bluuebunny/crossref_metadata_2025 huggingface dataset:
Vitrification of mouse oocytes results in aneuploid zygotes and malformed fetuses
Authors: Kola Ismail , Kirby Carol , Shaw Jillian , Davey Anna , Trounson Alan
Published: June 2005

Vitrification of mouse oocytes adversely affected the subsequent developmental potential of embryos and fetuses derived from the fertilization of such oocytes after thawing. Only 5% of oocytes vitrified formed viable fetuses on the 15th day of gestation as compared to 47% in the controls. The incidence of chromosomally aneuploid zygotes, derived from cryopreserved oocytes, was approximately threefold higher than the controls irrespective of whether the oocytes were cryopreserved by vitrification or DMSO slow‐freezing. Malformed fetuses were obtained from oocytes that had been vitrified as well as those that had been exposed to vitrification solutions only, whereas no malformed fetuses were obtained in oocytes slow‐frozen by DMSO or fresh controls–thus demonstrating that the exposure of oocytes to the vitrification chemicals was responsible for the fetal malformations. The data in this study suggest that the vitrification technique should be cautiously applied to human oocyte cryopreservation. Furthermore', the data also demonstrate that the exposure of female gametes to carcinogenic and/or teratogenic chemicals may result in malformed embryos when such oocytes are subsequently fertilized.
Histochemical, immunofluorescence, and ultrastructural differences in fetal cartilage among three genetically distinct chondrodystrophic mice
Authors: Seegmiller R. E. , Brown K. , Chandrasekhar S.
Published: June 2005

The severe lethal chondrodystrophies in man result in a common clinical syndrome including shortening of the face, mandible, and limbs. Studies of three lethal chondrodystrophic mutants in mice, viz., chondrodysplasia (cho), cartilage matrix deficiency (cmd), and disproportionate micromelia (Dmm), which share this syndrome, were performed with the aim of identifying histochemical, immunofluorescence, or ultrastructural differences which might exist among these hereditary cartilage disorders: We examined limb cartilage epiphyses from day 18 normal and mutant fetuses and observed repeatable, mostly qualitative differences. All observations were made relative to the normal control. Histochemical staining of matrix proteoglycan was moderately decreased in cho and Dmm cartilage and markedly decreased in cmd when compared to the normal control. Staining of matrix collagen was irregular in distribution in cho, increased in cmd, and decreased in Dmm. Immunofluorescence of proteoglycan was increased in the matrix of cho and Dmm and decreased in cmd. Immunofluorescence of type II collagen was heterogeneous and moderately decreased in the matrix of cho, increased in cmd, and markedly decreased in Dmm. Immunofluorescence of link protein in cho was localized in the cellular‐pericellular region as in the normal and appeared increased in the matrix of cmd and Dmm. Immunofluorescence of chondronectin was localized in the cellular‐pericellular region and appeared normal in all three mutants. Major differences in cellular and matrix ultrastructure were observed among the mutants, including a decreased frequency of small‐diameter collagen fibrils in cho and Dmm, increased density of collagen fibrils in cmd, and dilated RER in Dmm. These observations demonstrate that distinct structural and possibly molecular differences exist among the chondrodystrophies. In the case of cmd, the differences correlated with a previously reported molecular defect, viz., absence of core protein of cartilage specific proteoglycan in the cartilage of this mutant. It is anticipated that the methods used in the present study can be applied to humans in case classification and in identifying potential mouse‐human correlates.
Association of tracheoesophageal anomalies with visceral and parietal malformations in a human embryo (Carnegie stage 21)
Authors: Nebot‐Cegarra Josep , Domenech‐Mateu Josep M.
Published: June 2005

A human embryo (Carnegie stage 21) with tracheoesophageal malformations (esophageal atresia and tracheoesophageal fistula) and anomalies at the caudal end of the embryo (anorectal atresia, rectovesical fistula, vertebral and notochordal defects, and agenesis of the metanephroi) was studied. Other anomalies observed were: absence of right umbilical artery, fusion of spinal ganglia, and absence of cloacal outlet of mesonephric ducts. The possible pathogenesis of these associated malformations is discussed.
Embryotoxicity of phenyl ketone analogs of cyclophosphamide
Authors: Hales Barbara F. , Ludeman Susan M. , Boyd Victoria L.
Published: June 2005

Phenylketophosphamide and phenylketoisophosphamide are preactivated acyclic ketone analogs of cyclophosphamide and isophosphamide with antitumor activity. These compounds undergo an elimination reaction to yield phosphoramide or isophosphoramide mustard and phenyl vinyl ketone. In this study, the embryotoxicity of phenylketophosphamide, phenylketoisophosphamide, and phenyl vinyl ketone were determined. Embryotoxicity was assessed in vitro in whole rat embryos cultured on day 10.5 of gestation in the absence and presence of an activating system derived from maternal liver. Both phenylketophosphamide and phenylketoisophosphamide were embryotoxic in the absence of metabolic activation. Moreover, there was no enhancement of this embryotoxicity in the presence of an activating system. A 10‐μM concentration of phenylketophosphamide produced 100% malformed embryos, while this concentration of phenylketoisophosphamide was not teratogenic. At 25 μM phenylketoisophosphamide, all the surviving exposed embryos were malformed. Phenylketophosphamide was embryolethal to more than 50% of the exposed embryos at a concentration of 50 μM. In contrast, a concentration of phenylketoisophosphamide of 100 μM was required to produce significant embryolethality. Phenyl vinyl ketone was not embryotoxic at any of the concentrations tested. The major malformation observed, a hypoplastic prosencephalon, and the growth retardation effects were not only similar for phenylketophosphamide and phenylketoisophosphamide, but also similar to those previously reported for “activated” cyclophosphamide. Unlike the results with cyclophosphamide, where both phosphoramide mustard and the aldehydic metabolite of cyclophosphamide, acrolein, are toxic, the embryotoxic effects of pheynlektophosphamide and phenylketoisophosphamide are mediated only by the mustard metabolite.
Effects of human diabetic serum on the in vitro development of early somite rat embryos
Authors: Zusman Igor , Yaffe Perhija , Raz Itamar , Bar‐on Hanoch , Ornoy Asher
Published: June 2005

