Juvenile rats do not exhibit elevated sensitivity to acrylamide toxicity after oral administration for 12 weeks.

Acrylamide (AA), a neurotoxic, testicular toxic, genotoxic and carcinogenic chemical, has been reported to be formed in processed food, and sensitivity to AA intoxication in childhood is a concern. In the present study, to clarify the general toxicological profile of AA in juvenile rats, subchronic toxicity was evaluated in F344 rats administered AA in the drinking water at 0 (control), 10, 20 and 40 ppm, presented to the dams (three per group) immediately after the birth of their litters, through lactation (3 weeks), and directly to the offspring in their drinking water after weaning for a further 9 weeks (12 weeks total). Treatment with AA caused a decrease in body weights in 20 and 40 ppm F(1) females, compared with the controls. Average AA intake throughout the treatment period for the 10, 20 and 40 ppm groups after weaning was equivalent to 1.0, 2.1 and 4.4 mg kg(-1) body weight per day, respectively, in males and 1.2, 2.5 and 4.9 mg kg(-1) body weight per day, respectively, in females. No toxicologically significant organ weight changes were observed. AA-induced histopathological changes were limited to focal degeneration and necrosis of the seminiferous epithelium in the testes and desquamated epithelium in the ducts of epididymides, noted only in 40 ppm males. Taken together with previous reports, juvenile rats are not necessarily more susceptible to AA-induced toxicity as compared with young adults.

Low-dose carcinogenicity of 2-amino-3-methylimidazo[4,5-f ]quinoline in rats: Evidence for the existence of no-effect levels and a mechanism involving p21(Cip / WAF1).

The carcinogenicity of the low amounts of genotoxic carcinogens present in food is of pressing concern. The purpose of the present study was to determine the carcinogenicity of low doses of the dietary genotoxic carcinogen 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and to investigate mechanisms by which IQ exerts its carcinogenic effects. A total of 1595 male F344 rats were divided into seven groups and administered with IQ at doses of 0, 0.001, 0.01, 0.1, 1, 10 and 100 p.p.m. in the diet for 16 weeks. We found that IQ doses of 1 p.p.m. and below did not induce preneoplastic lesions in either the liver or the colon, while IQ doses of 10 and 100 p.p.m. induced preneoplastic lesions in both of these organs. These results demonstrate the presence of no-effect levels of IQ for both liver and colon carcinogenicity in rats. The finding that p21(Cip/WAF1) was significantly induced in the liver at doses well below those required for IQ mediated carcinogenic effects suggests that induction of p21(Cip/WAF1) is one of the mechanisms responsible for the observed no-effect of low doses of IQ. Furthermore, IQ administration caused significant induction of CYP1A2 at doses of 0.01-10 p.p.m., but administration of 100 p.p.m. IQ induced CYP1A1 rather than CYP1A2. This result indicates the importance of dosage when interpreting data on the carcinogenicity and metabolic activation of IQ. Overall, our results suggest the existence of no-effect levels for the carcinogenicity of this genotoxic compound.

Effect of cigarette smoke on mutagenic activation of environmental carcinogens by cytochrome P450 2A8 and inactivation by glucuronidation in hamster liver.

To elucidate the mechanism underlying suppression of N-nitrosobis(2-oxopropyl)amine (BOP)-induced hamster pancreatic carcinogenesis by cigarette smoke (CS), hepatic levels of microsomal cytochrome P450 (CYP) enzymes, mutagenic activation of environmental carcinogens and three types of uridine diphosphate-glucuronyltransferase (UDPGT) and sulphotransferase (ST) activities were assayed in male Syrian golden hamsters and F344 rats exposed to CS. Immunoblot analyses of microsomal CYP proteins revealed induction of constitutive CYP1A2 (2.6-fold increase) and 2A8 (4.0-fold increase) and induction of CYP1A1 and constitutive CYP1A2 (3.9-fold increase) in rats following exposure to CS for 4 weeks using a Hamburg type II smoking machine. CS exposure enhanced mutagenicities of four heterocyclic amines in the presence of liver S9 in both species, whereas the mutagenicities of aflatoxin B(1) (AFB(1)), 2-amino-3-methyl-9H-pyrido[2,3-b]indole (MeAαC) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) were significantly increased by CS in hamsters but not in rats. However, no CS-induced alterations in the mutagenic activities of other carcinogens, including BOP and other pancreatic carcinogens, were observed in either species. Application of several CYP inhibitors revealed that the mutagenic activities of MeAαC, AFB(1) and NNK in the hamster liver S9 were partly associated with CYP2A8, whereas those of the three pancreatic carcinogens were selectively associated with CYP2B. CS enhanced UDPGT activities towards 4-nitrophenol (4-NP) (1.9- to 2.0-fold) but did not affect those of bilirubin, testosterone UDPGTs and three STs in both species. Together with the previous findings that BOP does not induce tumourigenesis in rats and that the glucuronidation of ß-oxypropylnitrosamines is higher in rats than in hamsters, suppression of BOP-induced pancreatic carcinogenesis by CS might be attributed to increased detoxification by 4-NP UDPGT and not decreased CYP2B activation. This is the first demonstration of the induction of CYP2A protein by CS; CYP2A protein polymorphisms have been associated with oral and pulmonary carcinogenesis in smokers.

