bluuebunny/crossref_metadata_2025 huggingface dataset:
Evaluation of developmental toxicity of coniine to rats and rabbits
Authors: Forsyth Carol S. , Frank Anthony A.
Published: June 2005

Conium maculatum (poison hemlock, CM) is teratogenic in several domestic species, presumably due to its piperidine alkaloids, including coniine, which has been verified to be teratogenic in cattle. Coniine/CM teratogenicity culminates in production of arthrogryposis. The purpose of this study was to evaluate coniine‐induced teratogenicity in two laboratory animal species, Sprague‐Dawley rats and New Zealand white rabbits. Pregnant rats were given coniine (25 mg/kg body weight) by oral gavage at 8‐hour intervals on gestation days 16–18. Pregnant rabbits were given coniine (40 mg/kg body weight) by oral gavage at 8‐hour intervals on gestation days 20–24. Rats were killed on day 19 and rabbits on day 29. Fetuses were immediately removed, weighed, and examined for external abnormalities. Alternate fetuses were either stained for skeletal examinations with alizarin red‐S or fixed in Bouin's solution for visceral examination. Symptoms of maternal intoxication due to coniine administration were observed in both the rat and the rabbit, and higher doses were uniformly lethal. Rabbits treated with coniine appeared to lose more weight and eat less than controls, but there was no statistically significant difference between groups. Fetal weights were significantly lower in coniine‐exposed rat and rabbit fetuses indicating fetotoxicity. The only statistically significant treatment‐related visceral or skeletal malformation was a reduction of cranial ossification of rabbit fetuses, probably related to maternal toxicity. Coniine‐exposed rabbit litters tended to be affected by arthrogryposis (no bony deformities noted on skeletal exam) more than controls (2/6 vs. 0/9). The lesion was characterized as hyperflexion of the antebrachial‐carpal joint with supination. Although this was not statistically significant, the incidence rate was significantly (p < 0.01) increased when compared to historical controls. This study indicates coniine is not teratogenic to rats and weakly teratogenic to rabbits. The reason for the lack of sensitivity to teratogenicity in these laboratory animal species as compared to domestic species is currently unknown. © 1993 Wiley‐Liss, Inc.
Methyl methacrylate: Inhalation developmental toxicity study in rats
Authors: Solomon H. M. , McLaughlin J. E. , Swenson R. E. , Hagan J. V. , Wanner F. J. , O'hara G. P. , Krivanek N. D.
Published: June 2005

Methyl methacrylate (99.9% pure) was administered by vapor inhalation exposure to five groups (27 rats/group) of presumed pregnant rats (Crl:CD) at concentrations of 0 (control), 99, 304, 1,178, and 2,028 ppm for 6 hr/day on days 6–15 of gestation (G). Maternal body weight, feed consumption, and clinical signs were recorded throughout gestation. Dams were euthanized on day 20 G. Each uterus was weighed and corpora lutea, implantation sites and resorptions were counted. The number of fetuses per litter were counted and their location within the uterus recorded. All fetuses were weighed, sexed and examined for external and skeletal alterations. One half of the fetuses from each litter were examined for visceral alterations. No treatment‐related deaths were noted at any concentration tested. Treatment‐related effects on maternal body weight and feed consumption were noted at all exposure levels. The decreases in maternal body weight at 99 and 304 ppm were minimal and transient since they returned to control values by the next weighing period. When exposure was discontinued, body weight gain and feed consumption in all exposure groups returned to control values. There were no treatment‐related changes in the number of litters produced or in the mean number per litter of corpora lutea, implantations, resorptions, live or dead fetuses, or sex ratio. Fetal body weights were similar between the control and treated groups. There were no treatmentrelated increases in the type or incidence of external, visceral, or skeletal malformations, developmental variations, or variations indicative of retarded development. Exposure to methyl methacrylate concentrations up to 2,028 ppm resulted in no embryo or fetal toxicity or malformations even at exposure levels that resulted in maternal toxicity. © 1993 Wiley‐Liss, Inc.
Sodium valproate‐induced cardiovascular abnormalities in the Jcl:ICR mouse fetus: Peak sensitivity of gestational day and dose‐dependent effect
Authors: Sonoda Tohru , Ohdo Shozo , Ohba Ken‐Ichi , Okishima Takahiro , Hayakawa Kunio
Published: June 2005

Sodium valproate was administered to Jcl:ICR mice in order to determine its effect on cardiovascular development. A single dose of 600 mg/kg of sodium valproate was injected intraperitoneally on gestational days 6, 7, 8, or 9. In same animals, a single dose of 300, 400, 500, or 700 mg/kg was injected on gestational day 7. On day 18 of gestation, dams were laparotomized to observe number of live, dead and resorbed fetuses. In addition, live fetuses were examined for cardiovascular abnormalities. Although cardiovascular abnormalities were noted in 3% of live fetuses and in 26% of litters in the group treated on day 6 (600 mg/kg), there was no significant difference from the control group, suggests that there may have been a biologically significant increase, although not a statistically significant increase. Cardiovascular abnormalities were found in 30%, 11%, and 8% of live fetuses in the groups treated with 600 mg/kg on days 7, 8, and 9, respectively. These represented a statistically significant increase in effects as opposed to the control groups. Among the varying dosages administered on day 7 of gestation, cardiovascular abnormalities occurred in 2%, 6%, 16%, and 36% of live fetuses in groups treated with 300, 400, 500, and 700 mg/kg, respectively, showing a significant dose‐dependent increase. These cardiovascular abnormalities observed were divided into the following groups: ventricular septal defect, endocardial cushion defect, transposition of the great arteries, doubleoutlet right ventricle, tricuspid atresia, and hypoplastic left heart syndrome. Maternaldeath did not occur at any treatment level. © 1993 Wiley‐Liss, Inc.
Prevention by Tiron (sodium 4,5‐dihydroxybenzene‐1,3‐disulfonate) of vanadate‐induced developmental toxicity in mice
Authors: Domingo Jose L. , Bosque M. Angeles , Luna Margarita , Corbella Jacinto
Published: June 2005