High levels of glucose, β‐hydroxybutyrate (B‐HOB), and acetoacetate are known to have embryotoxic and teratogenic effects on rat embryos in culture, especially when added concomitantly to the culture medium. We studied the effects of human serum from different types of diabetes mellitus on the in vitro development of 10½‐day‐old rat embryos cultured for 48 hours. We used serum from type I diabetes with and without ketoacidosis and type II diabetes either untreated or treated with insulin or with daonil. Type I diabetes without ketoacidosis increased the rate of malformations to 27% vs. 11% in controls. Serum from type I diabetes with ketoacidosis further increased the malformation rate to 44%. The rate of malformations induced by serum of type II diabetes was dependent on the treatment. It was relatively low among embryos cultured on serum from untreated (16%) or treated with daonil (19%) and rose to 27% among embryos cultured on serum from type II diabetes treated with insulin. No significant correlation was found between the rate of malformations and the concentrations of glucose, B‐HOB, acetoacetate, and HbA1c in all diabetic sera except serum from type I diabetes with ketoacidosis. We may therefore conclude that for most types of diabetes in humans, neither the high blood glucose concentrations nor the high levels of ketone bodies seem to be the main reason for the high rate of malformations. However, we used cultured rat embryos, and the effects on the human embryo may be different. The results of studies on various experimental animal models in diabetes teratogenicity seem to have only partial relevance to the human situation.
Dexamethasone does not interfere with hormone‐sensitive PI hydrolysis
Authors: London Fredda S. , Caamano‐Haigh Rosa , Chepenik Kenneth P.
Published: June 2005

Serum, but not epidermal growth factor (EGF), stimulated the release of radiolabeled inositol phosphates from human embryo palate mesenchyme (HEPM) cells prelabeled with [3H]‐myoinositol. Pretreatment of cells with 10−6 M dexamethasone (DEX) for 48 h had no effect on the release of inositol phosphates in response to serum. Furthermore, although treatment of the glucocorticoid‐sensitive A/J strain of mouse embryo palate mesenchyme (MEPM) cells with 10−6 M DEX inhibited their proliferation by 40%, it had no effect on the activity of phospholipase(s) C. However, DEX did enhance the incorporation of [3H]‐myoinositol into membrane lipids. We interpret these data to mean that 1) serum factors enhance metabolism of inositol lipids in HEPM cells, 2) DEX does not interfere with the primary events by which agonists utilize metabolism of inositol lipids as a mechanism for transmembrane signaling, and 3) DEX may affect synthesis of phosphoinositides, as reported by Grove et al. (Biochem. Biophys. Res. Commun. 110:200–207, 1983; J. Craniofac. Genet. Dev. Biol. Suppl. 2:285–292, 1986).
Histological study of cell death in digital malformations induced by 5‐azacytidine: Suppressive effect of caffeine
Authors: Kurishita Akihiro
Published: June 2005

The present study investigated microscopically the process of 5‐azacytidine (5‐AC)‐induced digital teratogenesis and caffeine's suppressive effect on this process. Three distinct zones of programmed cell death were observed in control and caffeine‐treated embryos 3 hours after 5‐AC injection: the preaxial and postaxial ectodermal regions and the central part of the mesodermal regions. 5‐AC temporarily suppressed programmed cell death in the ectoderm and mesoderm 3 hours after it was injected. However, caffeine promoted programmed cell death; normal programmed cell death was observed in the limb buds of embryos whose dams were treated with 5‐AC and caffeine. The percentage of total cell death in hindlimb buds of embryos treated with 5‐AC and caffeine was higher than that from embryos treated with 5‐AC, whereas 5‐AC‐induced digital malformations were reduced by post‐treatment with caffeine. Cell death reached a maximum 12 hours after the injection in limb buds from 5‐AC and caffeine‐treated embryos and at 24 hours in the 5‐AC treated embryos. Furthermore, in the 5‐AC and caffeine‐treated embryos, the frequency of cell deaths at 12 hours increased almost linearly with the doses of caffeine in parallel with the reduction of 5‐AC‐induced malformation frequency by caffeine. These results suggest that although induced cell death may be one of the factors leading to digital malformations produced by 5‐AC, it is not essential, and the existence of other factors affecting the pattern formation of the limb bud is proposed.
Exposure of Japanese quail embryos to o,p′‐DDT has long‐term effects on reproductive behaviors, hematology, and feather morphology
Authors: Bryan T. E. , Gildersleeve R. P. , Wiard R. P.
Published: June 2005

