linfócitos T/metabolismoRESUMO
The first epithelial surface encountered by inhaled materials is the epithelium of the respiratory tract. The epithelium is lined by a fluid (ELF) that can be sampled by a saline wash (lavage) of the area of interest. This technique, known as bronchoalveolar lavage (BAL), provides a means of sampling a body fluid that can provide valuable information on the reaction of the lung to inhaled materials. The most common responses measured are indicators of an inflammatory response, the most sensitive of which is an influx of neutrophils. In the extracellular fluid, levels of beta-glucuronidase activity indicate activation of macrophages, and lactate dehydrogenase activity indicates cytotoxicity. Other pro- and anti-inflammatory soluble factors that can be measured in BAL fluid include secretory products of macrophages and epithelial cells, such as tumor necrosis factor alpha, fibronectin, interleukin-1, various chemotactic factors (including IL-8, MIP-2), growth factors, proteases, and antiproteases. Oxidative stress can be measured by the levels of reduced glutathione in ELF, and increased levels of alkaline phosphatase indicate increased Type II cell secretions. Allergic responses are indicated by increased eosinophils and factors such as histamine and arachidonate metabolites in BAL fluid. BAL analysis can be used as a complementary technique with more traditional measures of lung injury, such as histopathology or radiology. The advantage of BAL analysis is that one can pick up early indicators of biochemical changes leading to later morphological changes in a disease process. A second advantage is that the BAL fluid analyses are quantitative, and dose-response measures can be obtained. In large animals, one can do repeated lavages to follow a disease process; in small animals, one can use serial sacrifices in similarly exposed rodents to achieve the same goal. Research related to the use of BAL fluid analyses to detect lung damage has been conducted at the Lovelace Respiratory Research Institute with funding from various sources including the US Department of Energy and the US Environmental Protection Agency.
Assuntos
Líquido da Lavagem Broncoalveolar/citologia , Lavagem Broncoalveolar/métodos , Exposição por Inalação/efeitos adversos , Mucosa Respiratória/efeitos dos fármacos , Sistema Respiratório/efeitos dos fármacos , Doenças Respiratórias/induzido quimicamente , Animais , Animais de Laboratório , Líquido da Lavagem Broncoalveolar/química , Mucosa Respiratória/patologia , Sistema Respiratório/patologia , Doenças Respiratórias/patologiaRESUMO
1,3-Butadiene (BD) is a monomer produced in petrochemical production facilities and from several combustion sources. The United States Environmental Protection Agency has defined BD as a probable human carcinogen. Methods for assessing exposure and internal dose are therefore of critical interest, and one technique is the measurement of urinary metabolites. Here we describe methods for measuring two urinary metabolites, N-acetyl-S-(3,4-dihydroxybutyl)-L-cysteine (referred to as MI) and an isomeric mixture of the regio- and stereoisomers (R)/(S)-N-acetyl-S-(1-(hydroxymethyl)-2-propen-yl)-L-cysteine and (R)/(S)-N-acetyl-S-(2-hydroxy-3-butenyl)-L-cysteine (referred to as MII). The method is based on isolation of the metabolites by solid-phase extraction and measurement using liquid chromatography and triple quadrupole mass spectrometry (LC-MS(3)). The LC-MS(3) allowed good selectivity with minimal sample preparation. Assay accuracy was within 10% or better, with substantial improvement in accuracy accompanying the commercial availability of deuterated internal standards for both compounds. Assay precision and linearity passed rigorous validation criteria, and precision-based limits of quantitation values were 12 and 1 ng/mL for MI and MII, respectively. Data are shown from analysis of human urine from occupationally exposed individuals and rat urine from BD exposures conducted to investigate rodent metabolic profiles. Both of these data sets clearly show that this assay can discern previously described relationships between BD exposure and the production of MI/MII.
Assuntos
Acetilcisteína/análogos & derivados , Acetilcisteína/urina , Poluentes Ocupacionais do Ar/farmacocinética , Butadienos/farmacocinética , Administração por Inalação , Animais , Bioensaio , Biomarcadores/urina , Butadienos/administração & dosagem , Cromatografia Líquida/métodos , Feminino , Humanos , Isomerismo , Espectrometria de Massas/métodos , Exposição Ocupacional , Ratos , Ratos Endogâmicos F344 , Reprodutibilidade dos Testes , EstereoisomerismoRESUMO
This unit describes how bronchoalveolar lavage can be used in laboratory animals to sample the epithelial lining fluid of the lung for information on the degree of pulmonary inflammation induced by exposure to an airborne toxicant. The technique allows quantitative assessment of inflammatory responses and is valuable for providing dose-response information in exposed animals. Lavage fluid samples may also be used for proteomic analyses, and the protein expression profiles may be used to address specific mechanistic questions.
