bluuebunny/crossref_metadata_2025 huggingface dataset:
Toxic effects of cyclophosphamide in differentiating chicken limb bud cell culture using rat liver 9,000 g supernatant or rat liver cells as an activation system: An in vitro short‐term test for proteratogens
Authors: Wiger Richard , Trygg Bente , Holme Jørn A.
Published: June 2005

Cells from 4‐day chicken embryo limb buds plated as micromass cultures differntiate and form cartilage nodules after a 5‐ to 6‐day growth period. The innate ability of these cells to biotransform compounds, such as cyclophosphamide (CP), into reactive metabolites is apparently inadequate. Protocols used rat liver S9 from Aroclor 1254 pretreated (PCB) rats or hepatocytes from control rats or polychlorinated biphenyls (PCB)‐pretreated rats and were added to micromass cultures with CP causing concentration‐related toxicity in the cell cultures. Coculturing chicken limb bud cells with PCB‐hepatocytes was the most efficient method, yielding an IC50 of 2 μg CP per ml. Toxic CP metabolites accumulated in the medium of PCB‐hepatocyte cultures and were quite stable. The toxicity of bioactivated CP was nearly identical for both proliferation and cartilage proteoglycan accumulation.
Early postimplantation embryolethality in mice following in utero inhibition of adenosine deaminase with 2′‐deoxycoformycin
Authors: Knudsen T. B. , Gray M. K. , Church J. K. , Blackburn M. R. , Airhart M. J. , Kellems R. E. , Skalko R. G.
Published: June 2005

Adenosine deaminase (ADA) catalyzes the hydrolytic deamination of adenosine (or 2′‐deoxyadenosine) to inosine (or 2′‐deoxyinosine). Previously, we have shown that ADA activity is subject to strong cell‐specific developmental regulation in placental tissues of mice between days 6 and 11 of gestation (Knudsen et al.:Biology of Reproduction 39:937–951, 1988). In the present study, we examined the effects of intrauterine exposure to 2′‐deoxycoformycin (dCF; pentostatin), a potent irreversible inhibitor of ADA, on early postimplantation development. Deoxycoformycin was administered to pregnant ICR mice as a single intraperitoneal injection at a dose of 5 mg/kg on one of days 6 through 11 of gestation (plug day 0). A marked increase in the incidence of implantation site resorptions was observed following treatment specifically on days 7 (61% resorbed) or 8 (78% resorbed). No effect was observed following treatment on days 6, 9, 10, or 11. ADA‐immunoreactive protein was shown, by ABC‐immunoperoxidase staining on days 7 or 8 of gestation, to be present at high levels in decidual cells of the antimesometrial region but at below‐detectable levels in the embryo. Treatment of pregnant dams with dCF on day 7 produced a complete (>99%) inhibition of ADA activity in the antimesometrial decidua by 30 min, induced excessive cell death in the prospective neural plate and primary mesenchyme of the trilaminar disc by 6 h, and arrested embryonic development at an early somite stage. These results suggest that the antimesometrial decidua plays a protective role in preventing an inappropriate accumulation of endogenous ADA substrates in the implantation site.
Defects of skeletal morphology, density, and structure in mouse fetuses with trisomy 16
Authors: Sterz H. , Buselmaier W. , Bacchus C. , Gromier L. , Eppler E.
Published: June 2005

Skeletal anomalies present in trisomy 16 in the mouse—an animal model of human trisomy 21—are described. Altogether 27 fetuses with trisomy 16 and 118 chromosomally balanced siblings were examined radiographically and by alizarin staining on day 20 of gestation; the radiographs were analyzed by computer‐aided densitometry and structural differentiation. Extensive asymmetry or abnormal fusion of the vertebral centers and alterations of the vertebral arches were observed along with rib malformations (rib‐vertebra syndrome). The skull primarily exhibited anomalies of the occipital bone. Ossification of the humerus, femur, and tibia was characterized by reduced mineralization. Typical, fracture‐like alterations affecting only the tibia were also observed. Measurement of the lengths of the humeri of fetuses of comparable weight revealed a growth retardation not correlatable with the degree of mineralization. The significance of these skeletal abnomalies with regard to the trisomy 21 syndrome is discussed.
Effects of phorone and/or buthionine sulfoximine on teratogenicity of 5‐fluorouracil in mice
Authors: Naya Masato , Mataki Youichi , Takahira Hiroshi , Deguchi Takashi , Yasuda Mineo
Published: June 2005

Embryotoxicity and teratogenicity of 5‐fluorouracil (5‐FU) and modulation of its effect by the depletors of glutathione (GSH) were evaluated in mice. Pregnant ICR mice were intraperitoneally (i.p.) injected with 25 mg/kg of 5‐FU on day 11 of gestation (vaginal plug = day 0). Mice were pretreated i.p. with 250 mg/kg of phorone, a GSH depleting agent and/or 200 mg/kg of buthionine sulfoximine (BSO, an inhibitor of GSH biosynthesis) 4 hours before dosing with 5‐FU. Dams were killed on day 17 of gestation. Fetuses were examined for external malformations, especially limb malformations. Pretreatment with phorone or BSO decreased fetal weight and increased the frequency and severity of oligodactyly induced by 5‐FU, as well as the reduction of maternal GSH levels. Combined use of 125 mg/kg phorone and 100 mg/kg BSO i.p. augmented growth retardation induced with 5‐FU. Cotreatment with exogenous GSH, at a dose of 300 mg/kg injected intravenously, could not suppress the augmentative effects of phorone and/or BSO on 5‐FU teratogenicity under these experimental conditions. These results indicate that the level of endogenous GSH is one of the factors which significantly affects teratogenicity of 5‐FU.
Teratogenicity of carbamazepine in rats
Authors: Vorhees Charles V. , Acuff Karen D. , Weisenburger Walter P. , Minck Daniel R.
Published: June 2005

