Inferred inactivation of the Cftr gene in the duodena of mice exposed to hexavalent chromium (Cr(VI)) in drinking water supports its tumor-suppressor status and implies its potential role in Cr(VI)-induced carcinogenesis of the small intestines.

Carcinogenicity of hexavalent chromium [Cr (VI)] has been supported by a number of epidemiological and animal studies; however, its carcinogenic mode of action is still incompletely understood. To identify mechanisms involved in cancer development, we analyzed gene expression data from duodena of mice exposed to Cr(VI) in drinking water. This analysis included (i) identification of upstream regulatory molecules that are likely responsible for the observed gene expression changes, (ii) identification of annotated gene expression data from public repositories that correlate with gene expression changes in duodena of Cr(VI)-exposed mice, and (iii) identification of hallmark and oncogenic signature gene sets relevant to these data. We identified the inactivated CFTR gene among the top scoring upstream regulators, and found positive correlations between the expression data from duodena of Cr(VI)-exposed mice and other datasets in public repositories associated with the inactivation of the CFTR gene. In addition, we found enrichment of signatures for oncogenic signaling, sustained cell proliferation, impaired apoptosis and tissue remodeling. Results of our computational study support the tumor-suppressor role of the CFTR gene. Furthermore, our results support human relevance of the Cr(VI)-mediated carcinogenesis observed in the small intestines of exposed mice and suggest possible groups that may be more vulnerable to the adverse outcomes associated with the inactivation of CFTR by hexavalent chromium or other agents. Lastly, our findings predict, for the first time, the role of CFTR inactivation in chemical carcinogenesis and expand the range of plausible mechanisms that may be operative in Cr(VI)-mediated carcinogenesis of intestinal and possibly other tissues.

Assessment of the mutagenic potential of hexavalent chromium in the duodenum of big blue® rats.

A cancer bioassay on hexavalent chromium Cr(VI) in drinking water reported increased incidences of duodenal tumors in B6C3F1 mice at exposures of 30-180ppm, and oral cavity tumors in F344 rats at 180ppm. A subsequent transgenic rodent (TGR) in vivo mutation assay in Big Blue® TgF344 rats found that exposure to 180ppm Cr(VI) in drinking water for 28days did not increase cII transgene mutant frequency (MF) in the oral cavity (Thompson et al., 2015). Herein, we extend our analysis to the duodenum of these same TgF344 rats. At study termination, duodenum chromium levels were below either the limit of detection or quantification in control rats, but were 24.6±3.8µg/g in Cr(VI)-treated rats. The MF in control (23.2×10-6) and Cr(VI)-treated rats (22.7×10-6) were nearly identical. In contrast, the MF in the duodenum of rats exposed to 1-ethyl-1-nitrosourea for six days (study days 1, 2, 3, 12, 19, 26) increased 24-fold to 557×10-6. These findings indicate that mutagenicity is unlikely an early initiating event in Cr(VI)-induced intestinal carcinogenesis.

Reevaluation and Classification of Duodenal Lesions in B6C3F1 Mice and F344 Rats from 4 Studies of Hexavalent Chromium in Drinking Water.

Thirteen-week and 2-year drinking water studies conducted by the National Toxicology Program (NTP) reported that hexavalent chromium (Cr(VI)) induced diffuse epithelial hyperplasia in the duodenum of B6C3F1 mice but not F344 rats. In the 2-year study, Cr(VI) exposure was additionally associated with duodenal adenomas and carcinomas in mice only. Subsequent 13-week Cr(VI) studies conducted by another group demonstrated non-neoplastic duodenal lesions in B6C3F1 mice similar to those of the NTP study as well as mild duodenal hyperplasia in F344 rats. Because intestinal lesions in mice are the basis for proposed safety standards for Cr(VI), and the histopathology data are relevant to the mode of action, consistency (an important Hill criterion for causality) was assessed across the aforementioned studies. Two veterinary pathologists applied uniform diagnostic criteria to the duodenal lesions in rats and mice from the 4 repeated-dose studies. Comparable non-neoplastic intestinal lesions were evident in mice and rats from all 4 studies; however, the incidence and severity of intestinal lesions were greater in mice than rats. These findings demonstrate consistency across studies and species and highlight the importance of standardized nomenclature for intestinal pathology. The differences in the severity of non-neoplastic lesions also likely contribute to the differential tumor response.

