bluuebunny/crossref_metadata_2025 huggingface dataset:
Low birthweight in rats induced by prenatal exposure to testosterone combined with alcohol, pair‐feeding, or stress
Authors: McGivern Robert F.
Published: June 2005

Pregnant Sprague‐Dawley dams were exposed to a liquid ethanol diet (35% ethanol‐derived calories), an isocaloric pair‐feeding regimen, restraint stress, or no treatment during the last week of pregnancy. Dams in each group received injections of testosterone propionate (TP) or the oil vehicle from days 15 through 20 of gestation. Birthweights of pups from dams administered TP and also exposed to alcohol, pair‐feeding, or restraint stress were significantly depressed by as much as 40 percent compared to oil‐injected counterparts. Prenatal exposure to alcohol, pair‐feeding, or restraint stress in the absence of TP did not significantly depress birthweight, nor was birthweight depressed in animals from dams injected with TP but exposed to no other treatment. Results are discussed with respect to an inhibition of fetal growth produced by a possible synergism between activation of the hypothalmic‐pituitary‐adrenal axis and elevated androgen levels.
Digital defects induced by vasodilating agents: Relationship to reduction in uteroplacental blood flow
Authors: Danielsson Bengt R. G. , Reiland Sven , Rundqvist Eva , Danielson Margaretha
Published: June 2005

The effects of nifedipine (40–100 μmol/kg), nitrendipine (40 and 80 μmol/kg), hydralazine (381 and 763 μmol/kg), felodipine (12 μmol/kg), and the pharmacologically inactive first‐step metabolite of felodipine, H152/37 (80 μmol/kg) were studied in rabbits (New Zeeland White) after oral administration on day 16 of gestation. The vasodilating drugs—nifedipine, nitrendipine, felodipine, and hydralazine—all induced digital defects in the fetuses. The defects consisted of a reduction, absence, or abnormal structure of the distal phalanx of especially the fourth digit on the hind paw(s). Histologically, a disturbed differentiation of the cartilage, and secondarily also of the ossification centre and joint structure of the distal phalanx, was observed. In contrast, no digital abnormalities were observed after administration of vehicle or H152/37. The findings that vasodilators with different structures, like dihydropyridines and hydralazine, induced the same type of digital defects strongly suggest that the observed phalangeal defects are secondary to pharmacological action, and not related to chemical structure. A decrease in uteroplacental blood flow, caused by excessive hypotension, is discussed as the most probable mechanism underlying the observed defects.
Prenatal ossification as an indicator of exposure to toxic agents
Authors: Beck Sidney L.
Published: June 2005

Following exposure to bromodeoxyuridine (BUDR), acetazolamide (ACZM), trypan blue (TRBL), cortisone (CORT), or diphenylhydantoin (DPH), alizarin‐stained, cleared fetuses were examined at 18 days postcoitus for unossified cervical vertebral central; number of ossified caudal vertebrae; number of ribs; and ossification of sternebrae, metatarsals, metacarpals, and phalangeal rows. At all teratogenic doses, in no vehicle‐treated groups, and rarely in lower‐dose groups, there were significant increases in frequency of unossified cervical centra, the first vertera (C1) being most often affected, and C7 least often affected. In the high‐dose CORT group, there was a significant correlation between unossified C1 and cleft palate. No association between abnormality and reduced ossification of cervical vertebrae was seen in other series examined, nor was there any correlation between litter size and abnormality. With minor complications, the number of ossified caudal vertebrae was significantly reduced after exposure at teratogenic dose levels to all compounds except DPH. Although caudal and cervical ossification were correlated with each other in those series examined, neither was correlated with abnormality. Frequency of 14 ribs was increased in BUDR, ACZM, and TRBL but not CORT or DPH. Other parameters were essentially unaffected. Significantly increased frequency of abnormality, when contrasted with untreated or vehicle‐treated groups, was seen at high‐dose levels in all but DPH treatments, and mortality was increased in ACZM D9‐11, TRBL, and CORT. These studies show that reduced ossification of cervical centra is an excellent indicator of prenatal exposure to noxious substnces, and caudal vertebrae appear to be useful as well. Increased frequency of 14 ribs occurred for all strong teratogens utilized if they were administered on day 7 or day 8 postcoitus.
Effects of L‐asparaginase on rat embryonic development and yolk sac membrane in vitro
Authors: Sanfeliu C. , Nebot‐Cegarra J. , Calvet S. , Domenech‐Mateu J. M.
Published: June 2005

A comparative analysis of the teratogenic effects of L‐asparaginase on 10.5‐day rat embryos after 24 and 48 hours of exposure in vitro, respectively, were performed. Several medium concentrations of L‐asparaginase (0.05, 0.25, and 1.5 IU/ml) were tested in both embryo series. Resulting embryos were submitted to morphological studies in a search for a specific route of pathogenesis. Morphological alterations of the visceral yolk sac were also studied to investigate its contribution to L‐asparaginase teratogenicity in rats. Main embryonic malformations (open truncal neural tube, open encephalic vesicles, anophthalmia, lack of inversion, abnormal frontolateral protrusions, great vascular dilations at the cephalic level) and developmental retardation were already generated after the first 24 hours of culture (embryos of 10.5 days) and presented a dose‐response related to an early mesenchyme deficiency. Reduced number of mesenchymal cells was more evident in embryos of 10.5 days than those of 11.5 days, suggesting the existence of a later compensatory mechanism of cellular proliferation in the older embryo. Visceral yolk‐sac endodermal cells at both embryonic stages were greatly deformed and enlarged by an incerase of the high electron‐dense vacuolar system. Therefore, both a blockage of the processes of lysosomal digestion and derived trophic deficiencies probably existed. A double teratogenic mechanism for L‐asparaginase is postulated: a direct action mainly in younger embryos (before invagination of the embryo into the yolk sac) and a yolk sac‐mediated one.
Teratogenic activity of trichloroacetic acid in the rat
Authors: Smith M. K. , Randall J. L. , Read E. J. , Stober J. A.
Published: June 2005

