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ABSTRACT: Sequential 28-day and 90-day oral toxicity studies were performed in male and female rats with ammonium perfluorobutyrate (NH(4)(+)PFBA) at doses up to 150 and 30mg/kg-d, respectively. Ammonium perfluorooctanoate was used as a comparator at a dose of 30mg/kg-d in the 28-day study. Female rats were unaffected by NH(4)(+)PFBA. Effects in males included: increased liver weight, slight to minimal hepatocellular hypertrophy; decreased serum total cholesterol; and reduced serum thyroxin with no change in serum thyrotropin. During recovery, liver weight, histological, and cholesterol effects were resolved. Results of RT-qPCR were consistent with increased transcriptional expression of the xenosensor nuclear receptors PPARα and CAR as well as the thyroid receptor, and decreased expression of Cyp1A1 (Ah receptor-regulated). No observable adverse effect levels (NOAELs) were 6 and >150mg/kg-d for male and female rats in the 28-day study and 6 and >30mg/kg-d in the 90-dat study, respectively.
Reproductive Toxicology 08/2011; 33(4):513-30. · 3.23 Impact Factor
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ABSTRACT: Doxorubicin (Adriamycin) is a potent and broad-spectrum antineoplastic agent, the clinical utility of which is restricted by a cumulative and progressive cardiomyopathy that develops with repeated dosing. Fundamental to the cardiac failure is an interference with mitochondrial respiration and inhibition of oxidative phosphorylation. Global gene expression arrays in cardiac tissue indicate that inhibition of mitochondrial oxidative phosphorylation by doxorubicin (DOX) is accompanied by a decreased expression of genes related to aerobic fatty acid oxidation and a corresponding increase in expression of genes involved in anaerobic glycolysis, possibly as an alternate source for ATP production. The aim of this investigation was to determine whether this is also manifest at the metabonomic level as a switch in metabolic flux in cardiac tissue, and whether this can be averted by co-administering the cardioprotective drug, dexrazoxane (DZR). (13)C-isotopomer analysis of isolated perfused hearts from male Sprague-Dawley rats receiving 6 weekly s.c. injections of 2mg/kg DOX demonstrated a shift from the preferential oxidation of fatty acids to enhanced oxidation of glucose and lactate plus pyruvate, indicative of a compensatory shift towards increased pyruvate dehydrogenase activity. Substrate-selective isotopomer analysis combined with western blots indicate an inhibition of long-chain fatty acid oxidation and not MCAD activity or fatty acyl-carnitine transport. Co-administering DZR averted many treatment-related changes in cardiac substrate metabolism, consistent with DZR being an effective cardioprotective agent against DOX-induced cardiomyopathy. This switch in substrate metabolism resembles that described for other models of cardiac failure; accordingly, this change in metabolic flux may represent a general compensatory response of cardiac tissue to imbalances in bioenergetic demand and supply, and not a characteristic unique to DOX-induced cardiac failure itself.
Toxicology 04/2010; 270(2-3):92-8. · 3.68 Impact Factor
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ABSTRACT: Perfluorooctanesulfonate (PFOS), a persistent and accumulative compound, is widely distributed in humans and wildlife. Human exposure can occur early in development, as evidenced by the detection of PFOS in umbilical cord blood and breast milk. As part of a developmental neurotoxicology study for which developmental endpoints, including those related to the developing nervous system, have been reported separately, groups of 25 pregnant Sprague Dawley rats were given daily oral doses of either vehicle control or potassium PFOS (K(+)PFOS) at 0.1, 0.3, and 1.0mg/kg-d from gestation day (GD) 0 (day positive for mating) through postnatal day (PND) 20. An additional 10 pregnant females per treatment group were treated through GD 19 and sacrificed on GD 20 in order to obtain maternal and fetal serum and tissue samples at the end of gestation. The present paper reports the results of samples of serum, liver, brain, and thyroid glands taken at various times to evaluate: (1) serum, liver, and brain PFOS concentrations by LC-MS/MS to establish the relationship between PFOS concentrations and study outcomes; (2) serum thyrotropin (TSH) concentrations by RIA; (3) thyroid follicular cell proliferation index by Ki-67 immunohistochemical staining; (4) thyroid follicle epithelial cell height and colloidal area by histomorphometric analysis; (5) selected liver mRNA transcripts by quantitative RT-PCR. PFOS concentrations in dam and pup serum, liver, and brain increased across treatment groups in approximate proportion to the proportional increases in maternal K(+)PFOS dose, and sex differences in PFOS concentrations were not apparent in pups on PND 21. In pups from K(+)PFOS maternal dose groups on PND 72, serum PFOS had decreased to about 3 and 11% of PND 21 concentrations in males and females, respectively, and liver PFOS had decreased to about 17% of PND 21 concentrations in both sexes. Liver PFOS concentrations were approximately 0.6-0.8 times serum PFOS in GD 20 fetuses, and increased to about 2-4 times serum concentrations on PND 4 and 21. GD 20 fetal and PND 4 pup brain PFOS concentrations were approximately 33% of the corresponding serum concentrations, dropping to approximately 10% by PND 21, in contrast to dam brain PFOS concentrations, which were approximately 4-9% of serum PFOS concentrations. Compared to controls, Cyp2b2 mRNA was increased (2.8-fold) in the 1.0mg/kg-d treatment-group dams on GD 20. In male pups on PND 21, Cyp4A1, ACoA, and Cyp2b2 were increased 2.1-, 1.5-, and 1.8-fold, respectively, and Cyp7A1 was decreased 3.5-fold. Serum TSH and thyroid follicular morphology were not altered by K(+)PFOS treatment. The mean number of proliferating thyroid follicular cells was increased 2.1-fold over control in GD 20 female fetuses from 1.0mg/kg-d-treated dams, yet the highest individual count was similar to that of controls (116 versus 113 in controls).
