Inhibition of poly(ADP-ribose) polymerase-1 attenuates the toxicity of carbon tetrachloride.

Carbon tetrachloride (CCl(4)) is routinely used as a model compound for eliciting centrilobular hepatotoxicity. It can be bioactivated to the trichloromethyl radical, which causes extensive lipid peroxidation and ultimately cell death by necrosis. Overactivation of poly(ADP-ribose) polymerase-1 (PARP-1) can rapidly reduce the levels of ß-nicotinamide adenine dinucleotide and adenosine triphosphate and ultimately promote necrosis. The aim of this study was to determine whether inhibition of PARP-1 could decrease CCl(4)-induced hepatotoxicity, as measured by degree of poly(ADP-ribosyl)ation, serum levels of lactate dehydrogenase (LDH), lipid peroxidation, and oxidative DNA damage. For this purpose, male ICR mice were administered intraperitoneally a hepatotoxic dose of CCl(4) with or without 6(5H)-phenanthridinone, a potent inhibitor of PARP-1. Animals treated with CCl(4) exhibited extensive poly(ADP-ribosyl)ation in centrilobular hepatocytes, elevated serum levels of LDH, and increased lipid peroxidation. In contrast, animals treated concomitantly with CCl(4) and 6(5H)-phenanthridinone showed significantly lower levels of poly(ADP-ribosyl)ation, serum LDH, and lipid peroxidation. No changes were observed in the levels of oxidative DNA damage regardless of treatment. These results demonstrated that the hepatotoxicity of CCl(4) is dependent on the overactivation of PARP-1 and that inhibition of this enzyme attenuates the hepatotoxicity of CCl(4).

Toxicology and human health assessment of decabromodiphenyl ether.

We evaluated the available pharmacokinetic data and human and animal toxicity data for 2,2',3,3',4,4',5,5',6,6'-decabromodiphenyl ether (BDE-209) (CASRN 1163-19-5) with the objective of deriving a reference dose (RfD) based on the best available science. The available studies for deriving an RfD were first screened using the Klimisch criteria and further evaluated using the United States Environmental Protection Agency's general assessment factors for data quality and relevance (i.e., soundness, applicability and utility, clarity and completeness, uncertainty and variability, and evaluation and review). The chronic 2-year dietary feeding study conducted by the United States National Toxicology Program ( NTP, 1986 , Technical Report Series No. 309) was selected for RfD derivation. Hepatocellular degeneration in male rats was chosen as the critical endpoint in the development of an RfD. For dose-response characterization, we applied benchmark-dose modeling to animal data and determined a point of departure (the 95% lower confidence limit for a 10% increase in hepatocellular degeneration) of 419 mg/kg-day for oral exposures. Based on the similar pharmacokinetic characteristics of BDE-209 across species, this value was converted to a human equivalence dose of 113 mg/kg-day by applying a dosimetric adjustment factor based on body weight scaling to the (3/4) power. An oral RfD of 4 mg/kg-day was calculated by using a composite uncertainty factor of 30, which consisted of 10 for intraspecies uncertainty, 3 for interspecies uncertainty (i.e., 3 for toxicodynamics x 1 for toxicokinetics), and 1 for deficiencies with the database. We consider the RfD to be adequately protective of sensitive subpopulations, including women, their fetuses, children, and people with hepatocellular diseases.

Septin 3 gene polymorphism in Alzheimer's disease.

