Promotion of liver and kidney carcinogenesis by ethyl tertiary-butyl ether (ETBE) in male Wistar rats.

Tumor-promoting effects of ethyl tertiary-butyl ether (ETBE) were investigated in a 2-stage carcinogenesis bioassay with regard to hepatic and renal carcinogenesis in rats. Male 6-week-old Wistar rats were given drinking water containing N-ethyl-N-(2-hydroxyethyl)nitrosamine (EHEN), as an initiator, at a dose of 500 ppm for 2 weeks. Starting one week thereafter, the animals were administered ETBE at dose levels of 0 (control), 100, 300, 500 or 1,000 mg/kg/day by gavage for 19 weeks from week 4 to 22. Necropsy of all rats was performed at week 23, and livers and kidneys were examined histopathologically. Incidences of hepatocellular adenomas, and those of combined hepatocellular adenomas and carcinomas were significantly elevated in rats given 1,000 mg/kg/day ETBE, but not 100‒500 mg/kg/day ETBE, and there was a significant increase in the average numbers of lesions. No significant differences in incidences and average numbers of renal tubule neoplasms were found in rats administered 100‒1,000 mg/kg/day ETBE. However, the average numbers of atypical tubule hyperplasias, considered to be preneoplastic lesions, were significantly increased in rats given ETBE at 1,000 mg/kg/day, but not in rats given 500 mg/kg/day or lower doses. Thus, these results imply that ETBE has hepatic and renal tumor-promoting activities that affect EHEN-induced carcinogenesis in male rats, and the no-observed-effect level is 500 mg/kg/day under the present experimental conditions.

Multigenerational effects of whole body exposure to 2.14 GHz W-CDMA cellular phone signals on brain function in rats.

The present experimental study was carried out with rats to evaluate the effects of whole body exposure to 2.14 GHz band code division multiple access (W-CDMA) signals for 20 h a day, over three generations. The average specific absorption rate (SAR, in unit of W/kg) for dams was designed at three levels: high (<0.24 W/kg), low (<0.08 W/kg), and 0 (sham exposure). Pregnant mothers (4 rats/group) were exposed from gestational day (GD) 7 to weaning and then their offspring (F1 generation, 4 males and 4 females/dam, respectively) were continuously exposed until 6 weeks of age. The F1 females were mated with F1 males at 11 weeks old, and then starting from GD 7, they were exposed continuously to the electromagnetic field (EMF; one half of the F1 offspring was used for mating, that is, two of each sex per dam and 8 males and 8 females/group, except for all offspring for the functional development tests). This protocol was repeated in the same manner on pregnant F2 females and F3 pups; the latter were killed at 10 weeks of age. No abnormalities were observed in the mother rats (F0 , F1 , and F2 ) and in the offspring (F1 , F2 , and F3 ) in any biological parameters, including neurobehavioral function. Thus, it was concluded that under the experimental conditions applied, multigenerational whole body exposure to 2.14 GHz W-CDMA signals for 20 h/day did not cause any adverse effects on the F1 , F2 , and F3 offspring.

Immunohistochemical characteristics of surfactant proteins a, B, C and d in inflammatory and tumorigenic lung lesions of f344 rats.

Surfactant proteins (SPs), originally known as human lung surfactants, are essential to respiratory structure and function. There are 4 subtypes, SP-A, SP-B, SP-C and SP-D, with SP-A and SP-D having immunological functions, and SP-B and SP-C having physicochemical properties that reduce the surface tension at biological interfaces. In this experiment, the expressions of SP-A, SP-B, SP-C and SP-D in lung neoplastic lesions induced by N-bis (2-hydroxypropyl) nitrosamine (DHPN) and inflammatory lesions due to quartz instillation were examined and compared immunohistochemically. Formalin fixed paraffin embedded (FFPE) lung samples featuring inflammation were obtained with a rat quartz instillation model, and neoplastic lesions, hyperplasias and adenomas, were obtained with the rat DHPN-induced lung carcinogenesis model. In the rat quartz instillation model, male 10-week old F344 rats were exposed by intratracheal instillation (IT) to quartz at a dose of 2 mg/rat suspended in saline (0.2 ml) on day 0, and sacrificed on day 28. Lung tumorigenesis in F344 male rats was initiated by DHPN in drinking water for 2 weeks, and the animals were then sacrificed in week 30. Lung proliferative lesions, hyperplasias and adenomas, were observed with DHPN, and inflammation was observed with quartz. The expressions of SP-A, SP-B, SP-C and SP-D were examined immunohistochemically. SP-B and SP-C showed strong expression in lung hyperplasias and adenomas, while SP-A and SP-D were observed in mucus or exudates in inflammatory alveoli. These results suggest the possibility that SP-B and SP-C are related to lung tumorigenesis.

