bluuebunny/crossref_metadata_2025 huggingface dataset:
Vertical development of the secondary palate in hamster embryos following exposure to 6‐mercaptopurine
Authors: Burdett David N. , Waterfield J. Douglas , Shah Ravindra M.
Published: June 2005

Cellular aspects of vertical development of the secondary palate were examined in control and 6‐mercaptopurine (6MP)‐treated hamster embryos. Cross‐sectional area of the palatal shelf was measured and the numbers of both epithelial and mesenchymal cells counted. Also, in 6MP‐treated palates the damaged mesenchymal cells, characterized by the presence of dense bodies, were counted. DNA synthesis in both control and treated fetuses was measured by 3H‐thymidine incorporation. The results indicated that both the shelf area and cell numbers increased with age in control and 6MP‐treated palates. However, in controls the mesenchymal cell density and DNA synthesis showed two peaks that were absent following 6MP treatment. Unlike controls, in treated embryos the damage to mesenchymal cells became increasingly pronounced between days 10:00 and 10:12 but subsided by day 11:00 of gestation. It is suggested that a major force in the development of the initial primordia and early vertical development of the palatal shelf may be provided by a spurt of DNA synthesis in the mesenchymal cells resulting in their increased number. After 6MP treatment, depression of DNA synthesis and consequent reduction in the mesenchymal cell number and density followed by cell damage lead to retardation in the vertical development of the palatal shelves.
A method to increase the survival rate of early chick embryos in experiments involving surgical intervention
Authors: Bagnall K. M.
Published: June 2005

A method is presented by which the survival rate of early chick embryos following surgical procedure can be greatly improved. It was discovered that the embryos were dying at the stage of the operating procedure at which ink was being introduced under the embryo. Ink was used to visualize the embryonic features more easily. The use of a contrast medium in which the ink has been previously mixed with yolk in the ratio of 1:30 solved the problem.
Strain differences in the teratogenicity induced by sodium valproate in cultured mouse embryos
Authors: Naruse Ichiro , Collins Michael D. , Scott William J.
Published: June 2005

Strain differences in the teratogenicity of valproic acid (VPA) have been reported in mice. Finnell and Chernoff (Proc. Grnwd. Genet. Ctr. 5:162–163, 1985) showed that 300 mg/kg of VPA twice a day on days 6–8 of gestation induced exencephaly in 82% of SWV embryos but in 0% of C57BL/6J embryos. In the present experiment, we have collected similar results and investigated this strain difference using whole embryo culture in an attempt to determine whether maternal or embryonic factors are responsible for the difference. Mouse embryos were explanted on day 8.5 (plug day 0), and embryos at the 6–8‐somite stage were cultured for 48 hours in rat serum containing various doses of sodium valproate (NaVP). All the embryos died within 24 hours with 4.5‐mM and higher doses of NaVP in C57BL/6NCr1BR (C57) and with 3.0‐mM and higher doses in SWV. Unfused brain folds were recognized in embryos treated with 3.0‐mM and higher doses in C57, and with 1.0‐mM and higher doses in SWV. Irregular somite formation was observed in many embryos treated with 1.6‐mM and higher doses in C57 and with 1.0‐mM and higher doses in SWV. These results indicate that SWV embryos have 1.5–3 times the sensitivity of C57 embryos to the embryolethal and teratogenic effects of NaVP. Furthermore, the results suggest that the basis of the strain difference resides within the embryo rather than the mother.
Alterations in developing rat uterine cell populations after neonatal exposure to estrogens and antiestrogens
Authors: Branham William S. , Zehr David R. , Chen James J. , Sheehan Daniel M.
Published: June 2005

Exposure of rats to either estrogens or antiestrogens during early postnatal development reduces subsequent uterine growth as measured by uterine weight. However, individual uterine cell types respond differently to these agents and uterine weight alone cannot discern subtle or even large alterations in individual cell populations. Using a computerized planimetric technique, we estimated the prepubertal growth of the uterine luminal epithelium, endometrial stroma, glands, and circular and longitudinal muscle after exposure of neonatal rats (postnatal days 1–5) to the estrogens 17β‐estradiol (E2), diethylstilbestrol (DES), or ethynylestradiol (EE), and the antiestrogens tamoxifen or clomiphene citrate. On postnatal day 26, the cross‐sectional areas of the luminal epithelium, endometrial stroma, and circular muscle were reduced after estrogen exposure, compared to untreated controls, while longitudinal muscle cross‐sectional area was not affected. Since cell densities (cell number/unit area) were increased, these estrogen‐induced area reductions demonstrate a decrease in cell size. Total cell numbers, estimated as the product of cell type areas and their respective cell densities, were also reduced by neonatal estrogen exposure. The synthetic estrogens DES and EE were more potent than E2 with respect to reduction of uterine growth. Neonatal antiestrogen exposure caused large area reductions only in the uterine glands and luminal epithelium. Little change in cell density occurred in any cell population exposed to antiestrogen. These data demonstrate that the decreased uterine growth resulting from estrogen exposure during early postnatal development is a consequence of combined hypotrophy and hypoplasia in all cell types except longitudinal muscle while antiestrogen‐induced morphological alterations were limited to hypoplasia having epithelial cell specificity.
On “rates” of birth defects
Authors: Schulman Jane , Shaw Gary , Selvin Steve
Published: June 2005

