Dichlorodiphenyltrichloroethane (DDT), DDT metabolites and pregnancy outcomes

Organochlorine pesticides (OCPs) are persistent endocrine disruptors. OCPs cross the placenta; this prenatal exposure has been associated with adverse pregnancy outcomes. We investigated associations between prenatal exposure to OCPs and gestational age and birth weight in 600 infants born between 1960 and 1963. The primary OCP was 1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane (p,p′-DDT), its primary metabolite, 1,1′-dichloro-2,2′-bis(p-chlorophenyl)ethylene (p,p′-DDE) and the contaminant, 1,1,1-trichloro-2-(p-chlorophenyl)-2-(o-chlorophenyl)-ethane (o,p′-DDT). Regression analysis indicated that for each natural log unit increase in p,p′-DDT, birth weight increased by 274 g (95% CI: 122, 425) when controlling for p,p′-DDE and o,p′-DDT. At a given level of p,p′-DDT exposure, o,p′-DDT and p,p′-DDE were associated with decreased birth weight. p,p′-DDE was negatively associated with length of gestation, controlling for p,p′-DDT and o,p′-DDT. These findings suggest opposing associations between exposure to p,p′-DDT and p,p′-DDE and birth weight. We did not find evidence to support mediation by maternal thyroid hormone status nor that the association differed by sex.     

Distribution of dichlorodiphenyltrichloroethane (DDT) and its metabolites in an Indian waterway

This paper reports on the results of systematic analyses carried out to assess the distribution of dichlorodiphenyltrichloroethane (DDT) and its metabolites, o,p′-DDT, p,p′-DDE, and p,p′-DDD in a tropical waterway, the Cochin Estuarine System. Six sampling stations along the system were studied. Analyses were carried out during premonsoon, monsoon, and postmonsoon seasons. ΣDDT concentrations as high as 55.422 μg L^?1 were detected. The predominant DDT metabolite was found to be p,p′-DDE. During the monsoon season, o,p′-DDT was not detected at any of the sampling Stations. Station 1 (riverine) and station 6 (coastal zone barmouth) emerged as DDT-free zones and the analytical results have enabled the identification of DDT “hot spots” in the estuary. © 1994 by John Wiley & Sons, Inc..     

Distribution and biochemical effects of DDT, DDD and DDE in penned double-crested cormorants

Concentrations, distribution and chronic effects of DDT, DDD and DDE were determined in penned cormorants (Phalacrocorax a. auritus) treated with 2, 5 and 10 mg of a combination of these compounds daily in their diet. Birds stressed by a one-half decrease in food after the cessation of 9 weeks of treatment and birds that died of DDT toxicity showed a marked increase in brain and liver residues and a decrease in carcass residues. Higher brain residue levels were significantly correlated with decreased body weight (p < 0.01) and carcass lipid content (p < 0.05). DDD concentrations in the brain were found to be the best indicator of DDT toxicity. Brain concentrations ranged from 24 to 85 ppm in birds that died of toxic effects and from 0.4 to 29 ppm in survivors, indicating that 30 ppm was diagnostic of toxicity. Brain concentrations of DDT and metabolites in wild cormorants indicated no immediate danger of toxicity. Average residue levels in the carcass of wild adult cormorants was 1.00 ppm for DDD and 10.6 ppm for the total of DDT, DDD and DDE. This represents a total body burden of 2.2 mg of DDD and 23.3 mg of the DDT complex as adult cormorants averaged 2200 g in body weight. Analysis of total liver vitamin A indicated a significant decrease of vitamin A due to treatment. An analysis of variance showed a significant decrease in liver (p < 0.05) and heart weights (p < 0.05) due to treatment but not brain or spleen weights when analyzed as percentage of body weight. A significant negative correlation was found between liver weight and brain concentrations of DDE + DDD + DDT (p < 0.05). No significant effects due to treatment were found on blood chemistry values.     