Preventive effects of calcitriol on the development of capsular invasive carcinomas in a rat two-stage thyroid carcinogenesis model.

We have shown phosphoinositide 3-kinase (PI3K)/Akt signaling activation in thyroid capsular invasive carcinomas (CICs), which are highly induced by promotion with sulfadimethoxine (SDM) in a rat 2-stage thyroid carcinogenesis model. To examine the potency of calcitriol, a synthetic vitamin D3 analog, on the development or progression of CICs, male F344 rats were injected with calcitriol (0.1 µg/kg body weight) three times a week intraperitoneally, during an entire period of SDM-promotion for 13 weeks (Experiment 1) or during the last 2 weeks of a 15-week SDM-promotion (Experiment 2). Initiation with N-bis(2-hydroxypropyl)nitrosamine preceded all treatments. In Experiment 1, long-term calcitriol treatment reduced the multiplicity of CICs, while cell proliferation activity, estimated by Ki-67 cell index in the induced CICs, was unchanged with SDM-promotion alone. Considering the strong dependency of promotion with SDM during the early stages on thyroid-stimulating hormone, the reduced multiplicity in Experiment 1 may be due to the effect on an early stage of neoplastic proliferation. Although the magnitude was mild, cell proliferation activity was decreased in existing CICs after short-term calcitriol treatment in Experiment 2, which was associated with a mild decrease in cyclin-dependent kinase-2-positive cells, cytoplasmic immunolocalization of phosphorylated, inactive, Rb protein and a mild increase in nucleocytoplasmic expression of p27(kip1). Although the effect was mild at the late stage of SDM-promotion in this hypothyroidism-related thyroid carcinogenesis model, our results suggest that calcitriol targets cell proliferation via inhibition of a molecular cascade downstream of PI3K/Akt signaling, controlling G1/S transition.

Identification and metastatic potential of tumor-initiating cells in malignant rhabdoid tumor of the kidney.

PURPOSE: Malignant rhabdoid tumor of the kidney (MRTK) is a rare and highly aggressive malignancy of infanthood. In an effort to delineate MRTK progression, we investigated the metastatic fate of some MRTK cells using xenotransplantation animal models and the tumor-initiating potential of CD133(+) MRTK cells. EXPERIMENTAL DESIGN: We established two MRTK cell lines (JMU-RTK-1 and JMU-RTK-2) from patients with MRTK. We generated five luciferase-expressing MRTK cells for in vivo luminescent imaging and evaluated the metastatic fate in an orthotopic xenotransplantation model. Capacities of MRTK-initiating cells were examined in nonobese diabetic/severe combined immunodeficient mice after antibody-mediated magnetic bead sorting. Use of chemokine receptor CXCR4 expression as a metastatic marker was evaluated by flow cytometry and Western blotting. RESULTS: MRTK cell lines showed distant organ metastasis. JMU-RTK-1, JMU-RTK-2, and G401 cells showed considerable aggressiveness compared with SWT-1 and SWT-2 cells (P < 0.05). Moreover, as few as 1,000 CD133(+) MRTK cells initiated tumor development in nonobese diabetic/severe combined immunodeficient mice by 21 days (60-100%) in all examined cell lines, although the same number of CD133(-) MRTK cells could not form tumors (0%). Interestingly, the metastatic potential of the CD133(+) population remained unaffected compared with a nonenriched population. The potential metastatic marker CXCR4 was expressed in CD133(+) and CD133(-) MRTK cells, and CD133(-) cells seemed to play a cooperative role in terms of tumorigenicity and metastasis. CONCLUSIONS: These results suggest that CD133(+) cells may determine the metastatic fate of MRTK cells and that CD133(-) cells may play an auxiliary role in tumor progression and metastasis.

