Biol Trace Elem Res (2010) 136:320–336 DOI 10.1007/s12011-009-8552-1
Lipid Peroxidation and Changes of Trace Elements in Mice Treated with Paradichlorobenzene Wang Suhua & Lu Rongzhu & Yin Changqing & Xing Guangwei & Han Fangan & Jing Junjie & Xu Wenrong & Michael Aschner
Received: 30 July 2009 / Accepted: 14 October 2009 / Published online: 4 November 2009 # Humana Press Inc. 2009
Abstract Paradichlorobenzene (pDCB) has been used as a space deodorant and moth repellant, as well as an intermediate in the chemical industry. Given its broad applications and high volatility, considerable concern exists regarding the adverse health effects of pDCB in the home and the workplace. In this study, changes in lipid peroxidation, antioxidants, and trace element levels in the liver and kidney of pDCB-treated mice were investigated to determine their roles in toxicity. Mice were orally gavaged once daily for seven consecutive days with pDCB (0 (corn oil control), 450, and 900 mg/kg). The level of malondialdehyde (MDA), an end product of lipid peroxidation, markedly increased in the high-dose pDCB group in both the liver and kidney compared with the control group. Changes in hepatic levels of reduced glutathione (GSH) in the pDCB groups were indistinguishable from the control group, while renal levels of reduced GSH in the highdose pDCB group were significantly lowered in comparison to the control and the low-dose groups. Superoxide dismutase (SOD) activity in the liver of mice treated with pDCB showed a downward trend, whereas there was no consistent trend associated with changes W. Suhua : L. Rongzhu (*) : X. Guangwei : X. Wenrong Department of Preventive Medicine, School of Medical Science and Laboratory Medicine, Jiangsu University, 301 Xuefu Rd, Zhenjiang, Jiangsu 212013, China e-mail:
[email protected] e-mail:
[email protected] Y. Changqing Zhenjiang Environmental Monitoring Center, Zhenjiang, Jiangsu 212002, China H. Fangan Zhenjiang Center for Disease Control and Prevention, Zhenjiang, Jiangsu 212003, China J. Junjie Center of Chemical Analysis, School of Chemistry and Chemical Engineering, Jiangsu University, Zhenjiang, Jiangsu 212013, China M. Aschner Department of Pediatrics, Pharmacology and Center for Molecular Neuroscience, Vanderbilt University Medical Center, Nashville, TN 37232, USA
pDCB Affects Lipid Peroxidation and Trace Elements
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in SOD activity in the kidney. Additionally, renal iron levels in the high-dose pDCB group were significantly decreased compared with the low-dose group and the controls, whereas hepatic iron content in the low-dose pDCB group was significantly lower compared with the controls. Selenium and zinc levels in the kidney were both significantly decreased in the high-dose pDCB group vs. the control and low-dose groups. There were no treatmentinduced changes in copper levels in either the kidney or liver. However, a significant increase was found in the liver zinc/copper ratio in the high-dose pDCB group vs. the controls. In addition, blood zinc levels showed a downward trend with increased pDCB dosage. These results suggest that pDCB toxicity is mediated by oxidative damage and tissue-specific alterations in trace element levels both in the liver and the kidney of mice. Keywords Paradichlorobenzene . Oxidative stress . Trace elements . Liver . Kidney
Introduction Paradichlorobenzene (pDCB, synonym 1,4-dichlorobenzene, or 1,4-DCB) has been extensively used as a deodorant, moth repellent, and mildew control agent, as well as an intermediate in dyestuff, fungicides, pharmaceuticals, and other chemical products, including polyphenylene [1]. Due to its high volatility, a major fraction of pDCB used in the aforementioned products is emitted into the air [2–5]. Furthermore, trace amounts of pDCB have been found in marine and drinking water [6–8], food [9], and honey [10–12]. pDCB has also been detected in adipose tissues, blood, and maternal milk [13–15], indicating widespread human exposure to this compound. Therefore, the toxicity of pDCB presents a concern both to industrial workers and to the public at large. Animal responses to pDCB have been extensively studied. pDCB is a liver carcinogen in male and female B6C3Fl mice and a kidney carcinogen in male rats [16–18]. Increased cell proliferation has also been observed in the livers of both male and female mice and in male rat kidney, corroborating