bluuebunny/crossref_metadata_2025 huggingface dataset:
Prenatal Diagnosis of Fetal Chromosomes: Evaluation of Mi dtri mester Amniocetesis and Chorionic Villus Sampling in Early Pregnancy*
Authors: SUZUMORI Kaoru
Published: May 2008

Abstract The congenital anomaly resulting from chromosomal aberrations is serious and well‐known. The incidence in newborn infants is estimated to be 0.5‐1% (WHO). In many cases the mechanism causing a chromosomal aberration is unknown. There appear to be the protective procedures. The procedure that fetal cells obtained by midtrimester amniocentesis could be grown in culture and karyotyped has opened a new area to genetic counseling for chromosomal aberrations. In the past 15 years, 949 pregnant women have undergone amniocentesis in our hospital. Most institutes now employ ultrasonography for examinations of the fetal position and the localization of placenta prior to amniocentesis. The use of this device remarkably reduced the incidence of fetal loss, and the amniocentesis appears to be safe and efficient. A discrepancy has been noted between the rates of chromosomal aberration by diagnosed at amniocentesis and the rates that would be expected from observation in live births. Multiple regression analysis was performed based on the personal informations of mother, providing what may be the risk factors for the recurrence of sibling with a non‐inhetrited chromosomal aberration.

Recently, the technique to obtain chorionic villus in early pregnancy for the prenatal diagnosis of fetal defects, now being tested in the United States and Europe, may replace midtrimester amniocentesis within the next few years. The main advantage is that it can reduce the psychologic impact because emotional and physical stresses of a midtrimester abortion are serious problems. There are many possible approaches to obtain chorionic villus sample. Our experience has shown that biopsy forceps inserted via the cervix is most successful. The possibility was discussed that this may be developed in the future for clinical use as an alternative to amniocentesis.
Prenatal Diagnosis of the Hemoglobin Disorders by DNA Analysis*
Authors: FUKUMAKI Yasuyuki
Published: May 2008

Abstract Three types of approach to prenatal diagnosis by DNA analysis were described with drawing on examples from sickle cell anemia and thalassemias. The first approach is to detect the change of the restriction pattern of fetal DNA due to the mutation. One type of 6|3‐thalassemia mutation was detected by Southern blotting analysis of genomic DNA when probed with a radiolabeled globin gene because of a large deletion in the β‐globin gene cluster. Since the sickle mutation altered the recognition site of MstII in the β‐globin gene, Southern blot of MstII digest of DNA from a homozygote hybridized with a 1.15 kb MstII 0‐globin gene sequence yielded autoradiograms with a 1.35 kb fragment instead of a normal 1.15 kb fragment.

The second approach is based upon the feasibility of using synthetic oligonucleotide probes to detect single base mismatches. Two synthetic 19 nucleotide long oilgonucleotide probes, one complementary to the normal human 0‐globin gene sequence and one complementary to a β‐thalassemia gene sequence (a C‐T substitution at position 654 of IVS2) were used to distinguish between the normal and thalassemia genes in BamHI digests of genomic DNAs.

Linkage analysis using restriction fragment length polymorphisms (RFLPs) in the β‐globin gene cluster provides the third approach to prenatal diagnosis by DNA analysis. Allele‐linked RFLPs are only available for detection of the sickle mutation and one type of nonsense mutation of β‐thalassemia in limited populations. Familial‐linked RFLPs have been used for prenatal diagnosis of β‐thalassemias because 17 polymorphic restriction enzyme cleavage sites were identified in the β‐globin gene cluster. This approach is also applicable to prenatal diagnosis of genetic disorders in which the gene products responsible for pathogenesis have not been identified. However, a possibility of diagnosis by this approach depends on RFLPs of parents. Further technical development to simplify the DNA analysis will facilitate clinical use of DNA analysis in prenatal diagnosis.
Developmental Change of Hematological Features of the Anemic CFO Mouse (heal hea) with Hereditary Congenital Erythroblastic Anemia
Authors: SHIMIZU Koji , KEINO Hiromi , MIZUTANI Akira
Published: May 2008