Trichloroacetic acid (TCA) is a by‐product of the chlorine disinfection of water containing natural organic material. It is detectable in finished drinking water at levels comparable to the trihalomethanes (30–160 μg/L). TCA is also formed in vivo after ingestion of hypochlorite and has been identified as a major metabolite of chlorinated hydrocarbons such as trichloroethylene. The developmental effects of TCA were evaluated in the pregnant Long‐Evans rat. Animals were dosed by oral intubation on gestation days 6–15 (plug = 0) with 0, 330, 800, 1,200 or 1,800 mg/kg/day. The vehicle control was distilled water. Maternal observations included clinical signs, weight change, and gross evaluation of organ weights and uterine contents at necropsy (day 20). Live fetuses were examined for external, skeletal, and soft tissue malformations. There were no maternal deaths associated with toxicity prior to sacrifice. Weight gain during treatment was reduced at 800, 1,200, and 1,800 mg/kg. Spleen and kidney weights were increased in a dose‐related manner. The mean percent of resorbed implants per litter was 34, 62, and 90 at 800, 1,200, and 1,800 mg/kg, respectively. Live fetuses showed dose‐dependent reductions in weight and length. The mean frequency of soft tissue malformations ranged from 9% at the low dose to 97% at the high dose. These were principally in the cardiovascular system (interventricular septal defect, levocardia). Skeletal malformations were found only at 1,200 and 1,800 mg/kg and were mainly in the orbit. Based on these observations TCA was considered to be developmentally toxic in the pregnant rat at doses of 330 mg/kg and above.
Paternal alcohol consumption: Effects of age of testing and duration of paternal drinking in mice
Authors: Abel Ernest L.
Published: June 2005

Male mice consumed liquid alcohol diets containing 25, 20, 15, 10, 5, or 0% ethanol‐derived calories (EDC). Animals receiving the 20‐0% EDC diets were pair‐fed to those consuming the 25% EDC diet. After 7 weeks of consumption males were bred to nontreated females. Offspring were tested for activity at 16–20 and 75 days of age. Offspring sired by alcohol‐consuming males did not differ from controls in litter size, birth weight, or weight at weaning, but were less active than controls on several measures of activity. Many of these decreases were best defined in terms of linear trends. However, these differences were evident only for animals tested prior to 20 days of age for most activity measures. In a second experiment adult males continued to consume alcohol for another 7 weeks and were bred again. Offspring of this second breeding were tested for activity at 16 days of age and were compared with offspring sired by the same father from the previous breeding. Offspring sired after this longer duration of paternal alcohol consumption did not differ significantly from controls in any of the above‐mentioned variables.
Mitosis in embryonic trisomy 1 mouse eye
Authors: Smith B. S.
Published: June 2005

Trisomy 1 embryos consistently show eye defects (e.g., aphakia, microphakia, retention of lens stalk). To determine if the plane of division of mitotic figures is abnormal in the eyes of these animals, trisomic embryos (9.5 through 12 gestational days) were produced from mice doubly heterzygous for Robertsonian translocation chromosomes [Rb(1.3)/Rb(1.10)]. To accommodate for the known delay in trisomic embryo development, animals were grouped according to stages of eye development rather than to gestational age. Serial sections were evaluated without knowledge of karyotype for orientation of mitotic figures (parallel, perpendicular, oblique) in lens, optic cup, and diencephalon. Location of mitotic figures was scored as apical (nearest the lumen), middle, and basal. Numerous anomalies were noted in trisomic eye development. No difference was found between orientation of mitotic figures in the lens and optic cup of trisomy 1 and normal embyros. Location of mitotic figures in trisomy 1 lens was significantly different from that of normal littermates. The data confirm earlier studies that trisomy 1 affects the eye, and they tend to corroborate evidence that this trisomy affects the lens more than it affects the optic cup.
Case report of a boy with penoscrotal transposition, double urethra, pseudodiphallia, and ectopic scrotum
Authors: Larsen Jørn
Published: June 2005

The paper describes a 14 year old Saudi boy with penoscrotal transposition and double urethra placed side by side in the penis. Ectopic scrotal skin and a pseudopenis were seen over the right ischial tuberosity. The patient had a large lumbosacral lipoma, which probably caused paralysis and atrophy of the right leg through an intraspinal extension.
Ethanol treatment induces a delayed segmentation anomaly in the chick embryo
Authors: Sanders E. J. , Cheung E.
Published: June 2005

