Interplay of Obesity, Ethanol, and Contaminant Mixture on Clinical Profiles of Cardiovascular and Metabolic Diseases: Evidence from an Animal Study.

Obesity, ethanol, and contaminants are known risk factors of cardiovascular and metabolic diseases (CMD). However, their interplay on clinical profiles of these diseases remains unclear, and thus were investigated in this study. Male lean or obese JCR rats were given water or 10% ethanol and orally treated with or without a contaminant mixture (CM) dissolved in corn oil and loaded on two cookies at 0, 1.6, or 16 mg/kg BW/day dose levels for 4 weeks. The CM consisted 22 environmental contaminants found in human blood or serum of Northern populations. Over 60 parameters related to CMD were examined. The results revealed that obesity in JCR rats resembles the clinical profiles of non-alcoholic fatty liver disease in humans. Obesity was also associated with increased serum and organ retention of mercury, one of the chemical components of CM. Exposure to ethanol lightened hyperlipidemia, increased liver retention of mercury, and increased risk for hypertension in the obese rats. CM lessened hyperlipidemia and hyperenzymemia, worsened systemic inflammation and increased the risk for hypertension in the obese rats. CM markedly increased serum ethanol levels with or without ethanol exposure. Tissue total mercury contents significantly correlated with clinical parameters with altered profiles by both ethanol and obesity. These results suggest that obese individuals may be more prone to contaminant accumulation. Ethanol and CM exposure can alter clinical profiles associated with obesity, which may lead to misdiagnosis of CMD associated with obesity. CM can alter endogenous production and/or metabolism of ethanol, further complicating disease progression, diagnosis, and treatment.

Exposure to Low Doses of Dechlorane Plus Promotes Adipose Tissue Dysfunction and Glucose Intolerance in Male Mice.

The prevalence of type 2 diabetes (T2D) continues to increase worldwide. It is well established that genetic susceptibility, obesity, overnutrition and a sedentary life style are risk factors for the development of T2D. However, more recently, studies have also proposed links between exposure to endocrine-disrupting chemicals (EDCs) and altered glucose metabolism. Human exposure to environmental pollutants that are suspected to have endocrine disruptor activity is ubiquitous. One such chemical is Dechlorane Plus (DP), a flame retardant, that is now detected in humans and the environment. Here we show that exposure of mice to low, environmentally relevant doses of DP promoted glucose intolerance in mice fed a high-fat diet independent of weight gain. Furthermore, DP had pronounced effects on the adipose tissue, where it induced the development of hypertrophied white adipose tissue (WAT), and increased serum levels of resistin, leptin, and plasminogen activator inhibitor-1. In addition, DP exposure induced "whitening" of brown adipose tissue (BAT), and reduced BAT uncoupling protein 1 expression. Importantly, some of these effects occurred even when the mice were fed a regular, low-fat, diet. Finally, WAT adipogenic markers were reduced with DP treatment in the WAT. We also show that DP directly inhibited insulin signaling in murine adipocytes and human primary subcutaneous adipocytes in vitro. Taken together, our results show that the exposure to low and environmentally relevant levels of DP may contribute to the development of T2D.

Comparison of tris(2-ethylhexyl) phosphate and di(2-ethylhexyl) phosphoric acid toxicities in a rat 28-day oral exposure study.

Tris(2-ethylhexyl) phosphate (TEHP, CAS no. 78-42-2) is a plasticizer and a flame retardant, while di(2-ethylhexyl) phosphoric acid (DEHPA, CAS no. 298-07-7) is an oil additive and extraction solvent. Publicly-available information on repeated exposure to these two related organophosphate compounds is fragmentary. Hence, adult male and female Fischer rats were exposed to TEHP (300, 1000 and 3000 mg/kg body weight [BW]/day) or DEHPA (20, 60 and 180 mg/kg BW/day) by gavage for 28 consecutive days, to assess and compare their toxicities. Although significantly impaired BW gains and evidence of TEHP enzymatic hydrolysis to DEHPA were observed only in males, exposures to the highest TEHP and DEHPA doses often resulted in similar alterations of hematology, serum clinical chemistry and liver enzymatic activities in both males and females. The squamous epithelial hyperplasia and hyperkeratosis observed in the non-glandular forestomach of rats exposed to the middle and high DEHPA doses were most likely caused by the slightly corrosive nature of this chemical. Although tubular degeneration and spermatid retention were observed only in the testes of males exposed to the highest TEHP dose, numerous periodic acid-Schiff stained crystalline inclusions were observed in testis interstitial cells at all TEHP dose levels. No-observed-adverse-effect levels for TEHP and DEHPA are proposed, but the lower serum pituitary hormone levels resulting from TEHP and DEHPA exposures and the perturbations of testicular histology observed in TEHP-treated males deserve further investigation. Improved characterization of the toxicity of flame retardants will contribute to better informed substitution choices for legacy flame retardants phased-out over health concerns.

