Toxicologic Pathology Associates

Jefferson City, AR, United States

Toxicologic Pathology Associates

Jefferson City, AR, United States
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Liu F.,National Center for Toxicological Research (NCTR) | Patterson T.A.,National Center for Toxicological Research (NCTR) | Sadovova N.,Toxicologic Pathology Associates | Zhang X.,National Center for Toxicological Research (NCTR) | And 6 more authors.
Toxicological Sciences | Year: 2013

Ketamine, a noncompetitive N-methyl-d-aspartate (NMDA) receptor antagonist, is frequently used in pediatric general anesthesia. Accumulating evidence from animal experiments has demonstrated that ketamine causes neuronal cell death during the brain growth spurt. To elucidate the underlying mechanisms associated with ketamine-induced neuronal toxicity and search for approaches or agents to prevent ketamine's adverse effects on the developing brain, a primary nerve cell culture system was utilized. Neurons harvested from the forebrain of newborn rats were maintained under normal control conditions or exposed to either ketamine (10μM) or ketamine plus l-carnitine (an antioxidant; 1-100μM) for 24h, followed by a 24-h withdrawal period. Ketamine exposure resulted in elevated NMDA receptor (NR1) expression, increased generation of reactive oxygen species (ROS) as indicated by higher levels of 8-oxoguanine production, and enhanced neuronal damage. Coadministration of l-carnitine significantly diminished ROS generation and provided near complete protection of neurons from ketamine-induced cell death. NMDA receptors regulate channels that are highly permeable to calcium, and calcium imaging data demonstrated that neurons exposed to ketamine had a significantly elevated amplitude of calcium influx and higher intracellular free calcium concentrations ([Ca2+]i) evoked by NMDA (50μM), compared with control neurons. These findings suggest that prolonged ketamine exposure produces an increase in NMDA receptor expression (compensatory upregulation), which allows for a higher/toxic influx of calcium into neurons once ketamine is removed from the system, leading to elevated ROS generation and neuronal cell death. l-Carnitine appears to be a promising agent in preventing or reversing ketamine's toxic effects on neurons at an early developmental stage. Published by Oxford University Press 2012.


Yang X.,U.S. Food and Drug Administration | Greenhaw J.,U.S. Food and Drug Administration | Shi Q.,U.S. Food and Drug Administration | Su Z.,ICF International | And 4 more authors.
Toxicological Sciences | Year: 2012

Circulating microRNAs (miRNAs) have emerged as novel noninvasive biomarkers for several diseases and other types of tissue injury. This study tested the hypothesis that changes in the levels of urinary miRNAs correlate with liver injury induced by hepatotoxicants. Sprague-Dawley rats were administered acetaminophen (APAP) or carbon tetrachloride (CCl 4) and one nonhepatotoxicant (penicillin/PCN). Urine samples were collected over a 24 h period after a single oral dose of APAP (1250 mg/kg), CCl 4 (2000 mg/kg), or PCN (2400 mg/kg). APAP and CCl 4 induced liver injury based upon increased serum alanine and aspartate aminotransferase levels and histopathological findings, including liver necrosis. APAP and CCl 4 both significantly increased the urinary levels of 44 and 28 miRNAs, respectively. In addition, 10 of the increased miRNAs were in common between APAP and CCl 4. In contrast, PCN caused a slight decrease of a different nonoverlapping set of urinary miRNAs. Cluster analysis revealed a distinct urinary miRNA pattern from the hepatotoxicant-treated groups when compared with vehicle controls and PCN. Analysis of hepatic miRNA levels suggested that the liver was the source of the increased urinary miRNAs after APAP exposure; however, the results from CCl4 were equivocal. Computational analysis was used to predict target genes of the 10 shared hepatotoxicant-induced miRNAs. Liver gene expression profiling using whole genome microarrays identified eight putative miRNA target genes that were significantly altered in the liver of APAP- and CCl 4-treated animals. In conclusion, the patterns of urinary miRNA may hold promise as biomarkers of hepatotoxicant-induced liver injury. Published by Oxford University Press 2011.