No Promoting Effect of Ethyl Tertiary-butyl Ether (ETBE) on Rat Urinary Bladder Carcinogenesis Initiated with N-Butyl-N-(4-hydroxybutyl)nitrosamine.

The effects of ethyl tertiary-butyl ether (ETBE) on two-stage urinary bladder carcinogenesis in male F344 rats initiated with N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) were investigated at various dose levels with regard to possible promoting activity. Groups of 30 rats were given drinking water containing 500 ppm BBN, as an initiator, for 4 weeks and starting one week thereafter received ETBE by gavage (daily, 7 days/week) at dose levels of 0 (control), 100, 300, 500 or 1000 mg/kg/day until experimental week 36. No statistically significant differences in incidences of preneoplastic lesions, papillomas, and carcinomas of the urinary bladder were evident in rats treated with 100-1000 mg/kg/day ETBE as compared with control values. Furthermore, the average numbers of preneoplastic or neoplastic lesions per unit length of basement membrane in rats given 100-1000 mg/kg/day ETBE were also comparable to control values. However, papillomatosis of the urinary bladder was found in 4 out of 30 rats (13%) in the group given 1000 mg/kg/day ETBE, and soft stones in the urinary bladder were found in 3 out of these 4 rats. The results thus demonstrated that ETBE did not exert promotional activity on urinary bladder carcinogenesis. However, papillomatosis of the urinary bladder developed in small numbers of the rats given ETBE at 1000 mg/kg/day but not in rats given 500 mg/kg/day or lower doses.

Involvement of macrophage inflammatory protein 1alpha (MIP1alpha) in promotion of rat lung and mammary carcinogenic activity of nanoscale titanium dioxide particles administered by intra-pulmonary spraying.

Titanium dioxide (TiO(2)) is evaluated by World Health Organization/International Agency for Research on Cancer as a Group 2B carcinogen. The present study was conducted to detect carcinogenic activity of nanoscale TiO(2) administered by a novel intrapulmonary spraying (IPS)-initiation-promotion protocol in the rat lung. Female human c-Ha-ras proto-oncogene transgenic rat (Hras128) transgenic rats were treated first with N-nitrosobis(2-hydroxypropyl)amine (DHPN) in the drinking water and then with TiO(2) (rutile type, mean diameter 20 nm, without coating) by IPS. TiO(2) treatment significantly increased the multiplicity of DHPN-induced alveolar cell hyperplasias and adenomas in the lung, and the multiplicity of mammary adenocarcinomas, confirming the effectiveness of the IPS-initiation-promotion protocol. TiO(2) aggregates were localized exclusively in alveolar macrophages and had a mean diameter of 107.4 nm. To investigate the underlying mechanism of its carcinogenic effects, TiO(2) was administered to wild-type rats by IPS five times over 9 days. TiO(2) treatment significantly increased 8-hydroxydeoxy guanosine level, superoxide dismutase activity and macrophage inflammatory protein 1alpha (MIP1alpha) expression in the lung. MIP1alpha, detected in the cytoplasm of TiO(2)-laden alveolar macrophages in vivo and in the media of rat primary alveolar macrophages treated with TiO(2) in vitro, enhanced proliferation of human lung cancer cells. Furthermore, MIP1alpha, also detected in the sera and mammary adenocarcinomas of TiO(2)-treated Hras128 rats, enhanced proliferation of rat mammary carcinoma cells. These data indicate that secreted MIP1alpha from TiO(2)-laden alveolar macrophages can cause cell proliferation in the alveoli and mammary gland and suggest that TiO(2) tumor promotion is mediated by MIP1alpha acting locally in the alveoli and distantly in the mammary gland after transport via the circulation.

