Induction of hepatic cytochromes P450 in dogs exposed to a chronic low dose of polychlorinated biphenyls.
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Induction of cytochrome P450 isoforms, specifically CYP1A1, and their catalytic activities are potential biomarkers of environmental contamination by polychlorinated biphenyls (PCBs). In this study, dogs were exposed to 25 ppm or 5 ppm Aroclor 1248 (PCB mixture) daily in their diet for 10 or 20 weeks, respectively. Relative to controls, hepatic microsomes from dogs dosed with PCBs had higher levels of CYP1A1 detected in immunoblots and higher levels of EROD activity, but low levels of induction for CYP2B and PROD activity. Concentrations of 96 PCB congeners in serum and liver were evaluated using capillary chromatography. Results showed that all dogs exposed to PCB mixtures had higher levels of PCB in serum and liver. Dogs preferentially sequestered highly chlorinated PCB congeners in liver relative to serum. With these experiments, we demonstrated that EROD activity was a potentially sensitive marker of PCB exposure at 5 and 25 ppm. Furthermore, CYP1A1 and EROD activity were maximally induced in dogs consuming dietary concentrations only 2.5 times the maximal permissible level for human food (FDA). The value of CYP1A1 induction as a biomarker of PCB exposure was tenuous because neither CYP1A1 levels nor EROD activity correlated with total PCB body burden. However, a small subset of congeners were identified in liver that may strongly influence EROD and PROD induction. Finally, two dogs in the 25 ppm dose group were fasted for 48 h. After 24 h of fasting, several new congeners appeared in the serum and remained in the serum for the remainder of the fast. The fast caused a 293% increase in PCB concentration in serum. This increase has strong implications regarding mobilization of toxic PCBs in wildlife during fasting (e.g., migration, hibernation). (+info)
Stimulation of pregnant rat uterine contraction by the polychlorinated biphenyl (PCB) mixture aroclor 1242 may be mediated by arachidonic acid release through activation of phospholipase A2 enzymes.
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The polychlorinated biphenyl (PCB) mixture Aroclor 1242 (A1242) increases frequency of contractions of pregnant rat uteri, suggesting a possible mechanism for decreased gestational age and increased spontaneous abortion in women and animals exposed to PCBs. In the present study, we hypothesized that A1242-induced stimulation of uterine contraction is mediated by arachidonic acid released by phospholipase A2 (PLA2) enzymes. Isometric uterine contraction was measured in longitudinal uterine strips isolated from gestation day 10 rat. Pretreatment of uterine strips with the PLA2 inhibitor (E)-6-(bromomethylene)tetrahydro-3-(1-naphthalenyl)-2H-pyran-2-one (HELSS) or manoalide, or an inhibitor of the G protein of PLA2, isotetrandrine, completely prevented the increase of contractile frequency induced by 50 microM A1242. However, the phospholipase C inhibitors 2-nitro-4-carboxyphenyl-N,N-diphenylcarbamate (NCDC) and neomycin were unable to block stimulation of uterine contraction by A1242. In accordance, A1242 (100 microM) did not release inositol phosphates from myo-[3H]inositol-labeled myometrial cells, whereas myometrial cells prelabeled with [3H]arachidonic acid released arachidonic acid in a concentration- and time-dependent manner after exposure to A1242 (10-100 microM). A1242 significantly stimulated arachidonic acid release in the absence of extracellular calcium, although the release was attenuated. Analysis of the eicosanoids released by A1242 indicated that only 0.83% of released [3H]arachidonic acid was metabolized to eicosanoids and 99.07% remained as free arachidonate. Uterine contraction increased in strips exposed to exogenous arachidonic acid (1-100 microM). This study suggests that A1242 stimulates contraction in pregnant rat uterus by a mechanism involving PLA2-mediated arachidonic acid release, and that arachidonic acid, rather than eicosanoids, may mediate A1242 uterotonic action in the uterus. (+info)
Establishment of a polychlorinated biphenyl-dechlorinating microbial consortium, specific for doubly flanked chlorines, in a defined, sediment-free medium.
