An in vitro screening system for characterizing the cleft palate-inducing potential of chemicals and underlying mechanisms.

An in vitro organ culture system with developing mouse palates was improved to characterize the cleft palate (CP)-inducing potential of chemicals and underlying mechanisms. Palatal explants collected from gestation day 12 mouse fetuses were cultured with various concentrations of teratogens and examined for palatal development after 48 and 72 h of culture to assess effects of the chemicals on growth and/or fusion of palatal shelves. When the explants were exposed to diphenylhydantoin or 5-fluorouracil, palatal growth was inhibited in a concentration-dependent manner at 48 h. Suppression of the expression of proliferative cell nuclear antigen revealed poor cell proliferation. At 72 h, the incidence of explants with CP was significantly increased in the high-dose groups, suggesting that CP induction is mainly attributable to inhibition of palatal growth. By contrast, retinoic acid and hydrocortisone significantly lowered the rates of fused palates at 72 h in all treated groups, while they exhibited no effects on palatal growth at 48 h even at the highest concentration. Because no apoptosis was found in the epithelial cells at the tip of these palates, these chemicals are suggested to inhibit palatal fusion process by preventing apoptosis.

Morphogenetic study of the eyelids in NC-eob mice fetuses with an open-eyelid malformation at birth.

The NC-eob mice are mutants having open eyelids at birth with complete penetrance. Detailed scanning electron microscopy and light microscopy for study of the eyelid development of NC-eob embryos/fetuses were performed at days 13 to 17 of gestation and compared with normal NC mice, with the aim of investigating the etiopathogenesis of the open eyelids at birth. No difference was observed between NC and NC-eob embryos by scanning electron microscopy and histologic examination at days 13 and 14 of gestation. At day 15 the epithelium at the tip of the eyelids had 2 to 3 layers, and clumps of round periderm cells appeared on the outer surface of the margin of the lids in NC and NC-eob embryos. However, these clumps in NC-eob embryos were not as remarkable in number and in size compared with NC embryos. At day 16 the eyelids of NC fetuses extended over the cornea and began to fuse. The epithelium at the tip of the eyelids consisted of 4 to 5 layers with the following elongated area. The surface of the tip of the elongated epithelium was covered with a large number of round periderm cells. In NC-eob fetuses the epithelium remained stationary at 2 to 3 layers. Only a small number of round cells were noted at the tip of the eyelids, and fusion did not occur. At day 17, although the upper and lower eyelids of NC fetuses were entirely fused, no change occurred in NC-eob fetuses, either in the periderm or in the epithelium.(ABSTRACT TRUNCATED AT 250 WORDS)

Reduced fertility in females of the hereditary polydactyly rat.

The present studies assessed reproductive capacity of hereditary polydactyly(pd/pd) female rats that had genetically caused polydactyly in both fore- and hindlimbs. During an 8- or 10-day period of vaginal estrous cycle observation, persistent estrus was found in 30 to 39% of the pd/pd females. Although mating indices were comparable between polydactylous (pd/pd) and phenotypically normal (pd/+ or +/+) females, fertility indices were significantly decreased in pd/pd females. All impregnated pd/pd females delivered normally but had a significant decrease in the numbers of corpora lutea and pups delivered as compared with pd/+ or +/+ females. Examination of oviductal ova at 2 days after copulation revealed ovulation in only 75% of the pd/pd females and also a slight decrease in the numbers of ovulatory corpora lutea and oviductal ova in these females. In the nonovulating persistent estrus pd/pd females, the ovaries were polycystic. These results demonstrate a reduction in reproductive ability of pd/pd females.

A two-generation reproduction study in rats with methyl bromide-fumigated diets.

Crj:CD(SD) rats were fed basal (control) diet or methyl bromide-fumigated diet containing 80, 200 or 500 ppm total bromine for two successive generations (18 wk for each generation). Compared with controls, food consumption was significantly lower in F1 parental females of the 500-ppm group during the second half of the dosing period, and F2 female pups of the 500-ppm group showed lower body weights throughout the lactation period. No other treatment-related changes were found at any dietary level of total bromine in either parental animals (F0, F1) or their offspring (F1, F2) for any of the parameters studied (i.e. clinical signs, oestrous cycle, sperm count and morphology, mating, fertility, gestation, parturition, litter size, pup viability, organ weights, and gross or histopathological examination). It was concluded that residues of 200 and 500 ppm total bromine in diets fumigated with methyl bromide are the no-observed-effect level and the minimum toxic level, respectively, for both parental rats and their offspring, and that bromine residues do not affect reproductive performance at dietary levels of up to 500 ppm.

Effects of the Ay gene on susceptibility to hydrocortisone fetotoxicity and teratogenicity in mice.

Effects of the Ay gene, a coat color gene, on susceptibility to hydrocortisone fetotoxicity and teratogenicity were investigated by using the congenic strain of C57BL/6-Ay (Ay/a) which had been maintained by repeated back-crosses of the Ay gene to the C57BL/6 (a/a) background. Matings were conducted as follows (female x male): group I, a/a; group II, a/a x Ay/a; and group III, Ay/a x a/a. Pregnant females were subcutaneously given daily doses of 0, 12.5, 25, or 50 mg/kg of hydrocortisone on days 10-13 of pregnancy. On day 18 of pregnancy, fetuses were sexed, weighed, and examined for external abnormalities. In group I, the mean fetal weight was significantly decreased at a dose of 25 mg/kg or more. The incidences of cleft palate were 3.2 and 22.7% at 25 and 50 mg/kg, respectively. In group II, in which half of the fetuses were expected to carry the Ay gene, the mean fetal weight was decreased significantly at 12.5 mg/kg or more. The incidence of cleft palate in group II at 50 mg/kg was 44.2%, which was significantly higher than that in group I. In group III, in which maternal mice as well as half of their fetuses carried the Ay gene, a decrease in the mean fetal weight was greater than in group II. In addition, the mean percentage of fetal resorptions was significantly increased at 50 mg/kg. The incidence of cleft palate in group III was significantly increased at 25 mg/kg (10.5%) when compared with those in groups I and II. These results indicate that the Ay gene may be associated with susceptibility to hydrocortisone fetotoxicity and teratogenicity in mice.