A medium-term, rapid rat bioassay model for the detection of carcinogenic potential of chemicals.

The Ito Liver Model and the Ito Multi-organ Model are used in conjunction and constitute an efficient and rapid bioassay for the identification of both genotoxic and nongenotoxic carcinogenic chemicals. The Ito Liver Model is an 8-week bioassay system that uses the number and size of foci of hepatocytes positive for glutathione S-transferase placental form (GST-P) as the end-point marker. One hundred fifty-nine compounds were tested using the Ito Liver Model: 61 of 66 hepatocarcinogens tested positive, and 10 of 43 nonliver carcinogens were also positive. The false-positive detection of noncarcinogens was low; a single false-positive result was obtained from the 50 noncarcinogens tested. Since more than half of all known carcinogens are hepatocarcinogens in rodents, the initial 8-week bioassay is able to detect most carcinogens. The Ito Multi-organ Model is a 28-week bioassay system for the detection of carcinogens that were not identified by the Ito Liver Model. Results are evaluated by preneoplastic and neoplastic lesions in major organs. Forty-four compounds were tested using the Ito Multi-organ Model: 17 out of 17 liver carcinogens were positive, and 19 out of 22 (86%) nonliver carcinogens were positive. None of the five noncarcinogens tested positive.

Lack of enhancing effects of sodium nitrite on 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)-induced mammary carcinogenesis in female Sprague-Dawley rats.

A number of heterocyclic amines (HCAs) have been shown to exert enhanced carcinogenic and mutagenic potential when given simultaneously with sodium nitrite (NaNO(2)). In the present experiment, effects of combined treatment with NaNO(2) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), one of the most prevalent carcinogenic HCAs in the human environment, were assessed with regard to mammary tumor induction in female Sprague-Dawley (SD) rats. Animals at 6 weeks of age were given intragastric doses of 100mg/kg body weight of PhIP twice a week for 4 weeks, during which period 0 or 0.2% NaNO(2) was administered in the drinking water. Control rats received 0.2% NaNO(2) alone for the 4 weeks or non-supplemented water during the entire 48 week experimental period, without carcinogen treatment. The first tumor in the PhIP+NaNO(2) group appeared significantly later than with PhIP alone, and during the experimental period, the incidence, multiplicity and volume of mammary tumors in this group tended towards decreased, although values did not significantly differ at the terminal sacrifice. These results indicate that NaNO(2) does not enhance PhIP-induced rat mammary carcinogenesis, rather possessing some potential for inhibition.

Lack of potential of low dose N-nitrosodimethylamine to induce preneoplastic lesions, glutathione S-transferase placental form-positive foci, in rat liver.

Induction of liver lesions in male F344 rats by the genotoxic and carcinogenic N-nitrosodimethylamine (NDMA) was studied at a wide range of dose levels, i.e. from 0.001 to 10 ppm, in drinking water for 16 weeks. Dose related and statistically significant increase of glutathione S-transferase placental form-positive foci, endpoint markers for hepatocarcinogenesis in rats, at 1 and 10 ppm dose groups was obtained, but no increment in foci could be detected with the lower doses (0.001, 0.01, and 0.1 ppm). This finding of a no-observed effect level supports our hypothesis that a threshold, at least in practical terms, exists in carcinogenesis proposed on the basis of extensive wide range dose-dependence studies of other genotoxic carcinogens.

Potent chemopreventive agents against pancreatic cancer.

Development of pancreatic cancers is clinically so silent in general that at the time of diagnosis, the vast majority of cases are incurable with a very poor prognosis. Therefore, effective preventive approaches against this aggressive disease are urgently required. Experimentally, carcinogenesis process is assumed to consist of at least two stages named initiation and promotion. Using a two-stage model of hamster pancreatic carcinogenesis, we have reported stage-specific inhibitory effects by a number of potent cancer chemopreventive agents. Among them, phenethyl isothiocyanate (PEITC), a constituent of cruciferous vegetables, remarkably blocked the initiation phase of pancreatic as well as lung carcinogenesis in hamsters initiated with N-nitrosobis(2-oxopropyl)amine (BOP). However, PEITC failed to affect both pancreatic and lung carcinogenesis when given during the post-initiation (promotion) phase of carcinogenesis. In contrast, our recent study clearly demonstrated that a cyclooxygenase (COX)-2 inhibitor substantially protects against BOP-induced pancreatic tumors in hamsters in line with decrease in cell proliferative activity of pancreatic ducts when given in the post-initiation phase. Interestingly, trypsin inhibitors inhibited both initiation and post-initiation phases of BOP-induced pancreatic carcinogenesis although they are known to induce hyperplastic acinar lesions in the rat pancreas. Taken together with these data, our review is aimed at looking over mechanistic insights into potent chemopreventive agents against pancreatic cancer.