Shi Q.,U.S. Food and Drug Administration | Guo L.,U.S. Food and Drug Administration | Patterson T.A.,U.S. Food and Drug Administration | Dial S.,U.S. Food and Drug Administration | And 6 more authors.
Neuroscience | Year: 2010

Ketamine, a non-competitive N-methyl-d-aspartate (NMDA) receptor antagonist, is associated with accelerated neuronal apoptosis in the developing rodent brain. In this study, postnatal day (PND) 7 rats were treated with 20 mg/kg ketamine or saline in six successive doses (s.c.) at 2-h intervals. Brain frontal cortical areas were collected 6 h after the last dose and RNA isolated and hybridized to Illumina Rat Ref-12 Expression BeadChips containing 22,226 probes. Many of the differentially expressed genes were associated with cell death or differentiation and receptor activity. Ingenuity Pathway Analysis software identified perturbations in NMDA-type glutamate, GABA and dopamine receptor signaling. Quantitative polymerase chain reaction (Q-PCR) confirmed that NMDA receptor subunits were significantly up-regulated. Up-regulation of NMDA receptor mRNA signaling was further confirmed by in situ hybridization. These observations support our working hypothesis that prolonged ketamine exposure produces up-regulation of NMDA receptors and subsequent over-stimulation of the glutamatergic system by endogenous glutamate, triggering enhanced apoptosis in developing neurons.


Liu F.,U.S. Food and Drug Administration | Rainosek S.W.,University of Arkansas for Medical Sciences | Sadovova N.,Toxicologic Pathology Associates | Fogle C.M.,U.S. Food and Drug Administration | And 5 more authors.
NeuroToxicology | Year: 2014

Propofol is a widely used general anesthetic. A growing body of data suggests that perinatal exposure to general anesthetics can result in long-term deleterious effects on brain function. In the developing brain there is evidence that general anesthetics can cause cell death, synaptic remodeling, and altered brain cell morphology. Acetyl- l-carnitine (l-Ca), an anti-oxidant dietary supplement, has been reported to prevent neuronal damage from a variety of causes. To evaluate the ability of l-Ca to protect against propofol-induced neuronal toxicity, neural stem cells were isolated from gestational day 14 rat fetuses and on the eighth day in culture were exposed for 24. h to propofol at 10, 50, 100, 300 and 600. μM, with or without l-Ca (10. μM). Markers of cellular proliferation, mitochondrial health, cell death/damage and oxidative damage were monitored to determine: (1) the effects of propofol on neural stem cell proliferation; (2) the nature of propofol-induced neurotoxicity; (3) the degree of protection afforded by l-Ca; and (4) to provide information regarding possible mechanisms underlying protection. After propofol exposure at a clinically relevant concentration (50. μM), the number of dividing cells was significantly decreased, oxidative DNA damage was increased and a significant dose-dependent reduction in mitochondrial function/health was observed. No significant effect on lactase dehydrogenase (LDH) release was observed at propofol concentrations up to 100. μM. The oxidative damage at 50. μM propofol was blocked by l-Ca. Thus, clinically relevant concentrations of propofol induce dose-dependent adverse effects on rat embryonic neural stem cells by slowing or stopping cell division/proliferation and causing cellular damage. Elevated levels of 8-oxoguanine suggest enhanced oxidative damage [reactive oxygen species (ROS) generation] and l-Ca effectively blocks at least some of the toxicity of propofol, presumably by scavenging oxidative species and/or reducing their production. © 2014.


PubMed | U.S. Food and Drug Administration and Toxicologic Pathology Associates
Type: | Journal: Toxicological sciences : an official journal of the Society of Toxicology | Year: 2017

Exposure to cigarette smoke causes a multitude of pathological changes leading to tissue damage and disease. Quantifying such changes in highly differentiated in vitro human tissue models may assist in evaluating the toxicity of tobacco products. In this methods development study, well-differentiated human air-liquid-interface (ALI) in vitro airway tissue models were used to assess toxicological endpoints relevant to tobacco smoke exposure. Whole mainstream smoke solutions (WSSs) were prepared from two commercial cigarettes (R60 and S60) that differ in smoke constituents when machine-smoked under International Organization for Standardization (ISO) conditions. The airway tissue models were exposed apically to WSSs 4-h per day for one to five days. Cytotoxicity, tissue barrier integrity, oxidative stress, mucin secretion, and matrix metalloproteinase (MMP) excretion were measured. The treatments were not cytotoxic and had marginal effects on tissue barrier properties; however, other endpoints responded in time- and dose-dependent manners, with the R60 resulting in higher levels of response than the S60 for many endpoints. Based on the lowest effective dose, differences in response to the WSSs were observed for mucin induction and MMP secretion. Mitigation of mucin induction by co-treatment of cultures with N-acetylcysteine suggests that oxidative stress contributes to mucus hypersecretion. Overall, these preliminary results suggest that quantifying disease-relevant endpoints using ALI airway models is a potential tool for tobacco product toxicity evaluation. Additional research using tobacco samples generated under smoking machine conditions that more closely approximate human smoking patterns will inform further methods development.