Biodistribution and Systemic Effects in Mice Following Intravenous Administration of Cadmium Telluride Quantum Dot Nanoparticles.

Quantum dots (QDs) are engineered nanoparticles (NPs) of semiconductor structure that possess unique optical and electronic properties and are widely used in biomedical applications; however, their risks are not entirely understood. This study investigated the tissue distribution and toxic effects of cadmium telluride quantum dots (CdTe-QDs) in male BALB/c mice for up to 1 week after single-dose intravenous injections. CdTe-QDs were detected in the blood, lung, heart, liver, spleen, kidney, testis and brain. Most CdTe-QDs accumulated in the liver, followed by the spleen and kidney. At high doses, exposure to CdTe-QDs resulted in mild dehydration, lethargy, ruffled fur, hunched posture, and body weight loss. Histological analysis of the tissues, upon highest dose exposures, revealed hepatic hemorrhage and necrotic areas in the spleen. The sera of mice treated with high doses of CdTe-QDs showed significant increases in alanine aminotransferase (ALT), aspartate aminotransferase (AST), and total bilirubin levels, as well as a reduction in albumin. CdTe-QD exposure also led to a reduced number of platelets and elevated total white blood cell counts, including monocytes and neutrophils, serum amyloid A, and several pro-inflammatory cytokines. These results demonstrated that the liver is the main target of CdTe-QDs and that exposure to CdTe-QDs leads to hepatic and splenic injury, as well as systemic effects, in mice. By contrast, cadmium chloride (CdCl2), at an equivalent concentration of cadmium, appeared to have a different pharmacokinetic pattern from that of CdTe-QDs, having minimal effects on the aforementioned parameters, suggesting that cadmium alone cannot fully explain the toxicity of CdTe-QDs.

Toxicity of Flame Retardant Isopropylated Triphenyl Phosphate: Liver, Adrenal, and Metabolic Effects.

The use of organophosphates phosphate flame retardants, particularly isopropylated triphenyl phosphate (IPTPP), has increased in recent years as replacements for polybrominated diphenyl ethers. This is despite limited understanding of the hazards of IPTPP. To examine the general and endocrine toxicity of IPTPP, adult Wistar rats were fed for 90 days on diets containing IPTPP estimated to deliver daily doses of 5 to 140 mg/kg/d. Exposure to IPTPP caused a dose-related increase in liver and adrenal gland weight in both sexes. Cells in the zona fasciculate (ZF) of the adrenal cortex were observed to be filled with droplets that stained with Nile red, suggesting they contained neutral lipid. Despite marked structural changes, there was no change in basal or stress-induced serum levels of their major secreted ZF product corticosterone (B), suggesting cell function was not altered. There were no effects on responses to glucose or insulin challenge, but serum levels of fructosamine were elevated by IPTPP exposure, suggesting a slight tendency of exposed animals to be hyperglycemic. Serum levels of total cholesterol and high-density lipoprotein cholesterol were significantly elevated in both sexes at the 2 highest doses. This study demonstrates that IPTPP exposure causes hypertrophy and neutral lipid accumulation in adrenal cortex ZF cells but does not result in impaired B production.

A reproductive and developmental screening study of the fungal toxin ochratoxin A in Fischer rats.

The presence of the mycotoxin ochratoxin A (OTA) in cereal grains is due to the growth of toxigenic Penicillium mold on stored crops. Human exposure to OTA is higher in infants, toddlers, and children than in adolescents and adults, based on exposure assessments of ng OTA consumed/kg body weight/day. Ochratoxin A is nephrotoxic and teratogenic in animals, but its effects on juveniles exposed during the reproduction and development period have not been studied. To address this, Fischer rats were exposed to 0, 0.16, 0.4, 1.0, or 2.5 mg OTA/kg diet throughout breeding, gestation, and lactation and its adverse effects were assessed in adult rats and their offspring on postnatal day (PND) 21. There were no effects on implantation but post-implantation fetotoxicity was observed in the 2.5 mg/kg dose group, corresponding to a calculated dose of 167.0 µg/kg bw/day in dams. Adverse effects on body and kidney weights and on clinical parameters indicative of renal toxicity were significant in adult rats exposed to 1.0 mg OTA/kg diet (55.2 and 73.3 µg/kg bw/day in adult males and females, respectively) and in PND21 rats at the 0.4 mg/kg dose (33.9 µg/kg bw/day in dams), suggesting that weanling rats were more sensitive to OTA than adults. Overall, nephrotoxicity was the primary effect of OTA in weanling rats exposed throughout gestation and lactation at sub-fetotoxic concentrations in diet.

