Teratogenicity of vitamin B6 deficiency: omphalocele, skeletal and neural defects, and splenic hypoplasia.

Vitamin B(6) deficiency was induced in pregnant rats with a deficient diet and with 4-deoxypyridoxine, a B(6) antagonist. Treated animals developed typical skin changes of B(6) deficiency. Fetuses were small and appeared anemic. Major fetal malformations were omphalocele, exencephaly, cleft palate, micrognathia, digital defects, and splenic hypoplasia. This teratologic system was developed as a model for human syndromes that exhibit combined immunologic and neurologic or skeletal defects.

The ontogenesis of human fetal hormones. I. Growth hormone and insulin.

The content and concentration of immunoreactive growth hormone (GH) were measured in 117 human fetal pituitary glands from 68 days of gestation to term and in the pituitary glands of 20 children 1 month to 9 yr of age. Physicochemical and immunochemical properties of GH of fetal pituitary glands and GH from adult pituitary glands were indistinguishable by disc gel electrophoresis, immunoelectrophoresis, starch gel electrophoresis, and radioimmunoassay techniques. In the fetal pituitary gland, the GH content rose from mean levels of 0.44+/-0.2 mug at 10-14 wk of gestation, to 9.21+/-2.31 mug at 15-19 wk, to 59.38+/-11.08 mug at 20-24 wk, to 225.93+/-40.49 mug at 25-29 wk, to 577.67+/-90 mug at 30-34 wk, and to 675.17+/-112.33 mug at 35-40 wk. There was a significant positive correlation between growth hormone content of the pituitary and gestational age, crown-rump length, and the weight of the pituitary gland. The content and concentration (micrograms/milligram) of human growth hormone (HGH) in the fetal pituitary showed significant increments (P < 0.001) for each 4 wk period of gestation until 35 wk. Further increases in the HGH content were noted in pituitaries of children aged 1-9 yr (range of 832 to 11.211 mug). Immunoreactive GH was detected in fetal serum at a concentration of 14.5 ng/ml as early as 70 days gestation, the youngest fetus assayed. At 10-14 wk, the mean concentration of serum growth hormone was 65.2+/-7.6 ng/ml; at 15-19 wk 114.9+/-12.5 ng/ml; at 20-24 wk 119.3+/-19.8 ng/ml; at 25-29 wk 72.0+/-11.5 ng/ml; and 33.5+/-4.2 ng/ml at term. A significant negative correlation of serum growth hormone with advancing gestational age after 20-24 wk was observed (P < 0.001). In 17 fetuses paired serum and pituitary samples were assayed; no significant correlation between the concentration of serum GH and the pituitary content or concentration of GH was demonstrable. The serum concentration of chorionic somatomammotropin (HCS) in the fetus was unrelated to gestational age. Insulin (1-30 muU/ml) was detected in 42 of 46 fetal sera assayed. These data suggest that the appearance and development of the secretory capacity for GH by the human fetal pituitary gland coincides with developmental changes in the portal system and hypothalamus. Maturation of inhibitory central nervous system control mechanisms for secretion of GH may not occur until infancy.

Glucose absorption and utilization by rat embryos.

There is little doubt that glucose plays a significant nutritional role in early somite embryos. The high glucose utilization of anaerobic glycolysis drops as the activity of the Kreb's cycle and terminal electron transport increase. Concurrently, maturation of mitochondrial cristae and dependence on oxygen supply are taking place. The neuroepithelium of the early somite rat embryo responds in vitro during culture by microvilliar lengthening when exposed to glucose levels of 50 mg/dl or more. At lower glucose concentrations both in whole embryo culture and inside the closed neural tube the microvilli are shorter. Lengthening of the microvilli at room temperature is produced only by d-glucose and 2-deoxyglucose, two hexoses that are absorbed and phosphorylated. Cytochalasin D which disrupts actin polymerization causes ballooning of the microvilli. A role of this microvillar elongation in degenerative changes seen in uncontrolled diabetes and on function of the immune system is proposed. The amniotic cavity is one major portal of entry for glucose during the early somite embryo stage. The 7-fold increase in volume of the amniotic cavity after day 10 allows the rat embryo to convert its axis from dorsal to ventral flexion.