Distribution and metabolism of DDT in the catfish Heteropneustes fossilis in relation to the signs of poisoning

Adult male catfish, Heteropneustes fossilis (Bloch), were given a lethal dose of 800 mg DDT/kg as an im injection of 10% p,p′-DDT in peanut oil ($\text{w}\text{v}$) in the caudal region. The metabolites of DDT in various tissues were studied 24 hr after treatment. For DDT distribution studies, the treated animals were divided into 6 categories according to signs of poisoning: (1) no signs, (2) hyperexcitation, (3) tremors, (4) convulsions, (5) death, and (6) recovery. The concentration of DDT was determined in brain, spinal cord, liver, kidney and fat. Adequate replicates and controls were used in each case. p,p′-DDE was the only metabolite detected. The major proportion of DDE (10%), was found in the kidney, while liver and fat bodies contained slightly lesser amounts of DDE. The remainder was unchanged DDT. The concentration of DDT in the tissues increased with time and the severity of poisoning. The brain and spinal cord contained the lowest concentration of DDT, starting with 7.1 and 9.8 ppm when the fish showed no signs. The DDT content gradually rose to 50.5 and 58.7 ppm when the fish were dead, and declined to 9.1 and 19.0 ppm in fish which recovered completely. In fat, the DDT content continued to increase steadily and was maximal in fish which had recovered. The results indicate that signs of poisoning in the catfish were directly related to the concentration of DDT in the brain and spinal cord. The concentrations of DDT in brain and spinal cord of the catfish soon after death were similar to those found in some other vertebrates, e.g., rats, mice and a few species of birds, under identical circumstances.     

Comparison of p-ṕ-DDE and o-ṕ-DDE on Eggshell Thickness and Ca Binding Activity of Shell Gland in Ducks

Abstract: One of the main ecotoxicological effects of DDT and its stable metabolite DDE is to reduce the reproductive success in several species of birds by decreasing the thickness of the eggshell. In two different strains of ducks (Anas platyrhynchos var.) addition of 40 mg/kg dry weight of p-?-DDE to the diet for 45 days produced a significant and long-lasting reduction of the shell thickness, measured as eggshell index. The reduction was more marked in Indian Runner Ducks (IRD; 30±3%) than in a SwedishXRouen breed (SR; 10±2%. The p-?-DDE residues in the egg yolk + egg white were almost similar in the two strains; 38±2 and 51±6 p.p.m. (wet weight) respectively. In the IRD 40 mg/kg of o-?-DDE in the diet for 45 days was much less potent in reducing the eggshell index (8 ±3%) than thep-pisomer; the amount of residue of the o-? isomer in the eggs (2.2±0.1 p.p.m.) was also smaller. A homogenate of the shell gland from the SR breed accumulated ⁴⁵Ca²⁺ in the presence of ATP. The accumulation was inhibited about 50% by 5 mM sodium azide and 100% by 3 mM mersalyl. p-?- and o-?-DDE in the incubation medium both reduced the Ca²⁺ binding, in a similar and dose-dependent manner. The threshold concentration was about 20 p.p.m.; 40 p.p.m. reduced the accumulation by about 40%. DDE may reduce eggshell thickness by inhibiting the Ca binding to the membranes of the cells of the eggshell gland.     

Effect of a single oral dose of ddt on lipid metabolism in rat

A single oral dose of 1,1,1-trichloro-2,2-bis-(p-chlorophenyl)ethane (DDT) (600 $\text{mg}\text{kg}$ body weight) was given to rats and the levels of various lipids in adipose tissue, liver and plasma were studied. No alteration was observed in the levels of various lipid classes in these tissues except for a decrease in the phospholipid and triglyceride fractions of liver. Lipoprotein lipase activity of post-heparin plasma (protamine-sensitive and -resistant) was significantly decreased, whereas in liver and adipose tissue, the activity of this enzyme remained unchanged.     

Effects of environmental organochlorine pesticides on human breast cancer: Putative involvement on invasive cell ability

