Urinary mutagenicity and bladder cancer risk in northern New England.

The etiology of bladder cancer among never smokers without occupational or environmental exposure to established urothelial carcinogens remains unclear. Urinary mutagenicity is an integrative measure that reflects recent exposure to genotoxic agents. Here, we investigated its potential association with bladder cancer in rural northern New England. We analyzed 156 bladder cancer cases and 247 cancer-free controls from a large population-based case-control study conducted in Maine, New Hampshire, and Vermont. Overnight urine samples were deconjugated enzymatically and the extracted organics were assessed for mutagenicity using the plate-incorporation Ames assay with the Salmonella frameshift strain YG1041 + S9. Logistic regression was used to estimate the odds ratios (OR) and 95% confidence intervals (CI) of bladder cancer in relation to having mutagenic versus nonmutagenic urine, adjusted for age, sex, and state, and stratified by smoking status (never, former, and current). We found evidence for an association between having mutagenic urine and increased bladder cancer risk among never smokers (OR = 3.8, 95% CI: 1.3-11.2) but not among former or current smokers. Risk could not be estimated among current smokers because nearly all cases and controls had mutagenic urine. Urinary mutagenicity among never-smoking controls could not be explained by recent exposure to established occupational and environmental mutagenic bladder carcinogens evaluated in our study. Our findings suggest that among never smokers, urinary mutagenicity potentially reflects genotoxic exposure profiles relevant to bladder carcinogenesis. Future studies are needed to replicate our findings and identify compounds and their sources that influence bladder cancer risk.

The carcinogenic response of Tsc2 mutant Long-Evans (Eker) rats to a mixture of drinking water disinfection by-products was less than additive.

Cancer risk assessment methods for chemical mixtures in drinking water are not well defined. Current default risk assessments for chemical mixtures assume additivity of carcinogenic effects, but this may not represent the actual biological response. A rodent model of hereditary renal cancer (Eker rat) was used to evaluate the carcinogenicity of mixtures of water disinfection by-products (DBPs). Male and female Eker rats were treated with individual DBPs or a mixture of DBPs for 4 or 10 months. Potassium bromate, 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone, chloroform, and bromodichloromethane were administered in drinking water at low concentrations of 0.02, 0.005, 0.4, and 0.07 g/l, respectively, and high concentrations of 0.4, 0.07, 1.8, and 0.7 g/l, respectively. Low and high dose mixture solutions comprised all four chemicals at either the low or the high concentrations, respectively. Body weights, water consumption, and chemical concentrations in the water were measured monthly. All tissues were examined macroscopically for masses and all masses were diagnosed microscopically. Total renal lesions (adenomas and carcinomas) were quantitated microscopically in male and female rats treated for 4 or 10 months. A dose response for renal tumors was present in most treatment groups after 4 or 10 months of treatment. Treatment with the mixture produced on average no more renal, splenic, or uterine tumors than the individual compound with the greatest effect. This study suggests that the default assumption of additivity may overestimate the carcinogenic effect of chemical mixtures in drinking water.

Cholinesterase inhibition and depression of the photic after discharge of flash evoked potentials following acute or repeated exposures to a mixture of carbaryl and propoxur.