The proportion of children born with a particular defect is not a “birth defect rate” but, rather, a prevalence proportion. The implications of confusing a rate and a proportion are discussed in terms of the interpretation of birth defect data. It is recommended that “prevalence proportion” or “prevalence” be used to report the frequency of various defects rather than the often‐used “prevalence rate”.
Embryotoxicity studies of norfloxacin in cynomolgus monkeys: I. Teratology studies and norfloxacin plasma concentration in pregnant and nonpregnant monkeys
Authors: Cukierski M. A. , Prahalada S. , Zacchei A. G. , Peter C. P. , Rodgers J. D. , Hess D. L. , Cukierski M. J. , Tarantal A. F. , Nyland T. , Robertson R. T. , Hendrickx A. G.
Published: June 2005

Norfloxacin, a new orally active antibiotic, was investigated in cynomolgus monkeys for potential developmental toxicity. Fifty seven monkeys were administered a control vehicle or norfloxacin by nasogastric gavage during the major period of organogenesis on gestational days (GD) 21 through 50 at doses of 0, 50, 100, 150, or 200/300 mg/kg/day. There was no evidence of teratogenicity at any dose level. Maternotoxicity and a significant increase in embryolethality occurred following doses of 200/300 mg/kg/day. The maternotoxicity was not expected based on rangefinding studies in nonpregnant female monkeys, which showed no signs of toxicity in doses up to 500 mg/kg/day. Additional studies were conducted to determine if norfloxacin caused similar toxicity later in gestation. Forty‐six pregnant monkeys were dosed with a control vehicle or 200 mg/kg/day norfloxacin for one of three 10‐day periods on GD 36–45, 71–80, or 111–120. There were no maternotoxic, embryotoxic, or fetotoxic effecs observed. Plasma concentrations of norfloxacin in five cynomolgus monkeys following 50 and 200 mg/kg oral doses were not dose‐proportionate. However, at a given dose, administered in cross‐over fashion, plasma concentrations of norfloxacin were higher in nonpregnant females (± 20–40%) than during pregnancy when the same subject was compared. At the no‐observed‐effect dose for maternal and embryotoxicity (50 mg/kg), peak plasma concentrations of norfloxacin in pregnant cynomolgus monkeys are approximately threefold higher than those observed in human volunteers receiving norfloxacin at the maximum recommended therapeutic dose of 400 mg (5.7 mg/kg based on 70 kg body weight) twice per day.
Teratogenicity of 3,3‐dimethyl‐1‐phenyltriazene in the rat: Gross malformations including micrognathism
Authors: Frank Anthony A. , Thompson Daniel J. , Kazacos Evelyn A.
Published: June 2005

3,3‐Dimethyl‐1‐phenyltriazene (DMPT), an alkylating agent, has been reported to be teratogenic in chickens, mice, and rats. One of the most commonly affected structures is the mandible; however, a complete description and incidence rates of all malformations produced have not been published. Rats were treated on day 12 of gestation (day 0 = sperm in vaginal smear) with a single intraperitoneal injection of 30 mg DMPT/kg. Fetuses were examined on each subsequent day of gestation for external and skeletal abnormalities. Standard soft tissue examinations were performed on day‐20 fetuses. A high percentage (≥80%) of treated litters contained numerous fetuses with micrognathism, cleft palates, syndactyly, adactyly, and misshapen digits and limbs. Lordosis, cerebellar and cerebral hypoplasia, decreased fetal size, and generalized delayed ossification were also observed. Dams exposed to DMPT had decreased food consumption and weight gains, although clinicopathologic and histopathologic evaluations failed to indicate other evidence of maternal toxicity. DMPT caused numerous permanent structural alterations that were not attributed to maternal toxicity.
Studies of mechanisms of niridazole‐elicited embryotoxicity: Evidence against a major role for covalent binding
Authors: Fantel A. G. , Juchau M. R. , Tracy J. W. , Burroughs C. J. , Person R. E.
Published: June 2005

Studies reported here were designed to examine the hypothesis that covalent binding of reactive intermediates to macromolecules of the conceptus represents a major mechanism for the embryotoxicity of niridazole (NDZ). The roles of embryonic thiol content and oxygenation on: (1) malformation incidence; (2) reductive metabolism; and (3) covalent binding to embryonic macromolecules of metabolites resulting from reductive biotransformation of NDZ were studied. Results were compared with those from studies with the nondysmorphogenic analog of NDZ, 4′‐methylniridazole (MNDZ). Day 10 rat embryos were pretreated for 5 hours in vitro with either L‐buthionine‐S, R‐sulfoximine (BSO) or N‐acetylcysteine (NAC) to modulate their glutathione (GSH) content. BSO reduced GSH levels, but NAC was ineffective. Following pretreatment, embryos were cultured for an additional 15 hours in the presence of [14C]NDZ or [14C]NDZ with an initial oxygen concentration of 5%. At the end of the culture period (day 11, AM), those embryos with active heartbeat and vitelline circulation were examined for asymmetric malformations. Drug metabolites were subjected to multiple extractions from the culture medium and subjected to quantitative high‐performance liquid chromatography (HPLC) analysis. Homogenates of the embryos were extracted with trichloroacetic acid (TCA) to estimate the covalent binding of radiolabeled parent compound/metabolites. Autoradiographic analyses were performed on other embryos. BSO pretreatment, which reduces embryonic GSH tissue levels, dramatically increased both the conversion of NDZ to 1‐thiocarbamoyl‐2‐imidazolidinone (TCI) (generated via reductive metabolism of NDZ) and covalently bound label but failed to increase embryotoxicity. NAC, by contrast, did not significantly affect embryonic GSH levels, TCI generation, or covalent binding. Because both rates of metabolism of NDZ to TCI and covalent binding could vary independently of malformation incidence, we concluded that they do not represent critical mechanistic factors for the embryotoxicity of NDZ and related nitroheterocycles.
Cell cycle analysis in the cardiac and neuroepithelial tissues of day 10 rat embryos and the effects of phosphoramide mustard, the major teratogenic metabolite of cyclophosphamide
Authors: Mirkes Philip E. , Ricks Jerry L. , Pascoe‐Mason Julie M.
Published: June 2005