Chemoprevention of lung carcinogenesis by cacao liquor proanthocyanidins in a male rat multi-organ carcinogenesis model.

The effects of cacao liquor proanthocyanidins (CLPr) on tumorigenesis were investigated using a multi-organ carcinogenesis model in male F344 rats receiving combined treatment with a single i.p. injection of diethylnitrosamine (100 mg/kg body wt), four i.p. injections of N-methylnitrosourea (20 mg/kg body wt), four s.c. injections of dimethylhydrazine (40 mg/kg body wt), along with 0.05% N-butyl-N-(4-hydroxybutyl)nitrosamine and then 0.1% 2,2'-dihydroxy-di-n-propylnitrosamine, both in the drinking water, for 2 weeks each, during the initial 4-week period (DMBDD treatment). Starting 1 week thereafter, rats were administered CLPr at a dose of 0.025% or 0.25% and the experiment was terminated at week 36. The final survival rate for the DMBDD+0.25% CLPr group was significantly greater than for the DMBDD alone group. In the lung, significant reduction in the incidence and multiplicity of carcinomas was also observed, and in the thyroid, quantitative values for adenomas also tended to decrease in a CLPr dose-dependent manner. No significant modification in the small intestine, colon or kidney was evident. These results indicate that CLPr exerts chemopreventive effects in the lung without any promoting influence in other major organs.

Promoting effects of combined antioxidant and sodium nitrite treatment on forestomach carcinogenesis in rats after initiation with N-methyl-N'-nitro-N-nitrosoguanidine.

The effects of sodium nitrite (NaNO2), in combination with one of three antioxidants, tert-butylhydroquinone (TBHQ), alpha-tocopherol (alpha-Toc) and propyl gallate (PG), on N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) stomach carcinogenesis were investigated in F344 rats. Groups of 15 male rats were treated with an intragastric dose of 150 mg/kg body weight of MNNG, and starting 1 week later, were treated with 0.5% TBHQ, 1% alpha-Toc, 1% PG or basal diet with or without 0.2% NaNO2 in their drinking water until they were killed at the end of week 36. Macroscopically, in MNNG-treated animals, combined administration of alpha-Toc or PG with NaNO2 significantly increased the areas and numbers of forestomach nodules as compared with the respective antioxidant alone values. Microscopically, in MNNG-treated animals, treatment with TBHQ significantly increased the incidence and multiplicity of forestomach papillomas as compared with basal diet alone value. Combined administration of alpha-Toc with NaNO2 significantly raised the multiplicity of forestomach papillomas, with a tendency to elevation in the incidence as compared with the group given alpha-Toc alone. Incidences of forestomach moderate and/or severe hyperplasias were significantly higher in the TBHQ or PG plus NaNO2 groups than in the single compound groups. In rats without MNNG treatment, combined treatment of antioxidants with NaNO2 significantly increased the incidences of mild or moderate hyperplasia. In the glandular stomach, although the incidence of atypical hyperplasia showed a non-significant tendency for decrease with TBHQ treatment, additional administration of NaNO2 caused significant increase. These results indicate that co-administration of NaNO2 with alpha-Toc, TBHQ or PG and particularly the first, promotes forestomach carcinogenesis. Concurrent alpha-Toc, TBHQ or PG treatment with NaNO2 is likely to induce forestomach tumors in the long term.

Effects of cacao liquor proanthocyanidins on PhIP-induced mutagenesis in vitro, and in vivo mammary and pancreatic tumorigenesis in female Sprague-Dawley rats.