Vanadate is embryotoxic and fetotoxic in golden hamsters, mice and rats. Tiron (sodium 4,5‐dihydroxybenzene‐1,3‐disulfonate), a chelating agent widely used in analytical chemistry, is an effective antidote in the treatment of oral or parenteral vanadate poisoning. The present study evaluated the effect of administration of Tiron on sodium metavanadate (NaVO3)‐induced developmental toxicity in mice. NaVO3 (25 mg/kg, i.p.) was injected on day 12 of gestation, whereas Tiron was injected subcutaneously at 0, 24, 48, and 72 hr after NaVO3 administration. Tiron effectiveness was assessed at dosage levels of 0, 250, 500, and 1,000 mg/kg. Cesarean sections were performed on gestation day 18. All live fetuses were examined for external, internal, and skeletal malformations and variations. Amelioration by Tiron of NaVO3 developmental toxicity was evidenced by a significant decrease in the number of resorbed fetuses, an increase in the mean fetal weight, and a reduction in the incidence of the skeletal variations caused by NaVO3. According to these results, Tiron offers encouragement with regard to its therapeutic potential for pregnant women exposed to vanadate. However, further investigations, including the effect of increasing the time interval between acute vanadate exposure and initiation of Tiron therapy, are required. © 1993 Wiley‐Liss, Inc.
Retinoic acid induces changes in the rhombencephalic neural crest cells migration and extracellular matrix composition in chick embryos
Authors: Balbás J. A. Moro , Gato A. , Revuelta M. I. Alonso , Pastor J. F. , Represa J. J. , Barbosa E.
Published: June 2005

Chick embryos at 9–10 stages (Hamburger and Hamilton: J Morphol 88:49–82, 1951) have been treated with all‐trans retinoid acid (RA) (0.5 μg, 1.5 μg, and 2.5 μg) to determine the pattern and mechanism of RA‐induced effects on early cephalic development. We found that while 0.5 μg RA did not produce any significant dysmorphogenesis, 2.5 μg RA elicited wide malformation of both cephalic and trunk regions. However, 1.5 μg RA produced selective and specific changes at the cephalic level, which consisted of morphological alterations, changes in neural crest cells (NCC) migration and extracellular matrix (ECM) composition. Morphological alterations included hypoplasia of the first three branchial arches, swelling of either anterior cardinal veins or dorsal aortae, and atrophy of branchial arch arteries. Concurrently NCC did not migrate away, remaining clustered on the dorsal surface of the rhombencephalon, and in some cases they shifted into the neural tube cavity. Accordingly, the second branchial arch showed a reduction of the mesenchymal cellular population. The extracellular matrix in RA‐injected embryos showed changes in glycosaminoglycans (GAGs) concentration as compared with controls, that is, an increase in the non‐sulphated GAGs, stained with alcian blue 8GX at 2.5 pH, and a decrease in the sulphated GAGs stained with alcian blue 8GX at 1 pH. These quantitative changes reflected alterations in the pattern of distribution and composition of the GAGs within the cephalic ECM, which specifically consisted in an increase of the hyaluronic acid and a decrease of the chondroitin sulphate. Our findings indicate that RA is involved in abnormal cephalic development, suggesting that RA may effect neural crest cell migration via changes in the GAGs of the ECM. © 1993 Wiley‐Liss, Inc.
Cadmium‐induced inhibition of proliferation and differentiation of embryonal carcinoma cells and mechanistic aspects of protection by zinc
Authors: Piersma A. H. , Roelen B. , Roest P. , Haakmat‐Hoesenie A. S. , Van Achterberg T. A. E. , Mummery C. L.
Published: June 2005

Murine embryonal carcinoma cells have been used in in vitro models to study the effects of cadmium chloride on proliferation and differentiation of early embryonic cells. This approach allows the various cell types within the early embryo as well as several developmental mechanisms to be dissected and studied in isolation using larger numbers of cells than would be readily available from the embryo itself. The present study shows that both embryonal carcinoma cell proliferation and differentiation into parietal endoderm are inhibited by cadmium chloride. The effects are counteracted by the additional presence of zinc chloride. The uptake of cadmium into the cells is inhibited in the presence of zinc chloride, suggesting that competition between these metals for passage into the cells contributes to the mechanism underlying the protective effect of zinc. In addition, metallothionein gene expression is enhanced more rapidly after simultaneous incubation with zinc chloride, indicating that the attenuating effect of zinc on cadmium toxicity is also partly attributable to detoxification by metallothioneins. © 1993 Wiley‐Liss, Inc.
Real time micro‐fiberoptic monitoring of endogenous fluorescence in the rat conceptus during hypoxia
Authors: Thorsrud Bjorn A. , Harris Craig
Published: June 2005

