Melanie C. Coughlan

Bisphenol A activates the Nrf1/2-antioxidant response element pathway in HEK 293 cells.

Bisphenol A (BPA) is used in the production of polycarbonate plastics and epoxy resins for baby bottles, liners of canned food, and many other consumer products. Previously, BPA has been shown to reduce the activity of several antioxidant enzymes, which may contribute to oxidative stress. However, the underlying mechanism of the BPA-mediated effect upon antioxidant enzyme activity is unknown. Antioxidant and phase II metabolizing enzymes protect cells from oxidative stress and are transcriptionally activated by Nrf1 and Nrf2 factors through their cis-regulatory antioxidant response elements (AREs). In this work, we have assessed the effect of BPA on the Nrf1/2-ARE pathway in cultured human embryonic kidney (HEK) 293 cells. Surprisingly, glutathione and reactive oxygen species (ROS) assays revealed that BPA application created a more reduced intracellular environment in cultured HEK 293 cells. Furthermore, BPA increased the transactivation activity of ectopic Nrf1 and Nrf2 and increased the expression of ARE-target genes ho-1 and nqo1 at high (100-200 μM) BPA concentrations only. Our study suggests that BPA activates the Nrf1/2-ARE pathway at high (>10 μM) micromolar concentrations.

Exposure to a Northern Contaminant Mixture (NCM) Alters Hepatic Energy and Lipid Metabolism Exacerbating Hepatic Steatosis in Obese JCR Rats

Non-alcoholic fatty liver disease (NAFLD), defined by the American Liver Society as the buildup of extra fat in liver cells that is not caused by alcohol, is the most common liver disease in North America. Obesity and type 2 diabetes are viewed as the major causes of NAFLD. Environmental contaminants have also been implicated in the development of NAFLD. Northern populations are exposed to a myriad of persistent organic pollutants including polychlorinated biphenyls, organochlorine pesticides, flame retardants, and toxic metals, while also affected by higher rates of obesity and alcohol abuse compared to the rest of Canada. In this study, we examined the impact of a mixture of 22 contaminants detected in Inuit blood on the development and progression of NAFLD in obese JCR rats with or without co-exposure to10% ethanol. Hepatosteatosis was found in obese rat liver, which was worsened by exposure to 10% ethanol. NCM treatment increased the number of macrovesicular lipid droplets, total lipid contents, portion of mono- and polyunsaturated fatty acids in the liver. This was complemented by an increase in hepatic total cholesterol and cholesterol ester levels which was associated with changes in the expression of genes and proteins involved in lipid metabolism and transport. In addition, NCM treatment increased cytochrome P450 2E1 protein expression and decreased ubiquinone pool, and mitochondrial ATP synthase subunit ATP5A and Complex IV activity. Despite the changes in mitochondrial physiology, hepatic ATP levels were maintained high in NCM-treated versus control rats. This was due to a decrease in ATP utilization and an increase in creatine kinase activity. Collectively, our results suggest that NCM treatment decreases hepatic cholesterol export, possibly also increases cholesterol uptake from circulation, and promotes lipid accumulation and alters ATP homeostasis which exacerbates the existing hepatic steatosis in genetically obese JCR rats with or without co-exposure to ethanol.

Background bisphenol A in experimental materials and its implication to low-dose in vitro study.

In vitro low-dose studies are important to understand the mechanisms of bisphenol A (BPA) action. BPA doses used in current in vitro studies varied considerably, and doses as low as 10(-15)M have been reported. The actual doses of BPA used in the in vitro low-dose studies were rarely checked analytically, and the background BPA levels in experimental materials, which will determine the lowest BPA dose to be used, should be investigated or considered. In this study, the background BPA levels in various materials typically used in in vitro low-dose studies for BPA were investigated. Background BPA levels from the use of disposable pipettes and pipette tips were low (<0.20 ng mL(-1) or 0.88 nM). BPA was also detected in several commercial buffer solutions at levels close to the method limit of quantification (LOQ) (0.02 ng mL(-1); 0.088 nM). However, BPA was detected in all cell culture media obtained from various sources at levels ranging from 0.080 to 4.26 ng mL(-1) (or 0.35 to 19 nM) with an average of 0.83 ng mL(-1) (3.5 nM). We suggest that culture media used for low-dose BPA studies should be analysed for background BPA levels prior to use, and the medium with the lowest BPA levels should be used.

A Northern contaminant mixture impairs pancreas function in obese and lean JCR rats and inhibits insulin secretion in MIN6 cells.