Cao X.,U.S. Food and Drug Administration | Lin H.,U.S. Food and Drug Administration | Muskhelishvili L.,Toxicologic Pathology Associates | Latendresse J.,Toxicologic Pathology Associates | And 2 more authors.
Respiratory Research | Year: 2015

Background: The cadmium (Cd) present in air pollutants and cigarette smoke has the potential of causing multiple adverse health outcomes involving damage to pulmonary and cardiovascular tissue. Injury to pulmonary epithelium may include alterations in tight junction (TJ) integrity, resulting in impaired epithelial barrier function and enhanced penetration of chemicals and biomolecules. Herein, we investigated mechanisms involved in the disruption of TJ integrity by Cd exposure using an in vitro human air-liquid-interface (ALI) airway tissue model derived from normal primary human bronchial epithelial cells.Methods: ALI cultures were exposed to noncytotoxic doses of CdCl2 basolaterally and TJ integrity was measured by Trans-Epithelial Electrical Resistance (TEER) and immunofluorescence staining with TJ markers. PCR array analysis was used to identify genes involved with TJ collapse. To explore the involvement of kinase signaling pathways, cultures were treated with CdCl2 in the presence of kinase inhibitors specific for cellular Src or Protein Kinase C (PKC).Results: Noncytotoxic doses of CdCl2 resulted in the collapse of barrier function, as demonstrated by TEER measurements and Zonula occludens-1 (ZO-1) and occludin staining. CdCl2 exposure altered the expression of several groups of genes encoding proteins involved in TJ homeostasis. In particular, down-regulation of select junction-interacting proteins suggested that a possible mechanism for Cd toxicity involves disruption of the peripheral junctional complexes implicated in connecting membrane-bound TJ components to the actin cytoskeleton. Inhibition of kinase signaling using inhibitors specific for cellular Src or PKC preserved the integrity of TJs, possibly by preventing occludin tyrosine hyperphosphorylation, rather than reversing the down-regulation of the junction-interacting proteins.Conclusions: Our findings indicate that acute doses of Cd likely disrupt TJ integrity in human ALI airway cultures both through occludin hyperphosphorylation via kinase activation and by direct disruption of the junction-interacting complex. © 2015 Cao et al.; licensee BioMed Central.


PubMed | National Health Research Institute, Toxicologic Pathology Associates, University of Lisbon and National Center for Toxicological Research (NCTR)
Type: | Journal: Food and chemical toxicology : an international journal published for the British Industrial Biological Research Association | Year: 2016

Furan is a volatile organic chemical that is a contaminant in many common foods. Furan is hepatocarcinogenic in mice and rats; however, the risk to humans from dietary exposure to furan cannot be estimated accurately because the lowest tested dose of furan in a 2-year bioassay in rats gave nearly a 100% incidence of cholangiocarcinoma. To provide bioassay data that can be used in preparing risk assessments, the carcinogenicity of furan was determined in male F344/N Nctr rats administered 0, 0.02, 0.044, 0.092, 0.2, 0.44, 0.92, and 2mg furan/kg body weight (BW) by gavage 5 days/week for 2 years. Exposure to furan was associated with the development of malignant mesothelioma on membranes surrounding the epididymis and on the testicular tunics, with the increase being significant at 2mg furan/kg BW. There was also a dose-related increase in the incidence of mononuclear cell leukemia, with the increase in incidence being significant at 0.092, 0.2, 0.92, and 2mg furan/kg BW. Dose-related non-neoplastic liver lesions included cholangiofibrosis, mixed cell foci, basophilic foci, biliary tract hyperplasia, oval cell hyperplasia, regenerative hyperplasia, and cytoplasmic vacuolization. The most sensitive non-neoplastic lesion was cholangiofibrosis, the frequency of which increased significantly at 0.2mg furan/kg BW.


Delclos K.B.,900 NCTR Road | Camacho L.,900 NCTR Road | Lewis S.M.,Office of Scientific Coordination | Vanlandingham M.M.,900 NCTR Road | And 12 more authors.
Toxicological Sciences | Year: 2014