A micro‐fiberoptic methodology has been developed for noninvasive, real time measurement of endogenous pyridine nucleotide fluorescence from the surface of the visceral yolk sac (VYS) in intact, viable rat conceptuses. Gestational day (GD) 10–12 conceptuses are maintained in a customized perifusion system, which allows for control of oxygenation, as well as the continuous measurement of pH and oxygen concentration in the effluent perifusate. Miniaturized light guides were constructed by drawing 250 μm ESKA acrylic optical fibers through a stainless steel sheath with a high strength epoxy polymer. A single fiber supplied the excitation signal from a mercury arc lamp at a wavelength of 366 nm. The emission signal was returned via three additional fibers, electronically amplified, processed, and recorded, using a dual channel lamp‐compensated fluorometer, optimized for detection of reduced pyridine nucleotides at 455 nm. Endogenous fluorescence in the conceptus was monitored by placing the polished tip of the sensor directly on the surface of the VYS. Oxygen‐equilibrated conceptuses, exposed to 100% nitrogen, produced a reproducible biphasic surface fluorescence peak, which returned to baseline levels upon reoxygenation of the perifusate. This biphasic response consisted of an initial rapid rise in fluorescence (phase I), followed by an attenuated rate in fluorescence signal increase (phase II). The hypoxia produced age‐dependent rates of fluorescence change during phase I, while phase II remained relatively unchanged throughout GD 10–12. These results demonstrate the ability to monitor endogenous fluorescence, non‐invasively and in real time, during the period of organogenesis in the intact rat conceptus and will provide valuable information in studies of embryonic metabolism and response to chemical embryotoxicants. © 1993 Wiley‐Liss, Inc.
Effect of in utero exposure to diagnostic ultrasound on the postnatal survival and growth of mouse
Authors: Hande M. Prakash , Devi P. Uma
Published: June 2005

Pregnant Swiss albino mice were exposed to 3.5 MHz diagnostic ultrasound for 10 min (upper limit for ISPTP = 1 W/cm2 and for ISATA = 240 m W/cm2, acoustic power = ∼65 m W) on day 3.5 (preimplantation period), 6.5 (early organogenesis period), 11.5 (late organogenesis period), or 14.5 (early fetal period) of gestation. The offspring were observed for changes in litter size at birth, and sex ratio at 4 weeks of age, and postnatal mortality and growth retardation up to 6 weeks of age. No significant difference from control in litter size or sex ratio was observed in the offspring exposed to ultrasound on any of the gestation days studied. Exposure at the early organogenesis period produced a marginally significant increase in the postnatal mortality. A significant number of ultrasound exposed animals showed lesser body weights than the control group. However this growth retardation was transient and the normal growth pattern was restored by 6 weeks of age. It is concluded that the early stages in mouse gestation may be sensitive to the lethal effects of ultrasound than the later stages of pregnancy. Even though a small increase in the postnatal mortality was observed in the ultrasound exposed animals, the normal growth was not affected in the surviving animals except for a transient growth retardation. Low birth weight may be a characteristic effect of exposure only at the preimplantation period. © 1993 Wiley‐Liss, In.
In vitro embryotoxicity of N‐methyl‐N‐(7‐propoxynaphthalene‐2‐ethyl)hydroxylamine (QAB): Evidence for N‐dehydroxylated metabolite as a proximate dysmorphogen
Authors: Terlouw Gerrard D. C. , Namkung Moses J. , Juchau Mont R. , Bechter Rudolf
Published: June 2005

The rat conceptus biotransforms N‐methyl‐N‐(7‐propoxynaph‐thalene‐2‐ethyl) hydroxylamine (QAB) in vitro to 7‐propoxynaphthalen‐2‐ylacetic acid (QAA) and six more (M1 to M6) metabolites. Thus far, M4 has been identified as N‐demethyl‐QAB and M6 as N‐desoxy‐QAB. We investigated which of these two metabolites might be involved in QAB‐embryotoxicity in vitro. Conceptuses were cultured from day 9.5 to 11.5 post‐coitum, and were exposed to N‐demethyl‐QAB or N‐desoxy‐QAB either in the culture medium or by microinjection directly into the amniotic cavity. When added to the culture medium, N‐demethyl‐QAB (No Observed Adverse Effect Level, NOAEL, for growth 122 μM and for differentiation 41 μM) was less active than QAB itself (NOAEL for growth and differentiation 12 μM). N‐desoxy‐QAB caused severe growth retardation and an impairment of differentiation at a concentration of 11 μM (NOAEL 3.6 μM). As regards causing anomalies, the NOAEL of N‐demethyl‐QAB (41 μM) was 10‐fold higher than that of QAB (NOAEL 3.9 μM) and that of N‐desoxy‐QAB (NOAEL 3.6 μM). At an intraamniotic concentration of 0.7 mM, N‐demethyl‐QAB caused no effects on growth and differentiation and no increase of anomalies was observed, whereas QAB and N‐desoxy‐QAB each elicited an increase in dysmorphogenic embryos at equimolar concentrations without affecting growth and differentiation. It is, therefore, concluded that N‐desoxy‐QAB, but not N‐demethyl‐QAB, could be a proximate dsymorphogen responsible for the embryotoxicity/teratogenicity of QAB in vitro.© 1993 Wiley‐Liss, Inc.
Descriptive epidemiology of small intestinal atresia, Atlanta, Georgia
Authors: Cragan Janet D. , Martin M. Louise , Moore Cynthia A. , Khoury Muin J.
Published: June 2005