Using the autoradiographic technique of percent labeled mitoses (PLM), total cell cycle length (Tc) and lenght of the cell cycle phases G1, S, G2, and M have been estimated for the neuroepithelium and heart of day 10 rat embryos. The Tc for the neuroepithelium was estimated to be 9.45 hr, consisting of G1 of 1.24 hr, S of 6.15 hr, G2 of 1.34 hr, and M of 0.72 hr. The Tc for the heart was estimated to be 13.37 hr, consisting of G1 of 4.30 hr, S of 7.01 hr, G2 of 1.81 hr, and M of 0.25 hr. Comparison of cell cycle parameters in these two tissues indicates that the longer cell cycle in heart tissue is related primarily to an increase in G1. If embryos are exposed to the major teratogenic metabolite of cyclophosphamide, phosphoramide mustard, cell cycle analysis reveals that the S phase of the neuroepithelial cell cycle is lengthened and cells are either slowed or arrested in G2.
Evaluation of the bioeffects of prenatal ultrasound exposure in the cynomolgus macaque (Macaca fascicularis): II. Growth and behavior during the first year
Authors: Tarantal A. F. , Hendrickx A. G.
Published: June 2005

The extensive use of ultrasonography for the prenatal assessment of growth and development continues to present questions regarding biological effects. We are currently evaluating a nonhuman primate model (Macaca fascicularis) exposed to ultrasound from gestational day (GD) 21 to 152 ± 2. Exposures were performed with a commercial real‐time sector scanner (ATL, MK 600); animals were scanned five times weekly on GD 21–35 ± 2, three times weekly on GD 36–60 ± 2, and once weekly on GD 61–150 ± 2. The length of exposure was approximately the same as human exposure (GD 21–60 ± 2 = 10 min/exam and GD 61–150 ± 2 = 20 min/exam) although the frequency of the examinations was considerably greater. Initial reports indicated differences between control and treated animals including lower birth weight, higher simian Apgar scores, and changes in select hematologic parameters. Follow‐up evaluations of growth during the first year included measurements of body weight, hand and foot lengths, humerus and femur lengths, biparietal and occipitofrontal diameters, head circumference, arm circumference, chest circumference, skinfold thickness, and crown‐rump length. Results indicated a significant reduction in body weight in treated animals during the first three months, with nonsignificant differences during the following nine months. Hematologic analysis including complete blood counts (CBC) and clinical biochemistry at 6, 9, and 12 months of age were not significantly different. A series of behavioral evaluations including a neurobehavioral test battery (NBT) and tests assessing motor and cognitive skills were included. The NBT revealed increased muscle tone in treated animals at one, two, and four days. In an observation cage (week 1–14) more quiet activities were displayed by treated animals. Group differences in performance of motor and cognitive tasks were observed and may be attributable to agitation and difficulties in adjusting to test environments. There were no group differences observed in discrimination learning. When considering the possible implications to the human population, it is important to consider the amount of exposure these animals received, and the fact that most of the effects observed appeared to be transitory. Although human epidemiological studies have not revealed any significant bioeffects, the “prudent use” of diagnostic ultrasound should still be kept in mind. This is especially significant with the current rise in the use of endovaginal scanning and pulsed Doppler.
Teratogenicity of zinc deficiency in the rat: Study of the fetal skeleton
Authors: Da Cunha Ferreira R. M. C. , Marquiegui I. Monreal , Elizaga I. Villa
Published: June 2005

Zinc deficiency (ZD) is teratogenic in rats, and fetal skeletal defects are prominent. This study identifies fetal skeletal malformations that affect calcified and non‐calcified bone tissue as a result of gestational zinc deficiency in rats, and it assesses the effect of maternal ZD in fetal bone calcification. Pregnant Sprague‐Dawley rats (180–250 g) were fed 1) a control diet (76.4 μg Zn/g diet) ad libitum (group C), 2) a zinc‐deficient diet (0 μg/g) ad libitum (group ZD), or 3) the control diet pari‐fed to the ZD rats (group PF). On day 21 of gestation, laparotomies were performed. Fetuses were weighed, examined for external malformations, and stained in toto with a double‐staining technique for the study of skeletal malformations. Maternal and fetal tissues were used for Zn, Mg, Ca, and P determinations. Gross external malformations were present in 97% of the ZD fetuses. No external malformations were found in fetuses from groups C and PF. Ninety‐one percent of cleared ZD fetuses had multiple skeletal malformations, whereas only 3% of the fetuses of group PF had skeletal defects; no skeletal malformations were found in fetuses from group C. Some of the skeletal malformations described in the ZD fetuses, mainly affecting non‐calcified bone, were not mentioned in previous reports, thus stressing the importance of using double‐staining techniques. Examination of stained fetuses and counting of ossification centers revealed important calcification defects in ZD fetuses. These effects were confirmed by lower Ca and P concentrations in fetal bone with alteration of the Ca:P ratio.
Effects of dinocap on otolith development: Evaluation of mouse and hamster fetuses at term
Authors: Rogers John M. , Burkhead Lesley M. , Barbee Brenda D.
Published: June 2005