Japanese quail eggs were injected with 1‐(2‐chlorophenyl)‐1‐(4‐chlorophenyl)‐2,2,2‐trichloroethane p,p′‐DDT (1–10 mg), 1,1‐bis(4‐chlorophenyl)‐2,2,2‐trichloroethane p,p′‐DDT (1–10 mg), or, in one study, 0.5 mg chlordecone dissolved in 50 μl of corn oil on day 1 of incubation. Hatchability was not decreased by p,p′‐DDT or p,p′‐DDT, as compared to corn‐oil‐injected controls, but was reduced in progeny of parents injected in ovo with either isomer. Tremor was observed for up to 4 days posthatching only in birds injected with 1.75–10 mg p,p′‐DDT or chlordecone. Survivability to 5 weeks posthatch was reduced (± 50%) in birds injected in ovo with 6.25–7.5 mg, p,p′‐DDT or 1.75‐5 mg p,p′‐DDT as compared to corn oil (96%). Reproductive behaviors were attenuated in birds injected during development with p,p′‐DDT, both DDT isomers decreased the total number of ovipositions, and p,p′‐DDT increased the total number of eggshell malformations. Neither body weights nor reproductive organ weights at 12 weeks were affected by injection of either isomer. Exposure to DDT did not affect acquisition of a matched‐to‐sample food‐reinforced response or subsequent responding on a random interval schedule of reinforcement. In another experiment, total circulating erythrocyte numbers were reduced in females after injection in ovo with p,p′‐DDT but not after injection with p,p′‐DDT. A primary humoral immune response was not affected by in ovo exposure to either isomer of DDT. In ovo exposure to p,p′‐DDT but not to p,p′‐DDT had long‐term and estrogen‐like effects on behavior and hematology in Japanese quail. Posthatch primary feather morphology was also altered by embryonic exposure to o,p′‐DDT, p,p′‐DDT, and chlordecone.
Fetal pathology produced by ethylene oxide treatment of the murine zygote
Authors: Rutledge Joe C. , Generoso Walderico M.
Published: June 2005

Exposure of female mice to ethylene oxide by inhalation 1 or 6 h after mating produced not only multitemporal death of conceptuses but also high rates of abnormalities among surviving fetuses. In contrast, only marginal effects were observed when females were exposed 9 or 25 h after mating. The abnormalities found among 17 day gestation live fetuses were predominated by hydrops and eye defects, which, together, constitute 54% of all anomalies. Most of the remaining anomalies were distributed among 5 other types: small size, cleft palate, and cardiac, abdominal wall, or extremity and/or tail defects. In a follow‐up study, the fetuses of females treated 6 h postmating were examined at 11–15 days gestation and the progression of fetal death and of malformations was studied. Results indicate that the expression of most fetal anomalies does not become apparent until late in gestation. Several of these induced anomalies are similar to common human sporadic birth defects. This new class of experimentally induced fetal anomalies provides a new avenue for investigating zygotic biology and a system for studying the progression of aberrant development.
Concordance and discordance of congenital hydrocephalus in 107 twin pairs in Japan
Authors: Imaizumi Yoko
Published: June 2005

During the period from 1969 to 1985, 107 pairs of twins, of which at least one in each pair had congenital hydrocephalus, were ascertained in Japan. The rate of concordance for hydrocephalus among these twins was 15%, with a slightly higher rate in like‐sex male twins (28%) than in like‐sex female twins (14%). The rates were 21% in like‐sex and 0% in unlike‐sex pairs. Two twin pairs had one anencephalic twin and one hydrocephalic twin.
Comparative developmental toxicity of cationic and neutral rhodamines in mice
Authors: Hood Ronald D. , Jones Cathy L. , Ranganathan Sulabha
Published: June 2005

Rhodamines 123 and 6G (Rh 123 and Rh 6G) are cationic fluorescent dyes that inhibit oxidative phosphorylation following their selective accumulation within mitochondria. Neutral rhodamines (e.g., Rh 116 and Rh B) do not share these properties. To determine if cationic and neutral rhodamines differ in their effect on mammalian development, pregnant CD‐1 mice were injected i.p. with Rh 123, Rh B, or Rh 116 at doses of 15 mg/kg/day. The rhodamines were given alone or in combination with 500 mg/kg/day 2‐deoxy‐D‐glucose (2‐DOG), an inhibitor of glycolysis, daily on gestation days 7–10 (copulation plug = day 1). Additional pregnant mice were similarly treated with Rh 6G at a dose of 0.5 mg/kg/day. Controls were given saline equimolar to the dose of 2‐DOG. Treatment with Rh 6G, alone or in combination with 2‐DOG, significantly increased the incidences of prenatal mortality (17% and 35%, respectively) when compared with the control incidence (6%). Treatment with Rh 123 or Rh 6G, alone or with 2‐DOG, inhibited fetal growth. Treatment with the neutral rhodamines had little effect on prenatal survival or growth. Exposure to Rh 6G, with or without 2‐DOG, was associated with high incidences of gross malformations (41% and 61%, respectively). Rh 116 or Rh B, with or without 2‐DOG, and Rh 123 alone were not associated with statistically significant teratogenic effects, but results of the latter treatment were suggestive of such an effect (9.1% grossly malformed fetuses vs. 0% for controls). The incidences of skeletal malformations were significantly increased in the test groups given Rh 6G + 2‐DOG, Rh 123 + 2‐DOG, or Rh 6G alone. These results suggest a relationship between the charge on the rhodamine molecule and effects on the conceptus, and these effects may have been mediated at least in part by interference with mitochondrial metabolism.
Bendectin and human congenital malformations
Authors: Shiono Patricia H. , Klebanoff Mark A.
Published: June 2005