To describe the epidemiology of small intestinal atresia (SIA) in Atlanta, Georgia, from 1968 through 1989, we used the Metropolitan Atlanta Congenital Defects Program, an active, population‐based surveillance system for birth defects diagnosed during the first year of life. We identified 176 infants with SIA, a prevalence of 2.8 per 10,000 livebirths. Among black infants, the prevalence was 3.7 per 10,000 livebirths, significantly higher than the prevalence of 2.4 per 10,000 among white infants [relative risk (RR) = 1.6, 95% confidence interval (CI) = 1.1,2.1]. Nine infants were each one member of a unique pair of twins. The prevalence among twin infants was 7.3 per 10,000, significantly higher than the prevalence of 2.8 per 10,000 among singletons (RR = 2.7, 95% CI = 1.4,5.2). Forty‐nine percent of the infants had duodenal atresia, 36% had jejunal atresia, and 14% had ileal atresia. Two infants (1%) had atresia at an unspecified site in the small intestine. We grouped the infants by anatomic location of SIA into four categories: isolated SIA (53%), SIA with multiple unrelated defects (21%), sequences (16%), and syndromes (10%). We then compared the isolated and multiple unrelated defects groups by gender, race, maternal age, birth weight and one‐year mortality for each location of SIA. Among black infants the prevalence of isolated jejunal atresia was 1.4 per 10,000, significantly higher than the prevalence of 0.2 per 10,000 among white infants (RR = 6.3, 95% CI = 2.9, 13.5). The increased prevalence of these defects among twins was a particularly interesting finding. © 1993 Wiley‐Liss, Inc.

This article is a US Government work and, as such, is in the public domain in the United States of America.
Patterns of growth deficiency in rats exposed in utero to undernutrition, ethanol, or the neuroteratogen methylazoxymethanol (MAM)
Authors: Gavin Claire E. , Kates Barbara , Gerken Laurie A. , Rodier Patricia M.
Published: June 2005

Children and experimental animals exposed to ethanol (EtOH) in utero commonly have low birthweights, and many remain small at maturity. Low body weight or small stature in adulthood may reflect an inability to recover from in utero growth retardation, or it may reflect a separate, postnatal growth deficiency. In this study, daily body weights (postnatal days 1 to 60) were compared among the offspring of the following groups of Long Evans rats: dams fed liquid diet containing 35% EtOH‐derived calories; their pairfed and chow‐fed controls; and dams exposed to methylazoxymethanol (MAM) in two previous studies, in which offspring exhibited reduced numbers of growth hormone releasing factor (GRF) neurons. All treatments produced a number of offspring with weight deficits beginning after birth and persisting into maturity. Three distinct patterns of growth deficiency were observed: (1) weight loss relative to controls in the first weeks of life, seen in offspring exposed to EtOH, pair feeding, or MAM on gestation day 13 (G13); (2) a delay in the onset of the prepubertal growth spurt, seen in all EtOH‐exposed offspring and in G13 MAM‐exposed dwarfs; and (3) failure to sustain the prepubertal growth spurt, seen only after exposure to MAM on G14. The results of this study support the view that prenatal EtOH exposure is capable of affecting postnatal growth specifically; moreover, the pattern of growth deficiency seen in EtOH‐exposed offspring was distinct from that of the undernourished offspring of pair‐fed dams. © 1994 Wiley‐Liss, Inc.
Epidemiologic characteristics of phenotypically distinct neural tube defects among 0.7 Million California births, 1983–1987
Authors: Shaw Gary M. , Jensvold Nancy G. , Wasserman Cathy R. , Lammer Edward J.
Published: June 2005

Isolated neural tube defects (NTDs) appear to have different risk factors compared to nonisolated NTDs. To extend those observations, we explored routinely collected child and parental characteristics as possible risk factors among isolated versus nonisolated NTD cases, among high versus low spina bifida cases, and among open versus skin‐covered spina bifida cases. Fetuses and liveborn infants with anencephaly or spina bifida among the 1983‐‐87 cohort of births and fetal deaths (n = 712,863) were ascertained by the California Birth Defects Monitoring Program. One hundred and ninety‐three anencephalic cases and 272 spina bifida cases were compared to a random sample of 5,000 liveborn infants. Among anencephalic cases, 55% were livebirths and 85% were isolated. The proportion of males was similar to females across all subgroups. Increased risks were found for Hispanic whites, with risk estimates highest for nonisolated cases (odds ratio = 4.0, 95% confidence interval [1.5,10.5]). Among spina bifida cases, 92% were livebirths, 81 % isolated, 82% open, and 86% were low. More males were found among the group with isolated high open defects, and fewer males were found among the group of all closed defects. The proportion of males was similar to females in all other subgroups. Cases were more likely to be Hispanic with risks largest for high open defects (odds ratio = 2.9, [1.2,6.6]), particularly for nonisolated cases. This study provides some evidence that further classifications of NTDs may reveal subgroupings of cases with different etiologies. © 1994 Wiley‐Liss, Inc.
Role of oxygen free radicals in cocaine‐induced vascular disruption in mice
Authors: Zimmerman Ernest F. , Potturi Raju B. , Resnick Elisabeth , Fisher J. Edward
Published: June 2005