Human exposure to persistent organic pollutants (POPs) is a certainty, even to long banned pesticides like o,p′‐dichlorodiphenyltrichloroethane (o,p′‐DDT), and its metabolites p,p′‐dichlorodiphenyldichloroethylene (p,p′‐DDE), and p,p′‐dichlorodiphenyldichloroethane (p,p′‐DDD). POPs are known to be particularly toxic and have been associated with endocrine‐disrupting effects in several mammals, including humans even at very low doses. As environmental estrogens, they could play a critical role in carcinogenesis, such as in breast cancer. With the purpose of evaluating their effect on breast cancer biology, o,p′‐DDT, p,p′‐DDE, and p,p′‐DDD (50–1000 nM) were tested on two human breast adenocarcinoma cell lines: MCF‐7 expressing estrogen receptor (ER) α and MDA‐MB‐231 negative for ERα, regarding cell proliferation and viability in addition to their invasive potential. Cell proliferation and viability were not equally affected by these compounds. In MCF‐7 cells, the compounds were able to decrease cell proliferation and viability. On the other hand, no evident response was observed in treated MDA‐MB‐231 cells. Concerning the invasive potential, the less invasive cell line, MCF‐7, had its invasion potential significantly induced, while the more invasive cell line MDA‐MB‐231, had its invasion potential dramatically reduced in the presence of the tested compounds. Altogether, the results showed that these compounds were able to modulate several cancer‐related processes, namely in breast cancer cell lines, and underline the relevance of POP exposure to the risk of cancer development and progression, unraveling distinct pathways of action of these compounds on tumor cell biology. © 2013 Wiley Periodicals, Inc. Environ Toxicol 30: 168–176, 2015.     

Polychlorinated biphenyls and organochlorine pesticides in human milk samples from two regions in Croatia

We analyzed 20 polychlorinated biphenyls (PCBs) and seven organochlorine pesticides (OCPs) in milk samples collected during 2009–2011 from primiparae living in two different regions in Croatia. p,p′-DDE is the dominant organochlorine pesticide. α-HCH/γ-HCH and p,p′-DDE/p,p′-DDT ratios indicate that there is fresh input of γ-HCH in investigated population on both locations, while this is not applicable to p,p′-DDT. The PCB profile was dominated by higher chlorinated congeners. Non-ortho PCB congeners which have the highest TEF values were not detected in any of individual samples. Toxic equivalents for mono-ortho substituted PCB congeners indicated higher exposure to toxic PCBs in Zadar, but estimated daily intakes for both locations indicate that infants consuming mother's milk are not at risk of adverse effects caused by PCBs and OCPs. Our study builds on the previous research of human milk samples collected in Zagreb and reveals that over 10-year period, levels of investigated organochlorine compounds decreased significantly.     

Biphasic hormonal responses to the adrenocorticolytic DDT metabolite 3-methylsulfonyl-DDE in human cells

The DDT metabolite 3-methylsulfonyl-DDE (3-MeSO₂-DDE) has been proposed as a lead compound for an improved adrenocortical carcinoma (ACC) treatment. ACC is a rare malignant disorder with poor prognosis, and the current pharmacological therapy o,p′-DDD (mitotane) has limited efficacy and causes severe adverse effects. 3-MeSO₂-DDE is bioactivated by cytochrome P450 (CYP) 11B1 in mice and causes formation of irreversibly bound protein adducts, reduced glucocorticoid secretion, and cell death in the adrenal cortex of several animal species. The present study was carried out to assess similarities and differences between mice and humans concerning the adrenocorticolytic effects of 3-MeSO₂-DDE. The results support previous indications that humans are sensitive to the adrenocorticolytic actions of 3-MeSO₂-DDE by demonstrating protein adduct formation and cytotoxicity in the human adrenocortical cell line H295R. However, neither the irreversible binding nor the cytotoxicity of 3-MeSO₂-DDE in H295R cells was inhibited by the CYP11B1 inhibitor etomidate. We also report biphasic responses to 3-MeSO₂-DDE in cortisol and aldosterone secretion as well as in mRNA levels of the steroidogenic genes StAR, CYP11B1 and CYP11B2. Hormone levels and mRNA levels were increased at lower concentrations of 3-MeSO₂-DDE, while higher concentrations decreased hormone levels. These biphasic responses were not observed with o,p′-DDD or with the precursor DDT metabolite p,p′-DDE. Based on these results, 3-MeSO₂-DDE remains a viable lead compound for drug design, although the adrenocorticolytic effects of 3-MeSO₂-DDE in human cells seem more complex than in murine cells.     

