Terrance J Kavanagh

University of Washington Seattle, Seattle, Washington, United States

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Publications (156)609.98 Total impact

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    ABSTRACT: Doxorubicin is a substrate of murine Cbr3.•Gclm -/- hepatocytes express higher levels of Cbr3 than Gclm +/+ hepatocytes.•More doxorubicinol is made by Gclm -/- hepatocytes than Gclm +/+ hepatocytes.•Gclm -/- hepatocytes sensitize co-cultured C2C12 cells to doxorubicin toxicity.
    Chemico-Biological Interactions 11/2014; · 2.98 Impact Factor
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    ABSTRACT: Abstract Quantum dots (Qdots) are semiconductor nanoparticles with size-tunable fluorescence capabilities with diverse applications. Qdots typically contain cadmium or other heavy metals, hence raising concerns of their potential toxicity, especially in occupational settings where inhalation of nanomaterials may increase the risk of lung disease. Accordingly, we assessed the effects of tri-n-octylphosphine oxide, poly(maleic anhydride-alt-1-tetradecene) (TOPO-PMAT) coated CdSe/ZnS Qdots on mouse lung epithelial cells and macrophages. Mouse tracheal epithelial cells (MTEC), grown as organotypic cultures, bone marrow-derived macrophages (BMDM), and primary alveolar macrophages (AM) were derived from C57BL/6J or A/J mice and treated with TOPO-PMAT CdSe/ZnS Qdots (10-160 nM) for up to 24 h. Cadmium analysis showed that Qdots remained in the apical compartment of MTEC cultures, whereas they were avidly internalized by AM and BMDM, which did not differ between strains. In MTEC, Qdots selectively induced expression (mRNA and protein) of neutrophil chemokines CXCL1 and CXCL2 but only low to no detectable levels of other factors assessed. In contrast, 4 h exposure to Qdots markedly increased expression of CXCL1, IL6, IL12, and other pro-inflammatory factors in BMDM. Higher inflammatory response was seen in C57BL/6J than in A/J BMDM. Similar expression responses were observed in AM, although overall levels were less robust than in BMDM. MTEC from A/J mice were more sensitive to Qdot pro-inflammatory effects while macrophages from C57BL/6J mice were more sensitive. These findings suggest that patterns of Qdot-induced pulmonary inflammation are likely to be cell-type specific and genetic background dependent.
    Nanotoxicology 07/2014; · 7.34 Impact Factor
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    ABSTRACT: The mechanism by which acetaminophen (APAP) causes liver damage evokes many aspects drug metabolism, oxidative chemistry, and genetic-predisposition. In this study, we leverage the relative resistance of female C57BL/6 mice to APAP-induced liver damage (AILD) compared to male C57BL/6 mice in order to identify the cause(s) of sensitivity. Furthermore, we use mice that are either heterozygous (HZ) or null (KO) for glutamate cysteine ligase modifier subunit (Gclm), in order to titrate the toxicity relative to wild-type (WT) mice. Gclm is important for efficient de novo synthesis of glutathione (GSH). APAP (300 mg/kg, ip) or saline was administered and mice were collected at 0, 0.5, 1, 2, 6, 12, and 24 h. Male mice showed marked elevation in serum alanine aminotransferase by 6 h. In contrast, female WT and HZ mice showed minimal toxicity at all time points. Female KO mice, however, showed AILD comparable to male mice. Genotype-matched male and female mice showed comparable APAP-protein adducts, with Gclm KO mice sustaining significantly greater adducts. ATP was depleted in mice showing toxicity, suggesting impaired mitochondria function. Indeed, peroxiredoxin-6, a GSH-dependent peroxiredoxin, was preferentially adducted by APAP in mitochondria of male mice but rarely adducted in female mice. These results support parallel mechanisms of toxicity where APAP adduction of peroxiredoxin-6 and sustained GSH depletion results in the collapse of mitochondria function and hepatocyte death. We conclude that adduction of peroxiredoxin-6 sensitizes male C57BL/6 mice to toxicity by acetaminophen.
    Redox biology. 01/2014; 2:377-87.
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    ABSTRACT: The small size and heavy metal composition of quantum dots (QDs) combined with their growing consumer product and biomedical research applications have generated concern over their safety. In an occupational setting where QD-enabled products are being manufactured, inhalation is a likely route of exposure. Since current research indicates that QDs could cause inflammation and toxicity in the respiratory tract, it is important that a variety of methods be available to further characterize this potential respiratory hazard. This chapter focuses primarily on in vivo methods for modeling the inhalation and assessing the pulmonary toxicity of QDs.