To test the hypothesis that cocaine‐induced embryonic vascular disruption is mediated by oxygen free radicals, the antioxidants 2‐oxothiazolodine‐4‐carboxylate (OTC) and α‐phenyl‐N‐t‐butylnitrone (PBN) were employed. When cocaine (78 mg/kg) was administered on day 8 of gestation to ICR mice and embryos evaluated on day 10 (in vivo), 62.3% of cocainetreated embryos showed increased vasodilation compared to 4.9% for controls, and 33.1% of the cocaine‐exposed embryos showed marked hemorrhage compared to 3.3% for controls. In addition, cocaine increased the incidence of neural defects, in the form of open neural tube, hypoplastic prosencephalon, and microcephaly. Administration of OTC (0.25 and 0.5 mmol/kg) or PBN (300 mg/kg) prior to cocaine significantly reduced cocaine‐induced vasodilation and hemorrhage, while not preventing neural defects. When cocaine (78 mg/kg) was administered in vivo on day 8 of gestation and embryos were dissected 15 min later and subsequently cultured for 48 hr in the absence of cocaine (in vivo‐in vitro), marked vascular disruption was observed: normal yolk circulation/heartbeat was decreased to 26.6%, while edema/blisters and vasodilation/hemorrhorage were increased to 45.6% and 59.6%, respectively. Administration of PBN (300 mg/kg) prior to cocaine completely prevented cocaine‐induced vascular disruption. When cocaine was administered in vivo and PBN (300 μg/ml) was incubated with cultured embryos in vitro, the antioxidant only partially prevented cocaine‐induced cardiovascular defects in this model. Neural defects produced by cocaine were not significantly affected by PBN, administered either in vivo or in vitro. Cocaine (78 mg/kg) administered in vivo stimulated lipid peroxidation maximally after 3 hr in both day 8 and day 9 embryos. When cocaine was incubated in vitro during embryo culture at 33 μg/ml), a concentration that produces nonspecific inhibition of growth and development, embryonic lipid peroxidation on day 9 was not affected. Finally, when PBN (300 mg/kg) was administered prior to cocaine (78 mg/kg) on day 8 of gestation, stimulation of lipid peroxidation by cocaine was prevented. These results suggest that cocaine‐induced vascular disruption in early development is mediated by maternal production of oxygen free radicals. © 1994 Wiley‐Liss, Inc.
Anticonvulsant teratogenesis: I. A study design for newborn infants
Authors: Holmes Lewis B. , Harvey Elizabeth A. , Brown Kelly Sue , Hayes Ailish M. , Khoshbin Shahram
Published: June 2005

The basis for the apparent teratogenicity of maternal epilepsy is controversial. Is the critical factor the anticonvulsant drugs taken by the pregnant woman, or the genes which cause the mother's epilepsy? We describe a study design developed to assess these competing theories in a cohort study of newborn infants. We show the feasibility of ascertaining exposed and unexposed infants at several birthing hospitals in one urban area. Between 1986 and 1988, we identified 180 drug‐exposed and 218 epilepsy‐history infants among 49,403 infants. The rate of exposure to seizure medication was 0.36% and of maternal history of epilepsy was 0.44%. A significant number of infants could not be evaluated because they were missed, ineligible, or either the doctor, nurse or parent refused to participate. Overall, there was a significant increase in major malformations, microcephaly or growth retardation among the drug‐exposed infants in comparison to both the epilepsy‐history and the unexposed infants. The types of epilepsy and the apparent etiology were the same among women who took anticonvulsants and women with a history of epilepsy but no anticonvulsants during pregnancy. This study must be extended to include a sufficient number of infants exposed to each commonly used drug as mono‐therapy to allow for a comparison of the effect of each drug on pregnancy outcomes; to provide a comparison of infants whose mothers had a strong family history of epilepsy with infants whose mothers had trauma‐induced epilepsy; and to assess the possible impact of the severity of the mothers' disease on the infants. © 1994 Wiley‐Liss, Inc.
Reversed circulation in acardiac fetuses is associated with anatomic inversions in the aortic wall
Authors: Park Hyoung W. , Kapur Raj P. , Shepard Thomas H.
Published: June 2005

Blood vessel anatomy is probably related to hemodynamic factors which change during development. This principle has been postulated as the basis for differences in the numbers of elastic lamellae in the tunica media of the proximal and distal human aorta. Recent studies of human fetuses at varying stages confirmed that the number of elastin lamellae in the aortic wall varied along its length (spatially) and with age (temporally). These findings suggest that hemodynamic influences during prenatal development induce structural changes in the aortic wall. Acardiac fetuses provide a model for studying the effects of hemodynamic changes on the structure of elastic arteries because blood flow through the aorta in such fetuses is reversed and greatly reduced. Also, analysis of the vascular structure of acardiac fetuses would further define the characteristic features of this congenital disorder. In the present study, we have examined the gross and histological anatomy of the aorta from seven acardiac fetuses. In each case, the microscopic architecture of the aorta was deranged and the normal proximaldistal differences in arterial caliber and number of elastin lamellae were reversed. In the proximal aorta, medial lamellar units were thin, fragmented, and irregular. In some segments, only traces of medial lamellar units existed. We believe that these anatomic abnormalities represent degenerative or dysplastic responses to reversed flow and pressure gradients in the aorta. These findings are consistent with the hypothesis that aortic wall structure is influenced by hemodynamic factors during development. © 1994 Wiley‐Liss, Inc.
Methanol‐induced neural tube defects in mice: Pathogenesis during neurulation
Authors: Bolon Brad , Welsch Frank , Morgan Kevin T.
Published: June 2005