Previously, we reported that acute treatment with propoxur or carbaryl decreased the duration of the photic after discharge (PhAD) of flash evoked potentials (FEPs). In the current studies, we compared the effects of acute or repeated exposure to a mixture of carbaryl and propoxur (1:1.45 ratio; propoxur:carbaryl) on the duration of the PhAD and brain ChE activity in Long Evans rats. Animals were exposed (po) either to a single dose (0, 3, 10, 45 or 75 mg/kg), or 14 daily dosages (0, 3, 10, 30, 45 mg/kg), of the mixture. Acute and repeated treatment with 3mg/kg (or greater) of the mixture produced dose-related inhibition of brain ChE activity. Compared to controls, the PhAD duration decreased after acute administration of 75 mg/kg or repeated treatment with 30 mg/kg of the mixture. The linear relationship between the percent of control brain ChE activity and the PhAD duration was similar for both exposure paradigms. Dose-response models for the acute and repeated exposure data did not differ for brain ChE activity or the duration of the PhAD. Repeated treatment with the mixture resulted in slightly less (13-22%) erythrocyte ChE inhibition than acute exposure. Both acute and repeated treatment resulted in dose-additive results for the PhAD duration and less than dose-additive responses (6-16%) for brain ChE activity for the middle range of dosages. Acute treatment resulted in greater than dose-additive erythrocyte ChE inhibition (15-18%) at the highest dosages. In contrast, repeated treatment resulted in less than dose-additive erythrocyte ChE inhibition (16-22%) at the middle dosages. Brain and plasma levels of propoxur and carbaryl did not differ between the acute and repeated dosing paradigms. In summary, a physiological measure of central nervous system function and brain ChE activity had similar responses after acute or repeated treatment with the carbamate mixture, and brain ChE showed only small deviations from dose-additivity. Erythrocyte ChE activity had larger differences between the acute and repeated treatment paradigms, and showed slightly greater deviations from dose-additivity. Because these treatments utilized larger dosages than anticipated environmental exposures, concern for non-additive effects in humans is minimized. The small magnitude of the deviations from dose-additivity also suggest that in the absence of repeated exposure data, results from an acute study of readily reversible carbamate toxicity can be used to estimate the response to repeated daily exposures.

Inhibition of fried meat-induced colorectal DNA damage and altered systemic genotoxicity in humans by crucifera, chlorophyllin, and yogurt.

UNLABELLED: Dietary exposures implicated as reducing or causing risk for colorectal cancer may reduce or cause DNA damage in colon tissue; however, no one has assessed this hypothesis directly in humans. Thus, we enrolled 16 healthy volunteers in a 4-week controlled feeding study where 8 subjects were randomly assigned to dietary regimens containing meat cooked at either low (100°C) or high temperature (250°C), each for 2 weeks in a crossover design. The other 8 subjects were randomly assigned to dietary regimens containing the high-temperature meat diet alone or in combination with 3 putative mutagen inhibitors: cruciferous vegetables, yogurt, and chlorophyllin tablets, also in a crossover design. Subjects were nonsmokers, at least 18 years old, and not currently taking prescription drugs or antibiotics. We used the Salmonella assay to analyze the meat, urine, and feces for mutagenicity, and the comet assay to analyze rectal biopsies and peripheral blood lymphocytes for DNA damage. Low-temperature meat had undetectable levels of heterocyclic amines (HCAs) and was not mutagenic, whereas high-temperature meat had high HCA levels and was highly mutagenic. The high-temperature meat diet increased the mutagenicity of hydrolyzed urine and feces compared to the low-temperature meat diet. The mutagenicity of hydrolyzed urine was increased nearly twofold by the inhibitor diet, indicating that the inhibitors enhanced conjugation. Inhibitors decreased significantly the mutagenicity of un-hydrolyzed and hydrolyzed feces. The diets did not alter the levels of DNA damage in non-target white blood cells, but the inhibitor diet decreased nearly twofold the DNA damage in target colorectal cells. To our knowledge, this is the first demonstration that dietary factors can reduce DNA damage in the target tissue of fried-meat associated carcinogenesis. TRIAL REGISTRATION: ClinicalTrials.gov NCT00340743.

A mixture of ammonium perchlorate and sodium chlorate enhances alterations of the pituitary-thyroid axis caused by the individual chemicals in adult male F344 rats.