Lack of efficacy of blueberry in nutritional prevention of azoxymethane-initiated cancers of rat small intestine and colon.

BACKGROUND: Blueberries may lower relative risk for cancers of the gastrointestinal tract. Previous work indicated an inhibitory effect of consumed blueberry (BB) on formation of aberrant crypt foci (ACF) in colons of male Fisher F344 rats (inbred strain). However, effects of BB on colon tumors and in both genders are unknown. METHODS: We examined efficacy of BB in inhibition of azoxymethane (AOM)-induced colon ACF and intestine tumors in male and female Sprague-Dawley rats (outbred strain). Pregnant rats were fed a diet with or without 10% BB powder; progeny were weaned to the same diet as their dam and received AOM as young adults. RESULTS: Male and female rats on control diet had similar numbers of ACF at 6 weeks after AOM administration. BB increased (P < 0.05) ACF numbers within the distal colon of female but not male rats. There was a significant (P < 0.05) diet by gender interaction with respect to total colon ACF number. Colon and duodenum tumor incidences were less in females than males at 17 weeks after AOM. BB tended (0.1 > P > 0.05) to reduce overall gastrointestinal tract tumor incidence in males, however, tumor incidence in females was unaffected (P > 0.1) by BB. There was a tendency (0.1 > P > 0.05) for fewer adenocarcinomas (relative to total of adenomatous polyps plus adenocarcinomas) in colons of female than male tumor-bearing rats; in small intestine, this gender difference was significant (P < 0.05). BB favored (P < 0.05) fewer adenocarcinomas and more adenomatous polyps (as a proportion of total tumor number) in female rat small intestine. CONCLUSION: Results did not indicate robust cancer-preventive effects of BB. Blueberry influenced ACF occurrence in distal colon and tumor progression in duodenum, in gender-specific fashion. Data indicate the potential for slowing tumor progression (adenomatous polyp to adenocarcinoma) by BB.

N-Methyl-N-Nitrosourea (MNU): A positive control chemical for p53+/- mouse carcinogenicity studies.

This study evaluated the effects of a single intraperitoneal injection of N-methyl-N-nitrosourea (MNU) in citrate buffer (pH 4.5) at a dose of 75 mg/kg in thirty male and thirty female p53+/- mice followed by a six-month observation period. Fifteen control mice per sex received a single intraperitoneal injection of citrate buffer. Fifty-six of sixty mice treated with MNU died or were sacrificed before the end of the observation period. Twenty-four males and twenty-seven females treated with MNU developed malignant lymphoma of the thymus; of these, twenty-three males and twenty-seven females had corresponding enlargement or masses in the thymus at necropsy. Lymphoblasts in thymic lymphomas stained positively for mouse CD3 antigen, indicating a T-cell lineage. One control female mouse had malignant lymphoma of the spleen that did not involve the thymus. Nine males and five females treated with MNU had adenomas or adenocarcinomas of the small intestine, whereas no intestinal neoplasms were observed in control mice. These findings support the use of a single dose of MNU as a positive control chemical in six-month p53+/- mouse carcinogenicity studies and suggest that examination of the thymus alone is sufficient to evaluate the validity of the model system.

Dietary whey protein lowers serum C-peptide concentration and duodenal SREBP-1c mRNA abundance, and reduces occurrence of duodenal tumors and colon aberrant crypt foci in azoxymethane-treated male rats.

