Genotoxicity of phenacetin in the kidney and liver of Sprague-Dawley gpt delta transgenic rats in 26-week and 52-week repeated-dose studies.

Transgenic rat mutation assays can be used to assess genotoxic properties of chemicals in target organs for carcinogenicity. Mutations in transgenes are genetically neutral and accumulate during a treatment period; thus, assays are suitable for assessing the genotoxic risk of chemicals using a repeated-dose treatment paradigm. However, only a limited number of such studies have been conducted. To examine the utility of transgenic rat assays in repeated-dose studies, we fed male and female Sprague-Dawley gpt delta rats with a 0.5% phenacetin-containing diet for 26 and 52 weeks. A long-term feeding of phenacetin is known to induce renal cancer in rats. Phenacetin administration for 52 weeks in males significantly increased gpt (point mutations) mutant frequency (MF) in the kidney, the target organ of carcinogenesis. In the liver, the nontarget organ of carcinogenesis, gpt MFs were significantly elevated in phenacetin treatment groups of both genders during 26- and 52-week treatments. Furthermore, sensitive to P2 interference (Spi(-)deletions) MF increased in the liver of both genders following 52-week treatment. MFs were higher after treatment for 52 weeks than after treatment for 26 weeks. Frequencies of phenacetin-induced mutations were higher in the liver than in the kidney, suggesting that the intensity of genotoxicity does not necessarily correlate with the induction of tumor formation. Results from gpt delta rat assays of repeated-dose treatments are extremely useful to elucidate the relationship between gene mutations and carcinogenesis in the target organ induced by cancer-causing agents.

Evaluation of the genotoxicity of tamoxifen in the liver and kidney of F344 gpt delta transgenic rat in 3-week and 13-week repeated dose studies.

Transgenic rat gene mutation assays can be used to assess genotoxicity of chemicals in target organs for carcinogenicity. Mutations in transgenes are genetically neutral and accumulate during a treatment period; thus, the assays are suitable for assessment of the genotoxicity risk of chemicals using a repeated-dose treatment paradigm. However, few such studies have been conducted. To examine the utility of the transgenic rat assays in repeated-dose studies, we treated female F344 gpt delta rats with tamoxifen (TAM) at 20 and 40mg/kg, or toremifene (TOR) at 40mg/kg by gavage daily for 3 weeks. We also fed gpt delta rats with TAM at either 250ppm (15.4-17.6mg/kg) or 500ppm (30.0-32.9mg/kg) for 13 weeks. TAM is carcinogenic in the rat liver and TOR is not carcinogenic. TAM administration significantly increased gpt (point mutations) and Spi(-) (deletions) mutant frequencies (MFs) in the liver, the target organ of carcinogenesis; MFs were higher after treatment for 13 weeks than after treatment for 3 weeks. The MFs in the kidney did not increase in any of the TAM treatment groups. TOR had no effect on MFs (gpt and Spi(-)) in either the liver or the kidney. We conclude that the gpt delta rat assay in the repeated-dose treatment paradigm is sensitive enough to detect gene mutations induced by TAM in the target organ for carcinogenesis. Furthermore, the assay can be integrated into a 13-week dose-finding study for a 2-year cancer bioassay.

Integration of in vivo genotoxicity and short-term carcinogenicity assays using F344 gpt delta transgenic rats: in vivo mutagenicity of 2,4-diaminotoluene and 2,6-diaminotoluene structural isomers.

An important trend in current toxicology is the replacement, reduction, and refinement of the use of experimental animals (the 3R principle). We propose a model in which in vivo genotoxicity and short-term carcinogenicity assays are integrated with F344 gpt delta transgenic rats. Using this model, the genotoxicity of chemicals can be identified in target organs using a shuttle vector lambda EG10 that carries reporter genes for mutations; short-term carcinogenicity is determined by the formation of glutathione S-transferase placenta form (GST-P) foci in the liver. To begin validating this system, we examined the genotoxicity and hepatotoxicity of structural isomers of 2,4-diaminotoluene (2,4-DAT) and 2,6-diaminotoluene (2,6-DAT). Although both compounds are genotoxic in the Ames/Salmonella assay, only 2,4-DAT induces tumors in rat livers. Male F344 gpt delta rats were fed diet containing 2,4-DAT at doses of 125, 250, or 500 ppm for 13 weeks or 2,6-DAT at a dose of 500 ppm for the same period. The mutation frequencies of base substitutions, mainly at G:C base pairs, were significantly increased in the livers of 2,4-DAT-treated rats at all three doses. In contrast, virtually no induction of genotoxicity was identified in the kidneys of 2,4-DAT-treated rats or in the livers of 2,6-DAT-treated rats. GST-P-positive foci were detected in the livers of rats treated with 2,4-DAT at a dose of 500 ppm but not in those treated with 2,6-DAT. Integrated genotoxicity and short-term carcinogenicity assays may be useful for early identifying genotoxic and nongenotoxic carcinogens in a reduced number of experimental animals.

Protective effects of alpha1-acid glycoprotein and serum amyloid A on concanavalin A-induced liver failure via interleukin-6 induction by ME3738.

We examined whether the 22beta-methoxyolean-12-ene-3beta,24(4beta)-diol (ME3738)-mediated selective induction of interleukin-6 increased alpha1-acid glycoprotein and serum amyloid A expression, and whether these proteins protected against liver injury in vitro and in vivo. ME3738 treatment in male mice increased gene expression of alpha1-acid glycoprotein subtypes and serum amyloid A 2 genes, and plasma concentration of serum amyloid A. Treatment with alpha1-acid glycoprotein at 5 mg/animal or serum amyloid A at 0.03 and 0.1 mg/animal prior to concanavalin A administration reduced multifocal necrosis in the liver. Treatment with alpha1-acid glycoprotein and serum amyloid A, but not alpha1-antitrypsin, protected Hep G2 cells against cell injury. These results suggest that alpha1-acid glycoprotein and serum amyloid A, increased by ME3738-induced interleukin-6, might protect against concanavalin A-induced liver injury.