Acellular filtrate of a microbial-based cleaning product potentiates house dust mite allergic lung inflammation.

Microbial-based cleaning products (MBCPs) contain bacteria and chemical constituents. They are used in consumer applications such as odor reduction, unclogging drains, and surface cleaning. To determine the capacity of a model MBCP to contribute to acute allergic lung inflammation, a two-week repeated exposure regimen was used. Mice were exposed by endotracheal instillation to saline alone, MBCP alone, house dust mites (HDM) alone, or sequentially (i.e., MBCP followed by HDM, HDM followed by MBCP, or HDM + MBCP followed by HDM). Both whole MBCP and acellular MBCP filtrate were investigated, and showed minimal differences in the endpoints examined. HDM exposure caused pulmonary perivascular inflammation, bronchiolar mucous cell metaplasia, elevated bronchoalveolar lavage fluid (BALF) eosinophils, and HDM-specific IgG1. For MBCP, notable changes were associated with sequential exposures. MBCP/HDM caused elevated TH2 cytokines in BALF, and elevated neutrophils, eosinophils and IL-5 in peripheral blood. Co-administration of MBCP and HDM followed by HDM resulted in elevated blood and BALF eosinophils and HDM-specific IgE and IgG1. These results demonstrated that acellular MBCP filtrate, and not bacteria within MBCPs, potentiated the acute allergic inflammation to HDM. This methodology could be extended to investigate chronic allergic inflammation and inflammatory potential of other MBCPs and biotechnology products with complex compositions.

Gestational and Lactational Exposure to an Environmentally-Relevant Mixture of Brominated Flame Retardants: Effects on Neurodevelopment and Metabolism.

BACKGROUND: Developmental exposure to brominated flame retardants (BFRs), including polybrominated diphenyl ethers (PBDEs) and hexabromocyclododecane (HBCDD), has been associated with impaired neurodevelopment and some symptoms of metabolic syndrome. However, there are inconsistencies in studies reporting neurodevelopmental effects with studies of pure substances more likely to report effects than studies of technical products. In addition, the influence of early BFR exposures on later development of metabolic disease-like symptoms has not been investigated. This study examined the effects of perinatal exposure to an environmentally relevant mixture of BFRs based on relative levels observed in house dust, on several markers of neurodevelopment and metabolism in offspring. METHODS: Sprague-Dawley female rats were fed a diet estimated to deliver daily doses of 0, 0.06, 20, or 60 mg/kg of a mixture of PBDEs and HBCDD from before mating to weaning. Offspring were weaned to control diet and subjected to neurobehavioral and metabolic assessments. RESULTS: Exposure to BFRs decreased vertical movement in at postnatal day (PND) 32 and increased time to emerge to a lighted area on PND 105 in offspring of both sexes. Although early life exposure to the BFR mixture did not impact measures of glucose or insulin action, male offspring had significantly decreased fat pad weights at PND 46. Total cholesterol was increased in male and female offspring exposed to the highest dose at PND 21. CONCLUSIONS: These results suggest that gestational and lactational exposure to an environmentally relevant BFR mixture may induce changes in neurodevelopment and lipid metabolism in offspring. Birth Defects Research 109:497-512, 2017.© 2017 The Authors Birth Defects Research Published by Wiley Periodicals, Inc.

Murine MTHFD1-synthetase deficiency, a model for the human MTHFD1 R653Q polymorphism, decreases growth of colorectal tumors.