We evaluated partially hydrolyzed whey protein (WPH) for inhibitory effects on the development of colon aberrant crypt foci (ACF) and intestinal tumors in azoxymethane (AOM)-treated rats. Pregnant Sprague-Dawley rats and their progeny were fed AIN-93G diets containing casein (CAS, control diet) or WPH as the sole protein source. Colons and small intestines from the male progeny were obtained at 6, 12, 20 and 23 weeks after AOM treatment. At 6 and 23 weeks, post-AOM, WPH-fed rats had fewer ACF than did CAS-fed rats. Intestinal tumors were most frequent at 23 weeks, post-AOM. At this time point, differences in colon tumor incidence with diet were not observed; however, WPH-fed rats had fewer tumors in the small intestine (7.6% vs. 26% incidence, P=.004). Partially hydrolized whey protein suppressed circulating C-peptide concentration (a stable indicator of steady-state insulin secretion) at all four time points relative to the corresponding CAS-fed animals. The relative mRNA abundance for the insulin-responsive, transcription factor gene, SREBP-1c, was reduced by WPH in the duodenum but not colon. Results indicate potential physiological linkages of dietary protein type with circulating C-peptide (and by inference insulin), local expression of SREBP-1c gene and propensity for small intestine tumorigenesis.

Induction of duodenal tumors in F344 rats by continuous oral administration of N-ethyl-N-nitrosourea.

Forty male and 40 female inbred F344 rats were given a solution of 400 mg N-ethyl-N-nitrosourea/liter in their drinking water. Digestive tract tumors were induced in 32 males (an incidence rate of 80%) and in 28 females (an incidence rate of 70%). Among these digestive tract neoplasms, duodenal tumors occurred most frequently. Most were of the epithelial type, such as adenoma or adenocarcinoma. Tumors in hematopoletic organs were also found in 15 males (38% incidence) and in 17 females (43% incidence).

Effect of dioctyl sodium sulfosuccinate feeding on rat colorectal 1,2-dimethylhydrazine carcinogenesis.

The 1,2-dimethylhydrazine (DMH) rodent model for colorectal carcinogenesis was used to explore the effect of dietary dioctyl sodium sulfosuccinate (DSS) on carcinogenesis. Inbred male F344 rats were divided into two groups of 84 each and fed the following diets: ground chow and 5% corn oil (control group) and ground chow, 5% corn oil, and 1% DSS (experimental group). All rats received high-dose DMH base, 20 mg/kg/week sc for 20 weeks. Twenty rats per group were killed after 3, 4, 5, and 6 months. Duodenum, small intestine, colon, and rectum were dissected out. Each tumor was measured for size and location and evaluated histologically. The percentage of rats bearing tumors in the control and experimental groups did not differ significantly. In each rat there were fewer gastrointestinal tumors in the DSS-fed group of all histologic types combined, at all organ sites, at 5 and 6 months. This difference between the control and DSS-fed rats reached the level of statistical significance for tumors of the duodenum, colon, and rectum and for total gastrointestinal tumors at the 5th month.

Induction of Leukemias and digestive tract tumors in Donryu rats by 1-propyl-1-nitrosourea.

Three groups of female Donryu rats were continuously given 600, 300, or 150 ppm solution of 1-propyl-1-nitrosourea in their drinking water. Leukemias developed in 62 of 109 (57%) rats surviving for more than 17 weeks and tumors developed in the digestive tracts of 31 (28%) animals. Of the leukemias, the differentiated myelocytic type was the most frequent, followed by myeloblastic leukemia and erythroleukemia. Tumors in the digestive tract, predominantly in the glandular stomach and duodenum, were both epithelial and nonepithelial. The other induced tumors were mainly in the mammary glands, ear ducts, and thymus, though the incidence was less than 15%.

Enhancing effect of vitamin E on murine intestinal tumorigenesis by 1,2-dimethylhydrazine dihydrochloride.

The effect of the antioxidant vitamin E on the tumor-inducing ability of 1,2-dimethylhydrazine dihydrochloride (1,2-DMH) was investigated in randomly bred Swiss mice. Three groups of mice that were 6 weeks of age at the beginning of the experiment received the following treatments: a) vitamin E acetate [DL-alpha-tocopheryl acetate (TA)] at a 4% dose level in a powdered diet for life; b) 1,2-DMH, 10 weekly sc injections at 20 micrograms/g body weight; c) combination of a and b treatments. The administration of TA enhanced the tumorigenicity of 1,2-DMH, as evidenced by statistically significant incidences of tumors in the duodenum, cecum, colon, rectum, and anus. The present finding apparently is in contrast with the reported inhibitory effect of TA on colon carcinogenesis by 1,2-DMH.