Reproductive Toxicology 07/2009; 27(3-4):387-99. · 3.23 Impact Factor
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ABSTRACT: Perfluoroalkyl acids (PFAAs) are widely distributed and environmentally persistent agents whose potential toxicity is not yet fully characterized. Perfluorooctanoic acid (PFOA) and perfluorooctane sulfonic acid elicit a number of potential toxicities in rodents, the most prevalent of which are governed by activation of the peroxisome proliferator-activated receptor alpha (PPARalpha). The purpose of this investigation was twofold: (1) To conduct a structure-activity relationship study of the transcriptional activation of peroxisome proliferation in primary rat liver cell cultures for PFAA-related carboxylic and sulfonic acids of varying carbon chain length and (2) to explore whether this activity can be translated to human liver cells in culture. Exposure to PFOA caused a dose-dependent stimulation of the expression of acyl-CoA oxidase (Acox), Cte/Acot1, and Cyp4a1/11 transcripts that are indicative of peroxisome proliferation in primary rat hepatocytes. PFOA concentrations of 30 microM and above caused cell injury characterized by the expression of Ddit3. Perfluorobutanoic acid (PFBA), on the other hand, stimulated Acox, Cte/Acot1, and Cyp4a1/11 gene expression in primary rat hepatocytes only at concentrations of 100 microM and above. Neither PFOA nor PFBA at concentrations up to 200 microM stimulated PPARalpha-related gene expression in either primary or HepG2 human liver cells. These data demonstrate that (1) PFFAs cause a concentration- and chain length-dependent increase in expression of gene targets related to cell injury and PPARalpha activation in primary rat hepatocytes, (2) the sulfonates are less potent than the corresponding carboxylates in stimulating PPARalpha-related gene expression in rat hepatocytes, and (3) stimulation of PPARalpha-mediated gene transcription is a mechanism that is not shared by human liver cells, adding further substantiation that PPARalpha-dependent liver toxicity in rodents does not extrapolate to assessing human health concerns.
Toxicological Sciences 05/2009; 111(1):89-99. · 4.65 Impact Factor
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ABSTRACT: In utero exposure of laboratory rats to perfluorooctane sulfonate (PFOS, C(8)F(17)SO(3)(-)), a chemically stable surfactant that is widely disseminated in the environment and present in serum samples from wildlife and humans, is associated with decreased neonatal survival, and growth deficits as well as hepatomegaly. This hepatomegaly in newborn rats exposed to PFOS in utero resembles that observed in adults and is characterized by peroxisome proliferation and decreased liver triglycerides, both of which are suspected to be manifested through PPARalpha-mediated transcriptional regulation. The purpose of the present investigation was to determine whether these changes in metabolic status are a reflection of transcriptional changes in fetal rat liver using global gene expression array analyses. Gravid Sprague-Dawley rats were administered 3mg/kg PFOS by gavage daily from gestational day 2-20 and terminated on day 21. Although there was no treatment-related frank terata, there was a substantial effect of PFOS on the perinatal hepatic transcriptome-225 unique transcripts were identified as statistically increased and 220 decreased by PFOS exposure; few transcripts were changed by more than two-fold. Although the PPARalpha transcript (Ppara) itself was not affected, there was a significant increase in expression of gene transcripts associated with hepatic peroxisomal proliferation as well as those responsible for fatty acid activation, transport and oxidation pathways (both mitochondrial and peroxisomal). Additional metabolic pathways altered by in utero PFOS exposure were a stimulation of fetal hepatic fatty acid biosynthesis and a net reduction of Cyp7a1 transcript, which is required for bile acid synthesis. There were minimal effects on the expression of thyroid-related gene transcripts. In conclusion, gene expression analysis provides strong evidence indicating transcriptional control of the altered metabolic status of neonates following PFOS exposure in utero, much of which appears to be under the influence of a functional perinatal PPARalpha regulatory pathway.