The teratogenicity of carbamazepine (CBZ) was investigated in Sprague‐Dawley CD rats at doses of 0, 200, 400, and 600 mg/kg administered by gavage in corn oil on days 7–18 of gestation in a dosage volume of 2 ml/kg. The CBZ‐600 dose was maternally toxic in that dams in this group weighed 30.6% less than controls by E20. This group had significantly increased resorptions, reduced live fetal weight (51.6% less than controls), and increased skeletal and visceral abnormalities. The CBZ‐400 dose also significantly reduced maternal weight gain during gestation to 26.6% less than controls by E20. No significant increase in resorptions occurred in this group; live fetuses weighed 42.9% less than controls and showed an increase in visceral, but not skeletal, abnormalities. The CBZ‐200 dose did not significantly affect maternal weight gain or increase resorptions or fetal abnormalities but did reduce fetal body weight (20.3% less than controls). Maternal serum total CBZ concentrations 1 hr after the final dose were 22.9, 27.9, and 34.4 μg/ml for the 200, 400, and 600 mg/kg groups, respectively. These levels were little changed 6 h post‐treatment. CBZ was 65–70% serum protein bound across dose groups. Human therapeutic levels of CBZ are 4–12 μg/ml and the drug is typically 80% serum protein bound. This suggests that abnormalities in rats occur at concentrations well above the human therapeutic range. However, a no‐effect level was not found for fetal body weight. Further experiments will be required to determine how much lower doses will need to be in order to find a no‐effect level for fetal body weight. Nevertheless, the present data suggest that CBZ is not potent at inducing malformations in rats.
Morphogenesis of retinoic acid‐induced postaxial polydactyly in mice
Authors: Bynum S. V.
Published: June 2005

Previous experiments have demonstrated that oral administration of retinoic acid to ICR mice on day 10 of gestation results in an enhanced expression of postaxial polydactyly of the forelimbs. In the present investigations, the development of the limbs from the time of treatment until the appearance of the defect on day 14 were studied at 12‐ to 24‐hr intervals with histological and vital staining technics. A diffuse cytotoxicity in the central proximal mesoderm was noted with a safranin–toluidine blue stain 12 h after treatment which then peaked 24 hr after treatment. Treated limbs stained with Nile blue sulfate showed a significant increase of necrotic mesodermal cells in the foyer preaxial primaire, foyer marginal I and foyer marginal V when compared to the respective control group. The treated postaxial ectoderm was more hyperplastic than was the control ectoderm on day 12. The treated group developed a squarish bulge on the postaxial boundary of the fifth digital ray on day 14 that was larger than those of the controls. Most of the controls had bulges that were completely necrotic. The treated group also had some completely necrotic bulges but a large number of the treated fetuses had bulges characterized as being less than 60% necrotic.
Teratogenic effect of cocaine and diazepam in CF1 mice
Authors: Mehanny Sherif Z. , Abdel‐Rahman Mohamed S. , Ahmed Yehia Y.
Published: June 2005

This study was conducted to determine the teratogenic effect of cocaine hydrochloride, alone or in combination with diazepam. Pregnant mice were administered cocaine hydrochloride at 10 and 20 mg/kg body weight intravenously (tail), and/or diazepam at 20 or 40 mg/kg b.w. by gavage. Combinations of diazepam and cocaine (20/10 and 40/20, respectively) were used. Significant reductions of fetal weight and length were found in the group treated with diazepam and cocaine (40/20). The incidences of incomplete ossification of sternebrae and skull bones, as well as delayed ossification of the paws, were significantly increased in the combination groups. Hydronephrosis and cryptorchidism were observed in the cocaine‐treated groups, with the incidence of these malformations increasing significantly when diazepam was co‐administered.
Physiological consequences of early neonatal growth retardation: Effects of α‐difluoromethylornithine on renal growth and function in the rat
Authors: Gray Jacqueline A. , Kavlock Robert J.
Published: June 2005

The physiological consequences of early neonatal growth retardation in the kidney were investigated using α‐difluoromethylornithine (DFMO), a specific irreversible inhibitor of ornithine decarboxylase (ODC), a key enzyme in the biosynthesis of polyamines. We administered by s.c. 500 mg/kg/day DFMO, or saline, to Sprague‐Dawley rat pups from the day of birth through postnatal day (PD) 6 and evaluated renal function on PD 4, 7, 10, and 13 using tests of basal renal clearance and urinary concentrating ability. Kidney weights and gross pathology were also obtained. On PD 39, serum chemistries and organ weights were determined. In a second experiment, we evaluated concentrating ability on PD 7–10, and basal renal function, concentrating ability, diuretic response, serum chemistries, and organ weights on PD 132–140. DFMO selectively inhibited renal growth but did not inhibit glomerular and tubular functional maturation. In fact, the rates of filtration and reabsorption (per g renal tissue), and concentrating ability were increased in treated pups. These changes were associated with long‐term effects on renal function, including uremia, glucosuria, and male‐specific concentrating deficits in adulthood. Several hypotheses can be developed concerning the physiological mechanisms underlying these changes (e.g., altered renal urea metabolism), which in turn may reflect either a direct role of ODC in the regulation of maturation or secondary consequences of inhibition of ODC. In conclusion, the renal growth retardation induced by neonatal administration of DFMO (PD 2–6) was associated with an apparent precocious maturation of function in the early postnatal period (PD 4–13) and dysfunction in adulthood, thus demonstrating that the physiological consequences of growth retardation are not always obvious and predictable, and must be evaluted on a case by case basis.
Conceptus viability, malformation, and suspect mutagens or teratogens in humans. The Yule–Simpson paradox and implications for inferences of causality in studies of mutagenicity or teratogenicity limited to human livebirths
Authors: Hook Ernest B. , Regal Ronald R.
Published: June 2005