    Methods in molecular biology (Clifton, N.J.) 01/2014; 1199:179-190. · 1.29 Impact Factor
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    ABSTRACT: Advances in nanotechnology have produced a new class of fluorescent nanoparticles known as quantum dots (Qdots). Compared with organic dyes and fluorescent proteins, Qdots offer several unique advantages in terms of spectral range, brightness, and photostability. Relative to other imaging modalities, optical imaging with Qdots is highly sensitive, quantitative, and capable of multiplexing. Thus, Qdots are being developed for a wide range of applications, including biomedical imaging. Qdot production has also emerged in a number of industrial applications, such as optoelectronic devices and photovoltaic cells. This widespread development and use of Qdots has outpaced research progress on their potential cytotoxicity, engendering major concerns surrounding occupational, environmental, and diagnostic exposures. Given the extensive physicochemical heterogeneity of Qdots (size, charge, chemical composition, solubility, etc.), high-throughput in vitro cytotoxicity assays represent a feasible means of determining effects of multiple variables and can inform design of lower-throughput in vivo cytotoxicity studies. Here, we describe the application of two commonly used assays, lactate dehydrogenase (LDH) and 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS), for detection of Qdot-induced cytotoxicity.
    Methods in molecular biology (Clifton, N.J.) 01/2014; 1199:155-163. · 1.29 Impact Factor
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    ABSTRACT: Abstract Context: Inhalation of fine particulate matter (PM2.5) is associated with acute pulmonary inflammation and impairments in cardiovascular function. In many regions, PM2.5 is largely derived from diesel exhaust (DE), and these pathophysiological effects may be due in part to oxidative stress resulting from DE inhalation. The antioxidant glutathione (GSH) is important in limiting oxidative stress-induced vascular dysfunction. The rate-limiting enzyme in GSH synthesis is glutamate cysteine ligase and polymorphisms in its catalytic and modifier subunits (GCLC and GCLM) have been shown to influence vascular function and risk of myocardial infarction in humans. Objective: We hypothesized that compromised de novo synthesis of GSH in Gclm(-)(/+) mice would result in increased sensitivity to DE-induced lung inflammation and vascular effects. Materials and methods: WT and Gclm(-)(/+) mice were exposed to DE via inhalation (300 μg/m(3)) for 6 h. Neutrophil influx into the lungs, plasma GSH redox potential, vascular reactivity of aortic rings and aortic nitric oxide (NO•) were measured. Results: DE inhalation resulted in mild bronchoalveolar neutrophil influx in both genotypes. DE-induced effects on plasma GSH oxidation and acetylcholine (ACh)-relaxation of aortic rings were only observed in Gclm(-)(/+) mice. Contrary to our hypothesis, DE exposure enhanced ACh-induced relaxation of aortic rings in Gclm(-)(/+) mice. Discussion and conclusion: These data support the hypothesis that genetic determinants of antioxidant capacity influence the biological effects of acute inhalation of DE. However, the acute effects of DE on the vasculature may be dependent on the location and types of vessels involved. Polymorphisms in GSH synthesis genes are common in humans and further investigations into these potential gene-environment interactions are warranted.
    Inhalation Toxicology 07/2013; · 1.89 Impact Factor
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    ABSTRACT: Erythrocytes endure constant exposure to oxidative stress. The major oxidative stress scavenger in erythrocytes is glutathione. The rate-limiting enzyme for glutathione synthesis is glutamate-cysteine ligase, which consists of a catalytic subunit (GCLC) and a modifier subunit (GCLM). Here, we examined erythrocyte survival in GCLM-deficient (gclm(-/-)) mice. Erythrocytes from gclm(-/-) mice showed greatly reduced intracellular glutathione. Prolonged incubation resulted in complete lysis of gclm(-/-) erythrocytes, which could be reversed by exogenous delivery of the antioxidant Trolox. To test the importance of GCLM in vivo, mice were treated with phenylhydrazine (PHZ; 0.07 mg/g b.w.) to induce oxidative stress. Gclm(-/-) mice showed dramatically increased hemolysis compared with gclm(+/+) controls. In addition, PHZ-treated gclm(-/-) mice displayed markedly larger accumulations of injured erythrocytes in the spleen than gclm(+/+) mice within 24 h of treatment. Iron staining indicated precipitations of the erythrocyte-derived pigment hemosiderin in kidney tubules of gclm(-/-) mice and none in gclm(+/+) controls. In fact, 24 h after treatment, kidney function began to diminish in gclm(-/-) mice as evident from increased serum creatinine and urea. Consequently, while all PHZ-treated gclm(+/+) mice survived, 90% of PHZ-treated gclm(-/-) mice died within 5 days of treatment. In vitro, upon incubation in the absence or presence of additional oxidative stress, gclm(-/-) erythrocytes exposed significantly more phosphatidylserine, a cell death marker, than gclm(+/+) erythrocytes, an effect at least partially due to increased cytosolic Ca(2+) concentration. Under resting conditions, gclm(-/-) mice exhibited reticulocytosis, indicating that the enhanced erythrocyte death was offset by accelerated erythrocyte generation. GCLM is thus indispensable for erythrocyte survival, in vitro and in vivo, during oxidative stress.Cell Death and Differentiation advance online publication, 21 June 2013; doi:10.1038/cdd.2013.70.