A spectrum of cephalic neural tube defects was observed in near‐term (gestation day [GD] 17) mouse fetuses following maternal inhalation of methanol at a high concentration (15,000 ppm) for 6 hr/day during neurulation (GD 7–9). Dysraphism, chiefly exencephaly, occurred in 15% of fetuses, usually in association with reduction or absence of multiple bones in the craniofacial skeleton and ocular anomalies (prematurely open eyelids, cataracts, retinal folds). Measurements of cerebrocortical width in grossly normal, methanol‐exposed fetuses revealed significant semiquantitative differences in the thicknesses of the frontal cortex and its constituent layers (neuroepithelium, intermediate cortex/subventricular plate, and cortical layer 1) as well as apparent increases in subventricular plate cellularity relative to controls. Subsequently, the early morphogenesis of these neural changes was investigated in neurulating mouse embryos to define tissue‐specific patterns of methanol‐induced damage that lead to cephalic axial dysraphism. Following daily 6‐hr maternal inhalations of 15,000 ppm methanol during GD 7–8, the cephalic neural fold margins were swollen, blunted, and poorly elevated on GD 8.5 and 9 relative to controls. Histopathology of exposed GD 8.5 embryos revealed microcephaly in association with reductions in the cell density and mitotic index of at least 47% in the cranial mesoderm. The mitotic index in the embryonic neuroepithelium was also reduced by 55%, and groups of neural crest cells were displaced to the neural folds dorsal to the foregut (relative to the more ventral location in the facial regions of control embryos). When examined on GD 9.5 and 10.5, maternal methanol exposure (15,000 ppm for 6 hr/day) during GD 7–9 resulted in stunting, delayed rotation, and microcephaly in over 90% of the affected embryos. Persistent patency of the anterior neuropore and prosencephalic hypoplasia were seen in >40% and up to 90% of embryos, respectively. Shallow optic vesicles, stunted branchial arches, scoliosis, and hydropericardium were also observed. Many 10.5‐day‐old embryos were edematous. Occult dysraphism, recognized grossly by abnormally narrow cephalic conformation and histopathologically by the absence of mesoderm in the mesencephalon, was present in at least 21% of methanol‐exposed embryos on GD 9.5 and 10.5. Nile blue vital dye staining of methanol‐exposed embryos revealed no difference in dye accumulation between control and treated embryos on GD 8.5, 9.0, or 9.5. There were no apparent dysmorphogenic effects in control embryos at any stage of development. These data suggest 1) that exposure to a high concentration of methanol injures multiple stem cell populations in the neurulating mouse embryo and 2) that significant neural pathology may remain in older conceptuses even in the absence of gross lesions. © 1994 Wiley‐Liss, Inc.
Role of apoptosis in mediating phosphoramide mustard‐induced rat embryo malformations in vitro
Authors: Chen Beiyun , Cyr Daniel G. , Hales Barbara F.
Published: June 2005

Phosphoramide mustard, an active metabolite of the anticancer drug cyclo‐phosphamide, causes malformations in rat embryos undergoing organogenesis in vitro. The purpose of the present study was to investigate the hypothesis that apoptosis plays an important role in mediating the teratogenicity of phosphoramide mustard. Apoptosis is a process of active or programmed cell death which is characterized by internucleosomal DNA fragmentation and de novo RNA and protein synthesis. Sulphated glycoprotein‐2 (SGP‐2) or clusterin is induced in some models of apoptosis and is one of the proteins likely to be involved in the maintenance of cell integrity. In the present study, day 10 rat embryos were cultured for 6, 12, 24, and 45 hr, with or without the addition of 10 μM phosphoramide mustard. After culture for 24 or 45 hr with exposure to 10μM phosphoramide mustard, the embryos were both growth‐retarded and malformed. Exposure to phosphoramide mustard for 6 or 12 hr did not significantly alter the relative amounts of either the mRNA or protein for SGP‐2; this treatment also had no effect on DNA fragmentation in embryos or their yolk sacs. After 24 hr in culture, the relative amounts of SGP‐2 protein, but not mRNA, were increased 2‐fold in the yolk sacs of the phosphoramide mustard‐exposed embryos, but not in the embryos themselves. At this time, DNA fragmentation was detected in phosphoramide mustard‐exposed embryos, but not in their yolk sacs or in control embryos. After 45 hr in culture, SGP‐2 protein and mRNA levels were increased 2–4‐fold above the controls in the phosphoramide mustard‐exposed embryos and their yolk sacs. Immunohistochemical analysis revealed that in control embryos cultured for 45 hr, the SGP‐2 reaction product was localized in the heart, hindgut, and yolk sac. In contrast, in phosphoramide mustard‐treated embryos cultured for 45 hr, SGP‐2 immunostaining was found throughout the embryo, with a strong immunoreaction in the mesenchyme and ectoplacental cone. DNA fragmentation in the embryos exposed to phosphoramide mustard for 45 hr was more extensive than that found after 24 hr, but fragmentation was still not detected in the yolk sac. Thus exposure in vitro to a teratogenic concentration of phosphoramide mustard resulted in DNA fragmentation and an increased expression of SGP‐2 in the embryo. These data suggest that apoptosis is involved in mediating the teratogenicity of phosphormide mustard. © 1994 Wiley‐Liss, Inc.
A population‐based study of gastroschisis: Demographic, pregnancy, and lifestyle risk factors
Authors: Torfs Claudine P. , Velie Ellen M. , Oechsli Frank W. , Bateson Thomas F. , Curry Cynthia J. R.
Published: June 2005

Gastroschisis, an abdominal wall defect, most often occurs in infants of young mothers. To identify risk factors for gastroschisis, we conducted a case‐control study in the population surveyed by the California Birth Defects Monitoring Program (CBDMP). From structured questionnaire data, we compared sociodemographic, reproductive, and lifestyle factors for 110 mothers of infant with gastroschisis with those for 220 agematched mother of normal infants. Univariate matched‐pair analysis showed significant association of gastroschisis with mother's education, yearly family income, marital status, a history of mother's mother smoking, mothers fathers absence from home during the mother's youth, more than one elective abortion, a short interval between menarche and first pregnancy, siblings from different fathers, and use of either a recreational drug (either cocaine, amphetamine, marijuana, or lsd), alcohol, or tobacco during the trimester preceding pregnancy. For cocaine, amphetamine, and marijuana, use of more than one drug showed a stronger association than single drug use. The association was stronger if both parents used drugs. Although many variables were correlated, odds ratios (OR) were significant (95% confidence intervals) in multivariate conditional logistic analysis for; yearly family income <$10,000[OR = 4.34(1.54, 12.22)]or $10,000‐$49,999 [OR = 3.93(1.43, 10.80)]; mother's smoking status not known [OR = 3.99(1.66, 9.56)]; mother's fathers absence from home during her youth [OR = 3.11(1.14,8.46)]; and drug use by mother [OR = 2.21,(1.21, 4.03)], father [OR = [1.66(1.02, 2.69)], or both [OR = 3.05(1.48, 6.28)]. The best predictive model explained 32% of the deviance. Young, socially disadvantaged women with a history of substance use were at highest risk for a child with a gastroschisis. © 1994 Wiley‐Liss, Inc.
Development of the cerebellar defect in ataxic SELH/Bc mice
Authors: Harris M. J. , Juriloff D. M. , Gunn T. M. , Miller J. E.
Published: June 2005