If a genetic disorder or a congenital malformation is associated with embryonic or fetal death, then this may distort reported associations of environmental or biological factors with genetic and congenital disorders. We illustrate how such distortion may occur in humans as a manifestation of the Yule–Simpson (or Simpson) statistical paradox. We analyze the reported negative association of maternal smoking and Down syndrome in livebirths and demonstrate that under plausible conditions, valid negative associations both in livebirths and in embryonic or fetal deaths can occur, despite the fact that in all conceptuses there may be no association whatsoever. A similar analogous manifestation of the Yule–Simpson paradox may hold for positive associations. That is, positive relative risks may occur for an alleged mutagen or teratogen in separate studies both of livebirths, and of embryonic and fetal deaths, despite the absence of any effect in all conceptuses. These considerations illustrate the need for data on genetic disorders and malformations in embryonic and fetal deaths, and the need for a synthesis of such data with those from studies in livebirths before valid inferences may be made about mutagenic or teratogenic effects of environmental or biological factors.
Prenatal exposure to gamma/neutron irradiation: Sensorimotor alterations and paradoxical effects on learning
Authors: Di Cicco Daniela , Antal Sara , Ammassari‐Teule Martine
Published: June 2005

The effects of prenatal exposure on gamma/neutron radiations (0.5 Gy at about the 18th day of fetal life) were studied in a hybrid strain of mice (DBA/Cne males × C57BL/Cne females). During ontogeny, measurements of sensorimotor reflexes revealed in prenatally irradiated mice (1) a delay in sensorial development, (2) deficits in tests involving body motor control, and (3) a reduction of both motility and locomotor activity scores. In adulthood, the behaviour of prenatally irradiated and control mice was examined in the open field test and in reactivity to novelty. Moreover, their learning performance was compared in several situations. The results show that, in the open field test, only rearings were more frequent in irradiated mice. In the presence of a novel object, significant sex × treatment interactions were observed since ambulation and leaning against the novel object increased in irradiated females but decreased in irradiated males. Finally, when submitted to different learning tasks, irradiated mice were impaired in the radial maze, but paradoxically exhibited higher avoidance scores than control mice, possibly because of their low pain thresholds. Taken together, these observations indicate that late prenatal gamma/neutron irradiation induces long lasting alterations at the sensorimotor level which, in turn can influence learning abilities of adult mice.
A new method for explanting early postimplantation rat embryos for culture
Authors: Fujinaga Masahiko , Baden Jeffrey M.
Published: June 2005

Rat embryos were explanted either on day 8, 3 PM (primitive streak stage) or on day 9, 8 AM (presomite stage) and divided randomly into two groups (plug day = day 0). In one group, Reichert's membrane was removed starting from the side opposite to the ectoplacental cone which was left intact (standard method). In the second group, Reichert's membrane was removed starting from the same side as the ectoplacental cone after the latter structure was excised and the proamniotic cavity (day 8) or the ectoplacental cavity (day 9) destroyed (new method). Embryos were then cultured and examined at 10 AM on day 11. All embryos developed normally in size and general morphology. However, there was a high incidence of inverted heart, allantoic placenta and tail in all groups, except for day 9, new method group; this was the only group in which the ectoplacental cavity did not expand abnormally during culture. The new method, which is quick and easy to perform even when Reichert's membrane does not protrude well, lessens the chances of damaging embryonic ectoderm during the removal of Reichert's membrane. Furthermore, the absence of ectoplacental cone decreases embryo aggregation during culture. We hypothesize that the abnormally expanded ectoplacental cavity generates abnormal physical forces within the egg cylinder that disrupt the control of body asymmetry.
Sirenomelia associated with a “vanishing twin”
Authors: Kapur Raj P. , Mahony Barry S. , Nyberg David A. , Resta Robert G. , Shepard Thomas H.
Published: June 2005

A case is presented of twin gestation in which one gestational sac was completely resorbed and the remaining twin was subsequently found to be sirenomelic. First‐trimester prenatal ultrasound examination demonstrated a second gestational sac that disappeared 2 weeks later. The sonographic features that led to the diagnosis of sirenomelia in the remaining fetus included severe renal dysgenesis, persistently apposed lower extremities, and absence of fibulae. Postmortem examination, including angiographic studies of the fetus, revealed caudal dysgenesis and a single umbilical artery that arose from the abdominal aorta. Sirenomelia occurs more frequently in twin gestations than in singletons. This case suggests that the association between twinning and sirenomelia may be greater than is currently recognized. Two hypotheses are given to explain this association.
TCDD exposure of human embryonic palatal shelves in organ culture alters the differentiation of medial epithelial cells
Authors: Abbott B. D. , Birnbaum L. S.
Published: June 2005