A repeatable somite anomaly is described that results from the incubation of cultured chick embryos in the presence of ethanol. The anomaly comprises a misalignment of approximately five consecutive pairs of somites such that one of each pair is displaced cranially by up to one‐half a somite length. The appearance of the malformation is delayed by approximately six somite pairs after the beginning of treatment. These characteristics were shared by embryos treated at the stage of gastrulation (no somites yet present) up to embryos possessing ten pairs of somites at treatment time. The deleterious effect did not appear to result from a disruption in the mechanics of the segmentation process itself, since isolated segmental plates were able to form normal intersomitic clefts in the presence of ethanol. Similarly, there were apparently no alterations in the compaction process that occurs at the cranial end of the segmental plate, since both the contractile and adhesive components were unaffected, as judged by the distributions of actin and fibronectin. The potential mechanisms of the anomaly are discussed with reference to similar segmental defects produced by heat shock. In view of earlier results indicating that cells in the primitive streak at gastrulation are sensitive to the presence of ethanol, it is proposed that this somite anomaly is due to a disruption in the contribution of these mesoderm cells to the segmental plate.
Umbilical cord growth in human and rat fetuses: Evidence against the “stretch hypothesis”
Authors: Fujinaga Masahiko , Chinn Anne , Shepard Thomas H.
Published: June 2005

A total of 103 human fetuses between the 7th and 30th week of gestation were obtained from induced abortion (40 fetuses were normal and 63 were abnormal), and the umbilical cord length (UCL) was measured. The UCL increased almost linearly with gestational age among normal fetuses, contrary to the commonly held tenet that the UCL increases exponentially during the second trimester. When UCLs from the 63 abnormal fetuses were compared with those of normal fetuses, 15 fetuses were found to have short UCL and 10 fetuses to have long UCL. Among the 15 fetuses with short UCL, 6 had early amnion rupture syndrome. An unexpected finding among the 10 with long UCL was that 8 of them had oligohydramnios. It has been suggested that the UCL increases in response to tensile forces placed upon it (“stretch hypothesis”); however, our results are inconsistent with this hypothesis because fetuses with oligohydramnios should be less active and their umbilical cords be subject to less stress. In a separate experiment, we studied the normal development of the UCL in rat fetuses and observed an almost linear increase during the whole gestation similar to that seen in humans. This finding is also inconsistent with the “stretch hypothesis” because amniotic fluid volume decreases significantly from day 19 of gestation to term in rats.
Studies of the effect of retinoic acid on anterior neural tube closure in mice genetically liable to exencephaly
Authors: Tom C. , Juriloff D. M. , Harris M. J.
Published: June 2005

Previously we have shown that all SELH/Bc mouse embryos close their anterior neural tubes by an abnormal mechanism and that 10–20% of SELH/Bc embryos are exencephalic. The purposes of these studies were (1) to observe the effects of retinoic acid on the frequency of exencephaly in SELH/Bc embryos; (2) to compare the SELH/Bc response with those of normal strains and of other neural tube mutants; and (3) to compare, between SELH/Bc and a normal strain (SWV/Bc), the effects of retinoic acid on morphology of the closing anterior neural tube.

SELH/Bc was more liable to retinoic acid‐induced exencephaly than were normal strains. After maternal treatment with 5 mg/kg retinoic acid on day 8.5 of gestation, 53% of SELH/Bc embryos had exencephaly, compared with 22% in ICR/Bc and 14% in SWV/Bc. When these results were transformed according to the assumptions of the developmental threshold model, the effects of genotype and retinoic acid appeared to be additive.

Similar treatment on day 9 or 10 of gestation had little or no effect on the frequency of exencephaly in SELH/Bc mice. These results are similar to the reported responses of the curly‐tail and Splotch mutants, where frequencies of spina bifida but not exencephaly were decreased. This pattern suggests that studies of effects of periconceptional vitamin treatment on risk of human neural tube defects should consider anencephaly and spina bifida separately.

The study comparing the morphology of anterior neural tube closure in SELH/Bc and normal SWV/Bc embryos showed that retinoic acid delays the elevation of the mesencephalic neural folds. This results in a “stalling” of many embryos in the first steps of neural tube closure, with their neural folds remaining convex and splayed wide apart. The delay in fold elevation was superimposed on the different closure patterns of the two strains. The overall conclusion is that there is no nonadditive interaction in the parameters studied between retinoic acid treatment and the SELH/Bc genotype.
Fetal coronary thrombosis as a cause of single ventricular heart
Authors: Shepard Thomas H. , Fantel Alan G. , Kapur Raj P.
Published: June 2005

A fetus weighing 947 g was autopsied after prenatal echocardiographic diagnosis of a single ventricular heart. At autopsy a single ventricle was present with a pear‐shaped scar in the area presumed to represent the right ventricle. A small stoma from the single ventricle connected to the scar and the pulmonary outflow was obstructed. The left coronary artery was occluded and on histologic examination recanalization was seen. A chromatographic peak which closely eluted with the cocaine metabolite, benzoylecognine, was identified in fetal urine. We postulate that coronary spasm following possible cocaine exposure could have produced an infarct which destroyed the right ventricle. It is possible that this fetal pathology may be one mechanism that leads to single ventricle hearts.
Abnormal development of cardiovascular systems in rat embryos treated with bisdiamine
Authors: Tasaka Hiroyasu , Takenaka Hitoshi , Okamoto Naomasa , Onitsuka Toshio , Koga Yasunori , Hamada Minoru
Published: June 2005