Gene expression profiling and cellular distribution of molecules with altered expression in the hippocampal CA1 region after developmental exposure to anti-thyroid agents in rats.

To determine whether developmental hypothyroidism causes permanent disruption of neuronal development, we first performed a global gene expression profiling study targeting hippocampal CA1 neurons in male rats at the end of maternal exposure to anti-thyroid agents on weaning (postnatal day 20). As a result, genes associated with nervous system development, zinc ion binding, apoptosis and cell adhesion were commonly up- or down-regulated. Genes related to calcium ion binding were up-regulated and those for myelination were often down-regulated. We, then, examined immunohistochemical cellular distribution of Ephrin type A receptor 5 (EphA5) and Tachykinin receptor (Tacr)-3, those selected based on the gene expression profiles, in the hippocampal formation at the adult stage (11-week-old) as well as at the end of exposure. At weaning, both EphA5- and Tacr3-immunoreactive cells with strong intensities appeared in the pyramidal cell layer or stratum oriens of the hippocampal CA1 region. Although the magnitude of the change was decreased at the adult stage, Tacr3 in the CA1 region showed a sustained increase in expressing cells until the adult stage after developmental hypothyroidism. On the other hand, EphA5-expressing cells did not show sustained increase at the adult stage. The results suggest that developmental hypothyroidism caused sustained neuronal expression of Tacr3 in the hippocampal CA1 region, probably reflecting a neuroprotective mechanism for mismigration.

Carcinogenic potential of alizarin and rubiadin, components of madder color, in a rat medium-term multi-organ bioassay.

Madder color (MC), a food coloring extracted from roots of Rubia tinctorum L., has been proven to exert carcinogenicity in the rat kidney and liver. Furthermore, it induces DNA adducts in the kidney, liver, and colon. MC is in fact composed of anthraquinones such as lucidin-3-O-primeveroside and alizarin. To clarify which of these might be responsible for the carcinogenicity, a rat medium-term multi-organ carcinogenesis bioassay was performed focusing on the kidney, liver, and colon. Male 6-week-old F344 rats after receiving five different carcinogens were fed a diet containing either 0.008% or 0.04% of alizarin or rubiadin, a metabolite of lucidin-3-O-primeveroside, for 23 weeks. Treatment with 0.04% rubiadin significantly increased atypical renal tubules/hyperplasias and induced renal cell adenomas and carcinomas. Renal cell tumors were also increased with 0.04% alizarin, although at lower incidence than with rubiadin. In addition, glutathione S-transferase placental form-positive liver cell foci and large intestinal dysplasias were significantly increased with 0.04% rubiadin. These results indicate that both rubiadin and alizarin can increase renal preneoplastic lesions, the potential of the latter being weaker. Rubiadin may also target the liver and large intestine, suggesting a major role in madder color-induced carcinogenicity.

Cellular distributions of molecules with altered expression specific to thyroid proliferative lesions developing in a rat thyroid carcinogenesis model.

To identify differentially regulated molecules related to early and late stages of tumor promotion in a rat two-stage thyroid carcinogenesis model by an antithyroid agent, sulfadimethoxine, microarray-based microdissected lesion-specific gene expression profiling was carried out. Proliferative lesions for profiling were divided into two categories: (i) focal follicular cell hyperplasias (FFCH) and adenomas (Ad) as early lesions; and (ii) carcinomas (Ca) as more advanced. In both cases, gene expression was compared with that in surrounding non-tumor follicular cells. Characteristically, upregulation of cell cycle-related genes in FFCH + Ad, downregulation of genes related to tumor suppression and transcription inhibitors of inhibitor of DNA binding (Id) family proteins in Ca, and upregulation of genes related to cell proliferation and tumor progression in common in FFCH + Ad and Ca, were detected. The immunohistochemical distributions of molecules included in the altered expression profiles were further examined. In parallel with microarray data, increased localization of ceruloplasmin, cyclin B1, and cell division cycle 2 homolog A, and decreased localization of poliovirus receptor-related 3 and Id3 were observed in all types of lesion. Although inconsistent with the microarray data, thyroglobulin immunoreactivity appeared to reduce in Ca. The results thus suggest cell cycling facilitation by induction of M-phase-promoting factor consisting of cyclin B1 and cell division cycle 2 homolog A and generation of oxidative responses as evidenced by ceruloplasmin accumulation from an early stage, as well as suppression of cell adhesion involving poliovirus receptor-related 3 and inhibition of cellular differentiation regulated by Id3. Decrease of thyroglobulin in Ca may reflect dedifferentiation with progression.

