Statistical analysis for comparing the incidence of tumors in treated groups to historical control data is not generally performed. In the present study, a number of data exhibiting the lowest, low, moderate and highest incidence of tumors from long-term rodent bioassay studies has been compared with the historical control data. In studies exhibiting the lowest incidence (less than a few percent) of tumors, the Kastenbaum and Bowman test was found to be relevant since it takes into account the sample size of both the historical control data base and each treated group in the study. In studies where a wider range of tumor incidence was exhibited, a statistical method which employs a rejection limits based on the range of incidence in the historical data is recommended. When malignant tumors are evident in treated groups, no matter how low the incidence, they should be analyzed statistically and compared with the incidence in historical control data as well as those in the concurrent control group. Statistical analytical comparisons of study results to historical control data may contribute to more meaningful evaluations in carcinogenicity studies by eliminating possible false positive or false negative results.
Effects of itraconazole (ICZ) on the immune responses were studied in ICR mice. Mice were divided into 5 groups (10 mice/group), and ICZ at doses of 10, 20, 40 and 80 mg/kg were orally administered to mice once a day for 21 days. Mice were immunized and challenged with sheep red blood cells (SRBC). The body weight gains and the relative weights of spleen and thymus were dose-dependently increased following ICZ treatment. However, Plaque forming cells (PFC) and hemagglutination (HA) titers to SRBC were significantly suppressed in mice dosed at 80 mg/kg ICZ, as compared with those in controls. Delayed-type hypersensitivity (DTH) reaction to SRBC, phagocyte activity and circulating leukocytes also were significantly decreased in mice dosed at 40 and 80 mg/kg ICZ. These studies demonstrate that ICZ treatment results in a marked suppression in both humoral and cell-mediated immune responses to SRBC at concentrations producing embryotoxicity.
Braxin C (BC), ptaquiloside, is a toxin isolated from bracken fern. In order to determine the characteristics of the acute BC toxicity, BC was administered as a single subcutaneous (s. c.) injection to guinea pigs, rats, and mice. BC (5 and 10 mg/kg) induced hemorrhagic cystitis and hematuria in guinea pigs, whereas no changes were observed in rats and mice administered BC at a dose of 100 mg/kg. In guinea pigs administered BC (5 and 10 mg/kg), marked edema and hemorrhage were noted in the lamina propria of urinary bladder, though these inflammatory changes were also observed in the tissues such as perirenal adipose tissues or abdominal wall. These findings clearly demonstrate that BC is a potent toxin capable of inducing hemorrhagic cystitis in guinea pigs, and suggest that it is one of the toxic principles responsible for bracken poisoning.
A stepwise, simple screening test for skin and eye irritations, suitable for industrial chemicals or pesticides which are not required to be examined for their exact potential irritancy levels, was developed. The efficacy of the test was evaluated using 15 chemicals including typical irritants (acetic acid, ammonia, chloroacetic acid, dioxane ethanolamine, formaldehyde, formic acid, hydrogen peroxide, phenol, phosphoric acid, propionic acid, sodium hydroxide, sodium hypochlorite, sulfuric acid, and trichloroacetic acid). Chemicals were chosen so as to represent irritants which act by different mechanisms (i. e., strongly acidic, alkaline, reactive to protein, oxidizing etc.). The method consisted of physicochemical tests and animal tests using rats, mice or guinea pigs, namely, a skin irritation test, an intradermal reaction test and an eye irritation test in a sequential manner such that further tests are not required if a positive result is obtained in earlier steps. Results obtained between two laboratories using this method were very similar. Comparison of our results with the data obtained by the conventional method using rabbits showed fairly good coincidence. The method was shown to be useful in assessing skin and eye irritation of chemicals and causes minimal suffering to animals.
An inter-laboratory validation study was conducted to evaluate the potential of 4 chemicals to cause irritation with utilizing the Skin2 Dermal Model ZK1100 kit developed by Advanced Tissue Sciences, Inc. (formerly Marrow-Tech, Inc., La Jolla, California, USA). The chemicals tested were sodium dodecyl sulfate (SDS), 1-n-hexadecyl-pyridinium chloride monohydrate (CC), ethanol (EtOH), and dimethyl sulfoxide (DMSO). Eleven Japanese insititutions participated in this validation research to evaluate the usefulness of the Skin2 Model ZK1100 kit in accordance with an identical protocol. None of the participating laboratories had previously used the Skin2 Model ZK1100 kit. The MTT-50 value obtained in the individual institutions was 42 to 91 μg/ml for SDS, 2.7 to 8.6 μg/ml for CC, 2.0 to 9.3% for EtOH, and 11.5 to 21.9% for DMSO. Reproducibility was reasonably good as noted when one test chemical was repetitively tested by the same investigator. MTT-50 values obtained with the present method correlated with DS20 values obtained with Draize's method (r=0.9881) in one of the participant institutions. The irritation study using the Skin2 Model ZK1100 kit was easy to perform and generated quantitative data. When the test was repeated, reproducibility was demonstrated with a variation of less than 2 6. These data suggested that this newly developed in vitro method would be useful in toxicity screening studies in terms of both time and cost, and would serve as a useful alternative to the conventional methods of the eye irritation study.
An interrelation of Cd-contents with hepato- and nephrotoxicities, and a mechanism for Cd-exclusion from kidneys were investigated in rats that received a single intravenous injection of either CdCl2 or Cd-saturated metallothionein II (Cd-MT-II) with doses of 0.1 and 0.3 mg Cd/kg body weight (b.w.). Between the livers and kidneys, higher uptake of Cd was observed in the liver in terms of CdCl2, and in the kidney in terms of Cd-NT-II. The CdCl2 at the two doses hardly showed any histopathological alterations in the livers and kidneys. The 0.1 mg Cd/kg b.w. of Cd-MT-II showed a slight injury in the kidneys, while hardly in the livers. At Day 1 of the 0.3 mg Cd/kg b.w. of Cd-MT-II, the renal Cd-contents reached the maximal value of 8.22±0.36 μg/g wet tissue, and degeneration including necrosis and defluxion of proximal tubular cells were most highly observed. At Day 5 of the 0.3 mg Cd/kg b.w., the renal Cd-contents were lowered to 2.40±0.24 μg/g wet tissue, and fibrosis and regeneration of the proximal tubular cells were remarkably found. These findings strongly suggested that, in the case of administration of Cd-MT-II, the Cd taken into the kidneys was eliminated from there mainly by cellular death of the proximal tubulus and by their resultant defluxion.