Administration of N,N′‐bis(dichloroacetyl)‐1,8‐octamethylene‐diamine, bisdiamine, in pregnant Donryu rats on day 10 of gestation induces a high incidence of cardiovascular anomalies in fetuses. Bisdiamine administration induced aplasia of the sixth aortic arch artery, with both the right and left primitive pulmonary arteries being directly linked to the truncus, and resulting in four types of malformation of pulmonary arteries (PAs). When two primitive PAs shared a single root, the consequence was either pulmonary trunk hypoplasia, as is seen in tetralogy of Fallot, or type I persistent truncus arteriosus (PTA) as classified by Collet and Edwards. When root portions of two PAs did not fuse, either type II or type III PTA resulted. In controls, the right dorsal aorta (DA) between the right seventh intersegmental artery (IA) and the site where both DAs fuse degenerated and the left aortic arch (AA) and the right subclavian artery (SA) were formed. Bisdiamine administration induced two additional types of vascular anomalies. In one of these, the right DA between the right 4AA and the right 7IA degenerated and a left AA accompanied by an aberrant right SA resulted. In the other type, the left DA between the left 4AA and the left 7IA degenerated and a right AA accompanied by an aberrant left SA resulted. These results indicate that administration of bisdiamine induces malformation in the great blood vessels by disturbing persistency and degeneration of aortic arch arteries and DAs.
Effects of retinoic acid on chick tail bud development
Authors: Griffith C. May , Wiley Michael J.
Published: June 2005

The present study describes the teratogenic effects of retinoic acid (RA) on the development of the chick tail bud. Chick embryos were recovered 48 hours after treatment at HH stages 11 to 16 with various dosages of RA by subblastodermal injection. At the gross level, RA treatment resulted in varying degrees of caudal regression, scoliosis, limb malformations, and open posterior neuropores among the survivors. Histological examination of tail buds from treated embryos revealed defects which included total dysplasia of caudal structures, the presence of accessory neural tube and notochord tissue, and abnormal fusions of the notochord to the neural tube and tailgut. The incidence, severity, and location of the defects were dependent on the dose of the teratogen, and the stage of development at the time of treatment. The defects resembled those induced in previous studies by treatment with sialic acid binding lectins such as wheat germ agglutinin and limulus polyphemus lectin (Griffith and Wiley, '90b).
Comparative studies of embryotoxic action of ethylenethiourea in rat whole embryo and embryonic cell culture
Authors: Tsuchiya Toshie , Takahashi Atsushi , Asada Soichiro , Takakubo Fumie , Ohsumi‐Yamashita Noriko , Eto Kazuhiro
Published: June 2005

The effects of ethylenethiourea (ETU) were investigated using rat (Wistar‐imamichi) embryos cultured from days 11 to 13 of gestation or cultured rat embryonic cells extracted on day 11. Malformations in cultured embryos at the concentration of 30 μg/ml of ETU were found in the head and tail, which were severely affected, as well as the limb and face. All embryos exposed to 150 and 300 μg/ml of ETU had malformed heads, tails, limbs, and facial configurations. Protein contents of the cultured embryos were decreased dose‐dependently at the concentrations ranging from 30 to 300 μg/ml. In the histological studies of the cultured embryos with ETU, thinner neuroepithelium in head was observed. In the embryonic cells extracted on day 11 of gestation, ETU dose‐dependently inhibited the differentiation of midbrain (MB) cells into neurons and that of limb bud (LB) cells into chondrocytes at the concentrations ranging from 30 to 600 μg/ml of ETU. The concentrations of ETU that inhibited the production of differentiated foci by 50% (IC50) were 170 μg/ml in LB cells of day 11, and >600 μg/ml in LB cells on day 12 of development. Therefore, differentiation of MB cells was more sensitive to ETU than the differentiation of LB cells. These results indicated that there was a reasonable correlation of ETU induced changes in cultured whole embryos and embryonic cells.
Altered visceral yolk sac function produced by a low‐molecular‐weight somatomedin inhibitor
Authors: Hunter E. S. , Phillips L. S. , Goldstein S. , Sadler T. W.
Published: June 2005

A fraction from diabetic rat serum containing a low‐molecular‐weight (800–1000) somatomedin inhibitor (SI) alters growth and development in both neurulation and early limb bud staged mouse embryos in vitro. Previous studies suggested that an accumulation of serum proteins and morphological changes of the visceral yolk sac (VYS) were produced following exposure to the SI in early limb bud staged conceptuses. The morphological changes, characterized by the presence of large endosomes in the endodermal cells, suggested that the SI altered histiotrophic nutrition, whereby proteins are pinocytosed by the endodermal VYS cells and degraded to constituent amino acids. Therefore, the effects of the SI on pinocytosis and protein degradation by the VYS were evaluated using the whole embryo culture system. Results showed that the SI reduced fluid phase pinocytosis as determined by the uptake of [U‐14C]sucrose, but that accumulation of [3H]leucine‐labeled hemoglobin ([3H]Hb) by the VYS was greater following exposure to the SI than in controls. In contrast, the accumulation of 3H‐labeled amino acids in the embryo (produced from the degradation of [3H]Hb by the VYS) was reduced by the SI. The extent of amino acid reduction in embryonic accumulation is dependent upon the concentration of SI in the culture medium and correlates with the incidence of malformations produced by the SI, i.e., high rates of malformations occur with large reductions in embryonic 3H‐labeled amino acid accumulation. The apparent paradox of high [3H]Hb accumulation in the presence of decreased pinocytosis appears to be the result of altered processing of the [3H]Hb in the endodermal cells. The altered processing decreases the “elimination” of the proteins from the VYS and results in the decrease in 3H‐labeled amino acid present in the embryo proper. Therefore, the SI appears to alter two processes of VYS histiotrophic function: (1) decreased pinocytosis and (2) altered protein processing, ultimately resulting in a decreased availability of substrates for the embryo. During the early stages of embryogenesis in the human, the trophoblast cells of the placenta are responsible for the transport of nutrients from the maternal to embryonic systems. Since these cells show high phagocytic and pinocytotic activities, the SI may also disrupt these processes in the chorioallantoic placenta and contribute to diabetes‐induced embryopathies.
Maternal exposure to chemical and physical factors during pregnancy and cardiovascular malformations in the offspring
Authors: Tikkanen Jorma , Heinonen Olli P.
Published: June 2005

