Genotoxicity of the fungicide dichlofluanid in seven assays.
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Aniline Compounds↗
Biomedical subjects
J E Gallagher
Publications and source records attributed to J E Gallagher.
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↗
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↗
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↗
Colonization of the pharyngeal tonsil and respiratory tract of the gnotobiotic pig by a toxigenic strain of Pasteurella multocida type D.
Seven-day-old gnotobiotic pigs were inoculated intranasally with Pasteurella multocida and euthanatized 2, 5, 9, and 14 days after inoculation. Tissues from the oropharynx and respiratory tract of pigs were cultured quantitatively and analyzed microscopically. Pigs remained afebrile and alert, except one that died of acute fibrinopurulent pneumonia. Pasteurella multocida was isolated in greatest numbers from the pharyngeal tonsils, but only in low numbers from turbinate, trachea, lung, spleen, and liver. Significant histologic changes were limited to the tonsil. Infected pigs developed mild tonsillitis with lymphocytic hyperplasia, and accumulation of cell debris and bacteria in crypts. Capsular antigens of P. multocida, identified on tissue sections with rabbit anti-capsular polysaccharide antibody and immunocytochemical reagents, were confined to the crypt lumen. Ultrastructurally, bacteria were free within crypt material or within phagosomes of macrophages or neutrophils. In a second experiment, 5-day-old pigs were infected with Streptococcus suis type 2, followed by toxigenic Pasteurella multocida at 7 days of age; one pig died of streptococcal septicemia. Pigs developed a mild tonsillitis, and both bacteria were cultured from the tonsillar crypts for up to 14 days after infection. These studies show that a toxigenic strain of Pasteurella multocida, which is a causative agent of atrophic rhinitis, can colonize the tonsil and respiratory tract of gnotobiotic pigs for up to 14 days. In addition, colonization can occur concurrently with Streptococcus suis type 2.
Animals↗
1-Chloromethylpyrene: a reference skin sensitizer and genotoxin.
1-Chloromethylpyrene (1-CMP) has been evaluated as a model mutagen and toxin related to the ultimate electrophiles derived from benzo[a]pyrene and 1-nitropyrene. It was mutagenic to Salmonella (greater than 100 pg/plate) and exceptionally reactive to DNA when assessed by the 32P-postlabelling technique. 1-CMP was inactive in a mouse bone micronucleus assay when administered by gavage, probably due to hydrolysis, whose kinetics have been studied (t1/2 approximately 23 min at 37 degrees C). However, as expected, it was a potent skin toxin as determined by its activity as a mitogen to mouse skin and its contact allergenicity, as determined using the local lymph node proliferative assay. It is concluded that 1-CMP will probably be a potent human skin carcinogen and contact allergen.
Animals↗
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↗
Multiple DNA adducts in lymphocytes of smokers and nonsmokers determined by 32P-postlabeling analysis.
Identification of DNA adducts in peripheral lymphocytes could serve as a means of monitoring human exposure to potential genotoxic agents. In this study, DNA from peripheral lymphocytes of smokers and nonsmokers was examined for adducts by the P1 nuclease 32P-postlabeling technique. Thin layer chromatography (TLC) maps from both groups revealed multiple DNA adducts which ranged from no adducts for one individual to six adducts for a different individual. The total DNA adduct concentrations were approximately one adduct in 10(8)-10(10) normal nucleotides. Comparison of the adduct TLC profiles revealed individual variation in both pattern and level of DNA adducts. The type and amount of adduct was not influenced by smoking history and remained unchanged in four out of six subjects who were resampled after a 1 month interval. The capacity of lymphocytes to form BaP-derived DNA adducts after a 72 h incubation with 10(-6) M [3H]BaP was measured by both high-performance liquid chromatography (HPLC) and 32P-postlabeling analysis. The in vitro adduct values detected by [3H]nucleoside concentrations on HPLC ranged from 1 to 7 fmol adduct per micrograms DNA (3.3-23.3 adducts per 10(7) nucleotides). The [3H]nucleoside values were consistent with values obtained by 32P-postlabeling of the same sample (correlation coefficient of 0.88). No relationship was apparent between the capacity of lymphocytes to form a [3H]BaP-derived adduct in vitro and the concentration of any adduct, or total adducts present in untreated lymphocytes. These results suggest that multiple DNA adducts are present in lymphocytes from nonsmokers as well as smokers, although the profile and extent of these adducts can vary among individuals. The relationship of the lymphocyte DNA adducts detected in this study to human cancer susceptibility remains to be determined.