    Cell death and differentiation 06/2013; · 8.24 Impact Factor
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    ABSTRACT: Quantum dots (QDs) are unique semi-conductor fluorescent nanoparticles with potential uses in a variety of biomedical applications. However, concerns exist regarding their potential toxicity, specifically their capacity to induce oxidative stress and inflammation. In this study we synthesized CdSe/ZnS core/shell QDs with a tri-n-octylphosphine oxide, poly(maleic anhydride-alt-1-tetradecene) (TOPO-PMAT) coating and assessed their effects on lung inflammation in mice. Previously published in vitro data demonstrated these TOPO-PMAT QDs cause oxidative stress resulting in increased expression of antioxidant proteins, including heme oxygenase, and the glutathione (GSH) synthesis enzyme glutamate cysteine ligase (GCL). We therefore investigated the effects of these QDs in vivo in mice deficient in GSH synthesis (Gclm +/- and Gclm -/- mice). When mice were exposed via nasal instillation to a TOPO-PMAT QD dose of 6 µg cadmium (Cd) equivalents/kg body weight, neutrophil counts in bronchoalveolar lavage fluid (BALF) increased in both Gclm wild-type (+/+) and Gclm heterozygous (+/-) mice, whereas Gclm null (-/-) mice exhibited no such increase. Levels of the pro-inflammatory cytokines KC and TNFα increased in BALF from Gclm +/+ and +/- mice, but not from Gclm -/- mice. Analysis of lung Cd levels suggested that QDs were cleared more readily from the lungs of Gclm -/- mice. There was no change in matrix metalloproteinase (MMP) activity in any of the mice. However, there was a decrease in whole lung myeloperoxidase (MPO) content in Gclm -/- mice, regardless of treatment, relative to untreated Gclm +/+ mice. We conclude that in mice TOPO-PMAT QDs have in vivo pro-inflammatory properties, and the inflammatory response is dependent on GSH synthesis status. Because there is a common polymorphism in humans that influences GCLM expression, these findings imply that humans with reduced GSH synthesis capabilities may be more susceptible to the pro-inflammatory effects of QDs.
    PLoS ONE 05/2013; 8(5):e64165. · 3.53 Impact Factor
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    Li-Ping Liang, Terrance J Kavanagh, Manisha Patel
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    ABSTRACT: Depletion of glutathione has been shown to occur in autopsied brains of patients with Parkinson's disease (PD) as well as in animal models of PD. The goal of this study was to determine if chronic GSH deficiency per se resulted in complex I inhibition and/or dopamine depletion and whether these indices were further potentiated by aging or administration of paraquat, a redox-cycling herbicide that produces a PD-like neurodegeneration model in rodents (Brooks et al. 1999; McCormack et al. 2002). Deletion of the rate-limiting GSH synthesis gene, glutamate-cysteine ligase modifier subunit (Gclm) leads to significantly lower GSH concentrations in all tissues including brain. Gclm null (Gclm(-/-)) mice provide a model of prolonged GSH depletion to explore the relationship between GSH, complex I inhibition and dopamine loss in vivo. Despite ~60% depletion of brain GSH in Gclm(-/-) mice of ages 3-5 or 14-16 months, striatal complex I activity, dopamine levels, 3-nitrotyroine/tyrosine ratios, aconitase activity and CoASH remained unchanged. Administration of paraquat (10 mg/kg, twice/week, 3 weeks) to 3-5 month old Gclm(-/-) mice resulted in significantly decreased aconitase activity, complex I activity and dopamine levels but not in 3-5 month old Gclm(+/+) mice. Furthermore, paraquat-induced inhibition of complex I and aconitase activities in Gclm(-/-) mice was observed in the striatum but not cortex. The results suggest that chronic deficiency of GSH in Gclm(-/-) mice was not sufficient to result in complex I inhibition or dopamine depletion perhaps due to homeostatic mechanisms but required an additional oxidative stress insult as shown with paraquat exposure.
