Toxicological Sciences - current issue

2009-06-15

COVER

2009-06-15

Editorial Board

2009-06-15

SUBSCRIPTIONS

2009-06-15

Nitric Oxide Synthase: "Enzyme Zero" in Air Pollution-Induced Vascular Toxicity

Campen, M. J. — 2009-06-15

Chiral Toxicology: It's the Same Thing...Only Different

Smith, S. W. — 2009-06-15

Chiral substances possess a unique architecture such that, despite sharing identical molecular formulas, atom-to-atom linkages, and bonding distances, they cannot be superimposed. Thus, in the environment of living systems, where specific structure-activity relationships may be required for effect (e.g., enzymes, receptors, transporters, and DNA), the physiochemical and biochemical properties of racemic mixtures and individual stereoisomers can differ significantly. In drug development, enantiomeric selection to maximize clinical effects or mitigate drug toxicity has yielded both success and failure. Further complicating genetic polymorphisms in drug disposition, stereoselective metabolism of chiral compounds can additionally influence pharmacokinetics, pharmacodynamics, and toxicity. Optically pure pharmaceuticals may undergo racemization in vivo, negating single enantiomer benefits or inducing unexpected effects. Appropriate chiral antidotes must be selected for therapeutic benefit and to minimize adverse events. Enantiomers may possess different carcinogenicity and teratogenicity. Environmental toxicology provides several examples in which compound bioaccumulation, persistence, and toxicity show chiral dependence. In forensic toxicology, chiral analysis has been applied to illicit drug preparations and biological specimens, with the potential to assist in determination of cause of death and aid in the correct interpretation of substance abuse and "doping" screens. Adrenergic agonists and antagonist, nonsteroidal anti-inflammatory agents, SSRIs, opioids, warfarin, valproate, thalidomide, retinoic acid, N-acetylcysteine, carnitine, penicillamine, leucovorin, glucarpidase, pesticides, polychlorinated biphenyls, phenylethylamines, and additional compounds will be discussed to illustrate important concepts in "chiral toxicology."

Safety Assessment of Biotechnology Products for Potential Risk of Food Allergy: Implications of New Research

Selgrade, M. K., Bowman, C. C., Ladics, G. S., Privalle, L., Laessig, S. A. — 2009-06-15

Food allergy is a potential risk associated with use of transgenic proteins in crops. Currently, safety assessment involves consideration of the source of the introduced protein, in silico amino acid sequence homology comparisons to known allergens, physicochemical properties, protein abundance in the crop, and, when appropriate, specific immunoglobulin E binding studies. Recently conducted research presented at an International Life Sciences Institute/Health and Environmental Sciences Institute–hosted workshop adds to the scientific foundation for safety assessment of transgenic proteins in five areas: structure/activity, serum screening, animal models, quantitative proteomics, and basic mechanisms. A web-based tool is now available that integrates a database of allergenic proteins with a variety of computational tools which could be used to improve our ability to predict allergenicity based on structural analysis. A comprehensive strategy and model protocols have been developed for conducting meaningful serum screening, an extremely challenging process. Several animal models using oral sensitization with adjuvant and one dermal sensitization model have been developed and appear to distinguish allergenic from non-allergenic food extracts. Data presented using a mouse model suggest that pepsin resistance is indicative of allergenicity. Certain questions remain to be addressed before considering animal model validation. Gel-free mass spectrometry is a viable alternative to more labor-intensive approaches to quantitative proteomics. Proteomic data presented on four nontransgenic varieties of soy suggested that if known allergen expression in genetically modified crops falls within the range of natural variability among commercial varieties, there appears to be no need to test further. Finally, basic research continues to elucidate the etiology of food allergy.

Toxicity Testing in the 21st Century: A View from the Pharmaceutical Industry

MacDonald, J. S., Robertson, R. T. — 2009-06-15

Compensatory Induction of Liver Efflux Transporters in Response to ANIT-Induced Liver Injury Is Impaired in FXR-Null Mice

Cui, Y. J., Aleksunes, L. M., Tanaka, Y., Goedken, M. J., Klaassen, C. D. — 2009-06-15