The effects of cacao liquor proanthocyanidins (CLPr) on 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)-induced mutagenesis in vitro and on in vivo carcinogenesis in female Sprague-Dawley (SD) rats were investigated. In the Ames assay using Salmonella typhimurium TA98, CLPr showed strong antimutagenic effects against PhIP when assayed in the presence of S-9 mixture. For determination of the influence on initiation and subsequent development of lesions, CLPr (0.025% or 0.25%) were fed during the period of PhIP application (100 mg/kg given to rats via gastric tubes eight times over 4 weeks), or thereafter until the termination at 48 weeks. CLPr treatments did not affect body or organ weights. The incidences, multiplicities and volumes of mammary tumors in the 0.25% CLPr (post-initiation) group showed a tendency to decrease as compared to PhIP alone group values, although without statistical significance. The incidences of preneoplastic eosinophilic foci in the exocrine pancreas were significantly (P<0.05) decreased in a dose-dependent manner when CLPr were given during the initiation period. These results indicate that CLPr inhibit in vitro mutagenicity of PhIP, as well as rat pancreatic carcinogenesis in the initiation stage, but not mammary carcinogenesis induced by PhIP.

Chemopreventive effects of Aloe arborescens on N-nitrosobis(2-oxopropyl)amine-induced pancreatic carcinogenesis in hamsters.

The modification effects of freeze-dried aloe (Aloe arborescens) whole leaf powder during the initiation phase of carcinogenesis were investigated in hamsters treated with N-nitrosobis(2-oxopropyl)amine (BOP). Female Syrian hamsters were given four weekly subcutaneous injections of BOP at a dose of 10mg/kg and then given 0, 1 or 5% aloe in their diet for 5 weeks. At week 54 of the experiment, all surviving animals were sacrificed and development of neoplastic and preneoplastic lesions was assessed histopathologically. The incidences of pancreatic adenocarcinomas, atypical hyperplasias or total atypical hyperplasias plus adenocarcinomas were significantly (P<0.05) decreased with BOP+5% aloe, and that of adenocarcinomas were also significantly (P<0.05) reduced in the BOP+1% aloe as compared to the BOP alone group. Multiplicities of pancreatic adenocarcinomas, atypical hyperplasias or total lesions were also significantly (P<0.01 or P<0.05) lower in the BOP+5% aloe group than with the BOP alone. Quantitative data for neoplastic lesions in the lung, liver, gall bladder, kidney and urinary bladder of hamsters were not significantly different among the three groups. In a satellite experiment, pretreatment with aloe significantly (P<0.01) reduced the formation of O6-methyldeoxyguanosine in epithelial cells of pancreatic ducts as compared to the BOP alone value. Our results thus indicate that aloe prevents BOP-induced pancreatic neoplasia in hamsters in relation to decreased DNA adduct formation in the target tissue.

Lack of initiation activity in rat liver of low doses of 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline.

It has been generally accepted that genotoxic carcinogens have no threshold in exerting their potential for cancer induction. However, the non-threshold theory can be challenged for cancer risk assessment in humans. Here we examined low dose carcinogenicity of a food-derived, genotoxic hepatocarcinogen, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), using an in vivo medium-term bioassay to detect initiating activity for rat hepatocarcinogenesis. With MeIQx initiation at various doses followed by administration of phenobarbital, a well known hepatopromoter, no induction of glutathione S-transferase placental form-positive foci, assessed as preneoplastic lesions, was noted at doses of 0.001-1 ppm. The results imply a no-observed effect level for hepatocarcinogenicity with this genotoxic agent.

A 13-week subchronic toxicity study of paprika color in F344 rats.

The fruit of the paprika (Capsicum annuum) has been widely used in various countries as a spice and food-coloring additive. As a part of the safety assessment of paprika color (Paprika oleoresin), a 13-week subchronic toxicity study was performed in F344 rats. To establish a no-observed-adverse-effect level (NOAEL) for application in subsequent long-term studies, rats were fed powder diet containing paprika color at dose levels of 0 (basal diet), 0.62, 1.25, 2.5 and 5% (maximum) for 13 weeks. During the experiment, there were no remarkable changes in general appearance and no deaths occurred in any experimental group. Although serum total cholesterol was dose-dependently increased in both sexes, no related histopathological changes were observed in the liver. Slight inflammatory cell infiltration in the myocardium and vacuolation of hepatocytes were noted in both control and paprika color-treated animals, but there were no clear differences between groups. In conclusion, paprika color even at 5% in the diet (0.67 g/rat/day or 2948.4 mg/kg bw/day for male rats and 0.43 g/rat/day or 3197.4 mg/kg bw/day for female rats) did not cause remarkable adverse effects in F344 rats. Thus, the NOAEL and the maximum dose level for carcinogenicity testing of paprika color were concluded to be 5% in the diet.