The morphology of otoliths in CD‐1 mouse and Syrian hamster fetuses exposed to the fungicide dinocap were evaluated at the end of gestation. Pregnant mice were dosed by gavage with 0, 10, 15, 30, or 60 mg/kg/day dinocap in corn oil on days 7–16 of gestation. Pregnant hamsters were dosed by the same route with 0, 50, 100, or 200 mg/kg/day on days 7–14 of gestation. At the end of gestation (day 18 in mice, day 15 in hamsters) dams were killed and all fetuses were removed and fixed overnight in 70% ethanol. Fetal heads were then removed, left in 70% ethanol for at least 3 days, and then dehydrated in a graded ethanol series and cleared with methyl salicylate. Otoliths were examined by darkfield microscopy, and each otolith was scored for morphological completeness on a scale of 0 to 3. Otolith development was complete by day 18 of gestation in control mouse fetuses. Otolith development was complete in many, but not all, of the hamster fetuses by day 15 of gestation. In the mouse, dinocap exposure inhibited fetal otolith formation in a dose‐related manner, with a significant effect on total otolith score occurring at 10 mg/kg/day and above. Dinocap affected otolith formation in the hamster only at 100 mg/kg/day (200 mg/kg/day was embryolethal), concommitant with severe maternotoxicity and fetotoxicity. The results of this study indicate that the CD‐1 mouse is more sensitive than the Syrian golden hamster to the effects of dinocap on otolith development, and that otolith morphology can be easily examined in the mouse and hamster fetuses at term, although not by techniques currently used for fetal examination in standard teratology bioassays (e.g., Segment II studies).
Protective effect of ouabain on adriamycin‐induced cardiovascular anomalies in chick embryos
Authors: Takagi Yasuo , Bruyere Harold J. , Nishikawa Toshio
Published: June 2005

Adriamycin (2.5–10.0μg) was administered to 41/2 and 5 day embryonic chicks (Hamburger‐Hamilton developmental stages 24–26) to investigate the effect of the drug on cardiovascular morphogenesis. The drug produced dose‐related increases in both mortality rate and malformation frequency with a maximum incidence of 82% cardiovascular anomalies following a dose of 10.0 μg/egg (P < .001 relative to saline controls). Frequencies of embryos with ventricular septal defect (P < .005), dextroposition of the aorta (P < .005), or aortic arch anomalies (P < .05) were significantly higher than among controls. In a second study, embryos were pretreated with ouabain (12.2 μg), verapamil (0.5 μg), coenzyme Q10 (100 μg, 200μg), or vitamin E (1.0 mg, 5.0 mg)—agents previously shown to protect against adriamycin‐induced cardiotoxicity. Pretreatment of embryos with ouabain significantly reduced the incidence of cardiovascular malformations induced by adriamycin from 55 to 21% (P < .05). A major protective effect was observed relative to the induction of ventricular septal defect, the frequency of which was reduced from 45 to 14% (P < .05). However, administrations of verapamil, coenzyme Q10, or vitamin E did not have an appreciable effect on adriamycin‐induced frequencies of cardiovascular malformations. Negative inotropism is suggested as a mechanism for adriamycin‐induced cardiac anomalies but warrants further study.
Differential neuronal loss following early postnatal alcohol exposure
Authors: Pierce Dwight R. , Goodlett Charles R. , West James R.
Published: June 2005

Neonatal rats were exposed to 6.6 g/kg of alcohol each day between postnatal days 4 and 10 while artificial‐rearing procedures were used, in a manner which produced high peak and low trough blood alcohol concentrations each day. Gastrostomy controls were reared artificially with maltose/dextrin isocalorically substituted for alcohol in the milk formula, and suckle controls were reared normally by dams. The pups were sacrificed on day 10 and tissue sections (2 μm thick) were obtained in the sagittal plane through the cerebellum and in the horizontal plane through the hippocampal formation. Overall area measures were obtained for the hippocampus proper, area dentata, and cerebellum, along with areas of the cell layers of these regions. In the hippocampal formation, cell counts were made of the pyramidal cells of the hippocampus proper, the multiple cell types of the hilus, and the granule cells of the area dentata. In the cerebellum, cell counts of Purkinje cells, granule cells of the granular layer, granule cells of the external granular layer, and mitotic cells of the external granular layer were obtained from lobules I, V, VII, VIII, and IX. Alcohol selectively reduced areas and neuronal numbers in the cerebellum but had no significant effects on neuronal numbers in the hippocampal formation. Purkinje cells exhibited the greatest percent reductions, and cerebellar granule cells were significantly reduced in the granular layer but not in the external granular layer. All lobules showed these effects, but lobule I was significantly more affected than the other four lobules that were analyzed. The results demonstrate the differential vulnerability of selected neuronal populations to the developmental toxicity of alcohol exposure during the brain growth spurt.
Enhancement of murine phenytoin teratogenicity by the gamma‐glutamylcysteine synthetase inhibitor L‐buthionine‐(S, R)‐sulfoximine and by the glutathione depletor diethyl maleate
Authors: Wong Mayme , Helston Lori M. J. , Wells Peter G.
Published: June 2005