Fetal iron balance in the rat.

Maternal and fetal iron balance through pregnancy was examined in the rat. The 20th day was selected for detailed study because of the peak iron requirements at that time. The standard diet provided a borderline iron supply to the fetus due to the limited availability of its iron for absorption. When a more available form of iron was used, normal fetal development occurred over a range of dietary iron content from 16 to 2500 mg/kg. At a level of 5 to 8 mg/kg, there was attrition of placental tissues with frequent fetal death and resorption. When the iron-deficient pregnancy was sustained, both maternal and fetal iron deficiency were present. At progressively higher levels of dietary iron, feto-placental iron content was constant despite a progressive increase in maternal iron stores. Fetal iron supply appeared to be determined primarily by plasma iron concentration, and, at normal levels, about 25% of transferrin iron passing through the uterine vasculature, was removed by the intact placentas. Low levels of plasma iron resulted in damage to fetal tissues and reduced the capacity of placental tissues to take up iron. At high levels of plasma iron, plasma iron turnover initially increased 5-fold over basal levels in nonpregnant animals due to increased placental uptake. However, with continued hyperferremia, uptake was regulated so as to maintain fetal iron at a normal level. A comparison of these data with human iron requirements explained the occurrence of both maternal and fetal iron deficiency in the rat, but only maternal iron deficiency in the human.

Methods for detecting teratogenic agents in man.

At a multidiscipline international meeting sponsored by L'Institut de la Vie held at Guadeloupe in January 1974, current methods for detecting teratogenic agents were outlined and discussed. Recommendations of the participants of the conference were: recognize the limitations of the present defenses against teratogenic agents; educate the public and medical profession about the known human teratogenic agents; select for animal teratogenicity screening among new and existing agents by emphasizing substances to which the entire population will be exposed, agents to which pregnant women are exposed, viruses which are found to persist in the human fetus, and agents which have become suspect from clinical experience; recognize that nearly all compounds have a fetotoxic dose but that this does not imply teratogenicity; encourage the development of new, quick in vitro testing methods for detecting teratogenic agents; monitor for sudden increases in the frequency of specific malformations in newborn infants and in aborted fetuses; assure that expert multidiscipline committees are available to evaluate the threat when suspected teratogens are reported; improve teratology information storage and retrieval systems by record linkage of clinical data, linkage between computer systems, and universal identifier system for chemical compounds and congenical malformations; foster the exchange of data, particularly those held by the pharmaceutical industry.

Twin fetuses with abnormalities that overlap with three midline malformation complexes.

Twin fetuses aborted at an estimated gestational age of 145 days were concordant for oral, facial, skeletal, and central nervous system malformations. The twins were discordant for other anomalies including cardiac defects, polydactyly, and malrotated short bowel. The combination of malformations observed overlaps with that of the oral-facial-digital syndrome, hydrolethalus syndrome, and Pallister-Hall syndrome. The problem of phenotypic overlap between these syndromes is discussed.

Fetal renal malformation following treatment of Hodgkin's disease during pregnancy.

A case of human fetal renal maldevelopment following the administration of nitrogen mustard, vincristine, procarbazine, and prednisone in early pregnancy for therapy of Hodgkin's disease is reported. While these agents have been shown to be teratogenic in animal experiments, adverse fetal effects following their use during early human pregnancy have not been described. In spite of this previous experience, it has generally been recommended that these agents be withheld during the first trimester of pregnancy because of their presumed teratogenic potential. This case would seem to reinforce this recommendation.

Size of the fetal adrenal in bilateral renal agenesis.

Bilateral renal agenesis is a fetal malformation incompatible with extrauterine life. Accurate prenatal diagnosis is essential for patient counseling. False-negative diagnoses have been reported and were attributed to the sonographic misidentification of apparently hypertrophied fetal adrenal glands as fetal kidneys. To study the relationship between renal agenesis and adrenal size, we reviewed autopsy records from 11 affected fetuses that had undergone careful autopsy and organ weight determination in our laboratory. Anomalies of distant structures were present in five affected fetuses. A sonographic diagnosis of adrenal hypertrophy had been made in two cases. In four of 11 fetuses, the glands had taken on a flattened discoid appearance. The autopsy records of 240 normal fetuses were similarly reviewed, and regression lines were generated for adrenal weight based on foot length and crown-rump length. The adrenal weights from affected fetuses were well within normal limits when compared with these normal regression lines and with organ weight standards from the literature. We conclude that adrenal hypertrophy is not a common finding in this syndrome and that the reported false-negative diagnoses are more likely attributable to a change in adrenal shape rather than a true increase in adrenal mass.