In utero DDT exposure and breast density before age 50

Prior studies in the Child Health and Development Studies (CHDS) found in utero exposure to the pesticide, dichlorodiphenyltrichloroethane (DDT), increased breast cancer risk by age 52. Mammographic density is considered a primary risk factor for breast cancer. We conducted a study of 309 daughters from the CHDS to examine in utero DDT exposure and mammographic density in midlife. Among daughters with high (>75th percentile) exposure to p,p’-Dichlorodiphenyldichloroethylene (DDE), p,p’-DDT was significantly correlated with increased dense area and percent density regardless of her body mass in midlife. In the subset of women with lower (<75th percentile) p,p-DDE, p,p’-DDT was associated with increased non-dense breast area. This was explained by adjustment for midlife BMI suggesting that p,p’-DDT may be obesogenic. In aggregate our findings indicate that early life p,p’-DDT exposure impacts breast density in a complex way that depends on the hosts biological ability to sequester and process DDT and levels of exposure.     

Mechanisms of p,p′-DDE-induced apoptosis in human peripheral blood mononuclear cells

p,p′-Dichlorodiphenyldichloroethylene (DDE), the most stable metabolite of organochlorine insecticide p,p′-dichlorodiphenyltrichloroethane (DDT), has been detected in human populations living in malaria-endemic areas of México where this insecticide was used. DDE induces apoptosis in peripheral blood mononuclear cells (PMBC); however, the molecular mechanism of cell death induced by this compound is poorly understood. In the present study, PBMC isolated from healthy individuals (not exposed to DDE) were incubated in the presence of increasing concentrations of p,p′-DDE (0–80 μg/ml) over time. When PBMC were treated with low p,p′-DDE concentration (10 μg/ml) an antioxidant response and biomarkers of inflammation were induced, indicating a pro-inflammatory state. Moreover, when PBMC were treated with high p,p′-DDE concentration (80 μg/ml) several apoptotic biochemical events were triggered, such as activation of caspase-8, Bid, caspase-9 and caspase-3, as well as degradation of PARP and ubiquitination. The results described in this study show a possible inflammatory condition and the involvement of both extrinsic and intrinsic pathways in the induction of apoptosis in DDE-treated PBMC.     

DDT/DDE and breast cancer: A meta-analysis

The biological basis for investigating dichlorodiphenyltrichloroethane (DDT) exposure and breast cancer risk stems from in vitro and animal studies indicating that DDT has estrogenic properties. The objective of this study was to update a meta-analysis from 2004 which found no association between dichlorodiphenyldichloroethylene (DDE) and breast cancer. We searched PubMed and Web of Science for studies published through June 2012 assessing DDT/DDE exposure and breast cancer. Summary Odds Ratios (ORs) with 95% confidence intervals (CIs) were calculated for the prevalence of breast cancer in the highest versus the lowest exposed groups for DDT and DDE. Difference of means of exposure for cases versus controls was analyzed for DDT and DDE. From the 500 studies screened, 46 were included in the meta-analysis. Slightly elevated, but not statistically significant summary ORs were found for DDE (1.05; 95% CI: 0.93–1.18) and DDT (1.02; 95% CI: 0.92–1.13). Lipid adjusted difference of means analysis found a significantly higher DDE concentration in cases versus controls (11.30 ng/g lipid; p = 0.01). No other difference of means analysis found significant relationships. The existing information does not support the hypothesis that exposure to DDT/DDE increases the risk of breast cancer in humans.     

Avian liver organochlorine and PCB from South coast of the Caspian Sea,Iran

Liver samples (n = 43) of 9 avian species representing the families Phalacrocoracidae, Podicipedidae, Laridae, and Anatidae, were collected from the Iranian coast of the Caspian Sea. Samples were analyzed for organochlorine pesticides (OCPs), such as dichlorodiphenyltrichloroethane (DDT) and its metabolites, hexachlorobenzene (HCB), hexachlorocyclohexane isomers (HCHs), and seven PCB congeners. p,p′-DDE was predominantly found in all species, at concentrations ranging from the limit of quantification (LOQ) to 340 ng/g ww. Most frequently encountered PCB congeners, in all samples, were 118, 153 and 138; and birds in Phalacrocoracidae had the highest liver PCB (mean 90 ± 32; ranging from <LOQ to 106 ng/g ww) whereas Podicipedidae had the highest OCP (mean 147 ± 49; ranging from <LOQ to 340 ng/g ww) (P < 0.05). Differences in the diet, and migratory routes, were important species-specific factors that affected hepatic concentration of OCP and PCB in the species we studied. Range of OCP and PCB concentrations in the present study was lower than those reported for birds in other regions of the world. Hepatic PCB concentration found in our avian species was below toxic effect levels that have been previously reported in birds. To our knowledge this is the first report of persistent organochlorine pollutants in liver of birds from Iran.     