The common R653Q variant (∼20% homozygosity in Caucasians) in the synthetase domain of the folate-metabolizing enzyme MTHFD1 reduces purine synthesis. Although this variant does not appear to affect risk for colorectal cancer, we questioned whether it would affect growth of colorectal tumors. We induced tumor formation in a mouse model for MTHFD1-synthetase deficiency (Mthfd1S+/- ) using combined administration of azoxymethane (AOM) and dextran sodium sulfate (DSS) in male and female wild-type and Mthfd1S+/- mice. Tumor size was significantly smaller in MthfdS+/- mice, particularly in males. A reduction in the proliferation of MthfdS+/- mouse embryonic fibroblast cell lines, compared with wild-type lines, was also observed. Tumor number was not influenced by genotype. The amount of inflammation observed within tumors from male Mthfd1S+/- mice was lower than that in wild-type mice. Gene expression analysis in tumor adjacent normal (pre-neoplastic) tissue identified several genes involved in proliferation (Fosb, Fos, Ptk6, Esr2, Atf3) and inflammation (Atf3, Saa1, TNF-α) that were downregulated in MthfdS+/- males. In females, MthfdS+/- genotype was not associated with these gene expression changes, or with differences in tumor inflammation. These findings suggest that the mechanisms directing tumor growth differ significantly between males and females. We suggest that restriction of purine synthesis, reduced expression of genes involved in proliferation, and/or reduced inflammation lead to slower tumor growth in MTHFD1-synthetase deficiency. These findings may have implications for CRC tumor growth and prognosis in individuals with the R653Q variant. © 2016 Wiley Periodicals, Inc.

Toxicologic effects of 28-day dietary exposure to the flame retardant 1,2-dibromo-4-(1,2-dibromoethyl)-cyclohexane (TBECH) in F344 rats.

The brominated flame retardant TBECH is used as an additive to delay ignition and inhibit fires in construction materials and consumer goods. Trends in human exposure are not clear, although humans may be exposed to TBECH via indoor dust and air. In birds and fish there is some evidence of disruption in endocrine and reproductive parameters due to TBECH. In vitro studies indicate that TBECH is an androgen receptor agonist. In this study rats were exposed to 0, 10, 50, 250, 1250 or 5000mg/kg technical TBECH for 28days in diet, corresponding to 0, 0.9, 4.2, 21.3, 98.0 or 328.9mg TBECH/kg bw/d in males and 0, 0.8, 3.9, 19.4, 91.7 or 321.4mg TBECH/kg bw/d in females. Dose-dependent increases in α- and ß- TBECH were detected in serum, liver and adipose. Rats in the 5000mg/kg group lost weight rapidly and were euthanized after 15-18days. At study termination rats displayed dose-dependent clinical and histopathological changes consistent with mild hepatic and renal inflammation. In male rats, evidence of gender-specific alpha2u-globulin nephropathy was not considered predictive of renal toxicity in humans. Frank immunosuppression or inappropriate immunostimulation were not apparent, nor was there a primary effect of TBECH on adaptive immunity. Some evidence of hormone disruption was observed, including changes in serum testosterone levels in males and changes in serum T3 and T4 levels in females. Apparent increases in thyroid follicular cell hypertrophy and hyperplasia in male and female rats were not statistically significant. Benchmark dose (BMD) modelling indicated that clinical changes indicative of mild nephrotoxicity and increased blood monocyte numbers indicative of inflammation and tissue damage were the most sensitive outcomes of TBECH exposure that could be modelled. Preliminary evidence of hormone disruption supports the need for rodent studies using more sensitive models of growth, development and reproduction.

Immunomodulation by gastrointestinal carbon black nanoparticle exposure in ovalbumin T cell receptor transgenic mice.

Humans could become exposed to carbon black nanoparticles (CBNPs) in consumer products or an occupational setting. In rodent models, acute respiratory, subcutaneous, and direct immune cell exposure to CBNPs has been shown to enhance allergic sensitization to co-administered ovalbumin (OVA) protein from chicken egg. However, little is known about the effects of ingested CBNPs on immunological responses and oral tolerance to food antigens. We hypothesized that ingestion of CBNPs would enhance the development of food allergy to OVA. Allergy prone DO11.10 mice were orally exposed to CBNPs every second day for 2 weeks (total dose 10.8 (LOW) or 108 µg (HI)), with and without (±) co-administered OVA. Systemic immune parameters were measured at necropsy. Exposure to OVA resulted in significant increases in serum anti-OVA IgG1, anti-OVA IgM, and anti-OVA IgA antibodies relative to vehicle control. Immunophenotyping revealed a reduction in the number of OVA-specific CD4+ T helper cells upon OVA ± CBNPHI treatment in the spleen. Yet, secretion of the allergy-associated Th2 cytokines IL-4, IL-9, and IL-13 was greater in OVA323-339 peptide-pulsed splenocytes from OVA + CBNPHI-treated mice compared with control. Transcriptome analysis at necropsy of splenocytes from OVA + CBNPHI dose mice compared with OVA mice revealed increases in the allergy associated genes Il4 and Stat6 and decreases in Csf3r and Retnlg. Although oral exposure to high-dose CBNPs did not impact OVA-specific antibody production relative to OVA, we did observe increased expression of genes and cytokines associated with allergy in peripheral splenocytes. This work suggests that CBNP gastrointestinal exposure may potentiate allergy pathways.