7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗
Differences in detection of DNA adducts in the 32P-postlabelling assay after either 1-butanol extraction or nuclease P1 treatment.
The use of nuclease P1 treatment and 1-butanol extraction to increase the sensitivity of the 32P-postlabelling assay for DNA adducts have been compared. Although similar results were obtained with the two methods for standard adducts formed with benzo[a]pyrene diol epoxide I (BPDE-I), nuclease P1 treatment resulted in a significant reduction in detection of major adducts from 1-amino-6-nitropyrene (1-amino-6-NP), 1-amino-8-nitropyrene (1-amino-8-NP), 2-aminofluorene (2-AF), 2-naphthylamine (2-NA) and 4-aminobiphenyl (4-ABP) modified DNAs, but not following the 32P-postlabelling analysis of 2-acetylaminofluorene (2-AAF) modified DNA. These results suggest that, at least initially, both modifications of the 32P-postlabelling assay should be used for the detection of unknown adducts or for adducts derived from nitroaromatics and aromatic amines.
1-Butanol↗
Toxicological evaluation of complex industrial wastes: implications for exposure assessment.
We evaluated a variety of short-term bioassays to construct a battery of tests that could be used for assessing the biological effects of potentially hazardous complex industrial wastes. Ten samples were studied for hepatotoxicity; these samples and an additional 5 were studied for mutagenicity. Although the data are limited to these samples, the results suggest that the Salmonella assay (strain TA98) or a prophage-induction assay (both in the presence of S9) in combination with determination of relative liver weight and levels of a set of serum enzymes in rats may provide a battery of tests suitable to characterize complex industrial wastes for mutagenic and hepatotoxic potential. The biological activities exhibited by the wastes were not readily predicted by the chemical profiles of the wastes, emphasizing the importance of characterizing potentially hazardous complex industrial wastes by both chemical and biological means. DNA from liver, lung and bladder of rats exposed to some of the wastes was analyzed by the 32P-postlabeling technique for the presence of DNA adducts. A waste that produced mutagenic urine produced a DNA adduct in bladder DNA. The implications of this approach for assessment of exposure to complex hazardous waste mixtures are discussed.
Animals↗
Humeral medullary nailing--a new implant.
A new technique of medullary nailing of the humerus using a screw-headed nail is described. Eighteen patients with two-part fractures of the surgical neck of the humerus were reviewed. Fifteen had satisfactory results while three were unsatisfactory. Nine patients with fractures of the shaft of the humerus were also treated with satisfactory results in eight. Three patients with non-union of the humerus had unsatisfactory results.
Adolescent↗
Sialic acid mediates the initial binding of positively charged inorganic particles to alveolar macrophage membranes.
Pulmonary macrophages phagocytize inhaled particles and are postulated to play a role in the development of pulmonary interstitial fibrogenesis. The basic biologic mechanisms through which inhaled particles bind to macrophage membranes and subsequently are phagocytized remain unclear. We hypothesize that positively charged particles bind to negatively charged sialic acid (SA) residues on macrophage membranes. Alveolar Macrophages (AM) were collected by saline lavage from normal rat lungs. The cells adhered to plastic coverslips in serum-free phosphate buffered saline at 37 degrees C for 45 min and then were maintained at 4 degrees C for the binding experiments. Even distribution of SA groups on AM surfaces was demonstrated by scanning electron microscopy of wheat germ agglutinin (WGA) conjugated to 50 nm gold spheres. The WGA is a lectin that binds specifically to sialic acid, and pretreatment of AM with this lectin prevented the binding of positively charged carbonyl iron (C-Fe) spheres, aluminum (Al) spheres, and chrysotile asbestos fibers to AM surfaces. Limulus protein, another lectin with binding specificity for SA, similarly blocked the binding of positively charged spheres and chrysotile asbestos fibers but not negatively charged glass spheres or crocidolite asbestos fibers. Con A and ricin, lectins that bind to mannose and galactose residues, respectively, did not block particle binding. When both positively charged iron spheres and negatively charged glass spheres were prebound to AM membranes, subsequent treatment with WGA displaced only the positively charged spheres from macrophage surfaces. Con A and ricin had no effect on prebound positively charged C-Fe and Al spheres.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Humeral medullary nailing--a new implant.