Inhibitory effects of aminoguanidine on thyroid follicular carcinoma development in inflamed capsular regions of rats treated with sulfadimethoxine after N-bis(2-hydroxypropyl)nitrosamine-initiation.

We have reported that thyroid capsular thickening with inflammation induced by an antithyroidal agent, sulfadimethoxine (SDM), might play a role in the development of invasive follicular carcinomas in rats initiated with N-bis(2-hydroxypropyl)nitrosamine (DHPN). Inducible nitric oxide synthase (iNOS) expressed in the inflamed capsular regions further appeared to be implicated in the tumor progression. In the present study, the effects of an iNOS inhibitor, aminoguanidine (AG), on thyroid carcinogenesis were examined. F344 male rats were treated with SDM in drinking water (0.1%) with or without concomitant dietary administration of AG (0.2%) for 4 and 10 weeks after subcutaneous injection of DHPN at 2800 mg/kg bodyweight. At week 4, thyroid capsular thickening with inflammation was observed and iNOS-positive foci were found in the inflamed regions. In addition, single-strand DNA-positive inflammatory cells were scattered among neighboring follicular cells, indicating some cellular damage, at least partly in association with iNOS induction. Concurrent dietary administration of AG with SDM treatment slightly decreased the number of single-strand DNA-positive cells but did not alter the incidence and multiplicity of iNOS-positive foci in the inflamed capsular regions at week 4. At week 10, however, invasive follicular carcinomas predominantly arose in the thickened capsule in the DHPN-SDM-treated rats, and AG administration decreased (P < 0.05) their multiplicity. The carcinoma cells were partly positive for iNOS. These results thus suggested that iNOS induction in both inflammatory and tumor cells might play pivotal roles in tumor progression in this DHPN-SDM rat model.

The process behind the expression of mdr-1/P-gp and mrp/MRP in human leukemia/lymphoma.

There is a controversy over the link between phenotypes of multidrug resistance (MDR) and clinical outcome in leukemia/lymphoma patients. This may be because the process behind the induction and loss of expression of genotypes and phenotypes by which MDR develops and the role of MDR in fresh cells of human leukemia/lymphoma are not clearly defined. P-glycoprotein (P-gp) increased and decreased along with mdr-1 expression in three cell lines out of five vincristine (VCR)-resistant cell lines. MRP appeared with increased mrp expression in the other two cell lines. After the drug was removed from the culture system, mdr-1/P-gp changed in parallel with the level of VCR resistance, although mrp and MRP did not. It was concluded that P-gp is directly derived from mdr-1 and that mdr-1/P-gp supports the VCR-resistance but mrp/MRP is not directly linked to the VCR-resistance. These results should contribute to a better understanding of MDR phenomenon in cancer.

Modifying effects of prepubertal exposure to potassium perchlorate and tetrabromobisphenol A on susceptibility to N-bis(2-hydroxypropyl)nitrosamine- and 7,12-dimethylbenz(a)anthracene-induced carcinogenesis in rats.

Early life exposure to certain kinds of chemicals is of concern because of a possible increase in cancer risk, but relevant data are limited. In the present experiment, modifying effects of prepubertal administration of potassium perchlorate (KClO(4)) and tetrabromobisphenol A (TBBPA) on susceptibility to multi-organ carcinogenesis were evaluated. F344 dam rats were administered 0% (control), 0.01%, 0.1% or 1% TBBPA in diet or 0.01% KClO(4) in drinking water after parturition. Their weaned offspring in each group were treated for 2 weeks in the same manner. From 6 weeks of age, all offspring were treated with N-bis(2-hydroxypropyl)nitrosamine in drinking water for 4 weeks. In addition the females at 7 weeks of age were gavaged once with 7,12-dimethylbenz(a)anthracene. At weeks 39 and 47 of age, the males and females, respectively, were euthanized and the liver, kidney, lung, esophagus, thyroid, urinary bladder, testis, epididymis, ovary and mammary gland were histopathologically examined. The incidences of thyroid follicular adenomas in 1% TBBPA females (p<0.05) and of transitional cell papillomas in the urinary bladder of 0.01%, 0.1% and 1% TBBPA females were increased (p<0.05) as compared to the controls. These results indicate that prepubertal exposure to TBBPA raises susceptibility to thyroid and urinary bladder tumorigenesis in rats. Although causes of the effect on thyroid carcinogenesis might be direct and/or indirect hormonal actions, further studies are needed for confirmation.