Assuntos
Líquido da Lavagem Broncoalveolar , Lesão Pulmonar/etiologia , AnimaisRESUMO
A multiinstitutional, transitional epidemiologic study was conducted with a worker population in the Czech Republic to evaluate the utility of a continuum of non-disease biological responses as biomarkers of exposure to 1,3-butadiene (BD)* in an industrial setting. The study site included two BD facilities in the Czech Republic. Institutions that collaborated in the study were the University of Vermont (Burlington, Vermont, USA); the Laboratory of Genetic Ecotoxicology (Prague, the Czech Republic); Shell International Chemicals, BV (Amsterdam, The Netherlands); the University of North Carolina at Chapel Hill (Chapel Hill, North Carolina, USA); University of Texas Medical Branch at Galveston (Galveston, Texas, USA); Leiden University (Leiden, The Netherlands); and the Health and Safety Laboratory (Sheffield, United Kingdom). Male volunteer workers (83) participated in the study: 24 were engaged in BD monomer production, 34 in polymerization activities, and 25 plant administrative workers served as unexposed control subjects. The BD concentrations experienced by each exposed worker were measured by personal monitor on approximately ten separate occasions for 8-hour workshifts over a 60-day exposure assessment period before biological samples were collected. Coexposures to styrene, benzene, and toluene were also measured. The administrative control workers were considered to be a homogeneous, unexposed group for whom a series of 28 random BD measurements were taken during the exposure assessment period. Questionnaires were administered in Czech to all participants. At the end of the exposure assessment period, blood and urine samples were collected at the plant; samples were. fractionated, cryopreserved, and kept frozen in Prague until they were shipped to the appropriate laboratories for specific biomarker analysis. The following biomarkers were analyzed: * polymorphisms in genes involved in BD metabolism (Prague and Burlington); * urinary concentrations of 1-hydroxy-2-(N-acetylcysteinyl)-3-butene and 2-hydroxy-1-(N-acetylcysteinyl)-3-butene (M2 [refers to an isomeric mixture of both forms]) (Amsterdam); * urinary concentrations of 1,2-dihydroxy-4-(N-acetylcysteinyl)-butane (M1) (Amsterdam); * concentrations of the hemoglobin (Hb) adducts N-(1-[hydroxymethyl]-2-propenyl)valine and N-(2-hydroxy-3-butenyl)valine (HBVal [refers to an isomeric mixture of both forms]) (Amsterdam); * concentrations of the Hb adduct N-(2,3,4-trihydroxybutyl)valine (THBVal) (Chapel Hill); * T cell mutations in the hypoxanthine phosphoribosyltransferase (HPRT) gene (autoradiographic assay in Galveston with slide review in Burlington; cloning assay in Leiden with mutational spectra determined in Burlington); and * chromosomal aberrations by the conventional method and by fluorescence in situ hybridization [FISH]), and cytogenetic changes (sister chromatid exchanges [SCEs] (Prague). All assay analysts were blinded to worker and sample identity and remained so until all work in that laboratory had been completed and reported. Assay results were sent to the Biometry Facility in Burlington for statistical analyses. Analysis of questionnaire data revealed that the three exposure groups were balanced with respect to age and years of residence in the district, but the control group had significantly more education than the other two groups and included fewer smokers. Group average BD exposures were 0.023 mg/m3 (0.010 ppm) for the control group, 0.642 mg/m3 (0.290 ppm) for the monomer group, and 1.794 mg/m3 (0.812 ppm) for the polymer group; exposure levels showed considerable variability between and within individuals. Styrene exposures were significantly higher in the polymer group than in the other two groups. We found no statistically significant differences in the distributions of metabolic genotypes over the three exposure groups; genotype frequencies were consistent with those previously reported for this ethnic and national population. Although some specific genotypes were associated with quantitative differences in urinary metabolite concentrations or Hb adduct dose-response characteristics, none indicated a heightened susceptibility to BD. Concentrations of both the M2 and M1 urinary metabolites and both the HBVal and THBVal Hb adducts were significantly correlated with group and individual mean BD exposure levels; the Hb adducts were more strongly correlated than the urinary metabolites. By contrast, no significant relations were observed between BD exposures and HPRT gene mutations (whether determined by the auto-radiographic or the cloning method) or any of the cytogenetic biomarkers (whether determined by the conventional method or FISH analysis). Neither the mutational nor the cytogenetic responses showed any association with genotypes. The molecular spectrum of HPRT mutations in BD-exposed workers showed a high frequency of deletions; but the same result was found in the unexposed control subjects, which suggests that these were not due to BD exposure. This lack of association between BD exposures and genetic effects persisted even when control subjects were excluded from the analyses or when we conducted regression analyses of individual workers exposed to different levels of BD.