The highly toxic, polychlorinated aromatic compound 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin (TCDD) occurs as a contaminant throughout the environment. Epidemiology studies of populations accidentally exposed to TCDD have failed to identify TCDD as a human teratogen, but these studies are limited by the small numbers of exposed pregnancies and imprecise estimates of exposure. TCDD is highly teratogenic in mice, inducing cleft palate and hydronephrosis. TCDD exposure in vivo of embryonic mice alters the differentiation and expression of growth factors in the medial epithelial palatal cells. These alterations also occur in rat and mouse palates exposed to TCDD in organ culture. In the present study, human embryonic palatal shelves were cultured in the rodent organ culture system. In order to achieve in vitro the developmental stage at which fusion would normally occur, GD 52 shelves were cultured for 4 days, GD 53 shelves were cultured for 3 days, and GD 54 shelves were cultured for 3 days. Three of four palatal shelves exposed to 5 × 10−11 M TCDD were identical to their homologous controls (right shelf cultured with control medium; left shelf cultured with TCDD‐containing medium). TCDD at 1 × 10−7 M produced cytotoxicity detected by transmission electron microscopy (TEM). Exposure to 1 × 10−8 M TCDD resulted in continued incorporation of thymidine ([3H]‐TdR detected autoradiographically) by palatal medial cells, failure of the medial peridermal cells to degenerate as observed by scanning electron microscopy (SEM), and differentiation into a stratified, squamous epithelium. These alterations are identical to those induced by TCDD in vitro in rat and mouse palatal cells. The main difference between these species is the level of TCDD required to elicit the responses. Cultured mouse palates respond to 5 × 10−11 M TCDD with altered medial cell differentiation, and 1 × 10−10 M TCDD is cytotoxic. The rat shelves respond with altered differentiation at 1 × 10−8 M and cytotoxicity at 1 × 10−7 M. All the human shelves respond at 1 × 10−8 M TCDD with altered differentiation, 1 out of 4 responded at 5 × 10−11 M, and cytotoxicity occurred at 1 × 10−7 M. The present data suggest human embryonic palates are less sensitive than those of the C57BL/6N mouse, and that exposure to high levels of TCDD would be required to elicit altered differentiation in the palatal shelf.
Activity profiles of developmental toxicity: Design considerations and pilot implementation
Authors: Kavlock Robert J. , Greene Jacqueline A. , Kimmel Gary L. , Morrissey Richard E. , Owens Elizabeth , Rogers John M. , Sadler Thomas W. , Stack H. Frank , Waters Michael D. , Welsch Frank
Published: June 2005

The available literature was searched for quantitative test results from both in vitro and in vivo assays for developmental toxicity for five model compounds: cyclophosphamide, methotrexate, hydroxyurea, caffeine, and ethylenethiourea. These compounds were chosen on the basis of their extensive utilization in a variety of assay systems for developmental toxicity as evidenced by their representation in the ETIC database (each generally has 100–500 citations encompassing multiple test systems). Nine cellular‐based assays, six assays using whole embryos in culture, as well as Segment II and abbreviated exposure tests for mammalian test species are included in the database. For each assay, the critical endpoints were identified, each of which was then provided a three‐letter code, and the criteria for extraction of quantitative information were established. The extracted information was placed into a computerized reference file and subsequently plotted such that the qualitative (positive/negative) and quantitative (e.g.), IC50, highest ineffective dose (HID), lowest effective dose (LED)) results across all test systems could be displayed. The information contained in these profiles can be used to compare qualitative and quantitative results across multiple assay systems, to identify data gaps in the literature, to evaluate the concordance of the assays, to calculate relative potencies, and to examine structure‐activity relationships.
Isolated oral cleft malformations: Associations with maternal and infant characteristics in a California population
Authors: Shaw Gary M. , Croen Lisa A. , Curry Cynthia J.
Published: June 2005

Data on isolated oral cleft malformations from a birth defects registry covering a large population base were examined to describe potential associations with maternal and infant characteristics. Infants with cleft palate (CP) were analyzed separately from infants with cleft lip with or without cleft palate (CLP). The prevalence of isolated CLP per 1,000 births was 0.741, approximately twice the prevalence of isolated CP, which was 0.383. Male infants were more likely to be born with CLP (OR = 1.9) but less likely to be born with CP (OR = 0.56) than were females. Women 39 years or more of age were twice as likely as 25–29 year olds to have a child with either type of cleft. Black, non‐hispanic infants had a lowered risk for CLP compared to white, nonhispanics (OR = 0.40). These risks were found to be independent of each other based on multivariate analyses. Associations with either type of cleft malformation were not observed for plurality, number of previous live births, and maternal birthplace.
Role of glutathione and hsp 70 in the acquisition of thermotolerance in postimplantation rat embryos
Authors: Harris Craig , Juchau Mont R. , Mirkes Philip E.
Published: June 2005