Lack of adverse health effects following 30-weeks of dietary exposure to acrylamide at low doses in male F344 rats.

Understanding the health hazards following exposure to food-borne acrylamide, especially at low levels typified by human diets, is an ongoing food safety issue. We recently published results from a study that aimed to understand the effects of acrylamide short-term exposure at doses known to cause tumors in rodents, demonstrating that a number of key toxicological end points were altered by acrylamide exposure. Additionally, we reported that at much lower doses for 30 weeks of exposure, dietary acrylamide was 'not a complete carcinogen' to the colon in an organ-specific rodent carcinogenesis study but acted as a co-carcinogen along with azoxymethane (AOM, a colon-specific carcinogen). Here, we present toxicological data from a sub-set of this long-term exposure study from animals that received saline (instead of AOM). Briefly, male F344 rats were randomized to receive acrylamide at 0.5, 1.0 and 2.0 mg/kg diet (∼0.02, 0.04, and 0.09 mg/kg BW/day, respectively) or no acrylamide (control), for 30 weeks; all rats were then euthanized and their tissues harvested and processed for toxicological evaluation. We report that at the doses tested, acrylamide did not cause any changes in general well-being, body weight or food intake. Similarly, acrylamide did not cause any biologically relevant change in parameters associated with immunophenotyping, serum biochemistry or hematology. Histopathology assessment of tissues showed no changes except in the testis, where non-specific mild lesions were observed in all the groups, inclusive of the controls. No neuropathological effects of acrylamide were observed in the brain and nerve tissues. Together, these results suggest that acrylamide administered to rats through the diet at low doses for 30 weeks did not cause any toxicologically relevant changes. Given that the doses of acrylamide in the current study are low and are comparable to human dietary exposure, this null-effect study provides data that contribute to the body of scientific evidence relevant to understanding the health effects of acrylamide.

Toxicology of 3-epi-deoxynivalenol, a deoxynivalenol-transformation product by Devosia mutans 17-2-E-8.

Microbial detoxification of deoxynivalenol (DON) represents a new approach to treating DON-contaminated grains. A bacterium Devosia mutans 17-2-E-8 was capable of completely transforming DON into a major product 3-epi-DON and a minor product 3-keto-DON. Evaluation of toxicities of these DON-transformation products is an important part of hazard characterization prior to commercialization of the biotransformation application. Cytotoxicities of the products were demonstrated by two assays: a MTT bioassay assessing cell viability and a BrdU assay assessing DNA synthesis. Compared with DON, the IC50 values of 3-epi-DON and 3-keto-DON were respectively 357 and 3.03 times higher in the MTT bioassay, and were respectively 1181 and 4.54 times higher in the BrdU bioassay. Toxicological effects of 14-day oral exposure of the B6C3F1 mouse to DON and 3-epi-DON were also investigated. Overall, there were no differences between the control (free of toxin) and the 25 mg/kg bw/day or 100 mg/kg bw/day 3-epi-DON treatments in body and organ weights, hematology and organ histopathology. However, in mice exposed to DON (2 mg/kg bw/day), white blood cell numbers and serum immunoglobulin levels were altered relative to controls, and lesions were observed in adrenals, thymus, stomach, spleen and colon. Taken together, in vitro and in vivo studies indicate that 3-epi-DON is substantially less toxic than DON.

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 ß and α cell loss with or without exposure to alcohol. Studies conducted with cultured mouse insulin-secreting (MIN6) ß 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.

Toxicological effects of short-term dietary acrylamide exposure in male F344 rats.