Alpha-naphthyl isothiocyanate (ANIT) is a hepatotoxicant that produces acute intrahepatic cholestasis in rodents. Farnesoid X receptor (FXR) and pregnane X receptor (PXR) are two major bile acid sensors in liver. The purpose of this study was to characterize the regulation of hepatic transporters by FXR and PXR during ANIT-induced liver injury. Wild-type, FXR-null, and PXR-null mice were administered ANIT (75 mg/kg, po) and evaluated 48 h later for hepatotoxicity and messenger RNA (mRNA) expression of basolateral uptake (sodium taurocholate–cotransporting polypeptide, organic anion transporting polypeptide [Oatp] 1a1, Oatp1a4, Oatp1b2) and efflux transporters (organic solute transporter [Ost] , Ostβ, multidrug resistance–associated protein [Mrp] 3, Mrp4), as well as canalicular transporters (bile salt export pump [Bsep], Mrp2, multidrug resistance protein 2 [Mdr2], ATPase, class I, type 8B, member 1 [Atp8b1]). Livers from wild-type and PXR-null mice had comparable multifocal necrosis 48 h after ANIT. However, ANIT-treated FXR-null mice have fewer and smaller necrotic foci than wild-type mice but had scattered single-cell hepatocyte necrosis throughout the liver. Serum alanine transaminase, alkaline phosphatase (ALP), and direct bilirubin were increased in all genotypes, with higher ALP levels in FXR-null mice. Serum and liver unconjugated bile acids were higher in ANIT-treated FXR-null mice than the other two genotypes. ANIT induced mRNA expression of Mdr2, Bsep, and Atp8b1 in wild-type and PXR-null mice but failed to upregulate these genes in FXR-null mice. mRNA expression of uptake transporters declined in livers of all genotypes following ANIT treatment. ANIT increased Ostβ and Mrp3 mRNA in livers of wild-type and PXR-null mice but did not alter Ostβ mRNA in FXR-null mice. In conclusion, FXR deficiency enhances susceptibility of mice to ANIT-induced liver injury, likely a result of impaired induction of hepatobiliary efflux transporters and subsequent hepatic accumulation of unconjugated bile acids.

Resveratrol Inhibits Dioxin-Induced Expression of Human CYP1A1 and CYP1B1 by Inhibiting Recruitment of the Aryl Hydrocarbon Receptor Complex and RNA Polymerase II to the Regulatory Regions of the Corresponding Genes

Beedanagari, S. R., Bebenek, I., Bui, P., Hankinson, O. — 2009-06-15

The CYP1A family of cytochrome P450s (CYPs), comprising CYP1A1, CYP1A2, and CYP1B1, plays a role in bioactivation of several procarcinogens to carcinogenic derivatives, and also in detoxification of several xenobiotic compounds. Resveratrol (3,4,5-trihydroxystelbine) is a naturally occurring compound that has been shown in a number of studies to inhibit the induction of CYP1A1 and CYP1B1 by dioxin (2,3,7,8-tetrachloro-dibenzo-p-dioxin), but the mechanism(s) of resveratrol inhibition is controversial. In the current study, 100nM dioxin treatment for 24, 48, and 72 h induced CYP1A1, CYP1A2, and CYP1B1 mRNA levels in the human breast cancer cell line MCF-7, and CYP1A1 and CYP1A2 mRNA levels in the human hepatocellular carcinoma cell line, HepG2. Simultaneous treatment with 10µM resveratrol significantly inhibited dioxin-induced mRNA expression levels of these genes in both cell lines. Our studies are novel in that we used the chromatin immunoprecipitation assay to assay dioxin-induced recruitment of the aryl hydrocarbon receptor (AHR), and aryl hydrocarbon nuclear translocator (ARNT) to the enhancer regions and recruitment of RNA polymerase II to the promoter regions, of the CYP1A1 and CYP1B1 genes in their natural chromosomal settings. These recruitments were significantly inhibited in cells cotreated with resveratrol. Our studies thus indicate that resveratrol inhibits dioxin induction of the CYP1 family members either by directly or indirectly inhibiting the recruitment of the transcription factors AHR and ARNT to the xenobiotic response elements of the corresponding genes. The reduced transcriptional factor binding at their enhancers then results in reduced pol II recruitment at the promoters of these genes.

Discrimination of Tumorigenic Triazole Conazoles from Phenobarbital by Transcriptional Analyses of Mouse Liver Gene Expression

Nesnow, S., Ward, W., Moore, T., Ren, H., Hester, S. D. — 2009-06-15

Conazoles are fungicides used to control fungal growth in environmental settings and to treat humans with fungal infections. Mouse hepatotumorigenic conazoles display many of the same hepatic toxicologic responses as the mouse liver carcinogen phenobarbital (PB): constitutive androstane receptor (CAR) activation, hypertrophy, Cyp2b induction, and increased cell proliferation. The goal of this study was to apply transcriptional analyses to hepatic tissues from mice exposed to PB, propiconazole (Pro) or triadimefon (Tri) at tumorigenic exposure levels to reveal similarities and differences in response among these treatments. Mice were administered diets containing PB (850 ppm), Pro (2500 ppm), or Tri (1800 ppm) for 4 and 30 days. Targeted transcriptomic analyses were conducted at the gene level examining differentially expressed genes (DEGs), and subsets of DEGs: cell cycle genes, and transcription factors. Analyses were also conducted on function, pathway and network levels examining Ingenuity Pathway Analysis Tox Lists and Canonical Pathways, and Gene-Go MetaCore dynamic networks and their central hubs. Genes expressed by PB or the two conazoles were also compared with those genes associated with human hepatocellular cancer. The results from these analyses indicated greater differences between PB and the two conazoles than similarities. Significant commonalities between the two conazole treatments were also noted. We posit that the transcriptional profiles of tissues exposed to toxic chemicals inherently contain their mechanisms of toxicity. We conclude that although PB and these 2 conazoles induce mouse liver tumors and exhibit similar toxicological responses, their transcriptional profiles are significantly different and thus their mechanisms of tumorigenic action are likely to differ.