Production of adenocarcinomas of the small intestines of Wistar rats fed panfuran-S containing 3-di(hydroxymethyl)amino-6-(5-nitro-2-furylethenyl)-1,2,4-triazine.

Panfuran-S in the diet produced a high incidence of invasive and metastatic adenocarcinomas of the duodenum and the jejunum in outbred Wistar rats. Depressed growth and incidence of cancer production were dose-dependent. Adenocarcinomas of the duodenum were present in 2 of 18 (11%) rats that received 1,750 ppm 3-di(hydroxymethyl)amino-6-(5-nitro-2-furylethenyl)-1,2,4-triazine (DHNT) in the diet and in 11 of 19 (58%) rats that received 3,500 ppm DHNT. Adenocarcinomas of the jejunum were present in 10 of 18 (56%) and 16 of 19 (84%) rats that received 1,750 and 3,500 ppm DHNT, respectively, for 35-37 weeks. The only other tumors observed were squamous cell papillomas of the forestomach. Histologically, most of the adenocarcinomas appeared tubular or papillary and were similar to adenocarcinomas in the small intestine of man and also to human intestinal-type adenocarcinoma of the stomach. This system provided a useful experimental model for the study of the pathogenesis of carcinoma of the small intestine.

Ulcer formation and associated tumor production in multiple sites within the stomach and duodenum of rats treated with N-methyl-N'-nitro-N-nitrosoguanidine.

The effect of ulcers on the development of gastric tumors by N-methyl-N'-nitro-N-nitrosoguanidine in (MNNG) was studied in male Wistar rats. Ulcers were produced by the application of a steel rod, 5 mm in diameter and frozen at -78 degrees C, to the serosal surface of the forestomach, fundus, pylorus, or proximal duodenum. The existence of the ulcers at these areas was confirmed 1 week later in a preliminary experiment. Experimental groups were given MNNG in their drinking water at a concentration of 100 micrograms/ml for 16 weeks beginning 7 days after the ulcers developed. Administration of MNNG after ulceration resulted in a relative increase in the tumor incidences at each ulcer site, especially the proximal duodenum, which suggested that regenerating cells in the duodenum were the most susceptible cells among the cells of the four sites. The increase in tumor incidence following ulceration may be due to exposure of MNNG to a greater number of regenerating cells during the renewal process that seem to be more responsive to carcinogenic influences that normal cells.

Effect of dietary excess of inorganic selenium during initiation and postinitiation phases of colon carcinogenesis in F344 rats.

The effect of supplemental inorganic selenium given during the initiation or postinitiation phase of colon carcinogenesis induced by azoxymethane [(AOM)CAS:25843-45-2] was studied in male F344 rats. Weanling animals were raised on AIN-76A semipurified (control) diet. Starting at 4 wk of age, groups of animals intended for initiation study were fed the semipurified diets containing 0.5 and 2.5 ppm selenium in the form of sodium selenite, and those intended for postinitiation study were continued on the control diet. At 7 wk of age, all animals except the vehicle-treated controls were injected s.c. with AOM (15 mg/kg body weight, once weekly for 2 wk). One wk following AOM treatment, animals in the initiation study receiving the supplemental selenium were transferred to the control diet whereas those in the postinitiation study receiving the control diet were transferred to the diets containing 0.5 and 2.5 ppm selenium. These animals were continued on this regimen until the termination of the experiment at 34 wk post-AOM injection. Tissue and blood glutathione peroxidase activity was measured in vehicle-treated animals fed the control and selenium-supplemented diets. The results indicate that body weights were comparable among the various dietary groups. Feeding of diets containing 0.5 and 2.5 ppm selenium during the initiation phase had no effect on colon tumor incidence, but the multiplicity of adenomas was slightly inhibited in animals fed the 2.5 ppm selenium diet. The incidence and multiplicity of colon adenocarcinomas and the multiplicity of colon adenomas were inhibited in animals fed the 2.5-ppm selenium diet during the postinitiation phase of carcinogenesis. The incidence of small intestinal tumors was higher in animals fed the 2.5-ppm selenium diet during the initiation phase than in animals fed the control diet and 0.5-ppm selenium diet. Selenium-dependent glutathione peroxidase activity was increased in kidneys and small and large intestinal mucosae of animals fed the 2.5-ppm selenium diet compared to those fed the 0.5-ppm selenium and control diets.

