Bone age and growth in fetal alcohol syndrome.

We have found delayed mean bone age in 63 children with fetal alcohol syndrome (FAS). The mean bone age Z-score for boys (n = 31) was -2.12 SDs and for girls (n = 32) was -1.62 SDs. This might suggest that they have potential for catch-up growth. However, experience with children with intrauterine growth retardation suggests that this will not be the case and that FAS children will be of reduced height at maturity. Further support for this assumption was gained from a sample of 26 patients who were followed until at least the age of 14 years for females and 16 years for males. There was no significant change in height Z-scores from early childhood to early adulthood, the mean score being -2.16 SDs and -2.11 SDs at mean ages of 4.83 years and 18.69 years, respectively. On the other hand, there were significant changes in weight and head circumference. The mean weight Z-score changed from -2.10 SDs to -1.14 SDs (p < 0.001). The head circumference mean Z-score in 16 patients was -3.13 SDs at a mean age of 2.79 years and -2.63 SDs at a mean age of 17.37 years (p = 0.013). Short stature can continue to be used as a diagnostic criterion for FAS beyond childhood.

Effects of preconceptional and gestational exposure to Tordon 202c on fetal growth and development in CD-1 mice.

Female CD-1 mice were exposed to Tordon 202c (a picloram and 2,4-D combination herbicide) in the drinking water at concentrations of 0.21, 0.42, and 0.84% for 60 days prior to mating with untreated males. One-half of the pregnant females subsequently continued treatment throughout gestation while the remaining females were maintained on distilled water. Fetal weight, crown-rump length, placental weight, and maternal gestational weight gain were reduced in a dose-dependent manner following combined preconceptional and gestational exposure. The incidence of malformed fetuses (cleft palate, renal agenesis, hydronephrosis, unilateral testicular agenesis, and umbilical hernia) and fetuses with variants (especially incomplete ossification of the skeleton) were increased in a dose-dependent manner following combined exposure. Increased maternal mortality and decreased preconception weight gain were observed in the highest-dosage group. Relative maternal liver weight was increased in a dose-dependent manner. The results suggest that combined preconceptional and gestational exposure to Tordon 202c is required for teratogenesis and fetal growth depression. Preconceptional exposure alone is not effective in increasing the risk for embryotoxicity.

Effects of paternal subacute exposure to Tordon 202c on fetal growth and development in CD-1 mice.

Male CD-1 mice were exposed to Tordon 202c (a picloram and 2,4-D combination herbicide) in the drinking water at concentrations of 0.21, 0.42, and 0.84% solutions for 60 days prior to mating with untreated females. Subsequently there was no exposure to Tordon 202c during gestation. Fetal weight and crown-rump length were reduced in the highest dosage group. The incidence of malformed fetuses (e.g., ablepharon, cleft palate, and unilateral agenesis of the testes) was increased in the middle dosage group while the incidence of fetuses with variants was increased in the lowest (e.g., an extra pair of ribs) and the highest dosage groups (e.g., incomplete ossification of the skeleton). The frequency of pregnancy failure was increased in the middle dosage group. Indices of paternal toxicity included increased lethality and decreased water consumption in the highest dosage group and increased relative spleen weights in the lowest and middle dosage groups. The results suggest paternally mediated reproductive toxicity.

Effects of gestational exposure to Tordon 202c on fetal growth and development in CD-1 mice.

The teratogenic effects of Tordon 202c, a picloram and 2,4-D combination formulation, are unknown. Pregnant CD-1 mice were exposed to Tordon 202c in the drinking water at concentrations of 0.10, 0.21, and 0.42% from d 6 to 15 of gestation. Fetal growth parameters, including body weight and crown-rump length, were reduced in a dose-dependent manner, as was placental weight. The incidence of dead fetuses/resorptions and malformed fetuses (especially cleft palate) was increased in the highest dosage group. A subtle indication of maternal toxicity was noted in the highest dosage group as evidenced by decreased water consumption and increased relative liver weight. The present study suggests that Tordon 202c is embryotoxic and teratogenic in CD-1 mice when administered during organogenesis.

The effect of prenatal exposure to the n-butylester of 2,4-dichlorophenoxyacetic acid (2,4-D) on the immune response in mice.