Ochratoxin A-Mediated DNA and Protein Damage: Roles of Nitrosative and Oxidative Stresses

2009-06-15

Ochratoxin A (OTA) is a mycotoxin occurring in a variety of foods. OTA is nephrotoxic and nephrocarcinogenic in rodents. An OTA-mediated increase of the inducible nitric oxide synthase (iNOS) expression was observed in normal rat kidney renal cell line and in rat hepatocyte cultures, suggesting the induction of nitrosative stress. This was associated with an increased nuclear factor kappa-light chain enhancer of activated B cells activity. The potential consequences of iNOS induction were further investigated. A significant increase in the levels of protein nitrotyrosine residues was observed with OTA. In addition, OTA was found to increase the level of DNA abasic sites in both cell cultures system. This end point was used as an indirect measure of 8-nitroguanine formation. Treatment of the cells with L-N⁶-(1-iminoethyl) lysine, a specific inhibitor of iNOS activity, inhibited the OTA-mediated overnitration of proteins but did not reduce the level of DNA abasic sites. It was found previously that nuclear factor-erythroid 2 p45-related factor 2 (Nrf2) activators were able to restore the cellular defense against oxidative stress and could prevent DNA abasic sites in cell cultures. In the present study, pretreatment of the cells with activators of Nrf2 prevented OTA-mediated increase in lipid peroxidation, confirming the potential of Nrf2 activators to confer protection against OTA-mediated oxidative stress. In addition, it was found that Nrf2 activators could also prevent OTA-induced protein nitration and cytotoxicity. In conclusion, the present data further confirm oxidative stress as a key source of OTA-induced DNA damage and provide additional evidence for a role of this mechanism in OTA carcinogenicity. The exact role of nitrosative stress still remains to be established.

Effect of the Methoxychlor Metabolite HPTE on the Rat Ovarian Granulosa Cell Transcriptome In Vitro

Harvey, C. N., Esmail, M., Wang, Q., Brooks, A. I., Zachow, R., Uzumcu, M. — 2009-06-15

Ovarian granulosa cells play a central role in steroidogenesis, which is critical for female reproduction. Follicle-stimulating hormone (FSH) promotes cyclic adenosine monophosphate (cAMP)-mediated signaling to regulate granulosa cell steroidogenesis. We have shown previously that 2,2-bis-(p-hydroxyphenyl)-1,1,1-trichloroethane (HPTE) inhibits FSH- and dibutyryl cAMP-stimulated steroidogenesis and affects the messenger RNA levels of steroidogenic pathway enzymes in rat granulosa cells. However, HPTE showed a differential effect in FSH- and cAMP-stimulated cells in that HPTE more completely blocked FSH- when compared to cAMP-driven steroidogenesis. The objective of this study was to analyze the effects of HPTE on global gene expression profiles in untreated granulosa cells and those challenged with FSH or cAMP. Granulosa cells from immature rats were cultured with 0, 1, 5, or 10µM HPTE in the presence or absence of either 3 ng FSH/ml or 1mM cAMP for 48 h. Total RNA was isolated for real-time quantitative PCR and microarray analysis using the GeneChip Rat Genome 230 2.0 and ArrayAssist Microarray Suite. An investigation of changes in gene expression across all HPTE treatments showed that HPTE altered more genes in FSH- (~670 genes) than in cAMP-stimulated cells (~366 genes). Analysis confirmed that HPTE more effectively inhibited FSH- than cAMP-induced steroid pathway gene expression and steroidogenesis. Furthermore, expression patterns of novel genes regulating signal transduction, transport, cell cycle, adhesion, differentiation, motility and growth, apoptosis, development, and metabolism were all altered by HPTE. This study further established that HPTE exerts differential effects within the granulosa cell steroidogenic pathway and revealed that these effects include broader changes in gene expression.