Assuntos
Biomarcadores/análise , Butadienos/sangue , Butadienos/urina , Exposição Ocupacional/análise , Animais , Benzeno/análise , Benzeno/metabolismo , Butadienos/metabolismo , República Tcheca/epidemiologia , Genótipo , Hemoglobinas/efeitos dos fármacos , Humanos , Hipoxantina Fosforribosiltransferase/genética , Indústrias , Linfócitos/ultraestrutura , Masculino , Mutação , Exposição Ocupacional/estatística & dados numéricos , Polimorfismo Genético , Ratos , Estireno/análise , Estireno/metabolismo , Tolueno/análise , Tolueno/metabolismoRESUMO
Coexposure to subclinical levels of nerve gas and to heat stress may have induced some of the clinical symptoms of the Gulf War Syndrome. We tested the hypothesis that single or repeated subclinical exposure to sarin, particularly under conditions of heat stress, would impair regulation of body temperature and locomotor activity. Male F344 rats were housed at 25 degrees C or under mild heat stress at 32 degrees C and were exposed 1 h/day for 1, 5, or 10 days to 0, 0.2, or 0.4 mg/m(3) of sarin in a nose-only exposure system. Body temperature and activity were monitored continuously by telemetry during exposure and 1 month postexposure. Exposed rats showed no clinical symptoms of toxicity such as tremors, despite evidence of reduced red blood cell cholinesterase activity. Heat stress consistently elevated body temperature in unexposed animals, particularly during the dark period when animals are most active. Inhalation of sarin gas at the two subclinical levels did not affect body temperature acutely in a biologically meaningful manner after the first exposure nor after 5 or 10 repeated exposures, either at thermoneutral ambient temperature or during chronic heat stress. There were no consistent effects of sarin or housing temperature on activity. The data suggest that subclinical levels of sarin have minimal effects on temperature regulation and locomotor activity under these observation conditions.
Assuntos
Temperatura Corporal/efeitos dos fármacos , Substâncias para a Guerra Química/toxicidade , Inibidores da Colinesterase/toxicidade , Modelos Animais de Doenças , Atividade Motora/efeitos dos fármacos , Síndrome do Golfo Pérsico/etiologia , Sarina/toxicidade , Animais , Masculino , Ratos , Ratos Endogâmicos F344RESUMO
The purpose of this study was to determine whether exposure to levels of sarin causing no overt clinical signs would cause more subtle, adverse health effects that persisted after the exposure ended. Inhalation exposures of male Fischer 344 rats to 0, 0.2, or 0.4 mg/m(3) of sarin for 1 h/day for 1, 5, or 10 days under normal (25 degrees C) and heat-stressed (32 degrees C) conditions were completed and observations were made at 1 day and 1 month after the exposures. The sarin exposures had no observed effects on body weight, respiration rate, and minute volume during exposure nor in body temperature and activity during the 30-day recovery period. There was no evidence of cellular changes in brain determined by routine histopathology nor of any increase in apoptosis. Brain mRNA for interleukin (IL)-1beta, tumor necrosis factor-alpha, and IL-6 was increased in a dose-dependent manner. Autoradiographic studies demonstrated that M1 cholinergic receptor site densities were unchanged at 1 day after repeated exposures with or without heat stress. At 30 days, there was a decrease in M1 receptors in the olfactory tubercle (with and without heat), and, with heat stress, M1 sites also decreased in a dose-dependent manner in the frontal cortex, anterior olfactory nucleus, and hippocampus. M3 receptor sites were not affected by sarin exposure alone. In the presence of heat stress, there was an upregulation in binding site densities in the frontal cortex, olfactory tubercle, anterior nucleus, and striatum immediately after exposure, and these effects persisted at 30 days. Although red blood cell acetylcholinesterase (AChE) was not greatly inhibited by the 1-day exposure, there were 30 and 60% inhibitions after repeated exposures at the low and high doses, respectively. Histochemical staining for AChE demonstrated that sarin exposure alone reduced AChE in the cerebral cortex, striatum, and olfactory bulb. Sarin exposure under heat stress reduced AChE staining in the hippocampus, an area important for memory function. Thus, repeated exposures under heat-stress conditions, to levels of sarin that would not be noticed clinically, resulted in delayed development of brain alterations in cholinergic receptor subtypes that may be associated with memory loss and cognitive dysfunction.