The relationship between Bendectin exposure during the first trimester of pregnancy and the occurrence of congenital malformations was prospectively studied in 31,564 newborns registered in the Northern California Kaiser Permanente Birth Defects Study. The odds ratio for any major malformation and Bendectin use was 1.0 (95% confidence interval 0.8–1.4). There were 58 categories of congenital malformations; three of them were statistically associated with Bendectin exposure (microcephaly—odds ratio = 5.3, 95% confidence interval = 1.8–15.6; congenital cataract—odds ratio = 5.3, 95% confidence interval = 1.2–24.3; lung malformations (ICD‐8 codes 484.4–484.8)—odds ratio = 4.6, 95% confidence interval = 1.9–10.9). This is exactly the number of associations that would be expected by chance. An independent study (the Collaborative Perinatal Project) was used to determine whether vomiting during pregnancy in the absence of Bendectin use was associated with these three malformations. Two of the three (microcephaly and cataract) had strong positive associations with vomiting in the absence of Bendectin use. We conclude that there is no increase in the overall rate of major malformations after exposure to Bendectin and that the three associations found between Bendectin and individual malformations are unlikely to be causal.
Proboscis lateralis: A case report
Authors: Antoniades Kostas , Baraister Michael
Published: June 2005

A lateral proboscis usually occurs in the region of the inner canthus. We present a case of holoprosencephaly accompanied by an obliquy facial cleft and an anterior encephalocele in which a proboscis lateralis occurred in a very lateral location.
Sex difference in digit and palate formation of mouse embryos at midgestation
Authors: Watanabe Toshiaki , Endo Akira
Published: June 2005

We investigated whether there is a difference in timing between the sexes of mouse embryos with regard to digit and palatal formation at midgestation by using a simple method of sex chromatin analysis for rodent embryos. At day 14.4 of gestation, although the mean body weight of male embryos was greater than that of female embryos, digit and palatal formation of female embryos was found to be more advanced than in males when compared according to their body weight. This was contrary to our previous finding of digit formation at the earlier stage (day 12.0 of gestation): namely, that digit development was more advanced in male embryos than in female embryos even when two sexes were compared according to their body weight. Thus, the development of female embryos catches up with that of male embryos at the later stages of midgestation. The period for digit development must be longer in male embryos than in female embryos. If mothers are exposed to some teratogens, a sex difference in incidence of digital defects might be produced.
Interpretation of some median anomalies as illustrated by cyclopia and symmelia
Authors: O'Rahilly Ronan , Müller Fabiola
Published: June 2005

Anomalies that involve the median plane are heterogeneous, and their embryological basis varies widely. Cyclopia and symmelia present a number of similarities: 1) They would appear to arise by neither fusion nor merging but mainly through a failure in lateralization. 2) Mesenchymal deficiency is important in both: possibly disturbance of the prechordal plate in cyclopia and failure of the caudal eminence in symmelia. The caudal eminence is an important developmental feature that is only recently becoming clearer in the human embryo. 3) Disturbance of axial material seems to be essential in both. 4) The results of experimental teratogenesis and an analysis of normal human development confirm that these conditions arise early. The teratogenetic termination‐periods in the human are probably 21/2 weeks for cyclopia sensu stricto (a median eye in a single orbit) and 3 weeks for cyclopia sensu lato, i.e., synophthalmia (paired ocular structures in a single orbit); 2 1/2 weeks for symmelia of the upper limbs (e.g., in cephalothoracopagus) and 3 1/2 weeks for symmelia of the lower limbs in a single individual. It is pointed out that in symmelia the limb buds, upper or lower, have failed to separate at their postaxial margins. This is in contrast to dimelia, in which the preaxial borders are missing and the postaxial margins are duplicated (postaxial dominance).
Amelioration of the teratogenicity of cadmium by the metallothionein induced by bismuth nitrate
Authors: Naruse Ichiro , Hayashi Yukimasa
Published: June 2005

The participation of maternal hepatic metallothionein (MT) in the amelioration of cadmium teratogenicity in mice was examined. Pretreatnient with Lisrriulh nitrate (subcutaneously) ameliorated the teratogenicity, including exencephaly and abnormalities of the axial skeleton, caused by a single intraperitoneal injection of cadmium sulfate. Pretreatment with bismuth nitrate for 3 days induced MT drastically in maternal liver and kidney. Six and 24 hr after the injection of cadmium sulfate, the accumulation of maternal hepatic cadmium increased and that in the decidua, including embryos, decreased after pretreatment with bismuth nitrate.