Reproductive and Developmental Toxicity of a Pentabrominated Diphenyl Ether Mixture, DE-71, to Ranch Mink (Mustela vison) and Hazard Assessment for Wild Mink in the Great Lakes Region

2009-06-15

Polybrominated diphenyl ethers (PBDEs) are widespread persistent and bioaccumulative environmental contaminants. Recent scientific attention has focused on the developmental toxicity of PBDE commercial mixtures following perinatal exposure of rodents; however, these studies do not necessarily predict toxicity to highly exposed top predators, such as mink (Mustela vison). Here we assessed the effects of environmentally relevant doses (0, 0.1, 0.5, and 2.5 ppm [wt/wt] in feed) of a technical pentabrominated diphenyl ether mixture, DE-71, on reproductive performance of mink and on development of offspring exposed perinatally and post-weaning until 33 weeks. A dietary concentration that causes no effects on reproduction in rodents, 2.5-ppm DE-71, resulted in complete reproductive failure in these mink, while whelping rates were not affected at all lower does. Developmental effects in offspring were evident in 33-week-old juveniles, which were more sensitive to effects than their respective dams. Juvenile thyroid hormone homeostasis was also much more sensitive compared to rodents, and at 0.5-ppm DE-71, total triiodothyronine (T3) was significantly decreased in all males and females, even despite a compensatory increase of total thyroxine (T4) in females. T4-outer-ring deiodinase activity, mainly contributed by type II deiodinase, was not affected at any dose for any life stage, but thyroid follicular epithelium cell height was elevated in the 0.5-ppm–treated juveniles (p = 0.057). Ethoxyresorufin O-deethylase activity was significantly induced in all offspring at 33 weeks, most likely as a consequence of polybrominated dioxin, furan, or biphenyl impurities in DE-71. Biomonitoring of wild mink in the Great Lakes region indicated that most populations had lower concentrations than what are expected to affect thyroid hormone homeostasis, but margins of safety are small and mink around Hamilton Harbour exceeded the no observed adverse effect level for T3 disruption.

Relative Developmental Toxicity of Glycol Ether Alkoxy Acid Metabolites in the Embryonic Stem Cell Test as compared with the In Vivo Potency of their Parent Compounds

2009-06-15

The embryonic stem cell test (EST) has been proposed as an in vitro assay that might reduce animal experimentation in regulatory developmental toxicology. So far, evaluation of the EST was not performed using compounds within distinct chemical classes. Evaluation within a distinct class of chemically related compounds can define the usefulness of the assay for the chemical class tested. The aim of the present study was to evaluate the relative sensitivity of the EST for a selected series of homologous compounds and to compare the data to the relative developmental toxicity of the compounds in vivo. To this end a series of proximate developmentally toxic glycol ether alkoxy acid metabolites was tested in the EST. All glycol ether alkoxy acid metabolites tested showed a concentration-dependent inhibition of cardiomyocyte differentiation at noncytotoxic concentrations, with methoxyacetic acid as the most potent compound followed by ethoxyacetic acid, butoxyacetic acid, and phenoxyacetic acid, respectively. The potency ranking of the compounds in the EST corresponds with the available in vivo data. The relative differences between the potencies of the compounds appeared more pronounced in the in vivo studies than in the EST. A possible explanation for this discrepancy could be the difference in the kinetics of the compounds in vivo as compared with their in vitro kinetics. This study illustrates that the EST can be used to set priorities for developmental toxicity testing within classes of related compounds.

Interference of Endocrine Disrupters with Thyroid Hormone Receptor-Dependent Transactivation

Hofmann, P. J., Schomburg, L., Kohrle, J. — 2009-06-15

Thyroid hormones regulate critical developmental processes and key metabolic pathways. A number of natural and synthetic substances have been identified which adversely interfere with the endocrine system. These so-called endocrine disrupters (ED) have mainly been studied for their impact on the gonadal hormone axis. The aim of this work was to develop a novel sensitive and convenient in vitro screening assay for the detection and characterization of potential ED of thyroid hormone (TH)–dependent transactivation of gene transcription and to apply this tool to test relevant environmental and nutritive ED compounds. We constructed a TH-responsive luciferase–based reporter plasmid and established a reporter gene assay in a 96 well microplate format using the human hepatocarcinoma cell line HepG2 as host system. Both the synthetic TH receptor (TR) agonist GC-1 and the antagonist NH-3 were used to evaluate the assay. Concentration-response data of test compounds (food constituents, isoflavones, ultraviolet-absorbers, pesticides, industrial chemicals) were recorded in activation assays. In addition, interference with TH-mediated transactivation was tested by coincubation of the ED with triiodothyronine (T₃) in competition assays. Most ED tested affected T₃ reporter gene activity at concentrations of 1µM or higher and displayed either agonistic or mixed agonistic/antagonistic activities. Effects of relevant ED occurred only at relatively high concentrations compared with the endogenous TR ligand T₃. However, on basis of their high production volumes and potential bioaccumulation of some fat-soluble ED our data indicate the need to carefully monitor certain ED for potential disruption of the TH system in intact organisms and humans.