Toxicology 10/2008; 251(1-3):8-20. · 3.68 Impact Factor
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Shugo Suzuki,
Lora L Arnold,
David Muirhead,
Xiufen Lu,
X Chris Le, James A Bjork,
Kendall B Wallace,
Takamasa Ohnishi,
Satoko Kakiuchi-Kiyota,
Karen L Pennington,
Samuel M Cohen
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ABSTRACT: Based on epidemiological data, chronic exposure to high levels of inorganic arsenic in the drinking water is carcinogenic to the urinary bladder of humans. Recently, models have been developed involving transplacental administration of inorganic arsenic and subsequent administration of another substance that produces a low incidence of urogenital neoplasms. Administration of arsenite or arsenate in the diet or drinking water to five-to eight-week-old mice or rats rapidly induces urothelial cytotoxicity and regenerative hyperplasia. In mice administered arsenite, we observed eosinophilic intracytoplasmic granules present in the urothelial cells. These granules were not present in urothelial cells of untreated mice or in treated or untreated rats. By transmission electron microscopy, the granules were located within the mitochondrial matrix, that is, mitochondrial inclusions. Arsenic, primarily as arsenite, was present in partially purified mitochondria containing these granules. Cells containing the granules were not usually associated with degenerative changes. Lack of these granules in rats suggests that they are not necessary for inorganic arsenic-induced urothelial cytotoxicity or hyperplasia. These granules have also been observed with exposures to other metals in other tissues and other species, suggesting that they represent a protective mechanism against metal-induced toxicity.
Toxicologic Pathology 02/2008; 36(7):999-1005. · 1.91 Impact Factor
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ABSTRACT: Perfluorooctanesulfonate (PFOS) is widely distributed and persistent in humans and wildlife. Prior toxicological studies have reported decreased total and free thyroid hormones in serum without a major compensatory rise in thyrotropin (TSH) or altered thyroid gland histology. Although these animals (rats, mice and monkeys) might have maintained an euthyroid state, the basis for hypothyroxinemia remained unclear. We undertook this study to investigate the causes for the PFOS-induced reduction of serum total thyroxine (TT4) in rats.
We hypothesized that exposure to PFOS may increase free thyroxine (FT4) in the rat serum due to the ability of PFOS to compete with thyroxine for binding proteins. The increase in FT4 would increase the availability of the thyroid hormone to peripheral tissues for utilization, metabolic conversation, and excretion. We also hypothesized that PFOS does not directly interfere with the regulatory functions of the hypothalamic-pituitary-thyroid (HPT) axis in rats.
Three experimental designs were employed to test these hypotheses. (1) Female Sprague-Dawley (SD) rats were given a single oral dose of 15 mg potassium PFOS/kg body weight. At intervals of 2, 6, and 24h thereafter, measurements were made for serum FT4, TT4, triiodothyronine (TT3), reverse triiodothyronine (rT3), thryrotropin (TSH), and PFOS concentrations, as well as liver PFOS concentrations, UDP-glucuronosyltransferase 1A (UGT1A) family mRNA transcripts, and malic enzyme (ME) mRNA transcripts and activity. (2) To provide evidence for increased uptake and metabolism of thyroxine (T4), 125 I-T4 was given to male and female SD rats by intravenous injection, followed in 2h by a single oral dose of 15 mg potassium PFOS/kg body weight. 125 I radioactivity was determined in urine and feces collected over a 24-h period and in serum and liver collected at 24h. (3) To assess the potentials effect of PFOS on the hypothalamic-pituitary-thyroid axis, over an 8-day period, groups of male SD rats were given PFOS (3mg/kg-d), propyl thiouracil (PTU, 10 microg/mL in water), or PTU and PFOS in combination, with controls receiving 0.5% Tween 20 vehicle. On days 1, 3, 7, and 8, TT4, TT3, and TSH were monitored. On day 8, pituitaries were removed and placed in static culture for assessment of thyrotropin releasing hormone (TRH)-mediated release of TSH.