Ammonium perchlorate (AP) and sodium chlorate (SC) have been detected in public drinking water supplies in many parts of the United States. These chemicals cause perturbations in pituitary-thyroid homeostasis in animals by competitively inhibiting iodide uptake, thus hindering the synthesis of thyroglobulin and reducing circulating T(4) (thyroxine). Little is known about the short-term exposure effects of mixtures of perchlorate and chlorate. The present study investigated the potential for the response to a mixture of these chemicals on the pituitary-thyroid axis in rats to be greater than that induced by the individual chemicals. Adult male F-344 rats were exposed, via their drinking water, to the nominal concentrations of 0.1, 1.0, 10 mg/L AP or 10, 100, 1000 mg/L SC and their mixtures for 7 days. Serum T(4) levels were significantly (p < 0.05) reduced in rats following exposure to the mixtures, but not after exposure to the individual chemicals. Serum T(3) (triiodothyronine) was not altered by treatment and TSH (thyroid stimulating hormone) was only increased after the high-dose chlorate treatment. Histological examination of the thyroid gland showed colloid depletion and hypertrophy of follicular epithelial cells in high-dose single chemical and all mixture-treated rats, while hyperplasia was observed only in some of the rats treated with mixtures (AP 10 + SC 100, AP 0.1 + SC 1000, and AP 10 + SC 1000 mg/L). These data suggest that short-term exposure to the mixture of AP and SC enhances the effect of either chemical alone on the pituitary-thyroid axis in rats.

Identification and characterization of novel stable deoxyguanosine and deoxyadenosine adducts of benzo[a]pyrene-7,8-quinone from reactions at physiological pH.

Benzo[a]pyrene (B[a]P) is an archetypal member of the family of polycyclic aromatic hydrocarbons (PAHs) and is a widely distributed environmental pollutant. B[a]P is known to induce cancer in animals, and B[a]P-containing complex mixtures are human carcinogens. B[a]P exerts its genotoxic and carcinogenic effects through metabolic activation forming reactive intermediates that damage DNA. DNA adduction by B[a]P is a complex phenomenon that involves the formation of both stable and unstable (depurinating) adducts. One pathway by which B[a]P can mediate genotoxicity is through the enzymatic formation of B[a]P-7,8-quinone (BPQ) from B[a]P-7,8-diol by members of the aldo-keto-reductase (AKR) family. Once formed, BPQ can act as a reactive Michael acceptor that can alkylate cellular nucleophiles including DNA and peptides. Earlier studies have reported on the formation of stable and depurinating adducts from the reaction of BPQ with DNA and nucleosides, respectively. However, the syntheses and characterization of the stable adducts from these interactions have not been addressed. In this study, the reactivity of BPQ toward 2'-deoxyguanosine (dG) and 2'-deoxyadenosine (dA) nucleosides under physiological pH conditions is examined. The identification and characterization of six novel BPQ-nucleoside adducts obtained from the reaction of BPQ and dG or dA in a mixture of phosphate buffer and dimethylformamide are reported. The structures of these adducts were determined by ultraviolet spectroscopy, electrospray mass spectrometry, and NMR experiments including (1)H, (13)C, two-dimensional COSY, one-dimensional NOE, ROESY, HMQC, HSQC, and HMBC. The reaction of BPQ with dG afforded four unique Michael addition products: two diastereomers of 8-N(1),9-N(2)-deoxyguanosyl-8,10-dihydroxy-9,10-dihydrobenzo[a]pyren-7(8H)-one (BPQ-dG(1,2)) and two diastereomers of 10-(N(2)-deoxyguanosyl)-9,10-dihydro-9-hydroxybenzo[a]pyrene-7,8-dione (BPQ-dG(3,4)). The BPQ-dG(1,2)( )()adducts suggest a 1,6-Michael addition reaction of dG, an oxidation of the hydroquinone to the quinone, a 1,4-Michael addition of water, and an internal cyclization. The BPQ-dG(3,4)( )()adducts suggest a 1,4-Michael addition reaction of dG, an oxidation of the hydroquinone to the quinone, and a 1,6-Michael addition of water. Under similar but extended reaction conditions, the reaction of BPQ with dA produced only one diastereomeric pair of adducts identified as 8-N(6),10-N(1)-deoxyadenosyl-8,9-dihydroxy-9,10-dihydrobenzo[a]pyren-7(8H)-one (BPQ-dA(1,2)). The BPQ-dA(1,2)( )()adducts suggest a 1,4-Michael addition reaction of dA, an oxidation of the hydroquinone to the quinone, a 1,6-Michael addition of water, and an internal cyclization. As considerable efforts have been placed in documenting the genotoxic effects of BPQ, this first report of the identification and characterization of these stable adducts of BPQ formed under physiological pH conditions is expected to contribute significantly to the area of BPQ-mediated genotoxicity and carcinogenesis.