Inhibitory effects of dietary curcumin on forestomach, duodenal, and colon carcinogenesis in mice.

Curcumin (diferuloylmethane), a yellow pigment that is obtained from the rhizomes of Curcuma longa Linn., is a major component of turmeric and is commonly used as a spice and food-coloring agent. The inhibitory effects of feeding commercial grade curcumin (77% curcumin, 17% demethoxycurcumin, and 3% bisdemethoxycurcumin) in AIN 76A diet on carcinogen-induced tumorigenesis in the forestomach, duodenum, and colon of mice were evaluated. Administration p.o. of commercial grade curcumin in the diet inhibited benzo(a)pyrene-induced forestomach tumorigenesis in A/J mice, N-ethyl-N'-nitro-N-nitrosoguanidine-induced duodenal tumorigenesis in C57BL/6 mice, and azoxymethane (AOM)-induced colon tumorigenesis in CF-1 mice. Dietary commercial grade curcumin was given to mice at: (a) 2 weeks before, during, and for 1 week after carcinogen administration (during the initiation period); (b) 1 week after carcinogen treatment until the end of the experiment (during the postinitiation period); or (c) during both the initiation and postinitiation periods. Feeding 0.5-2.0% commercial grade curcumin in the diet decreased the number of benzo(a)pyrene-induced forestomach tumors per mouse by 51-53% when administered during the initiation period and 47-67% when administered during the postinitiation period. Feeding 0.5-2.0% commercial grade curcumin in the diet decreased the number of N-ethyl-N'-nitro-N-nitrosoguanidine-induced duodenal tumors per mouse by 47-77% when administered during the postinitiation period. Administration of 0.5-4.0% commercial grade curcumin in the diet both during the initiation and postinitation periods decreased the number of AOM-induced colon tumors per mouse by 51-62%. Administration of 2% commercial grade curcumin in the diet inhibited the number of AOM-induced colon tumors per mouse by 66% when fed during the initiation period and 25% when fed during the postinitiation period. The ability of commercial grade curcumin to inhibit AOM-induced colon tumorigenesis is comparable to that of pure curcumin (purity greater than 98%). Administration of pure or commercial grade curcumin in the diet to AOM-treated mice resulted in development of colon tumors which were generally smaller in number and size as compared to the control group of AOM-treated mice. These results indicate that not only did curcumin inhibit the number of tumors per mouse and the percentage of mice with tumors but it also reduced tumor size. Histopathological examination of the tumors showed that dietary curcumin inhibited the number of papillomas and squamous cell carcinomas of the forestomach as well as the number of adenomas and adenocarcinomas of the duodenum and colon.

Effect of flurbiprofen and 16,16-dimethyl prostaglandin E2 on gastrointestinal tumorigenesis induced by N-methyl-N'-nitro-N-nitrosoguanidine in rats: glandular epithelium of stomach and duodenum.

The effect of an exogenous synthetic prostaglandin analogue, 16,16-dimethyl prostaglandin E2 (16,16-dm-PGE2), as well as the effect of endogenous prostaglandin synthesis inhibition by a cyclooxygenase inhibitor, flurbiprofen, on chemically induced gastric carcinogenesis has been investigated in rats. Carcinogenesis was induced by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG; CAS:70-25-7). Animals were divided into six groups: Group I, treatment with MNNG alone; Group II, treatment with 16,16-dm-PGE2 plus MNNG; Group III, treatment with flurbiprofen plus MNNG; Group IV, treatment with 16,16-dm-PGE2 alone; Group V, treatment with flurbiprofen alone; and Group VI, controls. Treatment with high doses of MNNG resulted in rapid development of malignant tumors originating from the glandular epithelium of the stomach and duodenum in animals of all groups receiving the carcinogen. The first gastric adenocarcinoma infiltrating the muscularis proper was detected after 139 days in an animal treated with a combination of MNNG and flurbiprofen. The incidence of infiltrating adenocarcinoma and the incidence of all neoplastic lesions of the glandular stomach were both significantly higher in animals treated with a combination of MNNG and flurbiprofen compared with treatment by MNNG alone or in combination with 16,16-dm-PGE2 (P less than 0.05 and P less than 0.001). The difference in tumor incidence between the last two groups was not significant. The first duodenal adenocarcinoma was detected on Day 114 in another animal of the group treated with MNNG plus flurbiprofen. When compared with the group treated with MNNG plus 16,16-dm-PGE2, significantly more animals developed duodenal adenocarcinoma when treated with MNNG plus flurbiprofen (P less than 0.005) or with MNNG alone (P less than 0.05). Results of this study indicate that inhibition of endogenous prostaglandin synthesis favors development of adenocarcinoma in the glandular stomach of rats. Vice versa, the addition of an exogenous prostaglandin analogue inhibits the development of duodenal adenocarcinoma. This protective effect of prostaglandins may be due to an increase of the thickness of the mucus gel covering the glandular epithelium, thereby preventing access of carcinogen to the mucosa.