The possible effect of chemical and physical factors during pregnancy on the occurrence of cardiovascular malformations in the offspring was studied in 573 cases and 1,055 controls. The cases represented all verified cardiovascular malformations in Finland during 1982–1984. The controls were randomly selected from all babies born during the same period. Case and control mothers were interviewed by midwives approximately 3 months after delivery using a structured questionnaire. Maternal alcohol consumption during the first trimester of pregnancy was more common among the mothers of case infants (45.9%) than those of controls (39.6%). Exposure to organic solvents at work was slightly more prevalent among the ventricular septal defect group (12.1%) than the control mothers (7.8%). However, neither association was significant when adjusted for maternal age in logistic regression analysis. Moreover, one or both of these associations may be chance effects resulting from multiple comparisons.

The risk of cardiovascular malformations was not associated with maternal smoking, or coffee, tea, or cola consumption, and was equal in urban and rural areas. Maternal exposures to anesthetic gases, pesticides, wood preservatives, microwave ovens, and video display terminals at work or home were not associated with the risk of cardiovascular malformations.

It is concluded that some common environmental exposures during early pregnancy to physical and chemical factors should not necessarily be considered hazardous for the developing fetal heart. The causes of the majority of cardiovascular malformations remain unknown.
Abnormalities of neural tube formation in pre‐spina bifida splotch‐delayed mouse embryos
Authors: Yang Xiu‐Ming , Trasler Daphne G.
Published: June 2005

The splotch‐delayed homozygous mutant (Spd/Spd) develops spina bifida with or without exencephaly, has spinal ganglia abnormalities, and delays in posterior neuropore closure and neural crest cell emigration. The heterozygote (Spd/ +) has a pigmentation defect, and occasionally neural tube defects. To investigate the underlying mechanisms, we compared the neuroepithelium in the posterior neuropore region of cytogenetically identified 15–18 somite pair Spd/Spd, Spd/ +, and +/+ mouse embryos by transmission electron and light microscopy. The notochordal area and cell number in the non‐fused neuroepithelium region of Spd/Spd and Spd/ + embryos were significantly reduced compared to those of normal (+/+) embryos, which suggests an abnormality in notochord elongation. In the mesoderm, the mean cell number and mean ratio of cell number to area in the non‐fused region were significantly lower in the Spd/Spd compared with +/+ embryos. The distance of exposed neuroepithelium above the mesoderm in the just‐fused region was significantly lower in the Spd/Spd versus +/+ embryos, which may indicate an insufficient force exerted by the mesoderm during neural tube closure. Within the neuroepithelium, significantly more intercellular space was found in Spd/Spd than in +/+ embryos indicating disorganization. The basal lamina was poorly organized and the formation delayed around the neural tube in Spd/Spd and Spd/+ embryos. All together, these results suggest an early abnormality in interactions among the neuroepithelium, mesoderm, and notochord, which may lead to the delay or inhibition of neural tube closure observed in Spd/Spd mutants.
Normal mouse strains differ in the site of initiation of closure of the cranial neural tube
Authors: Juriloff D. M. , Harris M. J. , Tom C. , MacDonald K. B.
Published: June 2005

The scanning electron microscopic study of day 9 embryos reported here documents differences among normal mouse strains in morphology of cranial neural tube closure. The site of initiation of contact and fusion of the cranial neural folds, previously defined as Closure 2 (Macdonald et al., '89), is located in the region of the junction between the forebrain (prosencephalon) and midbrain (mesencephalon) in three normal strains: LM/Bc, AEJ/RkBc, and ICR/Bc. However in a fourth normal strain, SWV/Bc, Closure 2 is initiated much further rostral, in the prosencephalic region. In addition, the anterior neuropore, rostral to Closure 2, closes late in ICR/Bc embryos, relative to the posterior progress of development of the Closure 2 seam. Initiation of closure from the most rostral end of the neural tube (Closure 3) appears to be relatively delayed in ICR/Bc embryos. We hypothesize that the observed genetic polymorphism in location of the first site of fusion between the cranial neural folds in normal mouse embryos may be one basis for differences among normal strains in liability to exencephaly induced by teratogens.
Complexities of limb anomalies: The lower extremity in the “prune belly” phenotype
Authors: Genest David R. , Driscoll Shirley G. , Bieber Frederick R.
Published: June 2005