We recently reported that acrylamide, a known rodent and probable human carcinogen, does not increase the risk of azoxymethane (AOM)-induced rat colon precancerous lesions when administered through the diet. Here, we present toxicological data from non-AOM-injected rats. Briefly, male F344 rats were randomized into four dietary groups and received experimental diets based on AIN-93G formulation and containing acrylamide at 0 (control), 5, 10 or 50mg/kg diet (wt/wt) ad libitum for 10 weeks, after which they were killed and their blood collected for hematological and biochemical markers. Acrylamide at the higher doses (10 and 50mg/kg diet) significantly lowered (p<0.05) serum total high density lipoprotein and total testosterone and increased serum lipase in comparison to the control. Blood hematocrit values and lymphocyte counts were significantly lower (p<0.05) in the high dose acrylamide (50mg/kg diet) group compared to control, with a concomitant decrease in hemoglobin level, mean corpuscular volume and mean corpuscular hemoglobin. These results provide additional hazard characterization data and strengthen the notion that at high doses, acrylamide may involve systemic toxicity potentiating tumorigenesis in experimental animals. Further studies are required to understand the health effects of food-borne acrylamide, especially at the lower exposures typified by human diets.

Fermentable Carbohydrates Differentially Affect Colon Tumor Formation in Azoxymethane-Induced Male Fischer 344 Rats.

BACKGROUND: The role of fermentation compared with the source or type of the fermentable material in colon tumorigenesis remains an issue in refining the definition of dietary fiber (DF). OBJECTIVE: The aim of this study was to investigate the fermentation and source-specific effects of various carbohydrates in a medium-term colon tumorigenesis model. METHODS: Six-week-old male Fischer 344 rats were randomly allocated into 6 groups (n = 36/group) to receive either AIN-93G (control) or diets containing fructooligosaccharides, wheat bran (WB), oat bran (OB), polydextrose, or high-amylose maize starch (HAMS), each adjusted to contain a total DF concentration of 7% (wt:wt) and have a fermentability of 3% (wt:wt). After 2 wk, 24 rats/group received 2 subcutaneous doses of azoxymethane (at 15 mg/kg body weight) 1 wk apart while 12 rats/group were injected with a saline vehicle; all rats were maintained on the assigned diets for 24 wk postinjection and then killed. Colon tumor outcomes and pathology together with cecal short-chain fatty acid composition were assessed. RESULTS: No tumors were found in saline-injected rats, and all subsequent analyses were restricted to azoxymethane-injected rats. Colon tumor incidence was significantly lower in the polydextrose (21%) and WB (13%) groups than in the control group (63%; P < 0.05) but not different from the fructooligosaccharide (58%), HAMS (46%), and OB (33%) groups. In comparison to the control group (8 proximal/31 total tumors), fermentable materials reduced the number of tumors (P < 0.05) originating in the proximal colon: HAMS (5/15), polydextrose (2/7), OB (2/9), fructooligosaccharides (1/21), and WB (1/3). The mean ± SEM number of tumors/tumor-bearing rats was significantly lower in the WB (1.00 ± 0.00), OB (1.13 ± 0.13), and HAMS (1.36 ± 0.15) groups than in the control group (2.07 ± 0.27; P < 0.02); other groups did not differ. The mean ± SEM tumor burden/diet group was lower in the WB (1.2 ± 0.7 mm2), polydextrose (6.7 ± 3.2 mm2), and OB (7.0 ± 3.0 mm2) groups than in the control (21.4 ± 5.9 mm2) and fructooligosaccharide (22.1 ± 7.1 mm2; P < 0.05) groups but not significantly different from the HAMS group (15.1 ± 6.1 mm2). Total cecal SCFA concentrations did not differ among diet groups (overall mean ± SEM: 81 ± 4 µmol/g wet weight). CONCLUSION: The rate and extent of fermentation of the carbohydrate material as well as the characteristics of the material in the lumen of the lower gastrointestinal tract all appear to have an important role in tumor outcomes in the azoxymethane-induced rat colon tumorigenesis assay.

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 to 10% 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.

Negligible colon cancer risk from food-borne acrylamide exposure in male F344 rats and nude (nu/nu) mice-bearing human colon tumor xenografts.