Comparison of in vivo genotoxic and carcinogenic potency to augment mode of action analysis: Case study with hexavalent chromium.

Recent analyses-highlighted by the International Workshops on Genotoxicity Testing Working Group on Quantitative Approaches to Genetic Toxicology Risk Assessment-have identified a correlation between (log) estimates of a carcinogen's in vivo genotoxic potency and in vivo carcinogenic potency in typical laboratory animal models, even when the underlying data have not been matched for tissue, species, or strain. Such a correlation could have important implications for risk assessment, including informing the mode of action (MOA) of specific carcinogens. When in vivo genotoxic potency is weak relative to carcinogenic potency, MOAs other than genotoxicity (e.g., endocrine disruption or regenerative hyperplasia) may be operational. Herein, we review recent in vivo genotoxicity and carcinogenicity data for hexavalent chromium (Cr(VI)), following oral ingestion, in relevant tissues and species in the context of the aforementioned correlation. Potency estimates were generated using benchmark doses, or no-observable-adverse-effect-levels when data were not amenable to dose-response modeling. While the ratio between log values for carcinogenic and genotoxic potency was ≥1 for many compounds, the ratios for several Cr(VI) datasets (including in target tissue) were less than unity. In fact, the ratios for Cr(VI) clustered closely with ratios for chloroform and diethanolamine, two chemicals posited to have non-genotoxic MOAs. These findings suggest that genotoxicity may not play a major role in the cancers observed in rodents following exposure to high concentrations of Cr(VI) in drinking water-a finding consistent with recent MOA and adverse outcome pathway (AOP) analyses concerning Cr(VI). This semi-quantitative analysis, therefore, may be useful to augment traditional MOA and AOP analyses. More case examples will be needed to further explore the general applicability and validity of this approach for human health risk assessment.

[Intramural small-bowel hematoma due to anticoagulant therapy: a radiologic study; with regard to two cases]. / Hématome intramural du grêle dû aux anticoagulants: une etude radiologique a propos de deux observations.

We report two cases of spontaneous small bowel hematoma in two patients receiving long-term anticoagulant therapy. Plain abdominal film, ultrasound, CT-scan and oral barium examination were performed. Abdominal ultrasonography and CT-scan are in most cases relevant for the correct diagnosis of intra-mural small bowel hematoma. The diagnosis was based on the acknowledgment by the patient of anticoagulant drug consumption. Early diagnosis is crucial because most patients are treated nonoperatively with good outcome.

Induction of duodenal mucosal tumors of intestinal epithelial cell origin showing frequent nuclear ß-catenin accumulation similar to the concurrently induced colorectal tumors in rats after treatment with azoxymethane.