The so‐called “prune belly” syndrome (PBS) consists of megalocystis, cryptorchidism, and marked abdominal distension; in addition to these findings, many other abnormalities are commonly present, rarely including severe anomalies of the leg. We report two such individuals, in whom PBS coexisted with abnormal development of the lower extremity. The first, a stillborn male fetus, was delivered at 21 weeks gestation; generalized hypoplasia of both legs was present, more marked distally than proximally, and more severe on the left. The second case, a liveborn male infant, was the second of dichorionic twins, delivered at 33 weeks gestation; this infant died at two hours from respiratory insufficiency due to pulmonary hypoplasia. There was severe hypoplasia of the right leg, with gangrenous necrosis of all tissues distal to the knee. Additional findings included a single right umbilical artery, and a small congenital cystic adenomatoid malformation of the right lung. The findings in these cases are compared to other similar cases in the literature, and possible mechanisms for the etiology and pathogenesis of maldevelopment of the leg in PBS are discussed.
Results of prenatal alcohol exposure on the dimensions and binucleation of cardiac myocytes in neonatal and weanling rats
Authors: Alvarez Marvin R. , Clark Linda C. , Moore Jo Ann , Morales Martha C. , Gerdes A. Martin
Published: June 2005

Sprague–Dawley rats were exposed to ethyl alcohol in utero. The effect of chronic prenatal exposure was examined by giving mature females alcohol in isocaloric liquid diets which served as the sole source of liquid and caloric intake before mating and throughout gestation. Controls consisted of females maintained on laboratory chow or an isocaloric liquid diet minus alcohol before and during gestation. The offspring were sacrificed at 21 days of age (weanlings) and the hearts dissociated enzymatically to give purified cardiac myocytes. The effects of daily acute prenatal alcohol exposure were studied by gastric intubation of alcohol to chow‐fed females for the duration of pregnancy. The doses used approximated 4 and 5 shots of 80 proof liquor per day by a person weighing 150 lb. These offspring were sacrificed at 2, 6, and 21 days postnatal and cardiac myocytes prepared as above. Heart weights were determined and cardiac myocytes were analyzed for cell length, volume, cross‐sectional area, and percent binucleation. Additionally, nuclear DNA content was measured in all of the 21 day offspring. Statistical analysis of the data showed no significant differences between hearts exposed to prenatal alcohol and nonexposed controls with either regimen with the exception of percent binucleation which was significantly but only slightly higher in the 6‐day‐old hearts. These findings are discussed in relation to anatomical heart defects found in patients with full fetal alcohol syndrome.
Changes in the fetal tibial growth plate secondary to maternal zinc deficiency in the rat: A histological and histochemical study
Authors: da Cunha Ferreira R. M. C. , Gonzalez J. I. Rodriguez , Marquiegui I. Monreal , Elizaga I. Villa
Published: June 2005

Zinc deficiency (ZD) is teratogenic in rats, and fetal skeletal defects are prominent. To elucidate further the effects of maternal ZD in the fetal skeleton, we performed a morphological and histochemical study of tibial growth plate (GP) in ZD rat fetuses. The histochemical study included the identification of calcium, of hydrolytic enzymes associated with the process of calcification, and of oxidative enzymes related to energy production and to the synthesis of proteoglycans. Pregnant Sprague‐Dawley rats were fed (1) a control diet (76.4 μg Zn/g diet) ad libitum (group C), (2) a zinc‐deficient diet (0 μg/g) ad libitum (group ZD), or (3) the control diet pair‐fed to the ZD rats (group PF). On day 21 of gestation, laparotomies were performed, the fetuses were removed, and fetal tibiae obtained. Specimens were stained with hematoxylin—eosin (H&E) and Masson Trichrome and were processed for identification of alkaline phosphatase, adenosine triphosphatase, succinic dehydrogenase, NADH dehydrogenase, and calcium. The morphologic patterns found in ZD fetal tibiae indicated defects in various cell types implicated in bone metabolism. Staining for hydrolytic enzymes revealed alterations in the size and distribution of matrix vesicles and a weaker staining for ATPase in ZD fetuses. Staining for oxidative enzymes was overall more intense in ZD fetal tibiae. ZD fetuses also presented irregular and defective calcification. These findings indicate that severe maternal ZD in the rat results in structural and functional alterations in the GP of fetal bone, leading to a defective endochondral ossification.
Craniofacial abnormalities in mice with X‐Linked hypophosphatemic genes (Hyp or Gy)
Authors: Shetty Navin S. , Meyer Ralph A.
Published: June 2005

