James V. Bruckner

The Potential Adverse Health Effects of Dental Amalgam

There is significant public concern about the potential health effects of exposure to mercury vapour (Hg0) released from dental amalgam restorations. The purpose of this article is to provide information about the toxicokinetics of Hg0, evaluate the findings from the recent scientific and medical literature, and identify research gaps that when filled may definitively support or refute the hypothesis that dental amalgam causes adverse health effects.Dental amalgam is a widely used restorative dental material that was introduced over 150 years ago. Most standard dental amalgam formulations contain approximately 50% elemental mercury. Experimental evidence consistently demonstrates that Hg0 is released from dental amalgam restorations and is absorbed by the human body. Numerous studies report positive correlations between the number of dental amalgam restorations or surfaces and urine mercury concentrations in non-occupationally exposed individuals. Although of public concern, it is currently unclear what adverse health effects are caused by the levels of Hg0 released from this restoration material. Historically, studies of occupationally exposed individuals have provided consistent information about the relationship between exposure to Hg0 and adverse effects reflecting both nervous system and renal dysfunction. Workers are usually exposed to substantially higher Hg0 levels than individuals with dental amalgam restorations and are typically exposed 8 hours per day for 20–30 years, whereas persons with dental amalgam restorations are exposed 24 hours per day over some portion of a lifetime. This review has uncovered no convincing evidence pointing to any adverse health effects that are attributable to dental amalgam restorations besides hypersensitivity in some individuals.

Ontogeny of hepatic and plasma metabolism of deltamethrin in vitro: role in age-dependent acute neurotoxicity.

Deltamethrin (DLM) is a relatively potent and widely used pyrethroid insecticide. Inefficient detoxification has been proposed to be the primary reason for the greater sensitivity of immature rats to the acute neurotoxicity of DLM. The objective of this study was to test this hypothesis by characterizing the age dependence of DLM metabolism in vitro, as well as toxic signs and blood levels of the neurotoxic parent compound following administration of 10 mg DLM/kg p.o. in glycerol formal. Metabolism was quantified in vitro by monitoring the disappearance of the parent compound from plasma [via carboxylesterases (CaEs)] and liver microsomes [via CaEs and cytochromes P450 (P450s)] obtained from 10-, 21-, and 40-day-old male Sprague-Dawley rats. Mean (±S.E.) intrinsic clearances (Vmax/Km) in these respective age groups by liver P450s (4.99 ± 0.32, 16.99 ± 1.85, and 38.45 ± 7.03) and by liver CaEs (0.34 ± 0.05, 1.77 ± 0.38, and 2.53 ± 0.19) and plasma CaEs (0.39 ± 0.06, 0.80 ± 0.09, and 2.28 ± 0.56) increased significantly (p ≤ 0.05) with age, because of progressive increases in Vmax. Intrinsic clearance of DLM by plasma CaEs and liver P450s reached adult levels by 40 days, but clearance by liver CaEs did not. Hepatic P450s played the predominant role in DLM biotransformation in young and adult rats. The incidence and severity of neurotoxic effects varied inversely with age. Correspondingly, blood DLM areas under the concentration versus time curve (AUCs) and Cmax values progressively decreased with increasing age. Internal exposure to DLM (blood AUCs) was closely correlated with toxic signs (salivation and tremors). The present study provides evidence that the limited metabolic capacity of immature rats contributes to elevated systemic exposure and ensuing neurotoxic effects of DLM.

Oral Toxicity of Carbon Tetrachioride: Acute, Subacute, and Subchronic Studies in Rats

This investigation was conducted to characterize the acute, subacute, and subchronic toxic potency of ingested carbon tetrachloride (CCl4). In the first acute and subacute toxicity study, male Sprague-Dawley rats of 300-350 g were gavaged with 0, 20, 40, or 80 mg CCl4/kg once daily for 5 consecutive days, rested for 2 days, and dosed once daily for 4 additional days. Rats of 200-250 g were gavaged with 0, 20, 80, or 160 mg CCl4/kg according to the same dosage regimen in the second acute and subacute study. In the first and second studies one group of rats at each dosage level was sacrificed for clinical chemistry and histopathological evaluation at 24 hr, 4 days, and 11 days after initiation of dosing. Single 20- and 40-mg/kg doses had no apparent toxic effect at 24 hr, although 80 mg/kg caused mild hepatic centrilobular vacuolization and significant increases in some serum enzyme levels. In general, there was progressively severe hepatic injury at each dosage level over the 11-day period. CCl4 was more hepatotoxic to the 200-250-g rats than to the 300-350-g rats. In the subchronic study, rats initially 200-250 g were gavaged 5 times weekly for 12 weeks with 0, 1, 10, or 33 mg CCl4/kg. Body weight and clinical chemistry indices were monitored during the 12 weeks of dosing and 2 weeks after cessation of dosing. A dose of 1 mg/kg had no apparent adverse effect; 10 mg/kg produced slight, but statistically significant increases in sorbitol dehydrogenase activity and mild hepatic centrilobular vacuolization; 33 mg/kg caused marked hepatotoxicity. Serum enzyme levels remained elevated during the 12-week dosing period, but returned toward normal within 13 days of cessation of CCl4 exposure. Microscopic examination of livers of the 33-mg/kg rats revealed cirrhosis, characterized by bile duct proliferation, fibrosis, lobular distortion, parenchymal regeneration, hyperplastic nodules, and single-cell necrosis. The fibrosis was not reversed within the 13-day recovery period.