Azoxymethane (AOM) is a potent carcinogen used for induction of colon tumors in rats and mice. It is also known that AOM treatment induces small bowel tumors in addition to colorectal tumors in rats. The present study examined the histogenesis of AOM-induced rat duodenal tumors in comparison with concurrently induced colorectal tumors by histochemical and immunohistochemical approaches. Duodenal and colorectal tumors were positive for both periodic acid-Schiff reaction and Alcian blue staining. Immunohistochemically, duodenal tumors were positive for intestinal epithelial markers such as cytokeratin (CK) 20 (100%) and mucin (MUC) 2 (91.7%) but negative for pancreaticobiliary markers such as CK7 (100%) and MUC1 (100%). All colorectal tumors were also negative for CK7 and MUC1 but positive for CK20. Eighty percent of colorectal tumors were positive for MUC2. In addition, nuclear accumulation of ß-catenin was found in duodenal tumors (70.8%), which was similar to colorectal tumors (90.0%). These results indicate that duodenal tumors induced by AOM treatment of rats were derived from intestinal epithelium. Similar to colorectal tumors, nuclear accumulation of ß-catenin indicates activation of Wnt signaling as a driving force for tumor progression in AOM-induced duodenal tumors.

Synchrotron-based imaging of chromium and γ-H2AX immunostaining in the duodenum following repeated exposure to Cr(VI) in drinking water.

Current drinking water standards for chromium are for the combined total of both hexavalent and trivalent chromium (Cr(VI) and Cr(III)). However, recent studies have shown that Cr(III) is not carcinogenic to rodents, whereas mice chronically exposed to high levels of Cr(VI) developed duodenal tumors. These findings may suggest the need for environmental standards specific for Cr(VI). Whether the intestinal tumors arose through a mutagenic or non-mutagenic mode of action (MOA) greatly impacts how drinking water standards for Cr(VI) are derived. Herein, X-ray fluorescence (spectro)microscopy (µ-XRF) was used to image the Cr content in the villus and crypt regions of duodena from B6C3F1 mice exposed to 180 mg/l Cr(VI) in drinking water for 13 weeks. DNA damage was also assessed by γ-H2AX immunostaining. Exposure to Cr(VI) induced villus blunting and crypt hyperplasia in the duodenum--the latter evidenced by lengthening of the crypt compartment by ∼2-fold with a concomitant 1.5-fold increase in the number of crypt enterocytes. γ-H2AX immunostaining was elevated in villi, but not in the crypt compartment. µ-XRF maps revealed mean Cr levels >30 times higher in duodenal villi than crypt regions; mean Cr levels in crypt regions were only slightly above background signal. Despite the presence of Cr and elevated γ-H2AX immunoreactivity in villi, no aberrant foci indicative of transformation were evident. These findings do not support a MOA for intestinal carcinogenesis involving direct Cr-DNA interaction in intestinal stem cells, but rather support a non-mutagenic MOA involving chronic wounding of intestinal villi and crypt cell hyperplasia.

17Beta-estradiol modulates gastroduodenal preneoplastic alterations in rats exposed to the carcinogen N-methyl-N'-nitro-nitrosoguanidine.

Gastric cancers are a significant cause of morbidity worldwide. Epidemiological studies and animal models show that males have higher incidences of gastric cancers compared with females, suggesting that sex hormones may modulate gastric cancer risk. An animal model of the initiation phase of gastric cancer was used to determine the effects of systemic estrogen administration on morphological progression of preneoplastic lesions and to define cell populations at which estrogens may act. Preneoplastic progression in antral and duodenal mucosa was examined in male rats that received the chemical carcinogen, N-methyl-N'-nitro-nitrosoguanidine (MNNG), during treatment with implants containing 17beta-estradiol or oil vehicle. Histopathological changes in antral and duodenal gland morphology, numbers of proliferating cells and apoptotic bodies, and antral gastrin cell numbers and protein storage levels were determined 4 weeks later. With MNNG treatment, duodenal villous heights were significantly decreased, and epithelial cells displayed histological features of hyperplasia and dysplasia. Antral glands showed epithelial hyperplasia and dysplasia, increased mucosal height, and decreased mucin levels. Antral gastrin storage protein levels were decreased by MNNG. Systemic treatment with 17beta-estradiol significantly reversed MNNG-induced alterations in duodenal gland heights while increasing mucin and gastrin levels in antral glands. Cell proliferation and apoptosis rates were not significantly different between groups. The present results indicate that systemic 17beta-estradiol treatment influences antral and duodenal gland differentiation during the initiation phase of chemical gastroduodenal carcinogenesis in male rats. These results explain, in part, a potential pathway through which protective effects of estrogens on chemical carcinogenesis are mediated in the upper gastrointestinal tract.