The organochlorine p,p′-dichlorodiphenyltrichloroethane induces colorectal cancer growth through Wnt/β-catenin signaling

Dichlorodiphenyltrichloroethane (DDT), an organochlorine pollutant, is associated with several types of cancer. However, the relationship between DDT and colorectal cancer is uncertain. In this study, the impact of p,p′-DDT on colorectal cancer growth was evaluated using both in vitro and in vivo models. Our results indicated that the proliferation of human colorectal adenocarcinoma DLD1 cells was significantly promoted after exposed to low concentrations of p,p′-DDT ranging from 10⁻¹² to 10⁻⁷ M for 96 h. Exposure to p,p′-DDT from 10⁻¹⁰ to 10⁻⁸ M led to upregulation of phospho-GSK3β (Ser9), β-catenin, c-Myc and cyclin D1 in DLD1 cells. RNA interference of β-catenin inhibited the proliferation of DLD1 cells stimulated by p,p′-DDT. Inhibiting of estrogen receptors (ERs) had no significant effect on the action of p,p′-DDT. Treatment with p,p′-DDT induced production of intracellular reactive oxygen species (ROS) and inhibited superoxide dismutase (SOD) activity in DLD1 cells. Treatment with N-acetyl-L-cysteine (NAC), a ROS inhibitor, suppressed the induction of Wnt/β-catenin signaling and DLD1 cell proliferation by p,p′-DDT. Moreover, in a mouse xenograft model, 5 nmol/kg p,p′-DDT resulted in increased tumor size, oxidative stress and Wnt/β-catenin signaling. These results indicated that low concentrations of p,p′-DDT promoted colorectal cancer growth through Wnt/β-catenin signaling, which was mediated by oxidative stress. The finding suggests an association between low concentrations of p,p′-DDT exposure and colorectal cancer progression.     

p,p′‐Dichlorodiphenyltrichloroethane promotes aerobic glycolysis via reactive oxygen species–mediated extracellular signal‐regulated kinase/M2 isoform of pyruvate kinase (PKM2) signaling in colorectal cancer cells

Aerobic glycolysis is crucial to tumor cells to acquire energy for proliferation and metastasis. Dichlorodiphenyltrichloroethane (DDT), which is a persistent organic pollutant, has been associated with colorectal cancer (CRC) progressions, but the influence of p,p′‐DDT on CRC cell metabolism remains unclear. This study showed that exposure to low concentrations of p,p′‐DDT from 10^?11 to 10^?7M for 48 hours significantly increased glucose uptake and lactate production in colorectal adenocarcinoma cells, which were accompanied by the upregulation of proteins associated with aerobic glycolysis including glucose transporter1, lactate dehydrogenase A, and PDH kinase. We found p,p′‐DDT elevated the expression and nucleus translocation of M2 isoform of pyruvate kinase (PKM2), which was responsible for p,p′‐DDT–induced enhancement of aerobic glycolysis. Moreover, extracellular signal‐regulated kinase (ERK1/2) activation by p,p′‐DDT modulated the impacts of p,p′‐DDT on PKM2 and aerobic glycolysis. Treatment of p,p′‐DDT increased intracellular reactive oxygen species (ROS). N‐acetyl‐L ‐cysteine, an ROS inhibitor, prevented p,p′‐DDT–induced promotion of aerobic glycolysis, ERK1/2 activation, upregulation, and nucleus translocation of PKM2. Taken together, these results demonstrated that p,p′‐DDT promotes aerobic glycolysis via ROS‐mediated ERK/PKM2 signaling.     

Organochlorine pesticide residues in human breast tissue and their relationships with clinical and pathological characteristics of breast cancer