Mechanisms of Quantum Dot Nanoparticle Cellular Uptake

Zhang, L. W., Monteiro-Riviere, N. A. — 2009-06-15

Due to the superior photoemission and photostability characteristics, quantum dots (QD) are novel tools in biological and medical applications. However, the toxicity and mechanism of QD uptake are poorly understood. QD nanoparticles with an emission wavelength of 655 nm are ellipsoid in shape and consist of a cadmium/selenide core with a zinc sulfide shell. We have shown that QD with a carboxylic acid surface coating were recognized by lipid rafts but not by clathrin or caveolae in human epidermal keratinocytes (HEKs). QD were internalized into early endosomes and then transferred to late endosomes or lysosomes. In addition, 24 endocytic interfering agents were used to investigate the mechanism by which QD enter cells. Our results showed that QD endocytic pathways are primarily regulated by the G-protein–coupled receptor associated pathway and low density lipoprotein receptor/scavenger receptor, whereas other endocytic interfering agents may play a role but with less of an inhibitory effect. Lastly, low toxicity of QD was shown with the 20nM dose in HEK at 48 h but not at 24 h by the live/dead cell assay. QD induced more actin filaments formation in the cytoplasm, which is different from the actin depolymerization by cadmium. These findings provide insight into the specific mechanism of QD nanoparticle uptake in cells. The surface coating, size, and charge of QD nanoparticles are important parameters in determining how nanoparticle uptake occurs in mammalian cells for cancer diagnosis and treatment, and drug delivery.

Estrogen and Tamoxifen Protect against Mn-Induced Toxicity in Rat Cortical Primary Cultures of Neurons and Astrocytes

Lee, E.-S. Y., Yin, Z., Milatovic, D., Jiang, H., Aschner, M. — 2009-06-15

Chronic exposure to manganese (Mn) leads to a neurological disorder, manganism, which shares multiple common features with idiopathic Parkinson disease (IPD). 17β-Estradiol (E2) and some selective estrogen receptor modulators, including tamoxifen (TX), afford neuroprotection in various experimental models of neurodegeneration. However, the neuroprotective effects and mechanisms of E2/TX in Mn-induced toxicity have yet to be documented. Herein, we studied the ability of E2/TX to protect rat cortical primary neuronal and astroglial cultures from Mn-induced toxicity. Cell viability, Western blot, and reactive oxygen species (ROS) generation were assessed. Results established that both E2 (10nM) and TX (1µM) attenuated Mn-induced toxicity. The protective effects of E2/TX were more pronounced in astrocytes versus neurons. The E2-mediated attenuation of Mn-induced ROS generation in astrocytes at 6-h treatment (where no cell death was detected) was mediated by a classical estrogen receptor (ER) pathway and the TX-mediated effect on Mn-induced ROS generation was not mediated via classical ER-dependent mechanisms and likely by its antioxidant properties. The phosphatidylinositol-3 kinase (PI3K)/Akt signaling pathway was involved in both E2- and TX-induced attenuation of Mn-induced ROS formation (6 h) in astrocytes. Treatments with Mn for a longer duration (24 h) led to significant cell death, and the protective effects of E2 and TX were (1) not mediated by a classical ER pathway and (2) associated with activation of both mitogen-activated protein kinase/extracellular signal-regulated kinase and PI3K/Akt signaling pathways. Taken together, the results suggest that both E2 and TX offer effective therapeutic means for neuroprotection against Mn-induced toxicity.

Neurotoxicogenomic Investigations to Assess Mechanisms of Action of the Munitions Constituents RDX and 2,6-DNT in Northern Bobwhite (Colinus virginianus)

2009-06-15

Munitions constituents (MCs) including hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), 2,4,6-trinitrotoluene (TNT), and TNT derivatives are recognized to elicit aberrant neuromuscular responses in many species. The onset of seizures resulting in death was observed in the avian model Northern bobwhite after oral dosing with RDX beginning at 8 mg/kg/day in subacute (14 days) exposures, whereas affective doses of the TNT derivative, 2,6-dinitrotoluene (2,6-DNT), caused gastrointestinal impacts, lethargy, and emaciation in subacute and subchronic (60 days) exposures. To assess and contrast the potential neurotoxicogenomic effects of these MCs, a Northern bobwhite microarray was developed consisting of 4119 complementary DNA (cDNA) features enriched for differentially-expressed brain transcripts from exposures to RDX and 2,6-DNT. RDX affected hundreds of genes in brain tissue, whereas 2,6-DNT affected few (≤ 17), indicating that 2,6-DNT exposure had relatively little impact on the brain in comparison to RDX. Birds exhibiting RDX-induced seizures accumulated over 20x more RDX in brain tissues in comparison to non-seizing birds even within a common dose. In parallel, expression patterns were unrelated among seizing and non-seizing birds exposed to equivalent RDX doses. In birds experiencing seizures, genes related to neuronal electrophysiology and signal transduction were significantly affected. Comparative toxicology revealed strong similarity in acute exposure effects between RDX and the organochlorine insecticide dichlorodiphenyltrichloroethane (DDT) regarding both molecular mechanisms and putative mode of action. In a manner similar to DDT, we hypothesize that RDX elicits seizures by inhibition of neuronal cell repolarization postaction potential leading to heightened neuronal excitability and seizures facilitated by multiple molecular mechanisms.