The teratogenicity of phenytoin may result from its enzymatic bioactivation to a reactive intermediate, which, if not detoxified, can interact with embryonic tissues and alter development. Glutathione (GSH) is an important cofactor/substrate for many physiological processes and for the detoxification of xenobiotic reactive intermediates. This study examined the effects of the GSH depletor diethyl maleate (DEM) and the GSH synthesis inhibitor L‐buthionine‐(S,R)‐sulfoximine (BSO) on phenytoin embryopathy. Phenytoin, 55 mg/kg, was administered intraperitoneally (ip) to pregnant CD‐1 mice at 0900 hr on gestational days 12 and 13. Pretreatment with DEM, 150 or 300 mg/kg ip, enhanced the incidence of phenytoin‐induced cleft palates by 3.3‐fold and 2.3‐fold, respectively (P < 0.05), without affecting the incidence of resorptions, postpartum death, or mean fetal weight. BSO, 1,800 mg/kg ip, given 0.5 hr prior to phenytoin, resulted in a 2.4‐fold increase in postpartum lethality and a 5‐fold increase in fetal weight loss (P < 0.05), without altering the incidence of resorptions or cleft palates. In two subsequent studies, BSO, 680–1,018 mg/kg/day, was given in the drinking water on gestational days 9 to 13 in the first study and on days 10 to 14 in the second study. Phenytoin, 55 mg/kg ip, was given on days 11 and 12 and on days 11 to 13 in the respective studies. In the first drinking water study, BSO enhanced the incidence of phenytion‐induced fetal resorptions 3.8‐fold and cleft palates 3.3‐fold (P < 0.05) but did not affect postpartum death. In the second study, BSO enhanced the incidence of resorptions, cleft palates, and postpartum death by 2‐fold, 2.6‐fold, and 1.7‐fold, respectively (P < 0.05). In both of the latter two studies, phenytoin‐induced fetal weight loss was altered by BSO treatment (P < 0.05). BSO alone had no embryopathic effects. These results suggest that GSH may be involved in the detoxification of a reactive intermediate of phenytion and/or in fetal cytoprotection.
Growth of the secondary palate in the hamster following hydrocortisone treatment: Shelf area, cell number, and DNA synthesis
Authors: Shah Ravindra M. , Chen Yung Ping , Burdett David N.
Published: June 2005

The contribution made by mesenchymal cells during the later stages of palatal development was examined in control and hydrocorti‐sone‐treated hamster embryos. Cross‐sectional area of the palatal shelf was measured, and the numbers of both epithelial and mesenchymal cell were counted. DNA synthesis was measured by 3H‐thymidine incorporation and was usd as an index of growth by cell proliferation. The observations in controls indicated that, unlike development during the initial 24 hr, the later period of vertical palate development, followed by reorientation of shelves and their closure, was characterized by a steady level of mesenchymal cell number and palatal shelf area. An absence of corresponding growth in the epithelial cell number suggests that the cells may accommodate the growth either by increasing their size and/or by stretching along the basal lamina. Hydrocortisone treatment did not alter the growth pattern of cell numbers or shelf area. However, it prevented the fusion between the opposing shelves, perhaps by affecting the cytodifferentiation of the palatal tissues. Although a continuous increase in the number of mesenchymal cells during the latter half of vertical shelf development, i.e., between days 11:00 and 12:00 of gestation, is not required for reorientation and fusion of the shelves, it is not clear from the data from the present study whether a critical number of cells and/or cell density is essential for reorientation and fusion of the palate. It was suggested that, for normal palatal development, information on cell cycle and positioning of mesenchymal cells within the shelf during the vertical development may be crucial for further understanding of subsequent events of palatogenesis.
Developmental exposure of mice to pulsed ultrasound
Authors: Kimmel Carole A. , Stratmeyer Mel E. , Galloway W. Don , Brown Natalie T. , Laborde James B. , Bates Hudson K.
Published: June 2005

Exposure of pregnant mice on gestation day (gd) 8 to 1 MHz continuous‐wave ultrasound (0, 0.05, 0.50, or 1.00 W/cm2) was reported previously to result in a slight (nonsignificant) increase in malformations. The present study was conducted in a similar fashion using pulsed ultrasound but was designed to maximize the likelihood of finding effects of gd 8 ultrasound exposure on prenatal development. Pregnant ICR mice (approximately 60 animals/group) were exposed on gd 8 to pulsed ultrasound with a center frequency of 1 MHz at levels of 0 (sham control), 0.05, 0.50, or 1.00 W/cm2 (spatial average, temporal average intensities; ISATA) with a spatial peak, pulse average intensity (ISPPA) of 90 W/cm2 and pulse duration of 6.5 μsec. Anesthetized animals were placed in a degassed water bath (30°C) and exposed for two 10 min intervals during which the beam was centered 1 cm on either side of the abdominal midline. On gd 17, dams were killed; the uterus and its contents were weighed and examined; and live fetuses were weighed and examined for external, visceral, and skeletal malformations. Although one female in the 0.50 W/cm2 group and seven animals in the 1.00 W/cm2 group died following exposure, no other significant change from controls was seen in any maternal or fetal parameter evaluated. Thus the results of this study indicate that there was no detectable effect on prenatal development of mice following exposure to ultrasound on gd 8 (a time of maximal sensitivity), even at exposure intensities that were lethal to some maternal animals.
Prenatal dexamethasone administration disrupts the pattern of cellular development in rat lung
Authors: Navarro Hernan A. , Kudlacz Elizabeth M. , Eylers John P. , Slotkin Theodore A.
Published: June 2005