    Toxicological Sciences 05/2013; · 4.48 Impact Factor
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    ABSTRACT: Mitochondrial dysfunction plays a key pathogenic role in aging skeletal muscle resulting in significant healthcare costs in the developed world. However, there is no pharmacologic treatment to rapidly reverse mitochondrial deficits in the elderly. Here we demonstrate that a single treatment with the mitochondrial targeted peptide SS-31 restores in vivo mitochondrial energetics to young levels in aged mice after only one hour. Young (5 month old) and old (27 month old) mice were injected intraperitoneally with either saline or 3 mg/kg of SS-31. Skeletal muscle mitochondrial energetics were measured in vivo one hour after injection using a unique combination of optical and (31) P magnetic resonance spectroscopy. Age related declines in resting and maximal mitochondrial ATP production, coupling of oxidative phosphorylation (P/O), and cell energy state (PCr/ATP) were rapidly reversed after SS-31 treatment, while SS-31 had no observable effect on young muscle. These effects of SS-31 on mitochondrial energetics in aged muscle were also associated with a more reduced glutathione redox status and lower mitochondrial H2 O2 emission. Skeletal muscle of aged mice was more fatigue resistant in situ one hour after SS-31 treatment and eight days of SS-31 treatment led to increased whole animal endurance capacity. These data demonstrate that SS-31 represents a new strategy for reversing age-related deficits in skeletal muscle with potential for translation into human use. This article is protected by copyright. All rights reserved.
    Aging cell 05/2013; · 7.55 Impact Factor
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    ABSTRACT: Control of vascular insufficiencies due to various cardiovascular pathologies is important for developing specific and effective treatments. Fluctuations in oxidative stress significantly alter the progression of angiogenesis under physiological and pathological conditions. However, the precise amount of reactive oxygen species (ROS) required to influence subsequent signaling pathways for ischemic angiogenesis remain undefined. Here, we have determined the effect of ROS mediated molecular mechanisms on angiogenesis in a murine model of peripheral artery disease using Gclm mutant mice (a model of compromised glutathione synthesis, therefore reduced antioxidant capacity). Left femoral artery ligation and excision were performed in Gclm WT (+/+), heterozygous (+/-), and null (-/-) mice. Blood flow (laser Doppler), angiogenic index (CD31/DAPI) and proliferation index (Ki67/DAPI) were significantly increased in Gclm(+/-) mice but not in Gclm(+/+) or Gclm(-/-) mice. Measurements of reactive oxygen species suggest that the amount of superoxide required to stimulate angiogenesis following the induction of ischemia is 9.82pmol/mg of tissue. Protein carbonyl levels increased in a manner consistent with increasing oxidative stress. Superoxide and protein carbonyl levels were reduced by the addition of the nitroxide tempol, a known superoxide dismutase mimetic. Finally, restoration of blood flow in Gclm(+/-) mice was attenuated by a VEGF164 aptamer verifying that slightly elevated levels of ROS restore blood flow by stimulating endothelial cell proliferation through a VEGF dependent pathway. The results of this study reveal new information on the amount of ROS necessary for angiogenic activity and provide the foundation of critical redox parameters for vascular remodeling responses. The information obtained from this study on vascular ischemia, using a model of decreased antioxidant capacity, has provided insight into the control of revascularization and is a step forward in our ability to regulate angiogenic therapies.