X‐Linked hypophosphatemia is the most common cause of metabolic rickets in humans and is characterized by a reduced renal TmP/GFR and hypophosphatemia. Clinically, these changes are associated with growth retardation including attenuated craniofacial growth, femoral and tibial bowing, and radiologic and histomorphometric evidence of rickets and osteomalacia. Similar mutations occur in mice at the Hyp and Gy gene loci. Direct craniometric measurements were made on mouse skulls to investigate the pattern of craniofacial growth differences in the Hyp/ +, Hyp/Hyp and Gy/ + genotypes and to compare these to littermate normals in the C57BL/6J mouse strain. There was generalized attenuation in craniofacial growth in all mutants. The heterozygous Hyp and Gy mutants showed similar patterns of craniofacial growth with diminished neurocranial length, viscerocranial length, and mandibular height. The Gy/ + was significantly smaller than the Hyp/ + in neurocranial width. The homozygous Hyp mouse was not affected more severely than the heterozygous Hyp except in overall cranial length, nasal bone length, and mandibular length from mandibular foramen to third molar. In summary, the heterozygous Hyp and Gy mutant mice showed similar patterns of craniofacial growth. The homozygous Hyp mouse was not affected more severely than the heterozygous Hyp except in three of the 15 measured variables. Thus, these data demonstrate the almost complete dominance of the Hyp gene. In contrast, the Gy gene is incompletely dominant. The heterozygous Gy females survive, but the hemizygous Gy males do not, on a C57BL/6J background. This suggests that there is a family of closely linked genes on the X chromosome which, while similar in their effects on phosphate homeostasis, have differing mechanisms of action.
ACE inhibitor fetopathy and hypocalvaria: The kidney–skull connection
Authors: Barr Mason , Cohen M. Michael
Published: June 2005

Two cases of angiotensin converting enzyme inhibitor fetopathy with renal tubular dysgenesis and severely underdeveloped calvarial bone are described. Six additional, unexposed cases of hypocalvaria are described, and possible links between calvarial development and fetal hypotension and/or chronic hypoxia are explored.
Phenytoin teratogenicity and midgestational pharmacokinetics in mice
Authors: Roberts Linda G. , Laborde James B. , Slikker William
Published: June 2005

Mice of the A/J and C57BL/6J (C57) strains were dosed with phenytoin (PHT) every 48 hr throughout pregnancy by gastric intubation to test the hypothesis that maternal plasma PHT concentration may be the significant factor in determining PHT reproductive and developmental toxicity. Serial serum samples were obtained from each mouse from gestation day (GD) 10‐GD 12 for determination of individual dam PHT pharmacokinetics. Maximum PHT concentration and PHT AUC (area under‐the‐time—concentration curve) were regressed to laparotomy and fetal evaluation endpoints to determine whether significant association existed. Although serum PHT concentrations exceeded levels associated with teratogenicity (>10 μg/ml), few major malformations were induced in either strain. However, in the A/J strain, there was a significant increased incidence of hydrocephaly and open eyelid. Regression of pharmacokinetic parameters with embryo and maternal endpoints indicated significant associations between gestational weight gain and maximum concentration measured (Cmax) or AUC in both strains. This association was also found for fetal weight in the C57 strain. In the A/J strain, the induction of decreased ossification of the sternebrae was also associated with maternal PHT concentration; however, linear regression of hydrocephaly and open eyelid to PHT concentration was not statistically significant. These results suggest that maternal plasma PHT concentration may be a quantifiable determinant of certain aspects of PHT developmental toxicity in the mouse.
Additive incidence of developmental malformation for Xenopus embryos exposed to a mixture of ten aliphatic carboxylic acids
Authors: Dawson Douglas A.
Published: June 2005

A modified FETAX (Frog Embryo Teratogenesis Assay: Xenopus) protocol was used to assess the joint action of ten aliphatic carboxylic acids on Xenopus embryo development. Stock solutions of each acid alone, made up at twice the EC50 of the individual acids, were prepared for testing alone and in a mixture with an equal volume of each acid stock solution. For each treatment, five concentrations and a control dish, each with 25 embryos, were tested for 96 h, with solution renewal every 24 h. The embryos were then fixed and evaluated for gross malformations. For each dish, the number and types of malformations were recorded. An EC50 was calculated for each acid alone and this value was defined as 1.0 toxic unit (TU) for malformation induced by the acid. An EC50 was also calculated for the mixture. The concentration of each acid at the mixture EC50 and the TU values corresponding to these concentrations were then determined. A TU value of 0.990 (0.923–1.060) was obtained for the mixture by adding the TU values for each acid in the mixture. This represents a concentration additive rate of malformation. Microcephaly, TU = 1.09 (1.01–1.18), was the primary malformation, but did not completely account for the response. The concentration additive rate of malformation indicates that all ten acids are likely to induce malformation in Xenopus embryos in a similar manner. Quantitative structure‐activity relationship (QSAR) analysis revealed developmental malformation induced by the acids was highly correlated (r2 = 0.979) with hydrophobicity and molar refractivity (r2 = 0.949). The approach has potential application in determining compounds that induce developmental malformations in a similar manner, when metabolism and pharmacokinetic factors are considered.
A computer‐assisted image processing method for determining relative cardiac function in the chick embryo
Authors: Bruyere Harold J. , Xu Bin , James Thomas L. , Magee Michael J.
Published: June 2005