Septin 3 is a novel member of the septin subfamily of GTPase domain proteins that was recently identified in human neuronal cells. These proteins are involved in vesicle trafficking, neurite outgrowth, and neurofibrillary tangle formation; however, the expression and functional role of septin 3 in normal neuronal tissues and as an etiological agent in neurological disorders is currently unclear. To further characterize these parameters, the present study analyzed the expression of three isoforms of septin 3 (A, B, and C) in fetal and adult human brains and polymorphism of the septin 3 exon 11 microsatellite in control, pure Alzheimer's disease (AD), Lewy body variant (LBV) of AD, and Parkinson's disease. Septin 3 mRNAs for isoforms A and B, but not C, were detected in the frontal cortex of fetus and adult human samples, as measured by reverse transcription-coupled polymerase chain reaction. Genotype analyses indicated that polymorphic septin 3 alleles were distributed in two peaks of frequency in both control and disease groups. Categorization of the alleles into short (S) and long (L) types revealed a significant difference between AD patients and controls (p = 0.034 by chi-square test). Furthermore, the S-allele homozygosity was significantly underrepresented in AD compared with control (p = 0.015 by chi-square test). These results suggest that polymorphism in exon 11 of septin 3 may have a determinative role in the pathogenesis of AD.

Expression of septin 3 isoforms in human brain.

Septin 3 is a novel member of the septin subfamily of GTPase domain proteins. Human septin 3 was originally cloned during a screening of genes expressed in human teratocarcinoma cells induced to differentiate with retinoic acid. Alternative splicing of the septin 3 gene transcript produces two isoforms, A and B, in the human brain, though their regional expression and physiological function remain to be determined. The purpose of the present study was to identify the expression patterns of human septin 3 isoforms in normal human brain and a human neuroblastoma cell line, SH-SY5Y, after retinoic acid-induced differentiation. The expression and distribution patterns of septin 3 isoforms A and B were similar and resembled that of another septin, CDCrel-1. Septin 3A and 3B were expressed in normal human brain in a region-specific manner, with the highest level in the temporal cortex and hippocampus and the lowest level in the brainstem regions. Prominent immunoreactivity was observed diffusely in the neocortices in association with neuropils and punctate structures suggestive of synaptic junctions. Immunoprecipitation studies revealed that septin 3A, 3B, and CDCrel-1 form a complex in the frontal cortex of human brain. These findings, taken together, suggest that septin 3A and 3B, along with CDCrel-1, play some fundamental role(s) in synaptogenesis and neuronal development.

Overexpression of CYP2D6 attenuates the toxicity of MPP+ in actively dividing and differentiated PC12 cells.

Clonal pheochromocytoma cell lines overexpressing cytochrome P450 2D6 (CYP2D6) were established. CYP2D6 was localized in the endoplasmic reticulum, and its enzymatic activity in the microsomal fraction was confirmed by using high performance liquid chromatography analysis with [guanidine-14C]debrisoquine as a substrate. Overexpression of CYP2D6 protected both actively dividing and differentiated cells against the toxic effects of 1-methyl-4-phenylpyridinium ion at the concentration range of 20-40 microM, as assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The production of reactive oxygen species in the mitochondria was suppressed. The cytotoxicity of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine was unchanged in both actively dividing and differentiated cells overexpressing CYP2D6 versus mock-transfected controls at concentrations up to 500 microM. These results suggest that the lowered enzyme activity of CYP2D6 in individuals termed "poor metabolizers" may represent a risk factor from exposure to select neurotoxicants.

Activation of alpha(1)-adrenergic receptors potentiates the nephrotoxicity of ethylene dibromide.

Ethylene dibromide (EDB) has been used as a model compound for eliciting hepato- and nephrotoxicity. Conjugation with glutathione (GSH) has been shown to play a role in the bioactivation of EDB. The aim of this study was to determine whether activation of alpha(1)-adrenergic receptors, which causes a decrease in cellular GSH levels, could modulate the nephrotoxicity of EDB. For this purpose, male ICR mice were treated with EDB and/or the alpha-adrenergic agonist, phenylephrine (Pe), or the alpha-adrenergic antagonist, phentolamine (Phe). Animals treated with EDB (40 mg/kg, i.p.) had a 9.3-fold increase in urinary gamma-glutamyltranspeptidase (GGTP: EC 2.3.2.2) activity and a 38% decrease in renal non-protein bound sulfhydryl (NPSH) levels; however, animals co-treated with EDB and Pe (50 mg/kg, i.p.) exhibited a 27.8-fold increase in urinary GGTP activity and a 60% decrease in NPSH levels. The enhanced presence of urinary GGTP and decrease in cellular levels of NPSH was nearly blocked by treating animals concomitantly with EDB and Phe (10 mg/kg, i.p.) or EDB, Pe, and Phe. Histopathological examination revealed the enhanced degree of tissue damage and necrosis following treatment with EDB and Pe, and the protective effect of Phe at ameliorating EDB toxicity. These results indicate that factors that can influence alpha-adrenergic receptors may be critical in assessing dose-response data used in the risk assessment process.