Mouse embryos on day 7 of gestation were cultured for 48 hr. Exposure to cadmium sulfate in vitro induced unfused brain fold, which corresponds to exencephaly in vivo. From the in vitro experiment, it was suggested that the teratogenicity of cadmium on day 7 of gestation is a direct action against the mouse embryo. In the present experiment it was suggested that pretreatment with bismuth nitrate induced maternal hepatic and renal MT; cadmium was therefore trapped and detoxicated, and consequently embryos were exposed to a lower concentration of cadmium.
Congenital heart disease among spontaneous abortuses and stillborn fetuses: Prevalence and associations
Authors: Chinn Anne , Fitzsimmons Jack , Shepard Thomas H. , Fantel Alan G.
Published: June 2005

The prevalence, range, and associations of congenital heart disease (CHD) were studied among 400 spontaneous abortuses between 9 and 40 weeks' gestation. Fifty‐two (13.0%) cases of CHD were detected. To minimize selection bias the specimens were grouped by external appearance and the prevalence expressed accordingly. CHD was detected in 21 (7.3%) of 289 externally normal and 31 (27.9%) of 111 externally abnormal fetuses. Ventricular septal defect (VSD) was the most frequent CHD found in isolation as well as in combination with extracardiac malformations. Seventy‐five percent of isolated CHD was VSD. Forty (69.2%) of the 52 cases of CHD were associated with extracardiac malformations. Chromosomal syndromes were responsible for a minimum of 19.2% of the cases and suspected in up to 36.5%. The most frequent associations involved the musculoskeletal system, central nervous system, abdominal wall, and kidneys. In contrast, studies of liveborn infants have reported 70% of CHD as isolated defects, including many CHD infrequently seen among spontaneous abortuses. This suggests that fetuses with isolated CHD often survive to term, and CHD does not significantly affect the survival of the fetus in utero. Ventricular septum formation may be particularly susceptible to hemodynamic changes and may be indicative of an underlying pathologic condition that also leads to a spontaneous abortion.
Effect of smokeless tobacco on the development of the CD‐1 mouse fetus
Authors: Paulson Ruth , Shanfeld Joseph , Sachs Larry , Price Theresa , Paulson John
Published: June 2005

The objective of this study was to examine the effect of an aqueous extract of smokeless tobacco (ST) on the development of the CD‐1 mouse fetus. Three ST dosages were administered three times daily by gastric intubation during gestational days 1–17: 1 × ST equivalent to a dose of 4 mg nicotine/kg body weight, 3 × ST equivalent to 12 mg nicotine, and 5 × ST equivalent to 20 mg nicotine/kg body weight. Maternal plasma nicotine levels were determined 30 minutes after the second daily intubation at five different times during the gestational period.

At these ST dosages, the weight gain of ST‐treated dams was not significantly affected in comparison to treated controls, though the difference was significant (P<.05) in comparison to untreated controls. The mean maternal plasma nicotine level for the low dosage (1 ×) group was 99.0 ng/ml, which reasonably approximates human consumption levels. The 3 × ST and 5 × ST dosages produced higher nicotine plasma values, 398 ng/ml and 623 ng/ml, respectively, were considerably more toxic to the dams, and resulted in 18% and 31% maternal deaths. Fetal weights were reduced by 7.4% (P<.001) in the highest ST dosage group (5 ×), whereas at the 1 × and 3 × dosages fetal weight differences were not significantly different from treated controls. Resorptions increased in a dose‐related manner (P<.05), ranging from 4.7% in the 1 ×, to 6.4% in the 3 × and 8.9% in the 5 × dosage compared to 3.2% in treated controls. External malformations were few and minor in extent. Internal malformations increased in a linear, dose‐related manner (P<.05). Placental weights were unaffected by ST.

The results of skeletal examinations were inconclusive. Precocious ossification was seen in 60% and 70% of the parameters measured in the 1 × and 3 × dosage groups, respectively, in comparison to controls. In the 5 × ST group ossification levels were less than in controls for 30% of the parameters measured.

Under these experimental conditions the lowest ST dosage (1 ×) produced a negligible effect on the CD‐1 mouse fetus and the dam. The highest ST dose (5 ×) demonstrated embryotoxicity, growth retardation, few malformations, and maternal toxicity. The intermediate dose (3 ×) showed a range of effects between the highest and lowest doses to both the fetus and the dam.
Changes in red nucleus neuronal development following maternal alcohol exposure
Authors: Zajac C. S. , Bunger P. C. , Moore J. C.
Published: June 2005

The red nucleus of Swiss Webster mouse fetuses was examined for morphological changes following maternal ethanol exposure. Pregnant females were given a liquid diet containing 30% or 0% ethanol‐derived calories. Changes in numerical density of neurons and in neuronal nuclear volume were found in the rostral red (RR) nucleus of ethanol‐exposed pups but not in the caudal red (CR) nucleus. Because of the integrative nature of the RR, changes in neuronal morphology that might relate to synaptic connections could affect the behavioral response mechanisms of these off spring.
Discontinuity of primary and secondary neural tube in spina bifida induced by retinoic acid in mice
Authors: Yasuda Yoshiko , Konishi Hiroyoshi , Kihara Takahide , Tanimura Takashi
Published: June 2005

This report shows by light microscopy the appearance of secondary neurulation separated from primary neurulation and its developmental fate in the spinal cord of mice exposed to retinoic acid in utero. The embryos and fetuses were derived from pregnant mice (ICR strain) given 60, 40, or 0 mg/kg of retinoic acid in olive oil on day 8 of gestation orally and killed 1, 2, or 10 days later. Separation of the primary neural fold from the secondary neural tube was seen in 9‐ and 10‐day‐old embryos: the caudal part of the neuroepithelium of the primary neural fold was disarranged with non‐closed posterior neuropore, and underneath it the secondary neural tissue extended caudally with abnormal notochord. At term, fetuses showed spina bifida, including myeloschisis, myelocele, and diplomyelia (diastematomyelia) with abnormal distribution of ganglionic cells. These cord lesions were located between the third lumbar and second coccygeal levels. The former two cord anomalies were associated with diplomyelia and split the dorsal and ventral portions of the spinal cord with an overlapping zone between the third lumbar and third sacral levels. These findings suggest that the separation from primary neurulation is due to the lesions in both primary neural folds and notochord induced by retinoic acid and that the spinal cord caudal to the third lumbar level originates from both neuroectoderm and mesenchyme‐like cells while that caudal to the third sacral level originates from mesenchyme‐like cells only.
Processes involved in retinoic acid production of small embryonic palatal shelves and limb defects
Authors: Abbott B. D. , Hill L. G. , Birnbaum L. S.
Published: June 2005