Role of glutathione depletion in the cytotoxicity of acetaminophen in a primary culture system of rat hepatocytes

A primary culture system of postnatal rat hepatocytes was utilized to study the cytotoxicity of acetaminophen and the toxicological significance of glutathione (GSH) depletion. The relative time of onset and magnitude of GSH depletion, lipid peroxidation and cytotoxicity were contrasted in order to gain insight into their interrelationships. Exposure of the hepatocytes to acetaminophen resulted in time- and dose-dependent depletion of cellular GSH. The acetaminophen-induced GSH depletion and ensuing lactate dehydrogenase (LDH) leakage were quite modest and delayed in onset, in contrast to that caused by iodoacetamide (IAA) and by diethylmaleate (DEM), 2 well-known depletors of GSH. There was comparable LDH leakage, irrespective of drug treatment, when GSH levels decreased to about 20% of normal. Reduction of GSH levels below the 20% threshold by IAA treatment resulted in marked LDH leakage and loss of viability. Maximal LDH leakage in response to IAA and acetaminophen preceded maximal malondialdehyde (MDA) formation, suggesting that lipid peroxidation may be a consequence of cell damage as well as GSH depletion. IAA and DEM produced a comparable, modest accumulation of MDA, yet IAA was much more cytotoxic. These findings indicate that lipid peroxidation does not play a central role in hepatocellular injury by compounds which deplete GSH, although it may contribute to degeneration of the cell. As events in the cultured postnatal hepatocytes paralleled those reported in vivo, the system can be a useful and valid model with which to study mechanisms of chemical toxicity.

The Acute Exposure Guideline Level (AEGL) Program: Applications of Physiologically Based Pharmacokinetic Modeling

The primary aim of the Acute Exposure Guideline Level (AEGL) program is to develop scientifically credible limits for once-in-a-lifetime or rare acute inhalation exposures to high-priority, hazardous chemicals. The program was developed because of the need of communities for information on hazardous chemicals to assist in emergency planning, notification, and response, as well as the training of emergency response personnel. AEGLs are applicable to the general population, including children, the elderly, and other potentially susceptible subpopulations. AEGLs are the airborne concentrations of chemicals above which a person could experience notable discomfort or irritation (AEGL-1); serious, long-lasting health effects (AEGL-2); and life-threatening effects or death (AEGL-3). AEGLs are determined for five exposure periods (10 and 30 min and 1, 4, and 8 h). Physiologically based pharmacokinetic (PBPK) models can be very useful in the interspecies and time scaling often required here. PBPK models are used for the current article to predict AEGLs for trichloroethylene (TCE), based on the time course of TCE in the blood and/or brain of rats and humans. These AEGLs are compared to values obtained by standard time-scaling methods. Comprehensive toxicity assessment documents for each chemical under consideration are prepared by the National Advisory Committee for AEGLs, a panel comprised of representatives of federal, state, and local governmental agencies, as well as industry and private-sector organizations. The documents are developed according to National Research Council (NRC) guidelines and must be reviewed by the NRC Subcommittee on Acute Exposure Guideline Levels before becoming final. AEGLs for 18 chemicals have been published, and it is anticipated that 40 to 50 chemicals will be evaluated annually.