Profiling the Reproductive Toxicity of Chemicals from Multigeneration Studies in the Toxicity Reference Database

2009-06-15

Multigeneration reproduction studies are used to characterize parental and offspring systemic toxicity, as well as reproductive toxicity of pesticides, industrial chemicals and pharmaceuticals. Results from 329 multigeneration studies on 316 chemicals have been digitized into standardized and structured toxicity data within the Toxicity Reference Database (ToxRefDB). An initial assessment of data quality and consistency was performed prior to profiling these environmental chemicals based on reproductive toxicity and associated toxicity endpoints. The pattern of toxicity across 75 effects for all 316 chemicals provided sets of chemicals with similar in vivo toxicity for future predictive modeling. Comparative analysis across the 329 studies identified chemicals with sensitive reproductive effects, based on comparisons to chronic and subchronic toxicity studies, as did the cross-generational comparisons within the multigeneration study. The general pattern of toxicity across all chemicals and the more focused comparative analyses identified 19 parental, offspring and reproductive effects with a high enough incidence to serve as targets for predictive modeling that will eventually serve as a chemical prioritization tool spanning reproductive toxicities. These toxicity endpoints included specific reproductive performance indices, male and female reproductive organ pathologies, offspring viability, growth and maturation, and parental systemic toxicities. Capturing this reproductive toxicity data in ToxRefDB supports ongoing retrospective analyses, test guideline revisions, and computational toxicology research.

Pulmonary Nanoparticle Exposure Disrupts Systemic Microvascular Nitric Oxide Signaling

2009-06-15

We have shown that pulmonary nanoparticle exposure impairs endothelium dependent dilation in systemic arterioles. However, the mechanism(s) through which this effect occurs is/are unclear. The purpose of this study was to identify alterations in the production of reactive species and endogenous nitric oxide (NO) after nanoparticle exposure, and determine the relative contribution of hemoproteins and oxidative enzymes in this process. Sprague-Dawley rats were exposed to fine TiO₂ (primary particle diameter ~1 µm) and TiO₂ nanoparticles (primary particle diameter ~21 nm) via aerosol inhalation at depositions of 4–90 µg per rat. As in previous intravital experiments in the spinotrapezius muscle, dose-dependent arteriolar dilations were produced by intraluminal infusions of the calcium ionophore A23187. Nanoparticle exposure robustly attenuated these endothelium-dependent responses. However, this attenuation was not due to altered microvascular smooth muscle NO sensitivity because nanoparticle exposure did not alter arteriolar dilations in response to local sodium nitroprusside iontophoresis. Nanoparticle exposure significantly increased microvascular oxidative stress by ~60%, and also elevated nitrosative stress fourfold. These reactive stresses coincided with a decreased NO production in a particle deposition dose-dependent manner. Radical scavenging, or inhibition of either myeloperoxidase or nicotinamide adenine dinucleotide phosphate oxidase (reduced) oxidase partially restored NO production as well as normal microvascular function. These results indicate that in conjunction with microvascular dysfunction, nanoparticle exposure also decreases NO bioavailability through at least two functionally distinct mechanisms that may mutually increase local reactive species.

Differential Hepatic Effects of Perfluorobutyrate Mediated by Mouse and Human PPAR-{alpha}

2009-06-15

Perfluorobutryate (PFBA) is a short chain perfluoroalkyl carboxylate that is structurally similar to perfluorooctanoate. Administration of PFBA can cause peroxisome proliferation, induction of peroxisomal fatty acid oxidation and hepatomegaly, suggesting that PFBA activates the nuclear receptor, peroxisome proliferator–activated receptor- (PPAR-). In this study, the role of PPAR- in mediating the effects of PFBA was examined using PPAR- null mice and a mouse line expressing the human PPAR- in the absence of mouse PPAR- (PPAR- humanized mice). PFBA caused upregulation of known PPAR- target genes that modulate lipid metabolism in wild-type and PPAR- humanized mice, and this effect was not found in PPAR- null mice. Increased liver weight and hepatocyte hypertrophy were also found in wild-type and humanized PPAR- mice treated with PFBA, but not in PPAR- null mice. Interestingly, hepatocyte focal necrosis with inflammatory cell infiltrate was only found in wild-type mice administered PFBA; this effect was markedly diminished in both PPAR- null and PPAR- humanized mice. Results from these studies demonstrate that PFBA can modulate gene expression and cause mild hepatomegaly and hepatocyte hypertrophy through a mechanism that requires PPAR- and that these effects do not exhibit a species difference. In contrast, the PPAR-–dependent increase in PFBA-induced hepatocyte focal necrosis with inflammatory cell infiltrate was mediated by the mouse PPAR- but not the human PPAR-. Collectively, these findings demonstrate that PFBA can activate both the mouse and human PPAR-, but there is a species difference in the hepatotoxic response to this chemical.