    Free Radical Biology and Medicine 05/2013; · 5.27 Impact Factor
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    ABSTRACT: BACKGROUND: Engineered nanomaterials (ENMs) have potential benefits, but also present safety concerns for human health. Inter-laboratory studies in rodents using standardized protocols are needed for ENM toxicity assessment. METHODS: Four labs evaluated lung responses in C57BL/6 mice to ENMs delivered by oropharyngeal aspiration (OPA). Three labs evaluated Sprague-Dawley (SD) or Fisher (F)344 rats following intratracheal instillation (IT). ENMs tested were three forms of titanium dioxide (TiO2); anatase/rutile spheres (TiO2-P25), anatase spheres (TiO2-A), anatase nanobelts (TiO2-NB), and three forms of multiwalled carbon nanotubes (MWCNT); original (O), purified (P), and carboxylic acid "functionalized" (F). Bronchoalveolar lavage fluid was collected after 1 day for differential cell counts, lactate dehydrogenase (LDH), and protein. Lungs were fixed for histopathology. Responses were also examined at 7 days (TiO2) and 21 days (MWCNTs). RESULTS: TiO2-A, TiO2-P25, and TiO2-NB caused significant neutrophilia in mice at 1 day in 3 out of 4 labs, respectively. TiO2-NB caused neutrophilia in rats at 1 day in 2 out of 3 labs, while TiO2-P25 or TiO2-A had no significant effect in any of the labs. Inflammation induced by TiO2 in mice and rats resolved by day 7. All MWCNT types caused neutrophilia at 1 day in 3 out of 4 mouse labs and all rat labs. Three out of 4 labs observed similar histopathology to O-MWCNT or TiO2-NB in mice. CONCLUSIONS: ENMs produced similar patterns of neutrophilia and pathology in rats and mice. Although inter-laboratory variability was found in the degree of neutrophilia caused by the three types of TiO2 nanoparticles, similar findings of relative potency for the three types of MWCNTs were found across all laboratories, thus providing greater confidence in these inter-laboratory comparisons.
    Environmental Health Perspectives 05/2013; · 7.26 Impact Factor
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    ABSTRACT: Paraoxonase 2 (PON2), a member of a gene family that also includes PON1 and PON3, is expressed in most tissues, including the brain. In mouse brain, PON2 levels are highest in dopaminergic areas (e.g. striatum), and are higher in astrocytes than in neurons. PON2 is primarily located in mitochondria and exerts a potent antioxidant effect, protecting mouse CNS cells against oxidative stress. The aim of this study was to characterize PON2 expression and functions in the brains of male and female mice. Levels of PON2 (protein, mRNA, and lactonase activity) were higher in brain regions and cells of female mice. Astrocytes and neurons from male mice were significantly more sensitive (by 3-4-fold) to oxidative stress-induced toxicity than the same cells from female mice. Glutathione levels did not differ between genders. Importantly, no significant gender differences in susceptibility to the same oxidants were seen in cells from PON2(-/-) mice. Treatment with estradiol induced a time- and concentration-dependent increase in the levels of PON2 protein and mRNA in male (4.5-fold) and female (1.8-fold) astrocytes, which was dependent on activation of estrogen receptor alpha. In ovariectomized mice, PON2 protein and mRNA were decreased to male levels in brain regions and in liver. Estradiol protected astrocytes from wild-type mice against oxidative stress-induced neurotoxicity, but did not protect cells from PON2(-/-) mice. These results suggest that PON2 is a novel major intracellular factor that protects CNS cells against oxidative stress, and confers gender-dependent susceptibility to such stress. The lower expression of PON2 in males may have broad ramifications for susceptibility to diseases involving oxidative stress, including neurodegenerative diseases.
    Free Radical Biology and Medicine 01/2013; · 5.27 Impact Factor
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    ABSTRACT: Domoic acid (DomA) is a potent marine neurotoxin. By activating AMPA/kainate receptors, DomA induces oxidative stress-mediated apoptotic cell death in neurons. The effect of prolonged (10 day) exposure to a low, non-toxic concentration (5 nM) of DomA on acute (intermediate concentration) neurotoxicity of this toxin was investigated in cerebellar granule neurons (CGNs) from wild-type mice and mice lacking the glutamate cysteine ligase (GCL) modifier subunit (Gclm(-/-)). CGNs from Gclm(-/-) mice have very low glutathione (GSH) levels and are very sensitive to DomA toxicity. In CGNs from wild-type mice, prolonged exposure to 5 nM DomA did not cause any overt toxicity, but reduced oxidative stress-mediated apoptotic cell death induced by exposure to an intermediate concentration (100 nM for 24 h) of DomA. This protection was not observed in CGNs from Gclm(-/-) mice. Prolonged DomA exposure increased GSH levels in CGNs of wild-type, but not of Gclm(-/-) mice. Levels of GCLC (the catalytic subunit of GCL) protein and mRNA were increased in CGNs of both mouse strains, while levels of GCLM protein and mRNA, activity of GCL, and levels of GCL holoenzyme were only increased in CGNs of wild-type mice. Chronic DomA exposure also protected wild-type CGNs from acute toxicity of other oxidants. The results indicate that CGNs from Gclm(-/-) mice, which are already more sensitive to DomA toxicity, are unable to up-regulate their GSH levels. As Gclm(-/-) mice may represent a model for a common human polymorphism in GCLM, such individuals may be at particular risk for DomA-induced neurotoxicity.