Studies were initiated to determine the extent to which reduced glutathione (GSH) may be involved in the capacity of cultured rat embryos to develop heat‐induced tolerance to the deleterious effects of exposure to high temperatures (heat shock). Investigations of the modulation of dysmorpho‐genic responses of embryos to heat shock (43°C, 30 min) as well as to the expression of the hsp70 gene and subsequent formation of hsps indicated that the acquisition of thermotolerance by rat embryos could be significantly influenced by the inhibition of GSH synthesis. Treatment of conceptuses with L‐buthio‐nine‐S,R‐sulfoximine (BSO) reduced intracellular GSH concentrations and compromised the capacity of embryos to mount a thermotolerance response as assessed by alterations in indices of growth and development. Embryonic thermotolerance elicited by preexposure to 42°C for 30 min was accompanied by increases in GSH to levels greater than those measured in control embryos at 37°C just prior to the subsequent 43°C heat exposure. Expression of hsp70 mRNA was detectable soon after elevation of the temperature to 42°C and reached its highest level of accumulation 1.5 hr after the 43°C heat shock. BSO treatment had little if any effect on hsp70 message levels or on the synthesis of hsp70. The fact that BSO‐treatment attenuated the thermotolerance response but did not produce a decrease in hsp70 RNA or the synthesis of hsp70 suggests that hsp70 alone is not sufficient to confer thermotolerance upon cultured rat embryos.
Comparison of the behavioral teratogenic potential of phenytoin, mephenytoin, ethotoin, and hydantoin in rats
Authors: Minck Daniel R. , Acuff‐Smith Karen D. , Vorhees Charles V.
Published: June 2005

Pregnant Sprague‐Dawley CD rats were orally administered either phenytoin (PHT, 200 mg/kg), mephenytoin (MPH, 100 mg/kg), ethotoin (ETH, 600 mg/kg), hydantoin (HYD, 1,200 mg/kg) or vehicle (propylene glycol) on days 7–18 of gestation. Mean (±S.E.) maternal serum concentrations of PHT, MPH, and ETH 1 hour after dosing on gestational day 18 were 16.0 ± 3.3, 10.7 ± 3.0, and 65.2 ± 10.45, respectively, and free fractions were 16%, 18%, and 11%, respectively. The free fraction for PHT is similar, but was lower for both MPH and ETH than that seen in humans. Preweaning mortality for PHT, MPH, ETH, HYD, and controls was 25%, 6.3%, 12.5%, 2.0%, and 0.8%, respectively. The MPH and ETH‐exposed animals weighed approximately 6.6% less than controls throughout the study; the other groups did not differ significantly. PHT offspring showed increased early locomotor activity. Only PHT‐exposed animals (27%) exhibited abnormal circling behavior after weaning. PHT‐circlers accounted for higher levels of activity in an open‐field test and for longer straight channel swimming times. PHT‐circlers and noncirclers differed from one another and controls on performance of a complex (Cincinnati) maze and on the development of the air‐righting reflex. Offspring prenatally exposed to MPH showed an early delay in air‐righting. ETH and HYD offspring were not consistently different from controls in behavior. The data suggest the following ordinal relationship among the drugs for behavioral teratogenesis: PHT> >MPH>ETH≃HYD≃CON. The effects of PHT are consistent with previous findings. Data on the other drugs suggest that other hydantoins do not possess the behavioral teratogenic efficacy of PHT and that PHT may be unique in its effects on CNS development.
Fetal dexamethasone exposure impairs cellular development in neonatal rat heart and kidney: Effects on DNA and protein in whole tissues
Authors: Slotkin T. A. , Seidler F. J. , Kavlock R. J. , Bartolome J. V.
Published: June 2005

Fetal glucocorticoid exposure causes postnatal growth retardation. To examine the mechanisms underlying effects on specific organ systems, we administered 0.2 or 0.8 mg/kg of dexamethasone to pregnant rats on gestational days 17, 18, and 19 and assessed three biochemical markers of cell development in heart and kidney of the offspring: DNA content per organ as an index of total cell numbers, DNA per g tissue as an index of cell packing density, and protein/DNA ratio as an index of relative cell size. In both tissues, DNA content became markedly subnormal during the first postnatal week, the ontogenetic period of rapid cell division. Partial recovery occurred by the end of the first postnatal month. In the heart, cell packing density was subnormal initially and the cells were significantly enlarged. In contrast, packing density was slightly elevated in the kidney; protein/DNA was increased by the low dose of dexamethasone, but markedly decreased by the high dose. These results suggest that tissue growth impairment caused by prenatal dexamethasone treatment reflects primary deficits in cell proliferation that extend to a variety of different cell types; however, consequent effects on cell packing density and cell size are dose‐specific, possibly reflecting actions of glucocorticoids selective for certain cell types or phases of cell development.
Transplacental distribution of weak acids in mice: Accumulation in compartments of high pH
Authors: Srivastava M. , Collins M. D. , Scott W. J. , Wittfoht W. , Nau H.
Published: June 2005

The transplacental distribution of three weak acids was measured in mice on gestational day 11. The ratio of drug concentration in maternal, embryonic, and extraembryonic compartments compared to maternal plasma followed a consistent hierarchy such that amniotic fluid ⩾ exocoelomic fluid > embryo > embryo plasma > maternal skeletal muscle. This distribution pattern correlates well with pH in these compartments and suggests that pH gradients between compartments are an influential factor in determining weak acid drug disposition during pregnancy.
Morphogenetic and molecular correlates of teratogenesis in the amphibian embryo
Authors: Brennan Sean M.
Published: June 2005