Signal Transducer and Activator of Transcription 1 (STAT1) is Essential for Chromium Silencing of Gene Induction in Human Airway Epithelial Cells

Nemec, A. A., Barchowsky, A. — 2009-06-15

Hexavalent chromium (Cr(VI)) promotes lung injury and pulmonary diseases through poorly defined mechanisms that may involve the silencing of inducible protective genes. The current study investigated the hypothesis that Cr(VI) actively signals through a signal transducer and activator of transcription 1 (STAT1)–dependent pathway to silence nickel (Ni)–induced expression of vascular endothelial cell growth factor A (VEGFA), an important mediator of lung injury and repair. In human bronchial airway epithelial (BEAS-2B) cells, Ni-induced VEGFA transcription by stimulating an extracellular regulated kinase (ERK) signaling cascade that involved Src kinase–activated Sp1 transactivation, as well as increased hypoxia-inducible factor-1 (HIF-1) stabilization and DNA binding. Ni-stimulated ERK, Src, and HIF-1 activities, as well as Ni-induced VEGFA transcript levels were inhibited in Cr(VI)-exposed cells. We previously demonstrated that Cr(VI) stimulates STAT1 to suppress VEGFA expression. In BEAS-2B cells stably expressing STAT1 short hairpin RNA, Cr(VI) increased VEGFA transcript levels and Sp1 transactivation. Moreover, in the absence of STAT1, Cr(VI), and Ni coexposures positively interacted to further increase VEGFA transcripts. This study demonstrates that metal-stimulated signaling cascades interact to regulate transcription and induction of adaptive or repair responses in airway cells. In addition, the data implicate STAT1 as a rate limiting mediator of Cr(VI)-stimulated gene regulation and suggest that cells lacking STAT1, such as many tumor cell lines, have opposite responses to Cr(VI) relative to normal cells.

Continuous Electrocardiogram Reveals Differences in the Short-Term Cardiotoxic Response of Wistar-Kyoto and Spontaneously Hypertensive Rats to Doxorubicin

Hazari, M. S., Haykal-Coates, N., Winsett, D. W., Costa, D. L., Farraj, A. K. — 2009-06-15

Electrocardiography (ECG) is one of the standard technologies used to monitor and assess cardiac function, and provide insight into the mechanisms driving myocardial pathology. Increased understanding of the effects of cardiovascular disease on rat ECG may help make ECG assessments in rat toxicology studies routine, thus facilitating continuous measurement of functional decrements associated with cardiotoxicant exposure. These studies seek to test the hypothesis that hypertensive rats are more susceptible to the short-term cardiotoxic effects of doxorubicin (DOX) when compared with normotensive rats with respect to continuously measured ECG endpoints. Male Wistar-Kyoto (WKY) and spontaneously hypertensive (SH) rats surgically implanted with radiotelemeters were treated once a week for three weeks with either vehicle, 1.25 (low), 2.5 (medium), or 5 (high) mg/kg DOX (i.p.). ECG, heart rate (HR), and core body temperature (T_co) were continuously monitored during the 1-week baseline and throughout the experimental period until rats were sacrificed 24 h after the third injection. DOX prevented normal body weight gain in both strains and significantly decreased diurnal HR and T_co of high DOX SH rats. In the ECG, SH rats had prolonged baseline PR intervals and QT_c when compared with WKY rats. All DOX-treated WKY rats subsequently developed PR interval prolongation; however only those treated with high DOX had increased QT_c. DOX caused an increase in ST interval in SH rats, and resulted in ECG morphology changes. The number of arrhythmias due to DOX was increased in both strains. In conclusion, ECG analysis can reveal underlying cardiovascular disease as a risk factor in the heart's response to toxicant-induced injury in the rat; and be a valuable tool to evaluate baseline vulnerability and assess cardiotoxicity.