Comparative study of toxic effects of anatase and rutile type nanosized titanium dioxide particles in vivo and in vitro.

Two types of nanosized titanium dioxide, anatase (anTiO2) and rutile (rnTiO2), are widely used in industry, commercial products and biosystems. TiO2 has been evaluated as a Group 2B carcinogen. Previous reports indicated that anTiO2 is less toxic than rnTiO2, however, under ultraviolet irradiation anTiO2 is more toxic than rnTiO2 in vitro because of differences in their crystal structures. In the present study, we compared the in vivo and in vitro toxic effects induced by anTiO2 and rnTiO2. Female SD rats were treated with 500 ?g/ml of anTiO2 or rnTiO2 suspensions by intra-pulmonary spraying 8 times over a two week period. In the lung, treatment with anTiO2 or rnTiO2 increased alveolar macrophage numbers and levels of 8-hydroxydeoxyguanosine (8-OHdG); these increases tended to be lower in the anTiO2 treated group compared to the rnTiO2 treated group. Expression of MIP1??mRNA and protein in lung tissues treated with anTiO2 and rnTiO2 was also significantly up-regulated, with MIP1??mRNA and protein expression significantly lower in the anTiO2 group than in the rnTiO2 group. In cell culture of primary alveolar macrophages (PAM) treated with anTiO2 and rnTiO2, expression of MIP1??mRNA in the PAM and protein in the culture media was significantly higher than in control cultures. Similarly to the in vivo results, MIP1??mRNA and protein expression was significantly lower in the anTiO2 treated cultures compared to the rnTiO2 treated cultures. Furthermore, conditioned cell culture media from PAM cultures treated with anTiO2 had less effect on A549 cell proliferation compared to conditioned media from cultures treated with rnTiO2. However, no significant difference was found in the toxicological effects on cell viability of ultra violet irradiated anTiO2 and rnTiO2. In conclusion, our results indicate that anTiO2 is less potent in induction of alveolar macrophage infiltration, 8-OHdG and MIP1??expression in the lung, and growth stimulation of A549 cells in vitro than rnTiO2.

Arachidonate-enriched triglyceride oil does not promote tumor development in a rat medium-term multi-organ carcinogenesis model.

The modifying potential on tumor development of arachidonate-enriched triglyceride oil (ARA-oil) containing approximately 40% arachidonic acid was investigated in a medium-term multi-organ carcinogenesis bioassay using male and female F344 rats. The animals were sequentially given five carcinogens with different target sites in the first 4 weeks, and then administered ARA-oil for 24 weeks at dietary levels of 0% (control), 1.25%, 2.5% or 5.0%. No statistically significant differences in incidences and multiplicities of hyperplastic and neoplastic lesions were showed in the large intestine in either sex. In the liver, kidney, and lung in both sexes, and the mammary gland and uterus in females, tumor promoting potential was not evident with ARA-oil treatment. ARA-oil did not affect the quantitative data for glutathione S-transferase placental form positive foci of the liver. Increased induction of hyperplastic or neoplastic lesions in the urinary bladder and thyroid in ARA-oil-treated groups was without dose dependence. In addition, a second experiment with ARA-oil only administration for 8-week revealed no effects on cellular proliferation in the urinary bladder or thyroid in either sex. These results indicate that ARA-oil has no tumor promoting potential in any organs or tissues initiated with the five carcinogens applied in the present study.

Medium-term multi-organ carcinogenesis bioassay of ethyl tertiary-butyl ether in rats.