In attempting to develop a system to study molecular mechanisms of teratogenesis, examination of the effects of a teratogen (dimethyl sulfoxide) on both molecular and morphological aspects of embryonic development in the amphibian Xenopus laevis has been conducted. Characteristic morphological effects, which occur during the period from 7 to 16 hours after fertilization (i.e., gastrulation) are noted. Delays in gastrulation are accompanied by changes in the regulation of transcription of several genes known to be active during gastrulation in normal development. Later morphological effects are also observed, and these probably arise as a consequence of the changes occurring during gastrulation. Thus, molecular responses to a teratogen have been detected, and a correlation between molecular and morphological responses to a teratogen is observed. These findings represent the first demonstration of the effects of a teratogen on the transcription of specific genes, and invite speculation that one or more molecular events mediate teratogenesis. They further suggest that the amphibian system may be useful for studying early molecular responses to teratogens.
Glucocorticoid teratogenesis in the developing nephron
Authors: Crocker J. F. S. , Ogborn M. R.
Published: June 2005

Although the induction of polycystic kidney disease by neonatal glucocorticoid treatment has been extensively documented, there are no data on induction of polycystic kidney disease with fetal exposure to glucocorticoids. We injected groups of pregnant Swiss Webster albino mice subcutaneously with 250 mg/kg of hydrocortisone acetate on individual days from days 1 to 19 of gestation. A control group received an equal volume of saline. Histologic analysis of 1,522 kidneys from the offspring of these animals revealed no evidence of polycystic kidneys in the control group or in offspring of animals injected before day 11 of pregnancy. A bimodal distribution of cystic kidney disease was noted in the remaining animals, with highest prevalence after injection on day 12 (50.8%) and day 17 (34.3%). We conclude that 250 mg/kg of glucocorticoids may induce polycystic kidney disease in utero, but possibly only during critical periods of metanephric development.
Lack of attenuation of valproic acid‐induced effects by folinic acid in rat embryos in vitro
Authors: Hansen Deborah K. , Grafton Thomas F.
Published: June 2005

The anticonvulsant drug valproic acid (VPA) is suspected to be a developmental toxicant in humans, inducing primarily neural tube defects. The mechanism for this effect is unknown, but it has been suggested that the drug may act via a deficiency of the vitamin follic acid. We examined this possibility by concurrent treatment of rat embryos in a whole embryo culture system with VPA and folinic acid (FA), a folic acid derivative. Groups of CD rat embryos were treated with various concentrations of VPA, various concentrations of FA, or a combination of a teratogenic dose of VPA plus various levels of FA. At the end of the 44 hour culture period, each embryo was evaluated for viability (presence of a heartbeat), yolk sac circulation, presence of any malformations, morhological score, crown–rump and head lengths, as well as DNA and protein contents. The anticonvulsant did not decrease viability but did decrease yolk sac circulation and all growth and developmental endpoints in a dose‐responsive manner. There was also a dose‐related increase in the incidence of open neural tubes. The addition of FA alone had no significant effect on growth and development. When various concentratins of FA were added simultaneously with a teratogenic dose of VPA, there was no decrease in the incidence of open neural tubes. Growth and developmental endpoints were altered in a somewhat random fashion but were never increased to the control level. The lack of attenuation of FA was not due to instability of the compound in the culture system, nor was there a difference in the amount of FA, present in the exocoelomic fluid of VPA‐treated and control embryos. Overall, these results suggest that VPA‐induced open neural tubes in rat embryos cultured in vitro are not due to a deficiency of folinic acid.
Medical sonography: Reproductive effects and risks
Authors: Brent Robert L. , Jensh Ronald P. , Beckman David A.
Published: June 2005

While it is clear that the levels and types of medical sonography that have been used in the past have no measurable risks, it would be inaccurate to label the modality of ultrasound as totally safe regardless of exposure. Most agents have reproductive risks and even teratogenic risks if the exposure is raised sufficiently. Thus the prudent use of sonography means that clinicians and designers of equipment have to maintain exposures far below the risks that have been demonstrated in animal studies and from the knowledge obtained about the physical changes that can be produced in humans as the absorbed dose is elevated. The reproductive risks were evaluated using five criteria: 1) human epidemiology, 2) secular trend data, 3) animal experiments, 4) dose response relationships, and 5) biologic plausibility. The analysis reveals that the human epidemiology does not indicate that diagnostic ultrasound presents a measurable risk to the developing embryo or fetus. Animal studies also indicate that diagnostic levels of ultrasound are safe and do not elevate the fetal temperature into the region where deleterious embryonic and fetal effects will occur. Because higher exposures of ultrasound can elevate the temperature of the embryo, the use of diagnostic procedures and the design of sonographic equipment should take into consideration the hyperthermic potential of higher exposures of ultrasound and the hypothetical additional risk of performing sonography on pregnant patients who are febrile. It would appear that if the embryonic temperature never exceeds 39°C, then there is no measurable risk. We suggest that sonography (the field) and sonogram (the procedure) are the most appropriate and least anxiety provoking terms.
Development of the lung in mice with bromodeoxyuridine‐induced cleft palate
Authors: Bannigan John G. , Cottell David C.
Published: June 2005

Clinical and laboratory observations show that denial of free communication between the amniotic fluid and lung fluid results in pulmonary hypoplasia. Thus, cleft palate resulting from tongue obstruction to palatal shelf elevation might be associated with disturbed lung development. This association exists in the Pena‐Shokeir phenotype. The goal of these experiments was to see what effect bromodeoxyuridine (BUdR)‐induced cleft palate had on lung development. LACA mice were injected with 500 mg/kg BUdR on E11 or E11 and E12 of gestation, a treatment known to produce a 25% and 50% incidence of cleft palate, respectively. BUdR had a direct retarding effect on lung growth but, when cleft palate occurred as well, the lungs were more severely affected. Morphometry showed that lungs from fetuses with cleft palate had only one‐half the saccular volume of controls or of treated fetuses with normal palates. Although hypoplastic, lungs associated with cleft palate had type I and type II pneumocytes, and the latter were shown by electron microscopy to be capable of producing surfactant. Hence, cellular differentiation had not been affected by the treatment.