    Toxicological Sciences 01/2013; · 4.48 Impact Factor
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    ABSTRACT: Glutathione (GSH) is a critical intracellular antioxidant that is active in free radical scavenging and as a reducing equivalent in biological reactions. Recent studies have suggested that GSH can affect cellular function at the level of gene transcription as well, in particular by affecting NF-κB activation. Additionally, increased or decreased GSH levels in vitro have been tied to increased or decreased hepatocyte proliferation, respectively. Here, we investigated the effect of GSH on the liver's response to TNF-α injection and 2/3 partial hepatectomy (PH), using mice deficient for the modifier subunit of glutamate-cysteine ligase (GCLM), the rate-limiting enzyme in de novo GSH synthesis. We demonstrate that Gclm (-/-) mice have a delay in IκBα degradation after TNF-α injection, resulting in delayed NF-κB nuclear translocation. These mice display profound deficiencies in GSH levels both before and during regeneration, and after PH, Gclm (-/-) mice have an overall delay in cell cycle progression, with slower DNA synthesis, mitosis, and expression of cell cycle proteins. Moreover, there is a delay in expression of downstream targets of NF-κB in the regenerating liver in Gclm (-/-) mice. These data suggest that GSH may play a role in hepatic NF-κB activation in vivo, which is necessary for accurate timing of liver regeneration.
    Journal of liver: disease & transplantation. 01/2013; 1(2).
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    ABSTRACT: Polycyclic aromatic hydrocarbons (PAHs) such as benzo[a]pyrene (BaP) are ubiquitous environmental pollutants found in tobacco smoke, air pollution and grilled foods. Prenatal exposure to BaP causes premature reproductive senescence in mice, and other PAHs are transplacental ovarian carcinogens. Glutathione (GSH) is critical for detoxification of the reactive metabolites of PAHs. Therefore, we hypothesized that mice that are genetically deficient in GSH synthesis, due to deletion of the modifier subunit of glutamate cysteine ligase (Gclm), the rate-limiting enzyme in GSH synthesis, have increased destruction of oogonia, premature ovarian failure, and ovarian tumorigenesis after transplacental BaP exposure compared to Gclm+/+ females. Gclm+/- female and male mice were mated, and dams were treated with 0, 2, or 10 mg/kg/day BaP in sesame oil by gavage from gestational days 7-16. Compared to oil-treated F1 females of the same genotype, Gclm-/- prenatally BaP-treated females had significantly greater decrements in offspring production than Gclm+/+ BaP-treated females. Similarly, we observed significant BaP dose x Gclm genotype interactions on ovarian follicle counts and ovarian tumor multiplicity at 7.5 months of age, with Gclm-/- females having greater decrements in follicle numbers and more ovarian tumors in response to prenatal BaP exposure than Gclm+/+ females. The ovarian tumors were positive for the epithelial marker cytokeratin. Our results demonstrate that prenatal exposure of females to BaP causes premature ovarian failure and ovarian tumorigenesis and that embryonic GSH deficiency due to deletion of Gclm increases sensitivity to these transplacental ovarian effects of BaP.