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Adult↗
Binding of iron beads to sialic acid residues on macrophage membranes stimulates arachidonic acid metabolism.
Phagocytosis of carbonyl iron beads by rat alveolar macrophages induces the production of cyclooxygenase and lipoxygenase metabolites of arachidonic acid (AA). To study the relationship between the phagocytic event and AA metabolite production, rat alveolar macrophages were pretreated with cytochalasin D, which suppresses particle internalization, but not binding to the plasma membrane. The cells were then challenged with iron particles. Binding of the particles, without internalization, was a stimulus sufficient to initiate the AA metabolic cascade; this was true for cyclo- as well as for lipoxygenase pathways. To characterize the receptor responsible for iron bead binding to membranes, macrophages were pretreated with the lectin, wheat germ agglutinin. This treatment suppressed iron bead binding, phagocytosis, and the production of AA metabolites. Succinylated wheat germ agglutinin, which binds only to N-acetylglucosamine, prevented none of these events. Wheat germ agglutinin alone had a stimulatory activity on AA metabolism which was biphasic in nature, i.e., production of cyclooxygenase metabolites first and then leukotriene B4 upon rechallenge. Succinylated wheat germ agglutinin was devoid of such an effect on AA metabolism. Thus, AA metabolite production by alveolar macrophages during phagocytosis seems to be stimulated by initial binding of particles to the plasma membrane. Membrane-bound sialic acid residues appear to be instrumental in binding of carbonyl iron beads with consequent initiation of the AA metabolic cascade.
Acetylglucosamine↗
Irreducible trans-scaphoid perilunate dislocation.
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Adult↗
Development of a nonneuronal cell line derived from porcine spinal ganglia.
A cell line from porcine spinal ganglion (PSPG) tissue was initiated and characterized. The PSPG cell line contained epithelioid, fibroblast-like, and stellate-like cells. There were numerous intercellular communications between cells in the culture. Neuronal cells were not present in the PSPG cell line. The PSPG cell line was aneuploid at the 5th and 45th subculture levels. Seven porcine viruses replicated in cultures of PSPG cells, and 2 pseudorabies virus strains infected the cells after being diluted 10(-7).
Animals↗
A new guard for Kirschner wires.
Means of providing adequate protective cover for Kirschner wires protruding through the skin were investigated. A new guard was developed which satisfactorily abolishes the difficulties associated with other forms of cover.
Fracture Fixation, Internal↗
Effects of microbial isolates on porcine tracheal and bronchial explant cultures as observed by scanning electron microscopy.
Porcine tracheal and bronchial explant cultures exposed to log-phase cultures of Bordetella, Candida, Corynebacterium, Haemophilus, Klebsiella, Mycoplasma, Pasteurella, Proteus, Saccharomyces, Staphylococcus and Streptococcus were examined for surface sequential changes by scanning electron and light microscopy. Infected tissues, observed microscopically, had diminution or cessation of ciliary activity, and histologically had exfoliation of cilia, ciliocytophthoria, elevation of cellular borders, and cellular detachment. Treatment of these tissues with sterile medium containing penicillin and streptomycin did not prevent death or alteration of cells with increasing periods of incubation. The potential value of using scanning electron microscopy with explant cultures for studying organization of cellular surfaces in association with microbial growth and pathogenesis was demonstrated.
Bacteria↗