ABO genotyping of various hematopoietic cell lines to select model cells for research purposes.

Various cells from humans and animals have been established as cell lines, and their features, characteristics, and origins have been reported. Many laboratories use cell lines as model cells, which are selected to suit research purposes. We attempted to identify the ABO genotypes of 31 human leukemia and lymphoma cell lines stored in our laboratory using three methods: the PCR amplification of specific alleles (PASA), PCR-restriction fragment length polymorphism (RFLP), and the direct DNA sequencing of PCR products. We distinguished 31 human leukemia and lymphoma cell lines examined into six major ABO genotypes: A/O (A101/O01: n = 1, A101/O12: n = 4, A101/O26: n = 1, A101/O49: n = 1, A102/O01: n = 3), A/A (A101/A101: n = 1, A102/A102: n = 2), B/O (Bw29/O01: n = 1), B/B (B101/B101: n = 2), O/O (O01/O01: n = 9, O01/O02: n = 1, O01/O26: n = 1, O02/O03: n = 1), and A/B (A102/B101: n = 3). To the best of our knowledge, this is the first study to identify the ABO genotypes of various cell lines. The ABO genotypes of cell lines are important when selecting an experimental model cell for an ABO blood group study, and are essential information for cell lines. These results may be employed by research and clinical laboratories as well as in the forensic field.

Cellular distributions of molecules with altered expression specific to the tumor promotion process from the early stage in a rat two-stage hepatocarcinogenesis model.

A global gene expression profiling specific to the early process of tumor promotion by fenbendazole (FB) or phenobarbital (PB) in a rat two-stage hepatocarcinogenesis model revealed 33 genes to show altered expression in common with both chemicals. The immunohistochemical distribution of transferrin receptor (Tfrc), nuclear receptor subfamily 0, group B, member 2 (Nr0b2) and minichromosome maintenance deficient 6 (MCM6), included in the altered expression profile, were therefore examined in FB- and PB-induced proliferative lesions at both early and late stages of tumor promotion. In addition, immunoexpression of transforming growth factor beta receptor (TGFbetaR) I, TGFbetaRII, phosphatase and tensin homolog deleted on chromosome 10 (PTEN) and phosphorylated phosphatase and tensin homolog deleted on chromosome 10 (pPTEN) was also examined. In the early stage, most hepatocellular foci positive for glutathione S-transferase placental form (GST-P) showed co-expression of TGFbetaRI and lack of PTEN and pPTEN, some GST-P-positive foci co-expressing Tfrc and Nr0b2. In the late stage, selective expression of TGFbetaRI, but not TGFbetaRII, was also observed in many adenomas and carcinomas consistently expressing GST-P. Nr0b2 was variably expressed in the proliferative lesions, irrespective of the carcinogenic stage. Like the GST-P-positive foci, adenomas and carcinomas consistently lacked PTEN and pPTEN. Expression of Tfrc and MCM6 was increased in parallel with the carcinogenic stage. In conclusion, loss of PTEN and dysregulation of transforming growth factor beta signaling can be considered to be involved in rat hepatocarcinogenesis from early stages. Selective expression of Tfrc in proliferative lesions suggests an involvement of changes in iron homeostasis during the process of tumor promotion/progression driven by FB or PB.

Enhancement of esophageal carcinogenesis in acid reflux model rats treated with ascorbic acid and sodium nitrite in combination with or without initiation.