Fetuses with cleft palate had less amniotic fluid than controls but significantly more than those with normal palates after treatment. Thus, the pattern of abnormalities in this animal model bears some resemblance to that of the human Pena‐Shokeir phenotype.
Bias in congenital malformations information from the birth certificate
Authors: Hexter Alfred C. , Harris John A.
Published: June 2005

An analysis of 1983 data from California birth certificates, and from the California Birth Defects Monitoring Program case registry, showed that there is a bias in reporting of congenital malformations on the birth certificate. Hospitals with many births erroneously report lower malformation rates than do hospitals with few births. The bias is partly due to the source of information; larger hospitals are more likely to get their information about malformations from the obstetrician than from the pediatrician. Since malformation data recorded on the birth certificate is both incomplete and biased, at present it is advisable to use these data for epidemiologic analyses with great caution.
Ultrastructure and function of the rat yolk sac: Damage caused by teratogenic anti‐VYS serum and recovery
Authors: Beckman David A. , Ornoy Asher , Jensen Marcela , Arnon Judi , Brent Robert L.
Published: June 2005

It was hypothesized that heterologous anti‐rat visceral yolk sac serum (AVYS) exerts its teratogenic effect by reducing the endocytosis of serum proteins by the visceral yolk sac (VYS), thus reducing the supply of amino acids to the embryo and VYS. To evaluate this hypothesis, we studied the effect of teratogenic AVYS on the endocytic function of the VYS and the ultrastructure of the VYS and parietal yolk sac (PYS). Rat conceptuses were exposed to a teratogenic dose of AVYS on the 10th day of gestation in vivo or in vitro. Control and AVYS‐exposed specimens were collected 24–192 hr later and prepared for scanning and transmission electron microscopy (SEM and TEM, respectively) utilizing standard procedures. The Endocytic Index was calculated for the VYS utilizing standard procedures. Approximately 97% of the in vivo exposed and 94% of the in vitro exposed embryos were morphologically abnormal. Ultrastructural observations showed that exposure to AVYS in vivo or in vitro caused severe damage to the VYS endodermal epithelial cells with loss of cellular borders, reduction in the number and length of microvilli, and increased cellular inclusions; and some damage to PYS endodermal cells with increased blebbling and decreased cell number. Recovery was evident at 72 hr and complete by 96 hr. The Endocytic Index was significantly reduced in the VYS 24 and 48 hr after injecting AVYS into the pregnant rat but was not significantly different at 96 and 192 hr. Our results show that the AVYS antiserum damaged visceral endodermal epithelium experienced ultrastructural recovery with parallel functional recovery. These studies suggest that transient yolk sac placental ultrastructural damage and dysfunction was probably sufficient to cause irreversible damage to the developing embryo during early organogenesis. We conclude that the proximate effect of the AVYS was on the plasma membrane of the visceral endoderm and that decreased pinocytosis is a consequence of this effect.
Effects of in vivo exposure of pregnant hamsters to glucose. 1. Abnormalities in LVG strain fetuses following intermittent multiple treatments with two isomers
Authors: Gale Thomas F.
Published: June 2005

The increased frequency of congenital malformations including caudal regression syndrome, in infants of women with insulin‐dependent diabetes mellitus is well documented. Most of the related animal research has involved the in vitro embryo culturing methodology. This study involved the alternative in vivo approach in order to determine the effects of treatment of pregnant hamsters with the D‐ and L‐isomers of glucose at five times just before and during the period of embryonic organogenesis on maternal blood glucose levels and the rates and types of fetal abnormalities. One group of animals was injected with 5 doses (4 g/kg each) of D‐glucose, i.e., on gestation day (D) 6, 3 PM; D7, 8 AM and 3 PM; D8, 8 AM and 3 PM. Two other groups were treated the same way but with L‐glucose (4 g/kg per dose) and water (10 ml/kg per dose), respectively. The D‐glucose treatment produced alternating periods of hyperglycemia and normoglycemia in the pregnant hamsters, enlarged placentae and fetuses with small urinary bladders, microphthalmia and skeletal abnormalities of the sternum, caudal vertebrae, pelvic bones, and femora. The L‐glucose treatment did not produce changes in maternal blood D‐glucose levels but did produce fetuses with small urinary bladders, microphthalmia and abnormal ossification limited to the manubrium. Several interpretations of the D‐glucose‐induced fetal abnormalities involving the vertebrae, proximal hindlimb bones and urinary bladders are discussed, including the consideration that this cluster has interesting similarities to the spectrum of skeletal and soft tissue abnormalities of human diabetes‐related caudal regression syndrome.
Pathogenesis of ethanol‐induced hydronephrosis and hydroureter as demonstrated following in vivo exposure of mouse embryos
Authors: Gage Jennifer C. , Sulik Kathleen K.
Published: June 2005