Potential human teratogenicity of frequently prescribed drugs.

Published data regarding the human teratogenic potential of 157 drug components that are frequently prescribed to outpatients in the United States were evaluated according to a protocol developed for TERIS, an automated clinical teratology resource. This protocol stipulates that a bibliographic search be performed on each agent, a brief narrative summary of the available teratologic information prepared, and a risk rating assigned. The ratings are determined by consensus of five clinical teratologists, who independently assess the magnitude of teratogenic risk associated with each agent under usual therapeutic conditions as "none," "minimal," "small," "moderate," "high," and "undetermined." Forty-nine percent of the components of these frequently prescribed drugs had insufficient published information available to assess the risk of human teratogenicity. Of the agents that could be rated, the teratogenic risk in usual therapeutic doses was considered to be minimal or less in 92.5%. Many of these agents have also been assigned Pregnancy Categories by the United States Food and Drug Administration (FDA) according to a system designed to provide therapeutic guidance. There was no more agreement than that expected by chance between TERIS ratings and the FDA Pregnancy Categories for 83 agents that were classified according to both systems. We believe that the FDA Pregnancy Categories should not be used to provide counseling regarding the risk of teratogenic effects to women who have taken medication during pregnancy. Such counseling should be based on a more comprehensive evaluation of the teratologic literature and clinical situation, but need not involve consideration of the therapeutic benefit of the agent.

The teratogenicity of N(omega)-nitro-L-ariginine methyl ester (L-NAME), a nitric oxide synthase inhibitor, in rats.

Exposure of gravid rats to the nitric oxide synthase inhibitor N(omega)-nitro-L-arginine methyl ester (L-NAME) in drinking water or by implanted osmotic minipumps significantly elevates maternal blood pressure, reducing uteroplacental perfusion. Administration by either route causes fetal growth retardation, but oral exposure also causes hind limb reduction malformations. The present study employed both oral and intraperitoneal routes to determine the period of sensitivity to developmental toxicity, dose-response, and possible fetotoxic mechanisms. Hind limb hemorrhage occurred only in litters from dams exposed to oral doses of 1 to 2 mg/mL from gestational days 15 through 17. In contrast to oral exposure, single intraperitoneal injections caused both fore and hind limb reductions at doses of 25 mg/kg and above administered on gestational day 16 and later. Many other exposures that reduce uteroplacental perfusion have been associated with vascular disruptive dysmorphogenesis. These exposures include phenytoin, calcium channel inhibitors, cocaine, and uterine vascular clamping. Limb hemorrhage induced by these exposures is usually limited to distal structures, typically phalanges, and the incidence of affected fetuses rarely exceeds 50%. By contrast, hemorrhage caused by L-NAME frequently involves entire limbs, extending into adjacent flank in severe instances, and 100% of fetuses from treated dams may be affected. The basis of this difference and the differing defect patterns associated with the various routes of exposure are unclear, but the generation of reactive oxygen species during resumption of normal perfusion may play a role in this vascular disruption.

Assessment of reproductive disorders and birth defects in communities near hazardous chemical sites. I. Birth defects and developmental disorders.

Members of the workgroup on birth defects and developmental disorders discussed methods to assess structural anomalies, genetic changes and mutations, fetal and infant mortality, functional deficits, and impaired fetal and neonatal growth. Tier 1 assessments for all five adverse reproductive outcomes consist of questionnaires and reviews of medical records rather than laboratory testing of biologic specimens. The work-group members noted a role for neurodevelopmental testing and for limited genetic studies, such as karyotyping in Tier 2 assessments. Emerging methodologies to identify chromosomal aberrations, DNA adducts, and repair inhibition were reserved for Tier 3.

Uterine contraction in the development of Möbius syndrome.