Bisphenol A (BPA) is a high production volume industrial chemical to which there is widespread human oral exposure. Guideline studies used to set regulatory limits detected adverse effects only at doses well above human exposures and established a no-observed-adverse-effect level (NOAEL) of 5 mg/kg body weight (bw)/day. However, many reported animal studies link BPA to potentially adverse effects on multiple organ systems at doses below the NOAEL. The primary goals of the subchronic study reported here were to identify adverse effects induced by orally (gavage) administered BPA below the NOAEL, to characterize the dose response for such effects and to determine doses for a subsequent chronic study. Sprague Dawley rat dams were dosed daily from gestation day 6 until the start of labor, and their pups were directly dosed from day 1 after birth to termination. The primary focus was on seven equally spaced BPA doses (2.5-2700 μg/kg bw/day). Also included were a naïve control, two doses of ethinyl estradiol (EE. 2) to demonstrate the estrogen responsiveness of the animal model, and two high BPA doses (100,000 and 300,000 μg/kg bw/day) expected from guideline studies to produce adverse effects. Clear adverse effects of BPA, including depressed gestational and postnatal body weight gain, effects on the ovary (increased cystic follicles, depleted corpora lutea, and antral follicles), and serum hormones (increased serum estradiol and prolactin and decreased progesterone), were observed only at the two high doses of BPA. BPA-induced effects partially overlapped those induced by EE. 2, consistent with the known weak estrogenic activity of BPA. © Published by Oxford University Press on behalf of the Society of Toxicology 2014. This work is written by (a) US Government employee(s) and is in the public domain in the US.


Gamboa da Costa G.,National Center for Toxicological Research (NCTR) | Jacob C.C.,National Center for Toxicological Research (NCTR) | Von Tungeln L.S.,National Center for Toxicological Research (NCTR) | Hasbrouck N.R.,Center for Veterinary Medicine | And 4 more authors.
Toxicology and Applied Pharmacology | Year: 2012

The adulteration of pet food with melamine and derivatives, including cyanuric acid, has been implicated in the kidney failure and death of cats and dogs in the USA and other countries. In a previous 7-day dietary study in F344 rats, we established a no-observed-adverse-effect level (NOAEL) for a co-exposure to melamine and cyanuric acid of 8.6. mg/kg bw/day of each compound, and a benchmark dose lower confidence limit (BMDL) of 8.4-10.9. mg/kg bw/day of each compound. To ascertain the role played by the duration of exposure, we treated F344 rats for 28. days. Groups of male and female rats were fed diet containing 0 (control), 30, 60, 120, 180, 240, or 360. ppm of both melamine and cyanuric acid. The lowest dose that produced histopathological alterations in the kidney was 120. ppm, versus 229. ppm in the 7-day study. Wet-mount analysis of kidney sections demonstrated the formation of melamine cyanurate spherulites in one male and two female rats at the 60. ppm dose and in one female rat at the 30. ppm dose, establishing a NOAEL of 2.1. mg/kg bw/day for males and <. 2.6. mg/kg bw/day for females, and BMDL values as low as 1.6. mg/kg bw/day for both sexes. These data demonstrate that the length of exposure is an important component in the threshold of toxicity from a co-exposure to these compounds and suggest that the current risk assessments based on exposures to melamine alone may not reflect sufficiently the risk of a co-exposure to melamine and cyanuric acid. © 2012.


Jacob C.C.,National Center for Toxicological Research (NCTR) | Reimschuessel R.,Center for Veterinary Medicine | von Tungeln L.S.,National Center for Toxicological Research (NCTR) | Olson G.R.,Toxicologic Pathology Associates | And 4 more authors.
Toxicological Sciences | Year: 2011

The intentional adulteration of pet food with melamine and derivatives, including cyanuric acid, has been implicated in the kidney failure and death of a large number of cats and dogs in the United States. Although individually these compounds present low toxicity, coexposure can lead to the formation of melamine cyanurate crystals in the nephrons and eventual kidney failure. To determine the dose-response for nephrotoxicity upon coadministration of melamine and cyanuric acid, groups of male and female F344 rats (six animals per sex per group) were fed 0 (control), 7, 23, 69, 229, or 694 ppm of both melamine and cyanuric acid; 1388 ppm melamine; or 1388 ppm cyanuric acid in the diet for 7 days. No toxicity was observed in the rats exposed to the individual compounds, whereas anorexia and a statistically significant increase in blood urea nitrogen and serum creatinine levels was observed in the animals treated with 229 and 694 ppm melamine and cyanuric acid. The kidneys of these animals were grossly enlarged and pale yellow. Large numbers of crystalline structures deposited in the tubules were seen on sections in kidneys from all rats in these treatment groups. No significant changes were detected in the remaining treatment groups exposed to both melamine and cyanuric acid. In the melamine-only treatment group, 5 of 12 rats had scattered crystals present in renal tubules when examined by wet mount. These were not observed by histopathology. The observed adverse effect level (8.6 mg/kg bw [body weight]/day) and benchmark dose modeling data (8.4-10.9 mg/kg bw/day) determined in this study suggest that the tolerable daily intake values derived from studies conducted with melamine alone may underestimate the risk from coexposures to melamine and cyanuric acid. Published by Oxford University Press 2010.

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