Combined treatment with sodium nitrite (NaNO2) and ascorbic acid (AsA) has already been shown to promote rat forestomach carcinogenesis, possibly due to nitric oxide generation under acidic conditions. We hypothesized that a similar effect might occur in the esophagus when the luminal pH is decreased by acid reflux. To clarify this possibility, reflux esophagitis model rats (F344 male) were coadministered 0.2% NaNO2 in the drinking water and 1% AsA in the diet. After 32 weeks of the combined treatment, a significant increase in the incidence of epithelial hyperplasias of the lower-middle and lowest parts of the esophagus were observed compared with the basal-diet group, along with exacerbation of dysplasia and extension of the lesions. Additionally, one squamous cell papilloma was found only in the combined-treatment group. Subsequently, we confirmed the enhancing effects of NaNO2 and AsA cotreatment in the rat N-bis(2-hydroxypropyl)nitrosamine-initiated esophageal tumorigenesis model. The incidence of hyperplasia was enhanced in all segments, along with the incidence and multiplicity of squamous cell papillomas in the lowest segment of the esophagus. Thus, the data demonstrate that combined treatment with NaNO2 and AsA exerts promoting effects on rat esophageal carcinogenesis under acid reflux conditions, as in the forestomach. These findings suggest that the risk of excessive intake of a combination of nitrite and antioxidants for esophageal carcinogenesis is appreciable, particularly in patients with reflux esophagitis.

Induction of characteristic hepatocyte proliferative lesion with dietary exposure of Wistar Hannover rats to tocotrienol for 1 year.

Tocotrienol is an antioxidant which has found commercial application as a food additive and health supplement all over the world. Since there have been no reports regarding toxicological effects of long-term exposure, we performed a 52-week chronic study using Wistar Hannover rats of both sexes given the compound at doses of 0, 0.08, 0.4 or 2% in powdered basal diet. Since 6 animals in the 2% male group died of hemorrhage of several organs by week 50, the maximum dose level was changed to 1% in both sexes for the last 2 weeks. Decrease of body weight gain was observed in the 2% males from week 5 and females from week 10, this persisting to the end of the study. With the high dose, prolongation of prothrombin time and increase of serum ALT in males, and increase of serum ALP in both sexes were observed with statistical significance. In male and female rats receiving 0.4% or less, there were no toxicological changes in any of the parameters examined. At necropsy, multiple cyst-like nodules on the liver surface were macroscopically pronounced in both sexes receiving 2%. On histopathological examination, hepatocellular nodules were evident with distortion of hepatic cords and compression of the surrounding tissue, almost all including areas of spongiosis hepatis. The constituent hepatocytes were immunohistochemically stained with proliferation cell nuclear antigen at high rates. Nevertheless, they did not exhibit overt atypia and the basic lobular architecture remained intact. Additionally, they were consistently negative for glutathione S-transferase placental form (GST-P). Accordingly, we propose the newly categorized but previously used name 'nodular hepatocellular hyperplasia', which may not necessarily have a neoplastic or regenerative nature. However, quantitative GST-P analysis of the liver sections overall showed numbers of GST-P foci in the high dose females to be significantly elevated as compared to the control value. Based on the present data demonstrating nodular liver lesions only at the high dose of both sexes, we conclude that the no-observed-adverse-effect level (NOAEL) is 0.4% (303 mg/kg/day for males, and 472 mg/kg/day for females).

A new medium-term rat colorectal bioassay applying neoplastic lesions as end points for detection of carcinogenesis modifiers effects with weak or controversial modifiers.

We have established a two-stage, medium-term rat colorectal carcinogenesis model featuring induction of neoplastic lesions within ten weeks. In the present study, we examined the ability of this model to detect weak modifiers. F344 male rats were given three subcutaneous (sc) injections of 1,2-dimethyl-hydrazine (DMH, 40 mg/kg b.w.) in one week followed by drinking water containing 1% dextran sodium sulfate (DSS) for a second week. One week after this regimen, basal diet alone, or diets containing 10% perilla oil, 10% corn oil, 10% dextrin, or 0.1% indole-3-carbinol (I3C) were supplied. The perilla oil and corn oil groups did not show significant differences in the numbers of aberrant crypt foci (ACF) and incidences or multiplicity of proliferative lesions as compared to the controls at either time point. In the dextrin group, the total number of ACF at week ten was significantly increased. With I3C, the total number of ACF and incidence and multiplicities of adenocarcinomas at week ten and the incidence of invasive tumors at week twenty were significantly increased. These data essentially correspond with earlier reported results, except in the vegetable oil cases. Thus, the system is suitable for detection of colorectal carcinogenesis modifiers with advantages over previous models using ACF alone as end points.

A possible role of nrf2 in prevention of renal oxidative damage by ferric nitrilotriacetate.