Age, Dose, and Time-Dependency of Plasma and Tissue Distribution of Deltamethrin in Immature Rats

The major objective of this project was to characterize the systemic disposition of the pyrethroid, deltamethrin (DLT), in immature rats, with emphasis on the age dependence of target organ (brain) dosimetry. Postnatal day (PND) 10, 21, and 40 male Sprague-Dawley rats received 0.4, 2, or 10 mg DLT/kg by gavage in glycerol formal. Serial plasma, brain, fat, liver, and skeletal muscle samples were collected for up to 510 h and analyzed for DLT and/or 3-phenoxybenzoic acid (PBA) content by high-performance liquid chromatography. Toxicokinetic data from previous experiments of the same design with young adult (PND 90) rats (Kim, K.-B., Anand, S. S., Kim, H. J., White, C. A., and Bruckner, J. V. [2008]. Toxicokinetics and tissue distribution of deltamethrin in adult Sprague-Dawley rats. Toxicol. Sci. 101, 197-205) were used to compare to immature rat data. Plasma and tissue DLT levels were inversely related to age. Preweanlings and weanlings showed markedly elevated brain concentrations and pronounced salivation, tremors, choreoathetosis, and eventual fatalities. Plasma DLT levels did not reliably reflect brain levels over time. Plasma:brain ratios were time and dose dependent, but apparently not age dependent. Brain levels were better correlated with the magnitude of salivation and tremors than plasma levels. Hepatic intrinsic clearance of DLT progressively increased during maturation, as did the hepatic extraction ratio. Thus, limited capacity to metabolically inactivate DLT appeared primarily responsible for the inordinately high target organ doses and acute neurotoxicity in pups and weanling rats. Hepatic blood flow was not rate limiting in any age group. Limited DLT hydrolysis was manifest in vivo in the pups by relatively low plasma PBA levels. Elevated exposure of the immature brain to a pyrethroid may prove to be of consequence for long-term, as well as short-term neurotoxicity.

Physiological pharmacokinetic modeling of inhaled trichloroethylene in rats

The pharmacokinetics of trichloroethylene (TCE) was characterized during and following inhalation exposures of male Sprague-Dawley rats. The blood and exhaled breath TCE time-course data were used to formulate and assess the accuracy of predictions of a physiologically based pharmacokinetic (PB-PK) model for TCE inhalation. Fifty or 500 ppm of TCE was inhaled by unanesthetized rats of 325-375 g for 2 hr through a miniaturized one-way breathing valve. Repetitive samples of the inhaled and exhaled breath streams, as well as arterial blood, were collected concurrently during and for 3 hr following the exposures and analyzed for TCE by headspace gas chromatography. Respiratory rates and volumes were continuously monitored and used in conjunction with the pharmacokinetic data to delineate uptake and elimination profiles. Levels of TCE in the exhaled breath attained near steady-state soon after the beginning of exposures, and were then directly proportional to the inhaled concentration. Exhaled breath levels of TCE in rats were similar in magnitude to values previously published for TCE inhalation exposures of humans. Levels of TCE in the blood of the 50 ppm-exposed animals also rapidly approached near steady-state, but blood levels in the 500 ppm-exposed animals rose progressively, reaching concentrations 25- to 30-fold higher than in the 50 ppm group during the second hour of exposure. The 10-fold increase in inhaled concentration resulted in an 8.7-fold increase in cumulative uptake, or total absorbed dose. These findings of nonlinearity indicate that metabolic saturation ensued during the 500 ppm exposure. The PB-PK model was characterized as blood flow-limited with TCE eliminated unchanged in the exhaled breath and by saturable liver metabolism. The uptake and elimination profiles were accurately simulated by the PB-PK model for both the 50 and 500 ppm TCE exposure levels. Such a model may be quite useful in risk assessments in predicting internal (i.e., systemically absorbed) doses of TCE and other volatile organics under a variety of exposure scenarios.

Effect of oral dosing vehicles on the acute hepatotoxicity of carbon tetrachloride in rats

Although carbon tetrachloride (CCl4) is of concern as a drinking water contaminant, it has been necessary in most oral toxicity studies to give CCl4 in an oil vehicle due to its limited water solubility. The primary objective of our study was to assess the influence of dosing vehicles on the acute hepatotoxicity of CCl4. Fasted 200- to 230-g rats were generally found to be more susceptible to CCl4 hepatotoxicity than fasted 300- to 330-g rats. A time-course study revealed that corn oil did not delay the onset or time of maximal liver injury by an oral 100 mg/kg dose of CCl4, but did reduce the extent of injury relative to that when the chemical was given undiluted or as an aqueous emulsion. Fasted 200- to 230-g male Sprague-Dawley rats were given 0, 10, 25, 50, 100, 250, 500, or 1000 mg CCl4/kg body wt by gavage: in corn oil; as an aqueous emulsion; as the undiluted chemical; and in the 10 and 25 mg/kg doses only, in water. Blood and liver samples were taken 24 hr after dosing for measurement of serum and microsomal enzymes. Pathological examination of liver samples was also conducted. Dose-dependent increases in serum enzyme levels and pathological changes and dose-dependent decreases in microsomal P450 and glucose-6-phosphatase activity were observed in each vehicle group. Both the 10 and 25 mg/kg oral doses of CCl4 in water caused significant elevations in serum enzymes and hepatic centrolobular vacuolation. The study revealed that acute hepatotoxicity was less pronounced at each dosage level in rats given CCl4 in corn oil than in other vehicle groups. These findings demonstrate that dosing vehicles can significantly influence the acute hepatotoxicity of CCl4 in rats and are a cause for additional consideration and review of the practice of routinely using vegetable oils as a diluent in studies of volatile organic compound (VOC) toxicity. The use of aqueous Emulphor emulsions appears more appropriate in acute toxicity studies of VOC drinking water contaminants such as CCl4, in that the emulsion did not substantially alter the toxicity of CCl4 from that of undiluted CCl4 or CCl4 ingested in water.

