ABSTRACT Treatment of gravid mice (day 13 of gestation) with azosemide induces wavy ribs in their fetuses. The present study examined the morphological sequence of azosemide-induced wavy ribs, from their appearance through disappearance, by means of cartilage-bone double stain, hematoxylin and eosin stain, von Kossa's stain for calcium salts and alkaline phosphatase stain. Both endochondral and intramembranous ossifications of the ribs were inhibited in day 14 azosemide-treated fetuses. A curvature of the ribs occurred on day 15 of gestation, and excessive uncalcified osteoid was present on the outer surfaces of the bent region. In the region of curvature, endochondral and intramem- branous ossification, which is remarkable in normal, was not observed. Calcification, which began on day 14 in untreated fetuses, started including the bent region on day 16 of gestation. However, ribs in azosemide-treated fetuses exhibited alkaline phosphatase activity like in the control, during days 14-16 of gestation. These observations suggest that the defective ossification in azosemide-treated fetuses is caused by inhibition of cal- cium salts deposition on the uncalcified osteoid. During the lactation period, the bend of ribs was gradually normalized, and the bend was disappeared on day 9 postpartum.
ABSTRACT Tri-n-butyltin oxide (TBTO) is a biocide used in wood preservatives and ship paint system. Available literature data on maternal toxicity and embryotoxicity of TBTO in mice appears to be contradictory. The purpose of the study was to determine a no-observed-adverse-effect-level (NOAEL) for these parameters and to reevaluate terato- genic potency of the substance. Pregnant mice were treated orally by gavage daily with 0.5, 1.5, 4.5, 13.5, or 27 mg TBTOkg body weight (b.w.) on days 6-17 of gestation. The control group received 5 ml/kg b.w. peanut oil daily during the entire treatment period. Evidence of maternal toxicity was only observed at the dose level of 27 mg TBTO/kg. In this group 3 of 40 animals died during the treatment period. The number of implantation sites, number of viable fetuses, fetal sex ratio, number of dead fetuses as well as the number of resorptions showed no dose-related effects. The fetuses of dams exposed to the highest dose (27 mg TBTO/kg b.w.) revealed a high incidence of cleft palate (11.4%) when compared to the fetuses of the vehicle control group (0.8%). Additionally at this dose level 2 fetuses (1.2%) exhibited a bent radius; 8 fetuses (5%) were observed with a short mandible, and 5 fetuses (3%) showed a fusion of occipital bones with their basal parts. No signs of maternal and prenatal toxicity could be detected in mice under the experimental conditions chosen up to the dose of 13.5 mg TBTO/kg b.w. which represents a no-observed-adverse-effect level (NOAEL).