All‐trans‐retinoic acid (RA) is teratogenic to the embryonic mouse, producing malformations in many developing systems, including the limb bud and palate. High incidences of limb defects and cleft palate are induced at doses which are not maternally toxic and do not increase resorptions. Exposure to RA on gestational day (GD) 10 results in small palatal shelves, which fail to make contact on GD 14. The formation of small shelves could be a consequence of increased cell death, reduced proliferation, a combination of these effects, or some other effect such as inhibition of extracellular matrix production. After exposure to 100 mg RA/kg on GD 10, proliferation in mesenchymal cells of the palatal shelves was not reduced from GD 12 to GD 14 and the levels of cell death in control and treated shelves did not differ when observed by light and electron microscopy. The present study examines the effects of RA on cell death and proliferation from GDs 10–12 and compares the effects in palatal shelves and limb buds. Embryonic mice were exposed to RA suspended in corn oil (100 mg/kg on GD 10), a dose that was teratogenic but not maternally toxic or embryolethal. Embryos were collected at 4, 12, 24, 36, or 48 hr postexposure, and tissues which form the palate or limb were dissected from the embryos, stained by a modified Feulgen procedure, and whole mounted on slides. Mitotic index (MI) and percentage dead cells were determined for mesenchymal cells of the first visceral arch, maxillary process, or palatal shelf (depending on stage of development) and forelimb buds. In the palatal tissues from GD 10 to GD 12, RA did not significantly alter MI and percentage dead cells was significantly increased only at 4 hr postexposure. Some whole embryos were prepared for scanning electron microscopy (SEM). At 48 hr (GD 12) a reduction in the size of the shelves was not apparent on SEM. In the limb buds, RA did not increase percentage dead cells, but MI was significantly decreased. A decreasing rate of proliferation was detected in control facial tissues as development progressed, and this agrees with findings in rat and chick. Thus it appears that mesenchymal cell death and reduced proliferation are not responsible for the small palatal shelves seen on GD 14. RA did not increase cell death but inhibited proliferation in the limb bud, and this effect may contribute to the retarded development and malformations occurring in the limb.
Biochemical changes in the developing rat central nervous system due to hyperthermia
Authors: Rao G. Subba , Abraham Varkey , Fink Bradley A. , Margulies Nurit , Ziskin Marvin C.
Published: June 2005

Rat embryos at 10 days of gestation were exposed to 43°C for 8 minutes by submerging the exteriorized right uterine horn in heated saline solution and then reinserting the uterine horn into the abdominal cavity. At 15 days, the fetuses were removed, and cells from the cerebral hemispheres were dissociated and grown as primary cultures. Embryos from the left uterine horn served as controls. No morphological changes were observed between the cultures of cells from control and heat‐exposed embryos at different days in culture. However, exposure of embryos to hyperthermia at 10 days significantly affected the developmental pattern of activities of acetylcholine esterase associated with cholinergic neurons and of 2′,3′‐cyclic nucleotide phosphohydrolase associated with oligodendrocytes and myelin membrane formation. These results suggest that hyperthermia at 10 days of gestation in the rat may lead to an impairment in the development of neurons and oligodendrocytes in the central nervous system.
Parameters determining isotretinoin teratogenicity in rat embryo culture
Authors: Ritchie Helen , Webster William S.
Published: June 2005

At the in vitro threshold serum concentration of 500 ng/ml, isotretinoin induces defects of visceral arch development in 9.5‐day rat embryos grown in culture for 48 h. Experiments were performed to determine the minimum period of exposure necessary to induce these arch defects and whether an increase in concentration of isotretinoin could compensate for reduced exposure time. The results showed that a minimum 6‐h exposure to 500 ng/ml immediately prior to cranial neural crest migration was necessary to induce severe defects of the second visceral arch in a majority of embryos. Maximal increase in isotretinoin concentration to 16,000 ng/ml did not compensate for shorter exposure periods.

These results suggest that to cause malformations of the visceral arches, the embryo must be exposed to isotretinoin for a minimum period of time regardless of the concentration of isotretinoin above the threshold.
Preventive effects of phenoxybenzamine on nitrous oxide‐induced reproductive toxicity in Sprague‐Dawley rats
Authors: Fujinaga Masahiko , Baden Jeffrey M. , Suto Akira , Myatt John K. , Mazze Richard I.
Published: June 2005