Population-Based Discovery of Toxicogenomics Biomarkers for Hepatotoxicity Using a Laboratory Strain Diversity Panel

Harrill, A. H., Ross, P. K., Gatti, D. M., Threadgill, D. W., Rusyn, I. — 2009-06-15

Toxicogenomic studies are increasingly used to uncover potential biomarkers of adverse health events, enrich chemical risk assessment, and to facilitate proper identification and treatment of persons susceptible to toxicity. Current approaches to biomarker discovery through gene expression profiling usually utilize a single or few strains of rodents, limiting the ability to detect biomarkers that may represent the wide range of toxicity responses typically observed in genetically heterogeneous human populations. To enhance the utility of animal models to detect response biomarkers for genetically diverse populations, we used a laboratory mouse strain diversity panel. Specifically, mice from 36 inbred strains derived from Mus mus musculus, Mus mus castaneous, and Mus mus domesticus origins were treated with a model hepatotoxic agent, acetaminophen (300 mg/kg, ig). Gene expression profiling was performed on liver tissue collected at 24 h after dosing. We identified 26 population-wide biomarkers of response to acetaminophen hepatotoxicity in which the changes in gene expression were significant across treatment and liver necrosis score but not significant for individual mouse strains. Importantly, most of these biomarker genes are part of the intracellular signaling involved in hepatocyte death and include genes previously associated with acetaminophen-induced hepatotoxicity, such as cyclin-dependent kinase inhibitor 1A (p21) and interleukin 6 signal transducer (Il6st), and genes not previously associated with acetaminophen, such as oncostatin M receptor (Osmr) and MLX interacting protein like (Mlxipl). Our data demonstrate that a multistrain approach may provide utility for understanding genotype-independent toxicity responses and facilitate identification of novel targets of therapeutic intervention.

Nervous and Vestibular Toxicities of Acrylonitrile and Iminodipropionitrile

Llorens, J., Soler-Martin, C., Cutillas, B., Saldana-Ruiz, S. — 2009-06-15

On the Mechanism of Nitriles Toxicity

Khan, H. A., Alhomida, A. S., Arif, I. A. — 2009-06-15

ERRATUM

2009-06-15

Toxicological Sciences - current issue

CONTENTS

COVER

Editorial Board

SUBSCRIPTIONS

Nitric Oxide Synthase: "Enzyme Zero" in Air Pollution-Induced Vascular Toxicity

Chiral Toxicology: It's the Same Thing...Only Different

Safety Assessment of Biotechnology Products for Potential Risk of Food Allergy: Implications of New Research

Toxicity Testing in the 21st Century: A View from the Pharmaceutical Industry

Compensatory Induction of Liver Efflux Transporters in Response to ANIT-Induced Liver Injury Is Impaired in FXR-Null Mice

Resveratrol Inhibits Dioxin-Induced Expression of Human CYP1A1 and CYP1B1 by Inhibiting Recruitment of the Aryl Hydrocarbon Receptor Complex and RNA Polymerase II to the Regulatory Regions of the Corresponding Genes

Discrimination of Tumorigenic Triazole Conazoles from Phenobarbital by Transcriptional Analyses of Mouse Liver Gene Expression

Ochratoxin A-Mediated DNA and Protein Damage: Roles of Nitrosative and Oxidative Stresses

Effect of the Methoxychlor Metabolite HPTE on the Rat Ovarian Granulosa Cell Transcriptome In Vitro

Reproductive and Developmental Toxicity of a Pentabrominated Diphenyl Ether Mixture, DE-71, to Ranch Mink (Mustela vison) and Hazard Assessment for Wild Mink in the Great Lakes Region

Relative Developmental Toxicity of Glycol Ether Alkoxy Acid Metabolites in the Embryonic Stem Cell Test as compared with the In Vivo Potency of their Parent Compounds

Interference of Endocrine Disrupters with Thyroid Hormone Receptor-Dependent Transactivation

Mechanisms of Quantum Dot Nanoparticle Cellular Uptake

Estrogen and Tamoxifen Protect against Mn-Induced Toxicity in Rat Cortical Primary Cultures of Neurons and Astrocytes

Neurotoxicogenomic Investigations to Assess Mechanisms of Action of the Munitions Constituents RDX and 2,6-DNT in Northern Bobwhite (Colinus virginianus)

Profiling the Reproductive Toxicity of Chemicals from Multigeneration Studies in the Toxicity Reference Database

Pulmonary Nanoparticle Exposure Disrupts Systemic Microvascular Nitric Oxide Signaling

Differential Hepatic Effects of Perfluorobutyrate Mediated by Mouse and Human PPAR-{alpha}

Signal Transducer and Activator of Transcription 1 (STAT1) is Essential for Chromium Silencing of Gene Induction in Human Airway Epithelial Cells

Continuous Electrocardiogram Reveals Differences in the Short-Term Cardiotoxic Response of Wistar-Kyoto and Spontaneously Hypertensive Rats to Doxorubicin

Population-Based Discovery of Toxicogenomics Biomarkers for Hepatotoxicity Using a Laboratory Strain Diversity Panel

Nervous and Vestibular Toxicities of Acrylonitrile and Iminodipropionitrile

On the Mechanism of Nitriles Toxicity

ERRATUM