The modifying potential of ethyl tertiary-butyl ether (ETBE) on tumor development was investigated in a medium-term multi-organ carcinogenesis bioassay using male F344 rats. Animals were sequentially given 5 carcinogens with different target sites in the first 4 weeks for multi-organ initiation. After one week they received ETBE by gavage at dose levels of 0 (control), 300 or 1000mg/kg/day until experimental week 28. Further groups were also given ETBE at doses of 0 or 1000mg/kg/day without prior carcinogen application. Incidences and multiplicities of follicular cell hyperplasias and neoplasms in the thyroid were significantly increased at dose levels of more than 300mg/kg/day. Combined incidences of squamous cell hyperplasias and papillomas of the forestomach were also significantly increased at 300 and 1000mg/kg/day. Incidences and multiplicities of adenocarcinomas in the colon were increased at 1000mg/kg/day. The numbers and areas of glutathione S-transferase placental form (GST-P) positive foci per unit area of the liver sections, and the incidence of hepatocellular adenomas were also significantly increased at 1000mg/kg/day, along with multiplicities of atypical hyperplasias of renal tubules of the kidney and the incidence of papillomatosis of the urinary bladder. This latter lesion was also seen at low incidence at 1000mg/kg/day without initiation. Thus, the current results indicate that ETBE has tumor promoting potential for the thyroid and forestomach at dose levels of 300mg/kg/day and more, and for the colon, liver, kidney and urinary bladder at 1000mg/kg/day, under the present experimental conditions.

Concordance between Results of Medium-term Liver Carcinogenesis Bioassays and Long-term Findings for Carcinogenic 2-Nitropropane and Non-carcinogenic1-Nitropropane in F344 Rats.

This study was conducted to determine the concordance of results for a pair of structural isomers, 2-nitropropane (2-NP) and 1-nitropropane (1-NP), using the rat medium-term liver carcinogenesis bioassay (Ito test) and previously published long-term carcinogenicity tests. Male F344 rats were given a single intraperitoneal injection of DEN (200 mg/kg b.w.) to initiate hepatocarcinogenesis. After 2 weeks, they received per os 0, 0.8, 4 or 20 mg/kg/day of 2-NP or 1-NP six times a week and were subjected to two-thirds partial hepatectomy at week 3. Non-initiated groups receiving 0 or 20 mg/kg/day were also included. The animals were sacrificed for quantitative analysis of GST-P-positive foci at week 8. With the highest dose of 2-NP, significantly increased numbers and areas of GST-P-positive foci were demonstrated as compared with the respective control but were not noted with 1-NP. In the non-DEN-initiated groups, many small GST-P-positive foci of less than 0.2 mm in diameter were also induced in the rats treated with 2-NP at 20 mg/kg/day but were lacking with 1-NP. These results strongly support that 2-NP is a complete hepatocarcinogen with a potent initiation activity, whereas 1-NP is not.

Lack of skin carcinogenicity of topically applied titanium dioxide nanoparticles in the mouse.

This study was conducted to examine the post-initiation carcinogenic potential of coated and uncoated titanium dioxide nanoparticles (CTDN and UCTDN) using a mouse medium-term skin carcinogenesis bioassay. For this purpose, 5, 10 and 20mg/animal doses of CTDN or UCTDN were applied to mouse skin in the post-initiation phase (up to 20 weeks) in a two-stage skin carcinogenesis model using 7 week old CD1 (ICR) female mice. 7,12-dimethylbenz[a]anthracene (DMBA) and 12-o-tetradecanoylphorbol 13-acetate (TPA) were used as the initiator and a positive control promoter, respectively. Pentalan 408 served as a vehicle control. No changes in survival rate, general condition and body weight related to the test materials were observed. On macroscopic observation, 1-2 nodules/group on the skin were observed in each group applied CTDN and UCTDN as well as the control group after DMBA initiation. The nodules were histopathologically diagnosed as squamous cell hyperplasia, sebaceous gland hyperplasia, squamous cell papilloma and keratoacanthoma. CTDN and UCTDN experiments, while enlargement of the mandibular, pancreatic, lumbar region and inguinofemoral lymph nodes, spleen and thymus was observed in mice given 5 and 10mg but not 20mg, the lack of dose-dependence suggests no biological significance. In the present study, CTDN and UCTDN applied in post-initiation stages at doses of up to 20mg/mouse did not increase the development of nodules, and thus it was concluded that titanium dioxide nanoparticles do not possess post-initiation potential for mouse skin carcinogenesis.