Urinary tract abnormalities have been noted to occur in 10–27% of individuals diagnosed as having Fetal Alcohol Syndrome. Among a wide range of functional and structural abnormalities, renal agenesis/hypoplasia, hydronephrosis, and ureteropelvic obstruction feature most prominently. This study was designed to examine the pathogenesis of ethanol‐induced urinary tract abnormalities in a mouse model. C57Bl/6J mice were acutely exposed to two doses of ethanol (2.9 g/kg IP) administered 4 hours apart beginning on gestational day (GD) 9, hour 4. This resulted in an incidence of 40.7% urinary tract anomalies among GD18 fetuses. With the exception of duplicate ureter, urinary tract abnormalities consisted exclusively of hydroureter/hydronephrosis. Examination of GD13–17 fetuses revealed that the first grossly detectable differences in the urinary tracts of control vs. affected specimens occurred on GD16 and initially only involved ureteral changes. Hydronephrosis was first detected on GD17. A contributing factor to the development of hydronephrosis appears to be the abnormal location of the ureterovesicle junction which commonly involves duplicate ureteral lumens with resultant functional obstruction to urine flow at the distal end of the ureter. Study of the early pathogenetic changes which appear to result in the urinary tract malformations observed involved utilization of scanning electron microscopy, vital dye (Nile blue sulphate) staining of whole embryos, and analysis of histological sections. These studies revealed that 12 hours following initial maternal ethanol exposure, embryos have excessive amounts of cell death localized in the region of the developing mesonephric duct just proximal to the cloaca. Also affected were premigratory neural crest cells located just proximal to the posterior neuropore. We conclude that excessive amounts of ethanol‐induced cell death in these selectively vulnerable populations could account for the subsequently observed urinary tract malformations.
Microinjection of cultured rat embryos: Studies with retinol, 13‐cis‐ and all‐trans‐retinoic acid
Authors: Lee Qwihee P. , Juchau Mont R. , Kraft Joan Creech
Published: June 2005

Retinol, all‐trans‐retinoic acid or 13‐cis‐retinoic acid were intraamniotically microinjected in rat embryos on day 10 of gestation and cultured until day 11.5. A comparison of the concentration–effect relationships of the retinoids showed that the dysmorphogenic effects were qualitatively similar for all three, but were elicited by a low concentration of all‐trans‐retinoic acid (250 ng/ml), a 6‐ to 7‐fold higher concentration of 13‐cis‐retinoic acid and an approximately 16‐fold higher concentration of retinol. After microinjection of 2,000 ng/ml of retinol, no dysmorphogenesis was observed but instead an increase in all growth parameters as compared to the controls.
Asymmetric development of mitochondrial activity in rat embryos as a determinant of the defect patterns induced by exposure to hypoxia, hyperoxia, and redox cyclers in vitro
Authors: Fantel A. G. , Person R. E. , Burroughs‐Gleim C. , Shepard T. H. , Juchau M. R. , Mackler B.
Published: June 2005

Previous study has shown that midorganogenesis‐stage rat embryos exposed to strong redox cyclers under moderate hypoxia in vitro develop severe necrotic defects on the right side. Similar effects can be produced by exposure to severe hypoxia alone. Studies presented here indicate that exposure to severe but survivable hyperoxia induces comparable necrotic degeneration on the left sides of all embryos. We hypothesize that the basis of these axially asymmetric defects is relatively precocious mitochondrial maturity on the left side of the embryo. In order to investigate this hypothesis, we compared mitochondrial oxygen utilization (NADH oxidase activities) on either side of rat embryos between days 11 and 14 of gestation. Activities were consistently higher on the left side during this period and significantly higher on day 11. We also found that the asymmetric embryotoxicity induced by niridazole, a strong redox cycler, could be attenuated by prior culture under hyperoxic conditions. We propose that mitochondrial immaturity on the right results in inadequate energy generation under hypoxic conditions, either directly or as a result of redox cycling. On the other hand, necrosis associated with hyperoxic conditions results from “leakage” of superoxide from functionally mature mitochondria on the left side.
Maternal and developmental toxicity of low doses of cytosine arabinoside in mice
Authors: Ortega Arturo , Puig Misericòrdia , Domingo José L.
Published: June 2005

1‐β‐D‐Arabinofuranosylcytosine (Ara‐C), an effective drug for the treatment of leukemia and breast cancer, was evaluated for developmental toxicity in pregnant Swiss mice. Ara‐C was administered by intraperitoneal injection on gestational days 6–15 at doses of 0, 0.5, 2, and 8 mg/kg/day. Maternal observations included clinical signs, body weight change, food consumption, and gross evaluation of organs and uterine contents at necropsy (day 18). Live fetuses were examined for external, visceral, and skeletal alterations. Maternal toxicity was observed at 2 and 8 mg/kg/day, as evidenced by a significant decrease in body weight gain and food consumption during the treatment period. Significantly increased early and late resorptions and reduced number of live fetuses per litter as well as decreased fetal body weight were observed at 8 mg/kg/day. At 2 mg/kg/day, the incidence of cleft palate, renoureteral agenesis or hypoplasia, and poly‐ or oligodactyly was significantly increased, whereas fetal weight was reduced at 0.5 mg/kg/day. Thus, the developmental no‐observed‐adverse‐effect‐level (NOAEL) of Ara‐C in the pregnant mouse is lower than 0.5 mg/kg/day, while the NOAEL for maternal toxicity is 0.5 mg/kg/day. We believe that exposure to this agent ought to be avoided during organogenesis.