Acrylamide, a possible human carcinogen, is formed in certain carbohydrate-rich foods processed at high temperature. We evaluated if dietary acrylamide, at doses (0.5, 1.0 or 2.0 mg/kg diet) reflecting upper levels found in human foods, modulated colon tumorigenesis in two rodent models. Male F344 rats were randomized to receive diets without (control) or with acrylamide. 2-weeks later, rats in each group received two weekly subcutaneous injections of either azoxymethane (AOM) or saline, and were killed 20 weeks post-injections; colons were assessed for tumors. Male athymic nude (nu/nu) mice bearing HT-29 human colon adenocarcinoma cells-derived tumor xenografts received diets without (control) or with acrylamide; tumor growth was monitored and mice were killed 4 weeks later. In the F344 rat study, no tumors were found in the colons of the saline-injected rats. However, the colon tumor incidence was 54.2% and 66.7% in the control and the 2 mg/kg acrylamide-treated AOM-injected groups, respectively. While tumor multiplicity was similar across all diet groups, tumor size and burden were higher in the 2 mg/kg acrylamide group compared to the AOM control. These results suggest that acrylamide by itself is not a "complete carcinogen", but acts as a "co-carcinogen" by exacerbating the effects of AOM. The nude mouse study indicated no differences in the growth of human colon tumor xenografts between acrylamide-treated and control mice, suggesting that acrylamide does not aid in the progression of established tumors. Hence, food-borne acrylamide at levels comparable to those found in human foods is neither an independent carcinogen nor a tumor promoter in the colon. However, our results characterize a potential hazard of acrylamide as a colon co-carcinogen in association with known and possibly other environmental tumor initiators/promoters.

Dietary folate does not significantly affect the intestinal microbiome, inflammation or tumorigenesis in azoxymethane-dextran sodium sulphate-treated mice.

Inflammatory bowel disease (IBD) is a risk factor for the development of colon cancer. Environmental factors including diet and the microflora influence disease outcome. Folate and homocysteine have been associated with IBD-mediated colon cancer but their roles remain unclear. We used a model of chemically induced ulcerative colitis (dextran sodium sulphate (DSS)) with or without the colon carcinogen azoxymethane (AOM) to determine the impact of dietary folic acid (FA) on colonic microflora and the development of colon tumours. Male mice (n 15 per group) were fed a FA-deficient (0 mg/kg), control (2 mg/kg) or FA-supplemented (8 mg/kg) diet for 12 weeks. Folate status was dependent on the diet (P< 0·001) and colitis-induced treatment (P= 0·04) such that mice with colitis had lower circulating folate. FA had a minimal effect on tumour initiation, growth and progression, although FA-containing diets tended to be associated with a higher tumour prevalence in DSS-treated mice (7-20 v. 0%, P= 0·08) and the development of more tumours in the distal colon of AOM-treated mice (13-83% increase, P= 0·09). Folate deficiency was associated with hyperhomocysteinaemia (P< 0·001) but homocysteine negatively correlated with tumour number (r - 0·58, P= 0·02) and load (r - 0·57, P= 0·02). FA had no effect on the intestinal microflora. The present data indicate that FA intake has no or little effect on IBD or IBD-mediated colon cancer in this model and that hyperhomocysteinaemia is a biomarker of dietary status and malabsorption rather than a cause of IBD-mediated colon cancer.

Effects of long term exposure to the mycotoxin fumonisin B1 in p53 heterozygous and p53 homozygous transgenic mice.

The fungal toxin fumonisin B1 (FB1) is a potential human carcinogen based on evidence of renal carcinogenicity in rats and hepatocarcinogenicity in mice. The toxicity and carcinogenicity of FB1 is linked to ceramide synthase inhibition. Based on this mechanism of action and on lack of evidence of genotoxicity, FB1 is considered a non-genotoxic carcinogen. The p53 heterozygous (p53+/-) mouse is a cancer-prone model used for carcinogenesis. The effects of chronic dietary FB1 exposure were characterized in p53+/- mice to confirm non-genotoxicity using a model which is more sensitive to genotoxic than non-genotoxic carcinogens and to clarify the relationship between p53 expression, altered sphingolipid metabolism, and FB1-induced carcinogenesis. Responses to FB1 were similar in p53+/- and p53+/+ mice after 26 weeks exposure to 0, 5, 50 or 150 mg FB1/kg diet, supporting a non-genotoxic mechanism of action. Hepatic adenomas and cholangiomas were observed in mice exposed to 150 mg/kg FB1. For a 10% increase in hepatic megalocytosis, the estimated 95% lower confidence limit of the benchmark dose (BMDL10) ranged from 0.15 and 1.11 mg FB1/kg bw/day. Based on similar responses in p53+/- and p53+/+ mice, p53 and related pathways play a secondary role in responses to FB1 toxicity and carcinogenesis.