Assuntos
Substâncias para a Guerra Química/toxicidade , Inibidores da Colinesterase/toxicidade , Sarina/toxicidade , Acetilcolinesterase/metabolismo , Animais , Apoptose/efeitos dos fármacos , Temperatura Corporal/efeitos dos fármacos , Peso Corporal/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Encéfalo/enzimologia , Encéfalo/patologia , Citocinas/biossíntese , Pulmão/efeitos dos fármacos , Pulmão/fisiologia , Masculino , Ratos , Ratos Endogâmicos F344 , Receptores Muscarínicos/análise , Receptores Muscarínicos/efeitos dos fármacosRESUMO
The nerve gas sarin is a potent cholinergic agent, and exposure to high doses may cause neurotoxicity and death. Subclinical exposures to sarin have been postulated to contribute to the Gulf War syndrome; however, the biological effects of subclinical exposure are largely unknown. In this communication, evidence shows that subclinical doses (0.2 and 0.4 mg/m(3)) of sarin administered by inhalation to F344 rats for 1 h/day for 5 or 10 days inhibited the anti-sheep red blood cell antibody-forming cell response of spleen cells without affecting the distribution of lymphocyte subpopulations in the spleen. Moreover, sarin suppressed T cell responses, including the concanavalin A (Con A) and the anti-alphabeta-T cell receptor (TCR) antibody-induced T cell proliferation and the rise in the intracellular calcium following TCR ligation. These concentrations of sarin altered regional but not total brain acetylcholinesterase activity. Interestingly, serum corticosterone levels of the sarin-treated animals were dramatically lower than the control animals, indicating that sarin-induced immunosuppression did not result from the activation of the hypothalamus-pituitary-adrenal (HPA) axis. Pretreatment of animals with the ganglionic blocker chlorisondamine abrogated the inhibitory effects of sarin on spleen cell proliferation in response to Con A and anti-TCR antibodies. These results suggest that the effects of sarin on T cell responsiveness are mediated via the autonomic nervous system and are independent of the HPA axis.
Assuntos
Sistema Nervoso Autônomo/efeitos dos fármacos , Substâncias para a Guerra Química/toxicidade , Inibidores da Colinesterase/toxicidade , Sarina/toxicidade , Linfócitos T/efeitos dos fármacos , Animais , Células Produtoras de Anticorpos/efeitos dos fármacos , Sistema Nervoso Autônomo/fisiologia , Cálcio/metabolismo , Clorisondamina/farmacologia , Corticosterona/sangue , Sistema Hipotálamo-Hipofisário/efeitos dos fármacos , Sistema Hipotálamo-Hipofisário/fisiologia , Ativação Linfocitária/efeitos dos fármacos , Masculino , Sistema Hipófise-Suprarrenal/efeitos dos fármacos , Sistema Hipófise-Suprarrenal/fisiologia , Ratos , Ratos Endogâmicos F344 , Receptores de Antígenos de Linfócitos T/fisiologia , Linfócitos T/imunologiaRESUMO
BACKGROUND: Surfactant abnormalities have been described in bacterial pneumonia. OBJECTIVE: To determine the safety and effect of exogenous surfactant replacement in patients with ventilator-associated pneumonia (VAP). METHODS: Patients with VAP were randomized in a double-blind study to receive either an artificial surfactant (Exosurf) consisting mostly of disaturated phospholipids (DSPL) or saline via a continuous nebulizer system for 5 days. Patients underwent bronchoscopy and bronchoalveolar lavage (BAL) prior to and after 4 days of therapy. RESULTS: Twenty-two patients were randomized, with 8 receiving Exosurf. There was no detected difference in outcome between the saline- and Exosurf-treated patients in terms of days on ventilator, 30-day or hospital mortality. At the follow-up lavage, the patients treated with Exosurf had a significant rise in the level of DSPL (p < 0.05), while the saline group did not, suggesting delivery of drug. Also at the follow-up lavage, the percentage of neutrophils in the BAL fell in the Exosurf patients (p < 0.01), but not in the saline group. CONCLUSION: Exogenous surfactant replacement given to patients with VAP increased the amount of DSPL retrieved by BAL. This treatment was associated with a fall in the neutrophil response to pneumonia.