We report a patient with Möbius syndrome. The birth defect in this child is probably the result of both vasoconstriction and uterine contractions brought on by ergotamine taken during the sixth week of pregnancy. We propose that vasoconstrictive or mechanical effects, or both, of abortifacient drugs such as ergotamine and misoprostol may account for other cases of Möbius syndrome, and we suggest that uterine contraction from any cause, at about the sixth or seventh week of pregnancy, may cause this birth defect. Further observational studies are needed to verify this hypothesis.

Growth parameters in mid-trimester fetal Turner syndrome.

Growth failure is a consistent finding at birth in infants with Turner syndrome. However, the time of onset and pattern of growth deficiency is unknown. To determine the presence of growth failure in the second trimester in fetuses with Turner syndrome, second trimester fetuses that had a complete autopsy at the Central Laboratory for Human Embryology at the University of Washington were studied. A control group of specimens with normal findings was selected and compared with a study group with Turner syndrome documented by karyotype. Footlength and crown-rump length were measured directly with a ruler and femur, tibia, fibula, humerus, ulna and radius were measured from X-rays. Crown-rump length was used as the indicator of gestational age. Statistical comparisons between the normal and study groups were performed by multiple regression. Long bone measurements were made on 105 normal and 13 Turner fetuses. Footlength and the six long bones showed evidence of statistically significant growth failure. Fetuses with 45,X/46,XX mosaic Turner syndrome may demonstrate a lesser degree of growth retardation, at least for footlength, than those with a 45,X karyotype, but small numbers limited the analysis. We conclude that the growth failure consistently demonstrated in newborns with Turner syndrome begins early in gestation and is well-established by mid-pregnancy.

Brain growth in Down syndrome subjects 15 to 22 weeks of gestational age and birth to 60 months.

We have found similarities of skull shape, brain growth and brain maturation in 17 DS and 10 non-DS (control) fetuses, ages 15-22 weeks of gestational age (Group A), and differences in 101 DS and 80 non-DS cases, from birth to 60 months (Group B). Postnatally, the gross neuropathological differences between DS and control brains are more distinct after 3-5 months of age. The anterior posterior diameter fronto-occipital length of the brain hemispheres is shortened and that is secondary to reduction of frontal lobe growth. Also flattening of occipital poles, narrowing of the superior temporal gyruses and generalized retardation of brain growth were common findings. Standard morphometric methods indicate changes from birth [Wisniewski et al. 1984, 1986, 1990]. The cerebral cortex of the DS cases had a 20-50% reduction of neurons since birth, mainly in the granular layers [Wisniewski et al. 1984, 1986, 1990]. Changes in brain weight with age were greater in the non-DS than in the DS cases, and greater in males than in females. CHD and GI malformations were associated with less brain weight in both DS and non-DS cases. We suggest that the prenatal retardation of neurogenesis begins after 22 weeks' gestational age. The postnatal retardation of brain growth is secondary to pre- and postnatal abnormalities in synaptogenesis.

Embryonic and early fetal loss.

Embryonic and early fetal loss occurs in approximately one out of two pregnancies. Although this rate might appear unreasonably high, it is borne out by several types of studies and further confirmed by careful animal study. It now appears that karyotype deviation is common in early abortuses and probably underlies the high rate of gross abnormality seen in this group. In addition, environmental factors (radiation, smoking, anesthetics, and previous therapeutic abortion) may contribute to this wastage. Study and analysis of the abortus have allowed identification of a number of new embryonic and fetal syndromes. Careful description and classification combined with continued monitoring of abortuses shows promise both as an adjunct to reproductive counseling and as an important defense against teratogenic exposure. It is through this study that the etiology and ultimately the control of preventable pregnancy wastage will be understood and implemented.

Low-birth-weight and congenital rubella syndrome: effect of gestational age at time of maternal rubella infection.

Birth weights of 42 full-term patients with congenital rubella syndrome were analyzed. All of these infants were products of pregnancies in which the exact dates of the maternal first day of last menstrual period and of the time of onset of the mothers' rubella rash were known. The range of time of maternal rubella associated with low-birth-weight was in the gestational age interval from 16 to 100 days. Low-birth-weight may have a relationship with time of maternal rubella rather than with the type of defects, i.e., cataract, heart disease, and deafness.