To examine whether prenatal exposure to glucocorticoids could adversely affect subsequent cellular development of the lung, we administered 0.2 mg/kg of dexamethasone to pregnant rats on gestational days 17, 18, and 19. Lungs of the offspring were then examined for patterns of cell acquisition (DNA) and growth (protein). DNA concentration (a marker of cell packing density) and DNA content (a measure of total cell numbers) were reduced during gestation, and the shortfalls in concentration persisted past weaning. Disruption of development was also apparent in the protein/DNA ratio, which was consistently elevated, a finding consistent with cellular hypertrophy. In addition, lung, ODC became coupled to β‐adreñergic receptors prematurely in the dexamethasone group, suggesting that neural control of tissue differentiation is altered. These data indicate that prenatal glucocorticoids may compromise lung development through effects on cell replication and differentiation, which derive, in part, from alterations in the reception of trophic neural signals.
Teratogenicity of 3,3‐dimethyl‐1‐phenyltriazene in the rat: Histopathologic and ultrastructural alterations
Authors: Frank Anthony A. , Kazacos Evelyn A. , Hullinger Ronald L. , Thompson Daniel J.
Published: June 2005

3,3‐Dimethyl‐1‐phenyltriazene (DMPT) is a methylating agent which is teratogenic, carcinogenic, and mutagenic. A single intraperitoneal injection of 30 mg DMPT/kg given to pregnat rats on day 12 of gestation produces malformations with minimal maternal toxicity. Malformations include skeletal deformities such as micrognatism, cleft palate, and digital malformations, as well as central nervous system hypoplasia. The purpose of the present study was to characterize the light and electron microscopic alterations produced by DMPT. Electron microscopy (EM) revealed that at 4 hr postinjection of DMPT, rare cells of the neural tube contained few membrane‐bound aggregations of organelles and condensed chromatin; this change was consistent with apoptosis, a type of cell death characterized by morphologic and biochemical alterations distinct from necrosis. At 8 hr postinjection, apoptosis was more prominent in the neural tube and also observed in the mandibular process. At 16 hr postinjection, numerosu apoptotic cells were interspersed with unaffected cells that contained phagocytized apoptotic bodies. Light microscopic examination of DMPT‐exposed conceptuses showed apoptosis in the neural tube at 24 hr postinjection. Forty‐eight hours postinjection, apoptosis, in decreasing order of severity, was observed in the neural tube, craniofacial processes, limb buds, and somites and liver. Apoptosis was absent in all tissues by 72 hr postinjection. Nervous tissues failed to achieve proper histologic organization, but all other tissues appeared microscopically normal from 72 hr postinjection until the end of gestation. There appeared to be some degree of tissue specificity to the effects of DMPT.
Altered placental morphology associated with murine trisomy 16 and murine trisomy 19
Authors: Bersu Edward T. , Mossman Harland W. , Kornguth Steven E.
Published: June 2005

The morphology of placentas from trisomy 16 and trisomy 19 mouse conceptuses aged 12 to 18 gestational days was studied at the light microscopic level. Comparisons were made with placentas from normal littermate animals. Trisomy 16 placentas showed marked changes from normal: 1) the junctional zone showed little indication of normal morphologic differentiation throughout gestation; 2) clusters of germinal trophoblast cells persisted in the labyrinth throughout gestation, whereas these cells disappeared by gestational day 16 in the normal littermate placentas; 3) the labyrinth was reduced in size in the trisomic placentas, and the differentiation of the interhemal membranes was delayed. The size of the labyrinths from trisomy 19 placentas appeared to be decreased, but otherwise the placentas appeared to have normal morphology. These observations and others from the literature show that placental development is affected by the presence of a trisomic genome, and that different trisomies influence the development of the placenta differently. Fortrisomy 16, we propose that the striking changes of the junctional zone may be associated with the trisomy 16‐related gene dosage effect for alpha‐ and beta‐interferon cell surface receptors. Because of the homology for this and other genes on mouse chromosome 16 with genes on human chromosome 21, findings related to the altered development of the trisomy 16 mouse may be relevant to understanding some of the phenotypic variations associated with human trisomy 21, the Down syndrome.
Human achondroplasia: Defective mitochondiral oxidative energy metabolism may produce the pathophysiology
Authors: Mackler Bruce , Shepard Thomas H.
Published: June 2005

A summary is presented of previous studies by other investigators of human achondroplasia and dyschondroplastic animal models. In addition, studies previously reported from our laboratories are discussed, and they demonstrate that defective oxidative energy metabolism is present in mitochondrial preparations from achondroplastic human subjects and rabbits (ac/ac) with chondrodystrophy. The results of the studies support the hypothesis discussed fully in the manuscript that a partial defect in mitochondrial oxidative metabolism in achondroplastic subjects is expressed specifically in the growth plates of the long bones because this tissue has the lowest oxygen tension of any bodily organ undergoing active proliferation, thus leading to the achondroplastic phenotype in humans and the ac/ac rabbit. In the ac/ac rabbit phosphorylation at the cytochrome c oxidase region (site III) of the terminal respiratory system was shown to be absent in mitochondrial preparations from the livers of newborn ac/ac rabbits. Normal‐appearing littermates did not exhibit the defect. Studies of mitochondrial preparations from human skin fibroblasts (grown in tissue culture) from normal human subjects and subjects with homozygous achondroplasia demonstrated that concentrations of cytochrome a3 were decreased approximately 80% in preparations from homozygous achondroplastic cells. Levels of cytochrome a3 in heterozygous achondroplastic cells were intermediate between the levels in normal cells and homozygous achondroplastic cells demonstrating the effects of gene dosage. Determination of total heme a (as the pyridine hemochromogen) in the normal and achondroplastic preparations from human subjects showed that the observed decrease in concentration of cytochrome a3 in the achondroplastic preparations was due to an absence of cytochrome a3 and not to a change in its absorbancy (extinction coefficient).
Effects of retinoic acid on the distribution of glycoconjugates during mouse tail bud development
Authors: May Griffith C. , Wiley Michael J.
Published: June 2005