The effects of organic solvents on poly(ADP-ribose) polymerase-1 activity: implications for neurotoxicity.

Poly(ADP-ribose) polymerase-1 (PARP-1; EC 2.4.2.30), also termed as poly(ADP-ribose) synthetase, is a key enzyme in the recognition and repair of damaged DNA. Several conditions (e.g., ischemia-reperfusion or chemical-induced injury) have been shown to overactivate PARP-1, causing neurodegeneration and necrotic or apoptotic cell death from NAD+ and ATP depletion. In contrast, inhibitors of PARP-1 have been shown to have a neuroprotective effect by ameliorating this response. The purpose of this study was to determine the effects of three routinely used organic solvents (ethanol, methanol, and dimethyl sulfoxide (DMSO)) on the activity of purified PARP-1. A dose-response was examined with each of these solvents. A 112% and 82% increase in PARP-1 activity was observed with 15% ethanol and 20% methanol, respectively. In contrast, a near 20% decrease in the activity was observed with 4% DMSO. Kinetic analysis revealed that the maximal velocity remained unchanged with increasing concentrations of DMSO up to 20%, indicating that DMSO is a competitive inhibitor of PARP-1. Thus, PARP-1 inhibition by DMSO depends on NAD+ concentration and in some pathological processes might be significant even at low DMSO concentrations. Our findings suggest that the interpretation of data from dose-response studies obtained when using common organic solvents may be dramatically skewed, either exaggerating the inherent toxicity of the compound or masking its potential for damage.

Tissue metal concentrations and histopathology of rats gavaged with vitrified soil obtained from the former Charleston Naval Shipyard (SC, USA).

Male Sprague Dawley rats were administered a vitrified material obtained from the former Charleston Naval Shipyard (Charleston, SC, USA) by gavage once daily for 32 days. Group mean body weight of treated animals was within +/-5.4% of controls. No gross or histopathological changes were observed when animals were treated with 67, 174, or 370 mg/kg per day. Analysis of heavy metals revealed a statistically significant increase only in the concentration of arsenic in the livers of animals treated with 174 or 370 mg/kg per day versus controls. Although there was a statistically significant increase in liver arsenic levels, the concentrations were far below mean soil concentrations for western and eastern United States. If the standard assumption of 100% absorption is used, the concentrations observed in the present study are about 20 times less than the average background soil levels in these regions. Based on this, it is concluded that the vitrified material would not pose a public health risk for its intended use as an additive for asphalt and glass beams.

Natural inhibitors of poly(ADP-ribose) polymerase-1.