(1) PFOS transiently increased FT4 and decreased TSH within 6h, with values returning to control levels by 24h. TT4 was decreased by 55% over a 24-h period. TT3 and rT3 were decreased at 24h to a lesser extent than TT4. ME mRNA transcripts were increased at 2h and activity was increased at 24h. UGT1A mRNA transcripts were increased at 2 and 6h. (2) 125 I decreased in serum and liver relative to controls and consistent with a reduction in serum TT4. Concomitantly, 125 I activity was increased in urine and feces collected from PFOS-treated rats. (3) During the 8 days of dosing with PFOS, TSH was not elevated in male rats, while TT4 and TT3 were decreased. Pituitary response to TRH-mediated TSH release was not diminished after 8-daily oral doses of PFOS.
These findings suggest that oral dosing in rats with PFOS results in transiently increased tissue availability of the thyroid hormones and turnover of T4 with a resulting reduction in serum TT4. PFOS does not induce a classical hypothyroid state under dosing conditions employed nor does it alter HPT activities.
Toxicology 02/2008; 243(3):330-9. · 3.68 Impact Factor
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ABSTRACT: Increased generation of reactive oxygen species (ROS) is implicated in "glucose toxicity" in diabetes. However, little is known about the action of glucose on the expression of transcription factors in hepatocytes, especially those involved in mitochondrial DNA (mtDNA) replication and transcription. Since mitochondrial functional capacity is dynamically regulated, we hypothesized that stressful conditions of hyperglycemia induce adaptations in the transcriptional control of cellular energy metabolism, including inhibition of mitochondrial biogenesis and oxidative metabolism. Cell viability, mitochondrial respiration, ROS generation and oxidized proteins were determined in HepG2 cells cultured in the presence of either 5.5 mM (control) or 30 mM glucose (high glucose) for 48 h, 96 h and 7 days. Additionally, mtDNA abundance, plasminogen activator inhibitor-1 (PAI-1), mitochondrial transcription factor A (TFAM) and nuclear respiratory factor-1 (NRF-1) transcripts were evaluated by real time PCR. High glucose induced a progressive increase in ROS generation and accumulation of oxidized proteins, with no changes in cell viability. Increased expression of PAI-1 was observed as early as 96 h of exposure to high glucose. After 7 days in hyperglycemia, HepG2 cells exhibited inhibited uncoupled respiration and decreased MitoTracker Red fluorescence associated with a 25% decrease in mtDNA and 16% decrease in TFAM transcripts. These results indicate that glucose may regulate mtDNA copy number by modulating the transcriptional activity of TFAM in response to hyperglycemia-induced ROS production. The decrease of mtDNA content and inhibition of mitochondrial function may be pathogenic hallmarks in the altered metabolic status associated with diabetes.
Toxicology and Applied Pharmacology 01/2008; 225(2):214-20. · 4.45 Impact Factor
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ABSTRACT: Decreases in serum total thyroxine (TT4) and free thyroxine (FT4) without a compensatory rise in thyroid stimulating hormone (thyrotropin or TSH) or histological changes of the thyroid have been observed in studies with perfluorooctanesulfonate (PFOS) treatments in rats. Prior observations do not fit the clinical profile of a hypothyroid state. PFOS is known to compete with fatty acids for albumin binding, and serum free fatty acids (FFA) are known to interfere with FT4 measurement using analog methods due to competition for protein binding. Therefore, we hypothesized that measured decreases in serum FT4 by analog methods in the presence of PFOS were due to carrier protein binding interference. We compared FT4 analog assay methods with a reference method using equilibrium dialysis (ED-RIA) for FT4 measurement in rat sera in vitro and in vivo. We also measured hepatic malic enzyme mRNA transcripts and activity as a marker for hepatic thyroid hormone response. PFOS did not reduce serum TT4 and FT4 in vitro at concentrations up to 200 microM. After three daily 5mg/kg oral doses of potassium PFOS to female rats, serum TSH and FT4 by ED-RIA were unchanged (although FT4 determined by two common analog methods was decreased), and malic enzyme was not suppressed. These data suggest that prior reports of reduced free thyroid hormone in the presence of PFOS were due to negative bias in analog methods and that short-term PFOS treatment does not suppress the physiological thyroid status in rats. A reference method such as ED-RIA should be used for determination of serum FT4 in the presence of PFOS.