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Estuarine sediment from Charleston Harbor, South Carolina, was used as inoculum for the development of an anaerobic enrichment culture that specifically dechlorinates doubly flanked chlorines (i.e., chlorines bound to carbon that are flanked on both sides by other chlorine-carbon bonds) of polychlorinated biphenyls (PCBs). Dechlorination was restricted to the para chlorine in cultures enriched with 10 mM fumarate, 50 ppm (173 microM) 2,3,4, 5-tetrachlorobiphenyl, and no sediment. Initially the rate of dechlorination decreased upon the removal of sediment from the medium. However, the dechlorinating activity was sustainable, and following sequential transfer in a defined, sediment-free estuarine medium, the activity increased to levels near that observed with sediment. The culture was nonmethanogenic, and molybdate, ampicillin, chloramphenicol, neomycin, and streptomycin inhibited dechlorination activity; bromoethanesulfonate and vancomycin did not. Addition of 17 PCB congeners indicated that the culture specifically removes double flanked chlorines, preferably in the para position, and does not attack ortho chlorines. This is the first microbial consortium shown to para or meta dechlorinate a PCB congener in a defined sediment-free medium. It is the second PCB-dechlorinating enrichment culture to be sustained in the absence of sediment, but its dechlorinating capabilities are entirely different from those of the other sediment-free PCB-dechlorinating culture, an ortho-dechlorinating consortium, and do not match any previously published Aroclor-dechlorinating patterns. (+info)
Differential binding affinities of PCBs, HO-PCBs, and aroclors with recombinant human, rainbow trout (Onchorhynkiss mykiss), and green anole (Anolis carolinensis) estrogen receptors, using a semi-high throughput competitive binding assay.
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A comparative study was undertaken to assess the ability of 44 polychlorinated biphenyls (PCBs), 9 hydroxylated PCBs (HO-PCBs), and 8 aroclors at concentrations ranging from 1 nM to 10 microM to compete with [3H]17beta-estradiol (E2) for binding to bacterially expressed fusion proteins using a semi-high throughput competitive-binding assay. The fusion proteins consisted of the D, E, and F domains of human (alpha), cloned reptilian (Anolis carolinensis) and recloned rainbow trout (Onchorhynkiss mykiss) estrogen receptors (ER) linked to the glutathione S-transferase (GST) protein. GST-hERalphadef (human), GST-aERdef (reptile) and GST-rtERdef (rainbow trout) fusion proteins exhibited high affinity for E2 with dissociation constants (Kd) of 0.4+/-0.1 nM, 0.7+/-0.2 nM, and 0.6+/-0.1 nM, respectively. Of the 44 PCBs examined, only PCBs 104, 184, and 188 effectively competed with [3H]E2 for binding to the GST-rtERdef protein with IC50 values ranging from 0.4-1.3 microM. In contrast, these same congeners only caused a 30% displacement of [3H]E2 in GST-hERalphadef and GST-aERdef proteins. Several additional congeners were found to bind to the GST-rtERdef fusion protein, although the degree of interaction varied among congeners. Among the HO-PCBs, 2',3',4',5'-tetrachloro-4-biphenylol and 2,6,2',6'-tetrachloro-4-biphenylol bound to all three fusion proteins with IC50 values ranging from 0.1-0.3 microM. Dimethyl sulphoxide (DMSO) concentrations of 20% significantly increased the ability of PCBs 104, 184, and 188 to compete with [3H]E2 for binding to the GST-ERdef fusion proteins, whereas at 20% DMSO, a significant reduction in saturable binding was observed. These results demonstrate that ERs from different species exhibit differential ligand preferences and relative binding affinities for PCBs, which can be dramatically affected by DMSO concentration. (+info)
An assessment of neurotoxicity of aroclors 1016, 1242, 1254, and 1260 administered in diet to Sprague-Dawley rats for one year.
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As part of a comparative chronic toxicity/oncogenicity study of different Aroclors (1016, 1242, 1254, and 1260), neurotoxicity was assessed in male and female Sprague-Dawley rats using functional observational battery (FOB) and motor activity tests, and histopathologic evaluation of selected nervous system tissues. Doses varied by Aroclor and ranged from 25 to 200 ppm in the diet. Animals were evaluated prior to initiation of dosing and at 13, 26, 39, and 52 weeks of exposure. Clinical signs, body weights, and feed consumption were evaluated weekly. Data analysis of FOB and motor activity results revealed several instances where Aroclor-treated groups were different from control. However, these were considered incidental, as they lacked any consistent dose- or time-related pattern that would suggest Aroclor-induced neurotoxicity. The nonremarkable findings during each of the four assessments were supported by the absence of any treatment-related clinical signs or mortality. Decreased body weight gain was evident in the male 100 ppm Aroclor 1254 dose group and in all female Aroclor 1254 dose groups late in the study (when a linear relationship was assumed between body weight and time), correlating with decreased feed consumption. Although a variety of incidental, spontaneous, degenerative changes were found in nervous tissue evaluated histopathologically, these changes were seen with similar incidence and severity in treated and control groups. No lesions were found that could be attributed to Aroclor-related neurotoxicity. In summary, 52 weeks of exposure to Aroclors 1016, 1242, 1254, or 1260 mixed in the diet did not yield any functional or morphologic changes indicative of PCB-induced neurotoxicity. (+info)
Inhibition of rat testicular androgenesis by a polychlorinated biphenyl mixture aroclor 1248.