Retinoic acid (RA), a potent teratogen of caudal axial development in rodents, has been shown to alter glycoconjugates in a variety of embryonic tissues and teratocarcinomas. In this study, we examined its effects on the expression of cell surface and extracellular matrix glycoconjugates during tail bud development in mouse embryos by using lectin histochemistry. The lectins WGA, sWGA, and PNA showed striking differences in binding between RA‐exposed and control embryos. Computer‐assisted densitometry revealed a significant increase in binding of all three lectins to the extracellular material of the luminal and abluminal borders of the secondary neural tube and surrounding the notochord in RA‐exposed embryos. RA‐treated embryos also showed an increased binding affinity for the lectins sWGA and PNA to the cells of the notochord, while WGA showed increased binding to the neuroepithelial cells of the secondary neural tube. The results suggest that RA affects the expression of lectin binding sites during the early development of RA‐induced caudal axial defects.
Heat‐shock induced tolerance to the embryotoxic effects of hyperthermia and cadmium in mouse embryos in vitro
Authors: Kapron‐Brás Carolyn M. , Hales Barbara F.
Published: June 2005

Mammalian embryos growing in vitro are harmed by short elevations in the culture temperature. However, a relatively mild hyperthermic exposure can induce thermotolerance, a transient state of resistance to the effects of a subsequent heat exposure. The present study examines the induction of tolerance to heat and cross‐tolerance to another teratogen, cadmium, in day 8 CD‐1 mouse embryos in vitro. The ability of a mild heat pretreatment (5 min at 43°C) to partially protect embryos against an embryotoxic heat exposure (20 min at 43°C) was demonstrated. The frequency of death was reduced from 43% to 20%, abnormal branchial arches from 44% to 13.2%, and retarded turning from 22% to 5% in pretreated embryos. Other malformations, such as small forebrains and microphthalmia, were not affected, and the rate of exencephaly was significantly increased. The same heat pretreatment (5 min at 43°C) was also found to reduce the damaging effects of a subsequent exposure to 1.75 μM cadmium. In the absence of pretreatment, cadmium caused 55% embryo deaths and 87% malformations, but prior heat exposure caused significant reductions in these frequencies to 29% and 55%. The total morphological score was higher in the pretreated group, as were the measurements of the yolk sac diameter, crown‐rump length, and head length. Thus, embryos that have developed resistance to hyperthermia are also partially protected against the harmful effects of a second teratogen, cadmium. The response of the embryo to elevated temperatures may be involved in the development of tolerance to a variety of stresses.
Multiple antigens in the rat visceral yolk sac induce teratogenic antisera
Authors: Jensen Marcela , Lloyd J. B. , Vega Paz , Koszalka T. R. , Brent R. L.
Published: June 2005

Preparative isoelectric focusing was used to fractionate the supernatant from a homogenate of day 19 rat visceral yolk sac. Three fractions, of pI ranges 3.5–5.0, 5.0–7.0, and 7.0–9.0, were isolated and used to immunize rabbits, by four or six weekly injections, each containing 5 mg protein. The resulting antisera were all teratogenic when injected into rats on day 9 of gestation, but widely differing potencies were observed. The most potent antiserum was that against yolk sac components focusing in the pI 7.0–9.0 range: An optimum teratogenic dose of 50 mg protein per kg body weight was observed, and a dose of 100 mg/kg was shown to cause 100% embryonic resorption. Antiserum against the fraction focusing in the pI 3.5–5.0 range was the least teratogenic: A significant incidence of embryonic malformation and death was seen only at doses of 600 mg/kg and above. The two fractions that yielded the more teratogenic antisera were refocused over narrower pH ranges, yielding four subfractions in the pI 5.0–7.0 range and eight subfractions in the pI 7.0–9.0 range. Antisera against each of these 12 fractions were raised in rabbits; most of these antisera were shown to be teratogenic, although of differing potencies. It is concluded that the yolk sac contains many antigens that can elicit antibodies with teratogenic and yolk sac‐localizing properties.
Permanent neuronal deficits in rats exposed to alcohol during the brain growth spurt
Authors: Bonthius Daniel J. , West James R.
Published: June 2005

The purpose of this study was to determine whether developmental alcohol exposure could induce permanent neuronal deficits, whether the peak blood alcohol concentration (BAC) influences the severity of the effects, and whether the effects are gender related. Rat pups were reared artifically over postnatal days (PD) 4 through 11 (a period of rapid brain growth, comparable to part of the human third trimester). Alcohol treatments were administered on PD 4 through 9. Patterns of alcohol exposure that produce different peak BACs have been shown to affect differentially the amount of brain weight deficits and neuron loss shortly after the exposure period, so this study investigated whether the pattern of alcohol exposure was also effective in producing permanent deficits. Two groups received a daily alcohol dose of 4.5 g/kg, condensed into either four or two feedings. A third group received a higher daily alcohol dose of 6.6 g/kg administered in 12 uniformly spaced daily feedings. Pups were fostered back to dams on PD 11 and perfused on PD 90. Brain weights were measured, and Purkinje cells and granule cells were counted in each of the 10 lobules of the cerebellar vermis. In the hippocampal formation, cell counts were made of the pyramidal cells of fields CA1 and CA2/3, the multiple cell types of CA4 and the granule cells of the dentate gyrus. The groups receiving the lower daily dose (4.5 g/kg) condensed into either four or two feedings were exposed to higher peak BACs and suffered significant permanent brain weight deficits and neuronal losses, relative to controls. The group receiving the higher daily dose (6.6 g/kg) in continuous fractions had no significant brain weight reductions or neuronal loss. Vulnerability to alcohol‐induced neuronal loss varied among regions and cell populations and as a function of peak BAC. In the hippocampus, only the CA1 pyramidal cells were significantly reduced in number and only in group receiving the most condensed alcohol treatment. In the cerebellum, the severity of Purkinje cell and granule cell losses varied among lobules, and Purkinje cell vulnerability appeared to depend on the maturational state of the neuron at the time of the alcohol exposure, with the more mature Purkinje cells being the more vulnerable.
Involvement of the intestinal microflora in nitrazepam‐induced teratogenicity in rats and its relationship to nitroreduction
Authors: Takeno Satoshi , Sakai Takeshi
Published: June 2005