Toxicology 06/2007; 234(1-2):21-33. · 3.68 Impact Factor
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ABSTRACT: The cardiotoxicity associated with doxorubicin (DOX) therapy limits the total cumulative dose and therapeutic success of active anticancer chemotherapy. Cardiac mitochondria are implicated as primary targets for DOX toxicity, which is believed to be mediated by the generation of highly reactive free radical species of oxygen from complex I of the mitochondrial electron transport chain. The objective of this study was to determine if the protection demonstrated by carvedilol (CV), a beta-adrenergic receptor antagonist with strong antioxidant properties, against DOX-induced mitochondrial-mediated cardiomyopathy [Toxicol. Appl. Pharmacol. 185 (2002) 218] is attributable to its antioxidant properties or its beta-adrenergic receptor antagonism. Our results confirm that DOX induces oxidative stress, mitochondrial dysfunction, and histopathological lesions in the cardiac tissue, all of which are inhibited by carvedilol. In contrast, atenolol (AT), a beta-adrenergic receptor antagonist lacking antioxidant properties, preserved phosphate energy charge but failed to protect against any of the indexes of DOX-induced oxidative mitochondrial toxicity. We therefore conclude that the cardioprotective effects of carvedilol against DOX-induced mitochondrial cardiotoxicity are due to its inherent antioxidant activity and not to its beta-adrenergic receptor antagonism.
Toxicology and Applied Pharmacology 11/2004; 200(2):159-68. · 4.45 Impact Factor
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ABSTRACT: The cardiotoxicity associated with doxorubicin (DOX) therapy limits the total cumulative dose and therapeutic success of active anticancer chemotherapy. Cardiac mitochondria are implicated as primary targets for DOX toxicity, which is believed to be mediated by the generation of highly reactive free radical species of oxygen from complex I of the mitochondrial electron transport chain. The objective of this study was to determine if the protection demonstrated by carvedilol (CV), a β-adrenergic receptor antagonist with strong antioxidant properties, against DOX-induced mitochondrial-mediated cardiomyopathy [Toxicol. Appl. Pharmacol. 185 (2002) 218] is attributable to its antioxidant properties or its β-adrenergic receptor antagonism. Our results confirm that DOX induces oxidative stress, mitochondrial dysfunction, and histopathological lesions in the cardiac tissue, all of which are inhibited by carvedilol. In contrast, atenolol (AT), a β-adrenergic receptor antagonist lacking antioxidant properties, preserved phosphate energy charge but failed to protect against any of the indexes of DOX-induced oxidative mitochondrial toxicity. We therefore conclude that the cardioprotective effects of carvedilol against DOX-induced mitochondrial cardiotoxicity are due to its inherent antioxidant activity and not to its β-adrenergic receptor antagonism.
Toxicology and Applied Pharmacology.
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ABSTRACT: Doxorubicin (Adriamycin®) is a potent and broad-spectrum antineoplastic agent, the clinical utility of which is restricted by a cumulative and progressive cardiomyopathy that develops with repeated dosing. Fundamental to the cardiac failure is an interference with mitochondrial respiration and inhibition of oxidative phosphorylation. Global gene expression arrays in cardiac tissue indicate that inhibition of mitochondrial oxidative phosphorylation by doxorubicin (DOX) is accompanied by a decreased expression of genes related to aerobic fatty acid oxidation and a corresponding increase in expression of genes involved in anaerobic glycolysis, possibly as an alternate source for ATP production. The aim of this investigation was to determine whether this is also manifest at the metabonomic level as a switch in metabolic flux in cardiac tissue, and whether this can be averted by co-administering the cardioprotective drug, dexrazoxane (DZR). 13C-isotopomer analysis of isolated perfused hearts from male Sprague–Dawley rats receiving 6 weekly s.c. injections of 2 mg/kg DOX demonstrated a shift from the preferential oxidation of fatty acids to enhanced oxidation of glucose and lactate plus pyruvate, indicative of a compensatory shift towards increased pyruvate dehydrogenase activity. Substrate-selective isotopomer analysis combined with western blots indicate an inhibition of long-chain fatty acid oxidation and not MCAD activity or fatty acyl-carnitine transport. Co-administering DZR averted many treatment-related changes in cardiac substrate metabolism, consistent with DZR being an effective cardioprotective agent against DOX-induced cardiomyopathy. This switch in substrate metabolism resembles that described for other models of cardiac failure; accordingly, this change in metabolic flux may represent a general compensatory response of cardiac tissue to imbalances in bioenergetic demand and supply, and not a characteristic unique to DOX-induced cardiac failure itself.
Toxicology.