Agricultural pesticides are abundant environmental contaminants worldwide, prompting interest in studying their possible detrimental health effects. We examined organochlorine residues by quadrant (n = 245) in breast adipose tissues from 51 women with various stages of breast health to determine patterns of bioaccumulation within the breast and to assess relationships with patient clinical characteristics. Three organochlorine residues—2,2‐bis(p‐chlorophenyl)‐1,1‐dichloroethylene (p,p′‐DDE), hexachlorobenzene (HCB), and mirex—assayed by high resolution gas chromatography were abundant in breast tissue. p,p′‐DDE (745 ± 1054 ng/g lipid) was the most prevalent residue, comprising 97.5% of the total chemical burden. Mean levels of p,p′‐DDE and HCB were significantly correlated (P < .001) with patient age at mastectomy, and levels of p,p′‐DDE were correlated (P < .05) with BMI. Pesticide concentrations did not differ significantly by breast quadrant and were not different in the quadrant(s) where the primary tumor was located compared to other cancer‐free quadrants. In invasive cancer patients, organochlorine levels differed significantly based on clinical characteristics of the primary carcinoma, including stage, grade, ER status, and HER2 status, indicating that body burden of organochlorines may influence the development of specific subtypes of breast cancer. Potentially carcinogenic organochlorines were present at high levels within the human breast warranting further research to determine the impact of organochlorines in the etiology of breast cancer.     

Influence of dichlorodiphenylchloroethylene on vascular endothelial growth factor and insulin-like growth factor in human and rat ovarian cells

1,1-dichloro-2,2-bis(p-chlorophenyl)ethylene (p,p′-DDE, DDE), a metabolite of DDT is a persistent hormonally active environmental toxicant present in human serum and follicular fluid. The objective of this study was to investigate the effects of DDE on the expression of the ovarian vascular endothelial growth factor (VEGF) and insulin-like growth factor (IGF-1) in primary cultures of human granulosa cells and in the rat ovary. Granulosa cells were obtained at the time of oocyte retrieval for in vitro fertilization and cultured with environmentally relevant concentrations of DDE. Immature female rats were treated with 100 μg DDE/kg body weight or vehicle at 28 and 31 days of age and then euthanized at 50 days of age for collection of ovarian tissue. Expression of VEGF, the VEGF receptor fetal liver kinase (Flk-1) and IGF-1 were determined by Western blotting analysis of protein lysates from granulosa cell cultures and by immunohistochemistry in the rat ovary. DDE at concentrations of 100–1000 ng/mL increased the expression of VEGF, Flk-1 and IGF-1 in vitro in primary cultures of human granulosa cells, with the highest expression occurring at 1000 ng/mL. Similarly, acute administration of DDE resulted in a significant increase in immunoreactive VEGF, Flk-1 and IGF-1 in the rat ovary. We conclude that DDE, at levels, which have been detected in humans, alters the expression of the ovarian growth factors VEGF and IGF-1 both in vivo and in vitro. This alteration in expression of growth factors may lead to altered ovarian function as seen in polycystic ovaries and impaired fertility.     

p,p'-DDE induces apoptosis and mRNA expression of apoptosis-associated genes in testes of pubertal rats

One,1‐dichloro‐2,2 bis(p‐chlorophenyl) ethylene (p,p'‐DDE), the major metabolite of 2,2‐bis(4‐chlorophenyl)‐1,1,1‐trichloroethane (DDT), is a known persistent organic pollutant and male reproductive toxicant. It has antiandrogenic effect. However, the mechanism by which p,p'‐DDE exposure causes male reproductive toxicity remains unknown. To elucidate the mechanism underpinning the testicular effects of p,p'‐DDE, we sought to investigate apoptotic effects and mRNA expression of apoptosis‐associated genes in the testis of pubertal rats, including Fas, FasL, calpain‐1, cytochrome c, Bax, Bcl‐w, Bak, and caspase‐3, ‐8, ‐9, ‐12. Animals were administered with different doses of p,p'‐DDE (0, 20, 60, 100 mg/kg body weight) every other day by intraperitoneal injection for 10 days. The results indicated that p,p'‐DDE exposure at over 20 mg/kg body weight showed the induction of apoptotic cell death. p,p'‐DDE could induce decrease in SOD and GSH‐Px activity of serum in 60 mg/kg body weight group. Significant elevations in the mRNA levels of Fas, FasL, calpain‐1, cytochrome c, Bax, Bak, and caspase‐3, ‐8, ‐9, ‐12 were observed in testis of rat treated with p,p'‐DDE. Taken together, these results lead us to speculate that in vivo exposure to p,p'‐DDE might induce testicular apoptosis in pubertal rats through the involvement of Fas/FasL, mitochondria and endoplasmic reticulum‐mediated pathways. © 2011 Wiley Periodicals, Inc. Environ Toxicol 2013.