In previous studies, we have shown that the reproductive toxicity of N2O in rats is prevented by the co‐administration of either halothane or isoflurane, whereas treatment with folinic acid, which should reverse the effects of N2O on DNA production, does not prevent toxicity. These results cast doubt on the commonly held theory that inactivation of methionine synthase is the sole cause of N2O‐induced reproductive toxicity, and suggest the need for other hypotheses. One such possibility is that N2O causes adverse reproductive toxicity secondary to its sympathomimetic effects. As a first step to test this theory, we studied the effects of phenoxybenzamine (PX), an alpha‐1 adrenergic antagonist, on N2O‐induced reproductive toxicity using a well‐established in vivo rat model. On day 8 of gestation (plug day = day 0), 130 timed‐pregnant Sprague‐Dawley rats were injected s.c. with either 0.5 ml of either 0.9% saline (control and N2O alone groups) or PX (0.5, 5, or 50 μg/kg) in 0.9% saline, the latter the maximum tolerated PX dose. They were then exposed to either air (control) or 60% N2O for 24 hours (all other groups). On day 20 of gestation, cesarean sections were performed and the fetuses were removed and examined for either visceral of skeletal abnormalities. Compared with control, treatment with N2O alone resulted in increased incidences of fetal resorptions, major and minor visceral abnormalities, and minor skeletal abnormalities. The most common major abnormalities were various forms of altered body laterality (20/72 fetuses; 28%). The two highest PX doses partially reduced the incidence of resorptions but none of the PX doses prevented major teratogenic effects. These results suggest that stimulation of alpha‐1 adrenergic receptors accounts for some of the adverse reproductive effects of N2O. It should be noted that although the concentration of N2O used in the present study is in the clinical range, N2O is rarely administered to surgical patients for more than a few hours. Our main aim was to study the mechanisms involved in the reproductive toxicity of longterm N2O exposure and not for any concerns that short‐term (a few hours) exposure represented a reproductive risk.
Amelioration by leucovorin of methotrexate developmental toxicity in rabbits
Authors: DeSesso John M. , Goeringer Gerald C.
Published: June 2005

Methotrexate (MTX) is lethal or teratogenic to embryos of all species tested. New Zealand white rabbit embryos are relatively resistant to the embryolethal effects of MTX. However, when pregnant does were injected iv with 19.2 mg MTX/kg on gestational day 12, virtually all surviving fetuses exhibited multiple malformations of the head, limbs, and trunk. MTX is a structural analogue of folic acid that competitively inhibits dihydrofolate reductase, thereby preventing formation of folinic acid and essentially stopping one carbon metabolism. One carbon metabolism is important in the synthesis of methionine, histidine, glycine, and purine bases that are required for the de novo synthesis of DNA. Presumably these metabolic effects of MTX relate directly to its mechanism of developmental toxicity. An ameliorative treatment has been tested utilizing iv injection of pregnant rabbits with leucovorin (LV), a close structural analogue of folinic acid (the product of the inhibited enzyme), at various times after MTX exposure. When LV was injected at times up to 24 hours after MTX, fewer malformed fetuses resulted and the incidence of specific malformations was reduced. When given at times up to 20 hours after MTX administration, LV virtually eliminated the grossly apparent effects of MTX at term. In the forelimb bud, MTX increased the extracellular space surrounding limb bud mesenchymal cells within 8–10 hours; this process continued through 16 hours and remained unabated by 24 hours. Mesenchymal cell nuclei became hyperchromatic and pyknotic during this time period. By 24 hours, a moderate amount of cellular debris was observed in the mesenchymal compartment of limb buds from approximately one‐third of the embryos examined. Endothelial cell nuclei of the limb bud vasculature did not exhibit the histopathological alterations observed in the mesenchymal cells. Limb buds from embryos injected with LV at times up to 6 hours after MTX were histologically normal. When LV treatment was delayed until 16 or 20 hours after MTX, mesenchymal nuclei regained normal appearance within 2 hours of treatment; further, the abnormally large intercellular space began to decrease during the next 4 hours. Cellular debris was not a prominent feature of limb buds from LV‐treated embryos examined at any time. Embryos from rabbits injected with LV at 24 hours after MTX exhibited either typical MTX‐induced lesions or a sequence of reparative events similar to those described for the 16 and 20 hour LV‐treated embryos. These results demonstrate that LV, administered up to 24 hours after MTX, is able to ameliorate the developmentally toxic effects of MTX seen in rabbits at term. Histological examination revealed that LV is able to reverse the early sequelae of MTX exposure in limb bud mesenchymal tissue.
Effects of prenatal exposure to methylazoxymethanol (MAM) on brain weight, hypothalamic cell number, pituitary structure, and postnatal growth in the rat
Authors: Rodier Patricia M. , Kates Barbara , White Wendy A. , Muhs Ann
Published: June 2005