    Cancer Research 11/2012; · 9.28 Impact Factor
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    ABSTRACT: Semiconductor quantum dots (Qdots) are a promising new technology with benefits in the areas of medical diagnostics and therapeutics. Qdots generally consist of a semiconductor core, capping shell, and surface coating. The semiconductor core of Qdots is often composed of group II and VI metals (e.g., Cd, Se, Te, Hg) that are known to have toxic properties. Various surface coatings have been shown to stabilize Qdots and thus shield cells from the toxic properties of their core elements. In this study, HepG2 cells and primary human liver (PHL) cells were chosen as in vitro tissue culture models of human liver to examine the possible adverse effects of tri-n-octylphosphine oxide, poly(maleic anhydride-alt-1-tetradecene) copolymer (TOPO-PMAT)-coated CdSe/ZnS Qdots (TOPO-PMAT Qdots). The TOPO-PMAT coating is desirable for increasing aqueous solubility and ease of conjugation to targeting moieties (e.g., aptamers and peptides). HepG2 cells avidly incorporated these TOPO-PMAT Qdots into subcellular vesicles. However, PHL cells did not efficiently take up TOPO-PMAT Qdots, but nonparenchymal cells did (especially Kupffer cells). No acute toxicity or morphological changes were noted in either system at the exposure levels used (up to 40 nM). However, cellular stress markers and pro-inflammatory cytokines/chemokines were increased in the PHL cell cultures, suggesting that TOPO-PMAT Qdots are not likely to cause acute cytotoxicity in the liver but may elicit inflammation/hepatitis, demonstrating the importance of relevant preclinical safety models. Thus, further in vivo studies are warranted to ensure that TOPO-PMAT-coated Qdots used in biomedical applications do not induce inflammatory responses as a consequence of hepatic uptake.
    ACS Nano 10/2012; · 12.03 Impact Factor
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    ABSTRACT: Oxidative stress has been implicated in the development of vascular disease and in the promotion of endothelial dysfunction via the reduction in bioavailable nitric oxide (NO()). Glutathione (GSH) is a tripeptide thiol antioxidant that is utilized by glutathione peroxidase (GPx) to scavenge reactive oxygen species such as hydrogen peroxide and phospholipid hydroperoxides. Relatively frequent single-nucleotide polymorphisms (SNPs) within the 5' promoters of the GSH synthesis genes GCLC and GCLM are associated with impaired vasomotor function, as measured by decreased acetylcholine-stimulated coronary artery dilation, and with increased risk of myocardial infarction. Although the influence of genetic knockdown of GPx on vascular function has been investigated in mice, no work to date has been published on the role of genetic knockdown of GSH synthesis genes on vascular reactivity. We therefore investigated the effects of targeted disruption of Gclm in mice and the subsequent depletion of GSH on vascular reactivity, NO() production, aortic nitrotyrosine protein modification, and whole-genome transcriptional responses as measured by DNA microarray. Gclm(-/+) and Gclm(-/-) mice had 72 and 12%, respectively, of wild-type (WT) aortic GSH content. Gclm(-/+) mice had a significant impairment in acetylcholine (ACh)-induced relaxation in aortic rings as well as increased aortic nitrotyrosine protein modification. Surprisingly, Gclm(-/-) aortas showed enhanced relaxation compared to Gclm(-/+) aortas, as well as increased NO() production. Although aortic rings from Gclm(-/-) mice had enhanced ACh relaxation, they had a significantly increased sensitivity to phenylephrine (PE)-induced contraction. Alternatively, the PE response of Gclm(-/+) aortas was nearly identical to that of their WT littermates. To examine the role of NO() or other potential endothelium-derived factors in differentially regulating vasomotor activity, we incubated aortic rings with the NO() synthase inhibitor L-NAME or physically removed the endothelium before PE treatment. L-NAME treatment and endothelium removal enhanced PE-induced contraction in WT and Gclm(-/+) mice, but this effect was severely diminished in Gclm(-/-) mice, indicating a potentially unique role for GSH in mediating vessel contraction. Whole-genome assessment of aortic mRNA in Gclm(-/-) and WT mice revealed altered expression of genes within the canonical Ca(2+) signaling pathway, which may have a role in mediating these observed functional effects. These findings provide additional evidence that the de novo synthesis of GSH can influence vascular reactivity and provide insights regarding possible mechanisms by which SNPs within GCLM and GCLC influence the risk of developing vascular diseases in humans.