A study was undertaken to investigate the relationship between nitroreduction of nitrazepam and its teratogenic effects and the involvement of the intestinal microflora in Sprague‐Dawley rats. Incubation of bacterial suspensions from rat cecal contents with nitrazepam resulted in extensive reduction to 7‐aminonitrazepam. Rat liver homogenates also reduced nitrazepam but only under anaerobic conditions. Following oral administration of 300 mg/kg nitrazepam to pregnant rats, total excretion of reduced metabolites (7‐aminonitrazepam and 7‐acetylaminonitrazepam) in urine and feces accounted for approximately 30% of the administered dose. When antibiotics were administered to dams to deplete their intestinal microflora prior to administration to nitrazepam, the total excretion of the reduced metabolites in the urine and feces decreased to 2% of the dose. Nitroreductase activity of cecal contents was almost completely suppressed by antibiotic pretreatment, but the activity of liver homogenates was not significantly altered by the same treatment. The incidence of nitrazepam‐induced malformations was markedly decreased by antibiotic pretreatment. These results suggest that the intestinal microflora plays an important role in the reductive metabolism of nitrazepam and that the teratogenicity of nitrazepam may be related to its nitroreduction by the microflora.
Developmental toxicity of pentostatin (2′‐deoxycoformycin) in rats and rabbits
Authors: Dostal Lori A. , Brown Sandra , Bleck Jill , Anderson John A.
Published: June 2005

The developmental toxicity of the potent adenosine deaminase (ADA) inhibitor, pentostatin (2′‐deoxycoformycin), was investigated in pregnant rats and rabbits administered daily iv doses during organogenesis. Rats received 0, 0.01, 0.10, or 0.75 mg/kg on gestation days 6–15 and rabbits received 0, 0.005, 0.01, or 0.02 mg/kg on gestation days 6–18 and maternal and fetal parameters were evaluated on gestation day 21 (rats) or 30 (rabbits). Live fetuses were examined for external, visceral, and skeletal malformations and variations. In rats, maternal body weight gain and food consumption were significantly suppressed at doses of 0.10 and 0.75 mg/kg during the treatment period but returned to control levels during posttreatment. Increased postim‐plantation loss and decreased numbers of live fetuses, litter size, and fetal body weight were observed at 0.75 mg/kg. A statistically significant increase in the incidence of vertebral malformations occurred at 0.75 mg/kg. The incidence of certain skeletal variations (extra presacral vertebrae, extra ribs, hypoplastic vertebrae) was also increased at 0.75 mg/kg. Ossification of cervical centra was reduced at 0.75 mg/kg compared with controls. In rabbits, marked maternal toxicity (death, body weight loss, and decreased food consumption) and reproductive toxicity (abortion and premature delivery) occurred in all pentostatin‐treated groups. However, there were no significant effects on number of live fetuses, pre‐ or postimplantation loss, litter size, or fetal body weights in the animals with live litters. There was also no apparent increase in the incidence of malformations or variations in the live fetuses of pentostatin‐treated rabbits. Thus, these studies demonstrate developmental toxicity of pentostatin in rats and rabbits, and teratogenicity in rats, at maternally toxic doses.
Influence of paternal age, smoking, and alcohol consumption on congenital anomalies
Authors: Savitz David A. , Schwingl Pamela J. , Keels Martha Ann
Published: June 2005

The potential effects of paternal exposures on fetal development are of great public and scientific concern, yet few epidemiologic studies have examined this association. Single live births from 1959 to 1966 among 14,685 Kaiser Foundation Health Plan members who participated in the Child Health and Development Studies were analyzed to assess the impact of paternal age, cigarette smoking, and alcohol consumption on the occurrence of birth defects in the offspring. Prevalence odds ratios for anomalies identified by age 5 were analyzed, contrasting exposed to unexposed fathers with adjustment for maternal age, race, education, smoking, and alcohol use. Advanced paternal age was associated with increased risk of preauricular cyst, nasal aplasia, cleft palate, hydrocephalus, pulmonic stenosis, urethral stenosis, and hemangioma. Father's cigarette smoking was more common among children with cleft lip ± cleft palate, hydrocephalus, ventricular septal defect, and urethral stenosis. Alcohol use by the father was most positively related to the offspring's risk of ventricular septal defect. For both smoking and alcohol use, inverse associations were more common than positive associations. These data generally do not indicate strong or widespread associations between paternal attributes and birth defects. However, because of this study's imprecision, limited ability to isolate defects most likely to be of paternal origin, and the identification of several suggestive associations with age and smoking, further study of this issue would be of value.