In SELH/Bc mice, 5–10% of young adult are ataxic, due to a midline cleft in the cerebellum. An additional 10–20% of SELH/Bc embryos have exencephaly and die at birth. All SELH/Bc embryos omit a normal step in cranial neural tube closure, initiation of fusion at Closure 2. In the 80–90% that complete cranial neural tube closure, the last region of the closure, on late D9, is the region of prospective cerebellum, and its closure is late. We postulated that the cleft cerebellum in ataxic SELH/Bc mice derives from this delay in neural tube closure and predicted that we would see evidence of a cerebellar midline cleft in all earlier stages after cranial neural tube closure in normally complete.

In the present study we show that cerebellum in cleft in a 7–9% proportion of SELH/Bc D16 fetuses (2/28) and D11 embryos (15/167), and that the defect is detectable on D10. In these abnormal D16 fetuses, D11and D10 embryos, there is gap in midline continuity of cerebellar neuroepithelium, a finding consistent with our hypothesis that the neuroepithelium in this region fails to complete fusion in those embryos. We also show that cerebella of adult SELH/Bc ataxic mice have no obvious deficiency of lobules, or disorganization of tissue as in the Wnt−1 mutants. © 1994 Wiley‐Liss, Inc.
Anticonvulsant teratogenesis: 2. Statistical methods for multiple birth outcomes
Authors: Legler Julie M. , Ryan Louise M. , Harvey Elizabeth A. , Holmes Lewis B.
Published: June 2005

The purpose of this paper is to demonstrate the application of generalized estimating equations to assess an exposure effect using multiple birth outcomes. This multivariate approach provides the flexibility of regression modeling and improved power, as compared to series of univariate analyses or collapsing the multiple endpoints to a single indicator of affectedness. Motivating the discussion will be a large cohort study designed to assess the effects of anticonvulsant medications on a variety of birth outcomes, including major malformations, and growth and weight parameters, as well as a broad spectrum of minor physical anomalies. Because the study is still in progress, the aim here is not to present a definitive analysis, but to present and describe the application of these recently developed statistical methods to analyze studies with multiple outcomes. For simplicity, we will focus on the control and drug‐exposed groups only from that study (ignoring the seizure history group), and we will concentrate on an analysis of minor physical anomalies. © 1994 Wiley‐Liss, Inc.
Pathogenetic changes in the lung bud of mutant rats with heritable pulmonary lobation anomalies
Authors: Aoyama Hiroaki , Fujii Sakiko , Hojo Hitoshi , Teramoto Shoji
Published: June 2005

The present study aimed at investigating pathogenesis of pulmonary lobation anomalies in fpl/fpl mutant rats. Day 12–15 embryos were first examined for pulmonary lobation. Lung serial sections were then made and examined for bronchial branching and distribution of extracellular matrices (ECMs). The lung buds of both fpl/fpl embryos and their phenotypically normal fpl/+ littermates were formed as bilateral protrusion of the foregut on gestation day 12. In fpl/+ embryos, three processes appeared on the right lung bud on day 13, and fissures were completely formed by day 14. In fpl/fpl embryos, the right lung bud had no clear process on gestation day 13, and fissures were not formed on day 14 and thereafter, with the exception of incomplete separation between the cranial and middle lobes. Histological observation revealed that the right main bronchial bud of fpl/+ embryos ramified all lobar bronchial buds by gestation day 13. ECMs, a borderline between endodermal bronchial buds and surrounding mesenchyme, disappeared at the distal end of each lobar bronchial bud. By contrast, in fpl/fpl embryos, the right main bronchial bud did not ramify the middle and intermediate lobar bronchial buds at its lateral and ventral portions, but swelled on gestation day 13. It was covered with ECMs at the lateral side but not at the ventral region, from which the middle and intermediate lobar bronchial buds arose on gestation day 14. These observations suggest that altered distribution of ECMs causes branching abnormalities of the lobar bronchial buds and subsequent lobation anomalies in fpl/fpl embryos. © 1994 Wiley‐Liss, Inc.
12‐O‐tetradecanoyl‐phorbol‐13‐acetate‐induced rat embryo malformations in vitro are associated with an increased relative abundance of embryonic E‐cadherin mRNA
Authors: Chen Beiyun , Hales Barbara F.
Published: June 2005