The uptake and elimination of 1,1,1-trichloroethane during and following inhalation exposures in rats☆☆☆

The pharmacokinetics of 1,1,1-trichloroethane (TRI) was studied in male Sprague-Dawley rats in order to characterize and quantify TRI uptake and elimination oby direct measurements of the inhaled and exhaled compound. Fifty or 500 ppm TRI was inhaled for 2 hr through a one-way breathing valve by unanesthetized rats of 325-375 g. Repetitive samples of the separate inhaled and exhaled breath streams, as well as arterial blood, were collected concurrently both during and following TRI inhalation and analyzed for TRI by gas chromatography. Respiratory rates and volumes were continuously monitored during and following exposure and were used in conjunction with the pharmacokinetic data to characterize profiles of uptake and elimination. TRI was very rapidly absorbed from the lung, in that substantial levels were present in arterial blood at the first sampling time (i.e., 2 min). Blood and exhaled breath concentrations of TRI increased rapidly after the initiation of exposure, approaching but not reaching steady state during the 2-hr exposures. The blood and exhaled breath concentrations were directly proportional to the exposure concentration during the exposures. Percentage uptake of TRI decreased 30-35% during the first hour of inhalation, diminishing to approximately 45-50% by the end of the exposure. Total cumulative uptake in the 50 and 500 ppm groups over the 2-hr inhalation exposures was determined to be 6 and 48 mg/kg body wt, respectively. By the end of the exposure period, 2.1 and 20.8 mg, respectively, of inhaled TRI was eliminated from rats inhaling 50 and 500 ppm TRI. A physiological pharmacokinetic model for TRI inhalation was utilized to predict blood and exhaled breath concentrations for comparison with observed experimental values. Overall, values predicted by the physiological pharmacokinetic model for TRI levels in the blood and exhaled breath were in close agreement with measured values both during and following TRI inhalation.

Trichloroethylene risk assessment: A review and commentary

Trichloroethylene (TCE) is a widespread environmental contaminant that is carcinogenic when given in high, chronic doses to certain strains of mice and rats. The capacity of TCE to cause cancer in humans is less clear. The current maximum contaminant level (MCL) of 5 ppb (μg/L) is based on an US Environment Protection Agency (USEPA) policy decision rather than the underlying science. In view of major advances in understanding the etiology and mechanisms of chemically induced cancer, USEPA began in the late 1990s to revise its guidelines for cancer risk assessment. TCE was chosen as the pilot chemical. The final guidelines emphasized a “weight-of-evidence” approach with consideration of dose-response relationships, modes of action, and metabolic/toxicokinetic processes. Where adequate data are available to support reversible binding of the carcinogenic moiety to biological receptors as the initiating event (i.e., a threshold exists), a nonlinear approach is to be used. Otherwise, the default assumption of a linear (i.e., nonthreshold) dose-response is utilized. When validated physiologically based pharmacokinetic (PBPK) models are available, they are to be used to predict internal dosimetry as the basis for species and dose extrapolations. The present article reviews pertinent literature and discusses areas where research may resolve some outstanding issues and facilitate the reassessment process. Key research needs are proposed, including role of dichloroacetic acid (DCA) in TCE-induced liver tumorigenesis in humans; extension of current PBPK models to predict target organ deposition of trichloroacetic acid (TCA) and DCA in humans ingesting TCE in drinking water; use of human hepatocytes to ascertain metabolic rate constants for use in PBPK models that incorporate variability in metabolism of TCE by potentially sensitive subpopulations; measurement of the efficiency of first-pass elimination of trace levels of TCE in drinking water; and assessment of exogenous factors’ (e.g., alcohol, drugs) ability to alter metabolic activation and risks at such low-level exposure.