Poly(ADP-ribose) polymerases (PARPs) are enzymes that catalyze the transfer of ADP-ribose units from ß-nicotinamide adenine dinucleotide (NAD(+)) to acceptor proteins. PARP-1 is responsible for more than 90 % of protein poly-ADP-ribosylation in the brain and may play a role as a molecular switch for cell survival and death. The functional roles of PARP-1 are largely mediated by its activation after binding to damaged DNA. Upon binding, PARP-1 activity increases rapidly and cleaves NAD(+) into ADP-ribose and nicotinamide. Increased activity of PARP-1 can promote DNA repair and its interaction with several transcription factors, whereas hyperactivation of PARP-1 can result in poly(ADP-ribose) accumulation and depletion of NAD(+) and ATP which may lead to caspase independent apoptotic or necrotic cell death, respectively. Excessive PARP-1 activity has been implicated in the pathogenesis of numerous clinical conditions such as stroke, myocardial infarction, inflammation, diabetes, and neurodegenerative disorders. Therefore, it is not surprising that the search for PARP-1 inhibitors with specific therapeutic uses (e.g., brain ischemia, cancer) has been an active area of research. Beyond medicinal uses, naturally occurring PARP-1 inhibitors may also offer a unique preventative means at attenuating chronic inflammatory diseases through dietary supplementation. This possibility has prompted research for specific, naturally occurring inhibitors of PARP-1. Though fewer investigations focus on identifying endogenous inhibitors/modulators of PARP-1 activity in vivo, these activities are very important for better understanding the complex regulation of this enzyme and the potential long-term benefits of supplementation with PARP-1 inhibitors. With this in mind, the focus of this article will be on providing a state-of-the-science review on endogenous and naturally occurring compounds that inhibit PARP-1.

The application of non-default uncertainty factors in the U.S. EPA's Integrated Risk Information System (IRIS). Part I: UF(L), UF(S), and "other uncertainty factors".

The United States Environmental Protection Agency's Integrated Risk Information System (IRIS) includes hazard identification and dose-response assessment values developed by Agency scientists. Uncertainty factors (UFs) are used in the development of IRIS values to address the lack of information in five main areas. The standard UFs account for interspecies uncertainty (UF(A)) and intraspecies variability (UF(H)). The UF(A) addresses uncertainty related to the extrapolation of data from animals to humans, whereas the UF(H) addresses variability amongst individuals (i.e., intrahuman). Additional UFs have been employed to account for database incompleteness, extrapolations from a lowest-observed-adverse-effect level in the absence of a no-observed-adverse-effect level (UF(L)), and subchronic-to-chronic extrapolation (UF(S)). A sixth UF designated as "other uncertainty factors" (UF(O)) has also been applied in place of the UF(L) to account for uncertainty with the adversity of points of departure obtained using benchmark dose modeling. This review will discuss how UF(L), UF(S), and UF(O) have been applied in IRIS assessments, along with the rationale used to describe the choice of UF values that deviate from the standard default of 10.

Framework analysis for the carcinogenic mode of action of nitrobenzene.

Nitrobenzene (CASRN: 98-95-3) has been shown to induce cancers in many tissues including kidney, liver, and thyroid, following chronic inhalation in animals. However, with a few exceptions, genotoxicity assays using nitrobenzene have given negative results. Some DNA binding/adduct studies have brought forth questionable results and, considering the available weight of evidence, it does not appear that nitrobenzene causes cancer via a genotoxic mode of action. Nitrobenzene produces a number of free radicals during its reductive metabolism, in the gut as well as at the cellular level, and generates superoxide anion as a by-product during oxidative melabolism. The reactive species generated during nitrobenzene metabolism are considered candidates for carcinogenicity. Furthermore, several lines of evidence suggest that nitrobenzene exerts its carcinogenicity through a non-DNA reactive (epigenetic) fashion, such as a strong temporal relationship between non-, pre-, and neoplastic lesions leading to carcinogenesis. In this report, we first describe the absorption, distribution, metabolism, and excretion of nitrobenzene followed by a summary of the available genotoxicity studies and the only available cancer bioassay. We subsequently refer to the mode of action framework of the U.S. Environmental Protection Agency's 2005 Guidelines for Carcinogen Risk Assessment as a basis for presenting possible modes of action for nitrobenzene-induced cancers of the liver, thyroid, and kidney, as supported by the available experimental data. The rationale(s) regarding human relevance of each mode of action is also presented. Finally, we hypothesize that the carcinogenic mode of action for nitrobenzene is multifactorial in nature and reflective of free radicals, inflammation, and/or altered methylation.