Congenital brain damage syndromes typically are described in terms of behavioral symptoms. Many brain functions are not reflected in behavior, however, and prenatal injury to the developing nervous system could alter these functions, as well. To test the hypothesis that prenatal brain injury can result in postnatal endocrine malfunction, rats were exposed in utero to 20 mg/kg of methylazoxymethanol acetate, a potent neuroteratogen, at two stages of gestation when different sets of growth‐controlling neurons of the hypothalamus are forming. The growth hormone releasing factor (GRF) neurons stimulate release of growth hormone from the somatotropes of the anterior pituitary, contributing to rapid growth in the period between weaning and puberty. The somatotropin release inhibiting factor (SRIF) neurons have the opposite effect on the pituitary and can inhibit the GRF cells directly. Growth of treated animals was monitored daily from birth to 40 days and compared to that of controls Treatment on the 14th day of gestation produced a small number of dwarf animals characterized by normal weight at birth and a sudden decrease in growth rate at the beginning of the fourth postnatal week that led to a body weight about 50% of normal. Treatment on day 16 yielded an acceleration of postnatal growth (significant in males). In each group, most treated animals were like controls in adult size and pattern of growth. As adults, both treatment groups demonstrated massive reductions in brain weight which characterized all the subjects, whether or not they exhibited growth anomalies. The animals treated on day 14 were confirmed to have a significant, selective reduction in growth hormone releasing factor neurons. Reductions were greatest in the middle and posterior levels of the GRF cell distribution, the regions forming most actively at the time of exposure. Unexpectedly, the same group also had increased numbers of periventricular SRIF neurons. Neither type of neurons was significantly altered in the later treatment group. Examination of pituitary structure indicated that dwarfs had very small pituitaries, with an immature pattern of somatotrope distribution, and giants had very large pituitaries, with some hypertrophy of somatotropes. The results suggest that endocrine anomalies which manifest themselves long after birth may originate as birth defects of the nervous system.
Effects of magnetic resonance imaging on eye development in the C57BL/6J mouse
Authors: Tyndall Donald A. , Sulik Kathleen K.
Published: June 2005

An investigation was undertaken to ascertain the potential teratogenicity of magnetic resonance imaging (MRI) fields. The C57BL/6J mouse was chosen as the experimental model with eye malformations (microphthalmia and morphologic anomalies) designated as the biological end point. This mouse strain is genetically predisposed to this type of malformation as a 10% spontaneous incidence occurs. Dams in groups of 15 were subjected to MRI imaging conditions on gestational day (Gd) 7 for 36 minutes to a spin‐echo T‐2‐weighted scan by using a 1.5 Tesla magnetic field and a radiofrequency (RF) field of 64 MHz. One group was exposed at the magnetic isocenter while another was exposed at the entrance to the magnet lumen. There was also a sham control group. The dams were sacrificed at Gd 14. Assessment of eye abnormality was determined by, (1) a veterinary ophthalmologist, (2) a computer‐based method comparing eye areas, and (3) a methodology combining both the above subjective and quantitative methods. MRI fields were found to produce malformation rates (15–37%) higher than controls (2–19% P⩽.05, Kruskal‐Wallis Test) for both isocenter and lumen entrance groups. The malformation rates and degree of statistical significance varied somewhat with analytical methodology and the unit of measure (right eye, left eye, or fetus). The results suggest for the first time the potential of MRI fields to produce developmental malformations in an animal model utilizing clinically realistic exposure conditions. (However, the reader is reminded that the mouse strain utilized in this investigation was genetically prone to malformations.)
Explaining certain human limb anomalies and the limb—hematopoiesis community of syndromes using a model of determination
Authors: Lubinsky Mark S.
Published: June 2005

Disturbances of determination, the process of limitation of developmental potential, can cause structural as well as histologic anomalies. A polar coordinate model (PCM) developed from studies in animals, in which determination depends upon positional information arranged along polar coordinates, can be used to explain certain classes of human limb anomalies. Under the model, interference with this system primarily affects distal patterning. If the radial area is distal to the zone of polarizing activity in embryological development, as it appears to be, the PCM explains the teratologic equivalence of preaxial duplications and deficiencies in certain circumstances and the prevalence of ulnar dimelias in forearm duplications. Also, failures of hematopoiesis can be considered late problems with determination and may be markers for abnormalities of a determinative process that also has earlier developmental consequences. Abnormalities of retinoic acid morphogen receptors would be one possible mechanism. This would provide a rationale for the known association of postnatal hematologic problems with developmental anomalies preferentially affecting the radial area. Syndromes with limb and hematopoietic problems may well be a community of determinative disorders.
Lethal and teratogenic effects of long‐term low‐intensity radio frequency radiation at 428 MHz on developing chick embryo
Authors: Saito Kenichi , Suzuki Katsushi , Motoyoshi Shigekatsu
Published: June 2005

Exposure of developing chick embryos to 428 MHz radio frequency (RF) radiation at a power density of 5.5 mW/cm2 for more than 20 days resulted in embryolethal and/or teratogenic effects and delayed hatching. These adverse biological effects were not due to any thermal effect of the RF radiation. We have demonstrated teratogenicity in the chick embryo as a result of protracted low‐dose RF irradiation.
Developmental regulation for collagen II gene expression in transgenic mice
Authors: Bruggeman Leslie A. , Hou‐Xiang Xie , Brown Kenneth S. , Yamada Yoshihiko
Published: June 2005

In order to evaluate the involvement of the type II collagen regulatory sequences in development, we have injected a construct containing a toxin gene under the control of the rat type II collagen promoter and enhancer. The construct, pDAS10‐DTA, contained the diphtheria toxin A chain gene under the control of type II collagen sequences which had been used previously to target cartilagenous tissues in transgenics. Inspection of developing fetuses at various stages of gestation revealed a high number of aborted implants as well as abnormally developing fetuses. These abnormal fetuses were of small size, had shortened and underdeveloped limbs, cleft palates, and generally resembled a phenotype similar to chondrodystrophic mice. Histological comparisons of normal and abnormal fetuses indicated a reduced amount of extracellular matrix surrounding chondrocytes, and a disorganized appearance of the tissue. These results suggest that the expression of the toxin has occurred in chondrocytes and altered the survival and development of the transgenic mice. These results also indicate that the promoter and enhancer sequences contained in the transgene controlled the developmental expression of the type II collagen gene expression.