Epithelial‐cadherin (E‐cadherin) is a member of a family of Ca2+‐dependent cell adhesion molecules which are localized in zonulae adherens and play an important role during development. E‐cadherin is abundant in rat embryos and their yolk sacs during organogenesis. The phorbol ester, 12‐O‐tetradecanoyl‐phorbol‐13‐acetate (TPA), has been reported to disrupt the morphology and functional development of the rat embryonic visceral yolk sac. The present study investigated the possibility that the effect of TPA on yolk sac development may be due to the altered expression of E‐cadherin. Rat embryos, with their yolk sacs intact, were cultured on day 10 of gestation for 1 hr. At this time the vehicle, dimethyl sulfoxide (DMSO), or TPA (at different concentration) was added to the culture medium; the cultures were continued for up to 24 hr. Embryos and yolk sacs were collected separately at the end of each culture period. The relative abundances of E‐cadherin mRNA and protein were analyzed with Northern and Western blot analyses. Despite the TPA‐induced abnormalities in yolk sac development, the relative abundance of E‐cadherin mRNA or protein in the yolk sac was not altered by TPA exposure. However, in embryos exposed to dysmorphogenic concentration of TPA, the relative abundance of E‐cadherin mRNA was significantly increased after 24 hr in culture, compared to either controls or embryos exposed to non‐dysmorphogenic concentration of TPA. The magnitude of the increase in embryonic E‐cadherin mRNA appeared to correlate with the severity of the embryo malformations. Interestingly, despite this increase in embryonic E‐cadherin mRNA abundance, no increase in E‐cadherin protein concentrations was found in the embryo. The timing of the increased relative abundance of E‐cadherin mRNA in TPA‐exposed malformed would suggest an indirect relationship with the embryo malformations observed. Activation of protein kinase C is likely to be involved in both the embryotoxic effects of TPA and the induction of increased concentrations of E‐cadherin mRNA in the embryo since 4α‐phorbol‐12,13‐didecanoate (4α‐PDD), a phorbol ester analog which does not activate protein kinase C, had no effect. This study demonstrates that TPA‐induced embryo malformations are associated with an increased abundance of E‐cadherin mRNA in the embryo, but not in the yolk sac, and that these action of TPA depend on the activation of protein kinase C. © 1994 Wiley‐Liss, Inc.
Prevention of neural tube defects by and toxicity of L‐homocysteine in cultured postimplantation rat embryos
Authors: Vanaerts Leon A. G. J. M. , Blom Henk J. , Deabreu Ronney A. , Trijbels Frans J. M. , Eskes Tom K. A. B. , Peereboom‐Stegeman Jenny H. J. Copius , Noordhoek Jan
Published: June 2005

Mild hyperhomocysteinemia is frequently observed in mothers who gave birth to a child with a neural tube defect (NTD). In a previous study we showed L‐homocysteine was embryotoxic to gestational day 10 (GD 10) rat embryos in culture, however, no NTDs were observed. We therefore investigated the effect of L‐homocysteine on the development of neural plate stage (GD9. 5) rat embryos. Other objectives of this study were investigation into whether the embryotoxicity of L‐homocysteine could be attenuated by compounds related to its metabolism and clarification of the mechanism of L‐homocysteine embryotoxicity. In GD9. 5 rat embryos L‐homocysteine was not toxic at 1‐ and 2‐mM concentrations. Rather at these concentrations it promoted development of the rat embryos in serum that without supplementation caused NTDs in the embryos. L‐Methionine had the same preventive effect at even lower concentrations, but folinic acid (1mM) did not improve embryonic development. N5‐Methyltetrahydrofolate (5‐Ch3‐THF)(100 μ,M)L‐serine(6 mM), and L‐methionine(6 and 12 mM) attenuated the embryotoxicity of L‐homocysteine(6 mM) in GD10 rat embryos. Vitamin B12(10 μM) completely abolished the embryotoxicity of L‐homocysteine, which was shown to be mediated by catalysis of the spontaneous oxidation of L‐homocysteine, to the less toxic. L‐homocysteine. In GD 11 rat embryos, both L‐and D‐homocysteine were readily and taken up when added to the culture (3 mM)and increased embryonic S‐adenosylhomocysteine (SAH) levels 14‐and 3‐fold, respectively. This difference was shown to be caused by the stereospecific preference of SAH hydrolase. We propose the basis for L‐homocysteine embryotoxicity is an inhibition of transmethylation reactions by increased embryonic SAH levels. © 1994 Wiley‐Liss, Inc.
Immunohistochemical double‐staining for Ah receptor and ARNT in human embryonic palatal shelves
Authors: Abbott B. D. , Probst M. R. , Perdew G. H.
Published: June 2005

The aryl hydrocarbon receptor (AhR) and the AhR nuclear translocator protein (ARNT) are basic–helix–loop–helix‐PAS (HLH) proteins involved in transcriptional regulation. Polycyclic aromatic halogenated chemicals, of which 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin (TCDD) is the most potent, bind to the AhR. In the cellular cytoplasm, the AhR exists as a complex with the heat shock protein HSP90 and other small peptides. This complex dissociates following ligand binding and then the ligand‐bound AhR binds ARNT. The ligand–AhR–ARNT complex interacts with a specific, nuclear DNA sequence, the dioxin response elements (DRE), altering transcription of a regulated gene. Studies in hepatoma cell lines indicate that both proteins are required for regulation of transcription. In this study, AhR and ARNT were localized immunohistochemically in human embryonic palatal cells and specific patterns of expression were seen for each protein. A double‐staining protocol revealed that epithelial cells expressed both AhR and ARNT, but in mesenchyme and nasal spine cartilege individual cells were identified which expressed either AhR or ARNT. This heterogeneous pattern may be a means of suppressing transcriptional regulation and also suggests the existence of other, unidentified basic–helix–loop–helix partner(s). The heterogeneous expression pattern may also reflect a complex role for these HLH proteins as transcriptional regulators of embryonic development. © 1994 Wiley‐Liss, Inc.