    Free Radical Biology and Medicine 07/2012; 53(6):1264-78. · 5.27 Impact Factor
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    ABSTRACT: We have shown that diesel exhaust (DE) inhalation caused progression of atherosclerosis; however, the mechanisms are not fully understood. We hypothesize that exposure to DE upregulates cyclooxygenase (COX) expression and activity, which could play a role in DE-induced atherosclerosis. ApoE knockout mice (30-week old) fed with regular chow were exposed to DE (at 200 µg/m(3) of particulate matter) or filtered air (control) for 7 weeks (6 h/day, 5 days/week). The protein and mRNA expression of COX-1 and COX-2 were evaluated by immunohistochemistry analysis and quantitative real-time PCR, respectively. To examine COX activity, thoracic aortae were mounted in a wire myograph, and phenylephrine (PE)-stimulated vasoconstriction was measured with and without the presence of COX antagonists (indomethacin). COX-2 activity was further assessed by urine 2,3-dinor-6-keto PGF(1α) level, a major metabolite of prostacyclin I(2) (PGI(2)). Immunohistochemistry analysis demonstrates that DE exposure enhanced COX-2 expression in both thoracic aorta (p < 0.01) and aortic root (p < 0.03), with no modification of COX-1 expression. The increased COX-2 expression was positively correlated with smooth muscle cell content in aortic lesions (R(2) = 0.4081, p < 0.008). The fractional changes of maximal vasoconstriction in the presence of indomethacin was attenuated by 3-fold after DE exposure (p < 0.02). Urine 2,3-dinor-6-keto PGF(1α) level was 15-fold higher in DE group than the control (p < 0.007). The mRNA expression of COX-2 (p < 0.006) and PGI synthase (p < 0.02), but not COX-1, was significantly augmented after DE exposure. We show that DE inhalation enhanced COX-2 expression, which is also associated with phenotypic changes of aortic lesion.
    Inhalation Toxicology 07/2012; 24(8):518-27. · 1.89 Impact Factor
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    ABSTRACT: The concentration of glutathione (GSH), the most abundant intracellular free thiol and an important antioxidant, is decreased in the lung in both fibrotic diseases and experimental fibrosis models. The underlying mechanisms and biological significance of GSH depletion, however, remain unclear. Transforming growth factor β (TGF-β) is the most potent and ubiquitous profibrogenic cytokine and its expression is increased in almost all fibrotic diseases. In this study, we show that increasing TGF-β1 expression in mouse lung to a level comparable to those found in lung fibrotic diseases by intranasal instillation of AdTGF-β1(223/225), an adenovirus expressing constitutively active TGF-β1, suppressed the expression of both catalytic and modifier subunits of glutamate-cysteine ligase (GCL), the rate-limiting enzyme in de novo GSH synthesis, decreased GSH concentration, and increased protein and lipid peroxidation in mouse lung. Furthermore, we show that increasing TGF-β1 expression activated JNK and induced activating transcription factor 3, a transcriptional repressor involved in the regulation of the catalytic subunit of GCL, in mouse lung. Control virus (AdDL70-3) had no significant effect on any of these parameters, compared to saline-treated control. Concurrent with GSH depletion, TGF-β1 induced lung epithelial apoptosis and robust pulmonary fibrosis. Importantly, lung GSH levels returned to normal, whereas fibrosis persisted at least 21 days after TGF-β1 instillation. Together, the data suggest that increased TGF-β1 expression may contribute to the GSH depletion observed in pulmonary fibrosis diseases and that GSH depletion may be an early event in, rather than a consequence of, fibrosis development.
    Free Radical Biology and Medicine 05/2012; 53(3):554-63. · 5.27 Impact Factor

Publication Stats

3k Citations
609.98 Total Impact Points


  • 1987–2014
    • University of Washington Seattle
      • • Department of Environmental and Occupational Health Sciences
      • • Department of Comparative Medicine
      • • Department of Pathology
      • • Department of Medicine
      Seattle, Washington, United States
  • 2008–2013
    • Environmental and Occupational Health Sciences Institute
      Edison, New Jersey, United States
    • University of Colorado
      • Department of Pharmaceutical Sciences
      Denver, CO, United States
    • Universität Heidelberg
      Heidelburg, Baden-Württemberg, Germany
  • 2011–2012
    • University of British Columbia - Vancouver
      • Department of Anesthesiology, Pharmacology and Therapeutics
      Vancouver, British Columbia, Canada
    • University of Illinois at Chicago
      • Department of Psychiatry (Chicago)
      Chicago, IL, United States
  • 2001–2012
    • University of California, Irvine
      • • Department of Medicine
      • • Center for Occupational and Environmental Health
      Irvine, CA, United States
  • 2007
    • University Hospital of Parma
      Parma, Emilia-Romagna, Italy
  • 2002–2007
    • Fred Hutchinson Cancer Research Center
      Seattle, Washington, United States
  • 2000
    • Sapienza University of Rome
      Roma, Latium, Italy
  • 1991–1996
    • University of Wuerzburg
      Würzburg, Bavaria, Germany
  • 1984–1985
    • Michigan State University
      • • Department of Pediatrics and Human Development
      • • Department of Pediatrics
      East Lansing, MI, United States