Pregnant CD-1 mice were administered the n-butylester of 2,4-dichlorophenoxyacetic acid (2,4-D) by gastric intubation on day 11 of gestation at dosages ranging from 0 to 200 mg/kg (2,4-D content). The immune response in the female offspring was elevated at 6 weeks of age. The humoral immune response, antibody production against sheep red blood cells, was not altered by 2,4-D ester exposure during gestation. The mitogen responses of lymphocytes induced by concanavalin A, a T-lymphocyte mitogen, or by Escherichia coli lipopolysaccharide, a B-lymphocyte mitogen, were reduced in the highest exposure group (200 mg/kg), although the T-lymphocyte suppression was not statistically significant. A similar response pattern was observed in the background nonstimulated lymphocyte cultures, suggesting that the suppression was a generalized lymphocyte abnormality. Evaluation of the mitogen responses using stimulation indices to correct for the variable background responses demonstrated that 2,4-D produced no net suppressive effect in any of the treatment groups. Since in utero 2,4-D ester exposure produced no alterations in humoral immunity and only subtle effects on lymphocyte blastogenesis, it is unlikely to be of any immunotoxicological or immunoteratological significance. Further studies investigating commercial-grade 2,4-D formulations are necessary since these formulations contain other components that may potentially induce alterations in the immune system.

Effects of prenatal alcohol exposure on neural cells in mice.

Pregnant DBA/1J mice were treated orally with 1.0 or 2.0 g/kg ethanol from days 11-17 of gestation to determine whether ethanol can perturb normal brain development. On gestational day 18 the fetuses were removed and fetal growth parameters were determined. The cerebrums from one group of fetuses were subsequently analyzed for cell number and protein content. The remaining cerebrums were assayed for their ability to grow in an in vitro cell culture system. Prenatal ethanol exposure decreased fetal body and brain weights and crown-rump length. The brain was particularly affected as indicated by a decreased brain: body wt ratio. The percentage of affected and marginally affected fetuses increased in a dose-dependent manner. While the number of cells/brain was unaffected, the number of cells/g cerebrum and the number of cells/mg cerebral protein was increased. Prenatal ethanol exposure decreased the ability of cerebral cells to grow in culture as demonstrated by the reduced plating efficiency and reduced colony size. The data from the present study suggest that ethanol induces a two-fold effect on mouse brain development. First, since the total number of cells/brain was not appreciably affected by prenatal ethanol treatment, it is possible that the reduction in brain size is due to a decreased amount of neuropil. This putative effect on the neuropil was manifested in vitro by decreased colony area. Second, the decreased plating efficiency of cells from brains of affected fetuses suggests that these cells are not functionally normal. These effects may be important in the pathogenesis of central nervous system anomalies associated with the Fetal Alcohol Syndrome.

Determination of the proximate teratogen of the mouse fetal alcohol syndrome. 2. Pharmacokinetics of the placental transfer of ethanol and acetaldehyde.

CD-1 mice were treated ip on Day 10 of gestation with 4, 6, or 7 g/kg ethanol. Maternal and embryonic tissues were analyzed for ethanol and acetaldehyde levels by head-space gas chromatography 5 min to 24 hr after treatment. Dose-dependent ethanol concentrations were observed in maternal blood and liver. Ethanol rapidly crossed the placenta and appeared in the embryo 5 min after treatment. Acetaldehyde was detectable in maternal blood following all treatments and in maternal liver and embryos following treatment with 7 g/kg ethanol. Coadministration of 100 mg/kg 4-methylpyrazole, an alcohol dehydrogenase inhibitor, with 4 or 6 g/kg ethanol on Day 10 of gestation significantly reduced the rate of ethanol elimination in all tissues examined. This reduction was manifested as a prolongation in the half-life of ethanol detectable in maternal and embryonic tissues but not in an increase in maximum ethanol concentrations. Within 5 min of maternal ip treatment with 200 mg/kg acetaldehyde on Day 10 of gestation, acetaldehyde was detectable in the embryo. These data suggest that both ethanol and acetaldehyde are accessible to the embryo during a critical period of development.

Determination of the proximate teratogen of the mouse fetal alcohol syndrome. 1. Teratogenicity of ethanol and acetaldehyde.

The proximate teratogen of the fetal alcohol syndrome is unknown. CD-1 mice were treated ip on Day 10 of gestation with 2, 4, 6, or 7 g/kg ethanol. The percentage of resorptions and malformed fetuses was increased and mean fetal weight was decreased in a dose-related manner. Treatment with 7 g/kg ethanol ip on one of gestational Days 7, 8, 9, 10, or 11 significantly increased the percentage of malformed fetuses and decreased fetal weight. In addition, treatment on Days 10 or 11 significantly increased the percentage of resorptions. Coadministration of 100 mg/kg of 4-methylpyrazole, an inhibitor of alcohol dehydrogenase, orally with 6 g/kg ethanol ip on Day 10 of gestation dramatically increased the embryotoxicity of ethanol. Five ip treatments of 200 mg/kg acetaldehyde at 2-hr intervals on Day 10 of gestation did not significantly increase the percentage of resorptions and malformed fetuses or decrease fetal weight. These data suggest that ethanol is the proximate teratogen of the fetal alcohol syndrome in CD-1 mice.