Rates of obesity and diabetes mellitus of Arctic populations are increasing due to multiple reasons including a departure from traditional lifestyles and alcohol consumption patterns. These populations are also exposed to a variety of anthropogenic contaminants through consumption of contaminated country foods. We have previously shown that a Northern contaminant mixture (NCM), containing 22 organic and inorganic contaminants found in the blood of Canadian Arctic populations, induces endothelial cell dysfunction and exacerbates development of non-alcoholic fatty liver disease in experimental models. In order to determine if these contaminants affect pancreas function and physiology and if obesity and alcohol can influence contaminant toxicity and the development of diabetes, lean and obese JCR rats were orally treated with NCM at 0 (vehicle), 1.6 or 16mg/kg BW for four weeks in the presence or absence of 10% (v/v) alcohol. NCM treatment altered islet morphology, increased iron deposit in pancreas, and reduced circulating and pancreatic insulin levels and circulating glucagon levels as a result of direct islet injury with β and α cell loss with or without exposure to alcohol. Studies conducted with cultured mouse insulin-secreting (MIN6) β cells further demonstrated that NCM inhibited insulin release and induced cell death through oxidative stress and mitochondrial dysfunction. 2,3,4,6-Tetrabromophenol, a minor component of the NCM, alone also inhibited insulin release from MIN6 cells after 10min of exposure. These results suggest that Northern contaminants may contribute to pancreatic dysfunction, and possibly development of diabetes, in some of the highly exposed Arctic populations. The implications and relevance of these findings to Northern populations remains to be confirmed through epidemiological studies.

Bisphenol A exposure alters release of immune and developmental modulators and expression of estrogen receptors in human fetal lung fibroblasts

Bisphenol A (BPA) has been shown to exert biological effects through estrogen receptor (ER)-dependent and ER-independent mechanisms. Recent studies suggest that prenatal exposure to BPA may increase the risk of childhood asthma. To investigate the underlying mechanisms in the actions of BPA, human fetal lung fibroblasts (hFLFs) were exposed to varying doses of BPA in culture for 24hr. Effects of BPA on localization and uptake of BPA, cell viability, release of immune and developmental modulators, cellular localization and expression of ERα, ERβ and G-protein coupled estrogen receptor 30 (GPR30), and effects of ERs antagonists on BPA-induced changes in endothelin-1 (ET-1) release were examined. BPA at 0.01-100μmol/L caused no changes in cell viability after 24hr of exposure. hFLFs expresses all three ERs. BPA had no effects on either cellular distribution or protein expression of ERα, however, at 100μmol/L (or 23μmol/L intracellular BPA) increased ERβ protein levels in the cytoplasmic fractions and GPR30 protein levels in the nuclear fractions. These paralleled with increased release of growth differentiation factor-15, decreased phosphorylation of nuclear factor kappa B p65 at serine 536, and decreased release of ET-1, interleukin-6, and interferon gamma-induced protein 10. ERs antagonists had no effects on BPA-induced decrease in ET-1 release. These data suggest that BPA at 100μmol/L altered the release of immune and developmental modulators in hFLFs, which may negatively influence fetal lung development, maturation, and susceptibility to environmental stressors, although the role of BPA in childhood asthma remains to be confirmed in in vivo studies.

Dietary acrylamide exposure in male F344 rats: Dataset of systemic oxidative stress and inflammation markers

We previously reported that dietary acrylamide, at doses (10 and 50 mg/kg diet) known to cause rodent tumors, lowered serum total high density lipoprotein and total testosterone, increased serum lipase, and lowered lymphocytes levels together with other hematological parameters in male F344 rats exposed for 10 weeks (doi: 10.1016/j.etap.2014.11.009 [1]). Here we present data related to the role of food-borne acrylamide exposure (at 0, 5, 10 and 50 mg/kg diet) in the presence of low (7% wt/wt) or high (23.9% wt/wt) dietary fat on serum and urinary markers of oxidative stress and inflammation in F344 rats. Briefly, urine and serum samples were collected from the experimental animals a day prior to or at the time of necropsy, respectively and processed for enzyme-linked immunosorbent assay estimations of biochemical markers. Urine samples were analyzed for 8-hydroxydeoxyguanosine and isoprostane, and serum samples for total antioxidant capacity, paraoxonase 1 activity, c-reactive protein, homocysteine, oxidized low-density lipoprotein, intercellular adhesion molecule-1, thromboxin 2, and Nε-(carboxymethyl)lysine.