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Polychlorinated biphenyls (PCBs) are complex mixtures of congeners that exhibit carcinogenic and toxicant activities in a variety of mammalian tissues. Here, we studied the acute in vivo and in vitro effects of a commercially used PCB product, Aroclor 1248 (A1248), a mixture of tri-, tetra-, and pentachloro congeners. Single intraperitoneal (i.p.) or bilateral intratesticular (i.t.) injections of A1248 decreased serum androgen levels in both groups 24 h after injection. Chorionic gonadotropin-stimulated androgen production by acute testicular cultures from both groups was also reduced, and progesterone production was attenuated in cultures from i.t.-treated animals. The capacity of the postmitochondrial fractions from testes of i.t.-treated animals to convert pregnenolone to progesterone and progesterone to testosterone was reduced as well. In vitro studies revealed that a 10- to 15-min exposure of postmitochondrial testicular fractions and intact interstitial cells from normal animals to A1248 in a subnanomolar concentration range was sufficient to attenuate the conversion of pregnenolone to progesterone and progesterone to testosterone. At micromolar concentrations, A1248 added in vitro also inhibited the conversion of Delta(4)-androstendione to testosterone without affecting the viability of interstitial cells. These results indicate that A1248 down-regulates the testicular androgenesis by an acute inhibition of 3beta-hydroxysteroid dehydrogenase, 17alpha-hydroxylase/lyase, and 17beta-hydroxysteroid dehydrogenase activities. (+info)
Acute effects of polychlorinated biphenyl-containing and -free transformer fluids on rat testicular steroidogenesis.
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Polychlorinated biphenyl (PCB)-based transformer fluids belong to a class of environmentally persistent mixtures with known toxic effects. Here, we studied the acute effects of Askarel (which contains Aroclor 1260) and two substitute transformer fluids (the silicone oil-based DC561 and the mineral oil-based ENOL C) on rat testicular steroidogenesis. Single intraperitoneal (ip; 10 mg/kg body weight) or bilateral intratesticular (itt; 25 microg/testis) injections of Askarel markedly decreased serum androgen levels 24 hr after administration. In acute testicular cultures from these animals, chorionic gonadotropin-stimulated progesterone and androgen productions were severely attenuated. When itt was injected or added in vitro, Askarel inhibited 3ss-hydroxysteroid dehydrogenase (3ssHSD), stimulated 17[alpha]-hydroxylase/lyase (P450c17), and did not affect 17ss-hydroxysteroid dehydrogenase in testicular postmitochondrial fractions. The ip-injected Askarel did not affect 3ssHSD, but inhibited P450c17, suggesting that a more intensive metabolism of peripherally injected Askarel reduces the circulating levels of active ingredients below the threshold needed for inhibition of 3ssHSD and generates a derivative that inhibits P450c17. In contrast to Askarel, itt-injection (25 microg/testis) of DC561 and ENOL C did not affect in vivo and in vitro steroidogenesis. These findings show the acute effects of Askarel, but not silicone and mineral oils, on testicular steroidogenesis. (+info)
Effects of subchronic exposure to complex mixtures of dioxin-like and non-dioxin-like polyhalogenated aromatic compounds on thyroid hormone and vitamin A levels in female Sprague-Dawley rats.
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The aim of this study was to determine the effects of subchronic exposure to complex mixtures of polyhalogenated aromatic hydrocarbons (PHAHs) on the thyroid hormone and retinoid status in female Sprague-Dawley rats and to investigate the predictability of these effects by the toxic equivalency factor (TEF) concept. In the first experiment, the focus was on a complex dioxin-like PHAH mixture, which covered > 90% of the total toxic equivalents (TEQ) present in Baltic herring. In the second experiment, the contribution of non-dioxin-like polychlorinated biphenyls (PCBs) was investigated by testing the commercial PCB mixture Aroclor 1260, its 0-1 ortho and 2-4 ortho fractions and the reconstituted 0-4 ortho fraction. Hepatic retinoid levels were severely decreased ( approximately 70%) after treatment with the dioxin-like PHAH mixture, similar to the effect of a TEQ equivalent dose of 1 microg 2,3,7,8-TCDD/kg bw/week. However, the TEF concept failed to predict the effect on plasma retinol; a decrease (21%) was observed after treatment with the PHAH mixture, whereas an increase (21%) was found after treatment with TCDD. A more severe decrease of total thyroid hormone in plasma was observed after exposure to the PHAH mixture compared to treatment with TCDD ( approximately 60% vs. 38%). The discrepancy found between the predicted and observed effects for plasma retinol and thyroid hormone is possibly due to an additional effect of hydroxylated PCBs, formed from metabolizable PCBs present in the PHAH mixture. Aroclor 1260 and its fractions did not significantly alter the retinoid and thyroid hormone status at the dose levels tested, indicating that in case of exposure to complex PCB mixtures at environmental levels, no effects, or at best, only marginal effects can be expected on the retinoid and thyroid hormone status. (+info)