The general objective of this study was to develop a noninva‐sive method for efficiently and reproducibly determining relative cardiac function parameters in the chick embryo. The specific objectives of the study were 1) to develop several methods for computer‐assisted image processing and quan‐titation of relative intraventricular blood volumes in the 3‐day‐old embryonic chick heart and 2) to compare methods for precision and with a previously established manual processing method. Images of the embryonic chick heart in ovo were recorded on videocassette tape, digitized, and enhanced by computer‐aided histogram equalization. The area occupied by blood within the common ventricle was extracted by region‐growing and spurious region removal algorithms and defined by the determination of edge‐pixel coordinates. Edge‐pixel coordinates of the longitudinal and transverse axes of the common ventricular blood region were located by three different methods, the lengths of the axes calculated, and volumes computed from the equation for determining volume of a prolate spheroid. Twenty‐five images of the embryonic heart were randomly selected and processed. Volumes were calculated with each of the three methods on six different occasions. A coefficient of variation was determined for each image by each of the three methods. A mean coefficient of variation was calculated for each method. The intraobserver mean coefficient of variation for each method was 7.4%. When a 2‐way ANOVA was conducted, mean coefficients of variation did not differ significantly for the three methods. However, computer processing (in addition to significantly reducing the time required to generate data) reduced the coefficient of variation observed in manual processing by 56.5%.
Assessing developmental hazard: The reliability of the A/D ratio
Authors: Setzer R. Woodrow , Rogers John M.
Published: June 2005

The quantitative relationship between developmentally toxic exposure levels and adult toxic exposure levels has been used as an index of developmental hazard and has figured prominently in discussions of legal regulation of developmentally toxic agents. Perhaps the most frequently cited index of developmental hazard is the A/D ratio. This index, a ratio of marginally toxic adult and developmental dose levels (e. g., NOAELs or LOAELs), is attractive because it is easily calculated from published toxicity assays and because it has been argued that A/D is relatively constant across species for a given agent. We explored some quantitative aspects of the A/D ratio and of the concept of developmental hazard by simulating 661,500 mammalian developmental toxicity assays on 441 hypothetical compounds. In our simulations, A/D often varied substantially among replicate assays: the median ratio of the upper and lower limits of the distribution of A/D values that include about 95% of the observed A/D values is 16. In addition, A/D did a poor job of predicting the relative developmental and adult responses at dosages lower than those used to calculate the index: among simulated compounds with A/Ds of about 1.0, the developmental response at 1/100th of the NOAEL ranged from about 0.1% to 20,000% of the adult response. Finally, we measured the concordance between pairs of four different indices of developmental hazard, including A/D. Concordance was greatest when the indices were based on the same portion of the dose response, and was much weaker between indices that examined different portions of the dose response. Therefore, it seems likely that no single index can quantify “developmental hazard,” as defined by relative adult and developmental susceptibility, and more effort needs to be expended in refining the concept if it is to be useful for hazard assessment.
A teratologic evaluation of continuous‐wave, daily ultrasound exposure in unanesthetized pregnant rats
Authors: Vorhees Charles V. , Acuff‐Smith Karen D. , Weisenburger Walter P. , Meyer Richard A. , Smith Nadine B. , O'Brien William D.
Published: June 2005

Pregnant Sprague‐Dawley rats were trained to remain immobile when placed in water in an ultrasound exposure tank and exposed to 0, 0.1, 2.0, or 30.0 W/cm2 ISPTA (spatial peak, temporal average), 3.0‐MHz continuous wave (cw) ultrasound on embryonic (E) days 4–19 for approximately 15 min/ day. On E20 fetuses were removed; weighed; examined for external, skeletal, and visceral malformations; and uteri were examined for resorptions. Analyses revealed no increase in pre‐implantation loss and no effects on maternal body weight, food, or water consumption. No increase in skeletal or visceral malformations was found, in fact exposed groups had a lower incidence of defects than controls. A significant increase in resorptions in the lowest exposure group (0.1 W/cm2) was obtained, but the effect was isolated, non‐dose dependent and not credible as a treatment‐related effect. No reduction in fetal weight was obtained, in fact the lowest (0.1‐W/cm2) and middle (2.0‐W/cm2) exposure level groups weighed slightly more than controls. The immobility procedure succeeded in avoiding anesthetization or forced restraint of the dams, thereby eliminating these factors as potential confounders. The results demonstrated that in unanesthetized, unrestrained rats in utero exposure to incident intensities of ultrasound of up to 30.0 W/cm2 cw ultrasound (or estimated internal exposures of 4–21 W/cm2, depending on body orientation to the incident beam) produced no evidence of embryotoxicity based on fetal necropsy data.
Genetically determined variation in the azygos vein in the mouse
Authors: Biddle Fred G. , Jung Jacob D. , Eales Brenda A.
Published: June 2005

The normal mouse is expected to have a single and left‐sided azygos vein that develops from the paired embryonic cardinal venous system and drains most of the right and left thoracic walls into the left anterior vena cava. During routine autopsies of adult mice, most individuals of the C57BL/6J strain were found to have this pattern but a distribution of different azygos venous patterns was found in the WB/ReJ strain. In WB/ReJ the patterns varied from a single unpaired vein on the right side that connected to the right anterior vena cava through bilaterally symmetrical and paired veins to the expected unpaired vein on the left side. A classification scheme for the observed patterns of azygos veins was developed and the frequency distributions of C57BL/6J and WB/ReJ mice in these classes were compared. The strain difference in the azygos venous system between C57BL/6J and WB/ReJ can be interpreted as a genetically determined threshold trait of development. Beginning with a paired and symmetrical cardinal venous system, the C57BL/6J genotype shifts to a left‐sided azygos pattern but the WB/ReJ genotype remains with a more bilateral azygos pattern. Genetic study of this azygos trait will be useful for the study of lateral asymmetries in mammalian development and for the interpretation of venous heterotaxies (anomalous placement of veins) in the mouse that are found in association with mutations such as situs inversus viscerum (iv) and dominant hemimelia (Dh).