To ascertain the possible roles of nuclear erythroid 2 p45-related factor 2 (Nrf2), a key transcription factor of phase 2 drug-metabolizing enzymes, in renal cellular defense against oxidative stress, wild-type and Nrf2-knockout -/- mice were treated with ferric nitrilotriacetate (Fe-NTA) at doses of 3 or 6 mg iron/kg body weight. After Fe-NTA treatment, Nrf2 -/- mice consistently showed lower levels of glutathione (GSH) in the kidney at the low dose and the liver at the high dose than the wild-type mice. Gamma-glutamylcysteine ligase (GCL) activity in the kidney and liver of Nrf2-/- mice was also consistently lower than in wild-type mice after the Fe-NTA treatment. Histopathological examination revealed that nephrotoxicity of Fe-NTA, reflected in necrosis of renal tubule epithelial cells following nuclear damage, was more severe in the Nrf2-/- mice than in their wild-type counterparts. Overall, the data suggest that Nrf2 -/- mice are unable to compensate for depletion of renal GSH because of oxidative stress, being more susceptible to Fe-NTA-induced nephrotoxicity. In conclusion, the present study showed that Nrf2 might play an important role in protecting cells from oxidative stress in the kidney through its regulation of antioxidant enzymes.

Evaluation of subchronic toxicity of dietary administered Cry1Ab protein from Bacillus thuringiensis var. Kurustaki HD-1 in F344 male rats with chemically induced gastrointestinal impairment.

Bacillus thuringiensis (Bt) proteins are developed for genetically modified crops and the Bt proteins demonstrate no evidence of toxicity by the oral route in traditional animal models. However, the possible toxicity of Bt proteins under conditions of reduced gastric acid secretion and/or small intestinal damage has not been investigated. In the present study, we therefore evaluated following four F344 rat groups with a purified Bt protein Cry1Ab from B. thuringiensis var. Kurustaki HD-1. Gastrointestinal impairment (GI) alone and GI+Bt protein fed (GI+Bt) groups were given i.p. injections of famotidine to reduce gastric acid secretion twice a day at 30mg/kg body weight in weeks 2 and 4. GI and GI+Bt groups were additionally fed diets containing 80ppm indomethacin for induction of intestinal damage during weeks 1 and 3. Bt alone and GI+Bt groups were also fed diet containing Bt protein Cry1Ab at a concentration of 10ppm in weeks 2 and 4. A no treatment control group was also included. At the end of week 4, all animals were euthanized under ether anesthesia, blood samples were collected for hematology and serum biochemistry and a complete necropsy was performed. No significant changes indicative of toxicity of the Bt protein Cry1Ab used here were noted with any of the parameters investigated. In conclusion, no significant toxicological effects were detected in this subchronic gastrointestinal impairment rat model.

Modifying effects of chitin, chitosan and their related compounds on 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) in a rat medium-term hepatocarcinogenesis model, and their post-initiation effects in a female rat 2-stage multi-organ carcinogenesis model.

The modifying effects of chitin, chitosan, chitin-oligo sugar, chitosan-oligo sugar and chlorophyllin-chitosan on 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) was investigated in a rat medium-term hepatocarcinogenesis model. Male F344 rats were injected with diethylnitrosamine (DEN) and starting 2 weeks later, received 0.03% MeIQx alone, MeIQx plus each chemical (0.4%), or each chemical alone (0.1%) in diet for 6 weeks. Three weeks after the DEN injection, animals were subjected to 2/3 partial hepatectomy. The numbers and areas of glutathione S-transferase placental form (GST-P) positive foci given MeIQx plus test chemicals were similar to the MeIQx alone values. In a second experiment, post-initiation effects of chitin and chitosan on major organs were examined in female F344 rats after initiation with 1,2-dimethylhydrazine (DMH), 7,12-dimethylbenz[a]anthracene (DMBA) and 2,2'-dihydroxy-di-n-propylnitrosamine (DHPN). In rats fed a diet containing 1.0% chitin for 36 weeks, the development of palpable mammary tumors tended to be delayed and the final incidence and multiplicity of adenocarcinomas, were significantly lowered. However, in the colon, the number of aberrant crypt foci (ACF) in the chitin and chitosan groups was significantly increased. These results indicate that chitin, chitosan and related compounds do not exert unequivocal chemopreventive effects on heterocyclic amine-induced hepatocarcinogenesis, and that effects in other organs may be tissue specific with possible inhibitory action in the mammary gland being offset by promotion of colon lesion development.