the reported carcinogenic effects of pDCB in these tissues [19]. Results obtained from chronic carcinogenesis bioassays concluded that pDCB produced clear evidence of kidney carcinogenicity in male rats and liver tumors in male and female mice [16]. In male rats, the kidney carcinogenicity caused by pDCB was attributed to the protein specific to male rats, alpha-2u-globulin [20]; the nature of mechanisms of pDCBinduced liver tumors have yet to be fully explored. pDCB is oxidized by cytochrome P450 to 2,5-dichlorophenol (DCP) and 2,5-dichlorohydroquinone (DCHQ), resulting in the formation of a meta-stable epoxide intermediate and a reactive benzoquinone species. The nonenzymatic oxidation of DCHQ to 2,5dichloro-p-benzoquinone (DCBQ) produces reactive oxygen species (ROS) [19, 21–26]. Mizutani et al. also found that depletion of glutathione (GSH), one of the strongest nonenzymatic antioxidants, enhanced acute hepatotoxicity of pDCB in mice [27]. In calf thymus DNA, both DCHQ and DCBQ in the presence of NADH and copper (Cu), led to 8-oxodeoxyguanosine (8-oxodG) generation [28], attesting to their pro-oxidant potential during biotransformation. Several studies have also demonstrated that 1,2-DCB, a dichlorobenzene analog of pDCB, leads to the formation of ROS [29, 30]. Several studies have demonstrated that pDCB or its metabolites produce oxidative stress evidenced by oxidative DNA damage [28], in addition to activation of the antioxidant enzymes in freshwater fish (Jenynsia multidentata and Carassius auratus) and a South American aquatic plant species (Ceratophyllum demersum) [31–33].
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Many trace metals play crucial roles in multiple biological processes by activating or inhibiting enzymatic reactions and by affecting the permeability of cell membranes. Some transition metals, such as iron (Fe) and Cu are directly involved in the initiation of free radical formation and carcinogenicity [34–36]. Another trace metal, zinc (Zn), is a component of more than 3,000 Zn-associated transcription factors, including multiple zinc finger-containing DNA-binding proteins, and more than 300 enzymes, including Cu/Zn superoxide dismutase (Cu, Zn-SOD), the latter representing one of the most important enzymatic antioxidants in scavenging free radicals. Several Zn-containing proteins also contribute to DNA repair [37, 38]. Selenium (Se), an antioxidant metalloid, is a key cofactor in regulating GSH peroxidase activity, thus maintaining cellular redox status [39, 40]. Cu is a component of more than 30 enzymes, including ceruloplasmin, cytochrome oxidase, and lysine oxidase [37]. Notably, Cu-mediated DNA damage by metabolites of pDCB has previously been purported to mediate its pathological effects [28, 34]. Furthermore, Fe, Cu, Zn, and Se may also be involved in carcinogenesis given that concentrations of Fe, Zn, Cu, and other essential metals are significantly lower in cancerous stomach tissue than in normal tissues [41, 42]. The present study was designed to test the hypothesis that pDCB-induced toxicity is mediated via lipid peroxidation and/or imbalance in trace elements involved in maintaining cellular redox status. Given earlier reports that 2-year administration of pDCB by oral gavage induces only liver (and not kidney) tumors in mice [16], we also compared liver and kidney levels of oxidative stress biomarkers (Fe, Se, Zn, and Cu) in response to oral pDCB treatment, to determine the tissue specificity of its effects.
Materials and Methods Reagents pDCB was purchased from Shanghai Chemical Reagents Company. Fe, Cu, Se, and Zn standard stock solutions, 1,000 ppm, were supplied by the Jiangsu Center of Environmental Monitoring. Corn oil was purchased from the commercial market. Animals Male and female Kunmin mice weighing 18±1.0 g were purchased from the Laboratory Animal Center of Jiangsu University. All animals were allowed at least 1 week of acclimation prior to use and were maintained in environmentally controlled rooms (22±2°C and 50±10% relative humidity) with a 12:12-h light-dark cycle. They received food and tap water ad libitum. Animal chow was purchased from Nanjing Ailimo (Animal) Science and Technology Company, Ltd. The chow was produced according to National Standard of Laboratory Animal Fodders (GB14924-94). The content of general nutrients in 100 g of fodders were as follows: water, 18%; crude fat, >3%; crude fiber,