ABSTRACT Developmental changes of hematological features of anemic CFO mice (hea/hea) were studied in comparison with those of normal mice (+/+ or hea/+). Anemia of newborn hea/hea mice was not severe, but was easily distinguishable from normal littermates by their yellow body color and also by their blood smear pictures which showed abundant erythroblasts. This disease proceeded rather progressively though many died around 10 days after birth and none survived after aged 25 days. Erythrocyte count, hematocrit percentage and hemoglobin content of hea/hea mice became to be nearly 50% of those of normals during development. On the other hand, leukocyte number of hea/hea mice was 3 to 8 times as many as that of normals. Although hepatomegaly was not evident, splenomegaly was remarkable in hea/hea mice after aged 15 days: Platelet number of hea/hea was about 3 times as many as that of normals. On the contrary, megakaryocytes in the bone marrow of hea/hea mice were much fewer than those of normals. Nucleated cells, mainly orthochromatophilic erythroblasts, in the circulation of hea/hea mice were strikingly 80 to 100 times as many as those of normals. This progressively developed anemia caused by hea seemingly affected the differentiation of erythroid cells, but also that of myeloid cells and that of megakaryocytes.
Alterations in Amniotic Fluid Volume and Associated Reduction Deformities of the Limbs in Rat Induced by Cyclophosphamide
Authors: SINGH Shamer , JEYASEELAN Nadankutty
Published: May 2008

ABSTRACT Maternal administration of a single dose (20 mg/kg) of cyclophosphamide on day 12 of gestation in CF rats (Sperm += day zero) resulted in the reduction deformities of the limbs of the fetuses and increase in the volume of amniotic fluid at term. Chronological observations of 535 fetuses at 24 hour intervals from day 13 through day 20 revealed variations in the volume of amniotic fluid. On day 13 and 14 it was markedly reduced as compared to corresponding controls (P < 0.001). Though there was a gradual rise on day 15 and 16 it was again decreased during day 17 to 19. However, on day 20 it was found to be markedly increased. The associated oligohydramnios observed during the organo‐genetic period probably has a role in the causation of reduction deformities of the fetal limbs.
Abnormal Ultrastructural Features of Circulating Erythroblasts of the Laboratory Mouse with Hereditary Congenital Erythroblastic Anemia (hea/hea)
Authors: SHIMIZU Koji , YAMAZAKI Yoshiro , KEINO Hiromi , MIZUTANI Akira , ESAKI Kozaburo
Published: May 2008

ABSTRACT Abnormalities of erythroid cells in the bone marrow of anemic CFO mice (hea/hea) with hereditary congenital erythroblastic anemia were not evident, while abundant erythroblasts, especially orthochromatophilic erythroblasts, were observed in the circulation and many showed abnormalities. Among them by electron microscopy, cytoplasmic aggregations of ribosomes and cytoplasmic lattice‐like inclusions were characteristic of the hereditary congenital erythroblastic anemia.
Clinical Delineation of Multiple Congenital Anomalies*
Authors: KUROKI Yoshikazu , YASUDA Mineo
Published: May 2008

Abstract The evaluation of multiple congenital anomalies requires several points.

1. Definite diagnostic criteria of each malformation and its precise description are of importance in the diagnosis of malformation syndromes.

2. Complete family studies improve the accuracy of diagnosis and provide us with valuable information in genetic counselling.

3. The use of good reference books and computer‐aided diagnosing systems are effective measures.

4. It is of great importance to circulate case documents, and to exchange information on the diagnosis, among clinical dysmorphologists.

5. It is crucial to combine the alert clinician's eye and the epidemiologic approach for seeking etiologic associations.

6. Animal models of human malformation syndromes contribute much toward deep understanding of deviated morphogenesis.
Effects of Mitomycin C Treatment before Implantation on Development of Mouse Embryo
Authors: NAGAO Tetsuji , ISHIZUKA Yasuo , MIZUTANI Masahiro
Published: May 2008

Abstract ICR mice were treated intraperitoneally with mitomycin C (MMC) at a single dose of l‐5mg/kg on day 0, 1, 2 or 3 of pregnancy (vaginal plug = day G). Embryotoxicity and teratogenicity were examined at term.

A dose‐dependent increase in fetal mortality and decrease in the number of implants, fetal weight or number of ossified sacral and caudal vertebrae were observed. The incidence of external malformation, umbilical hernia, significantly increased among the fetuses of dams treated with MMC at 5mg/kg on day 1, 2 or 3. Skeletal malformation failed to show significant increase though some fetuses from MMC‐treated mice were attended with multiple defects such as fusion of ribs or axial skeleton.

Maternal contribution to and the direct action of MMC on embryonic development were investigated by embryo transfer. In regard to malformation inducement and decrease in implants, the maternal environment was found to have more significant effect than the direct action of this drug but both these factors are significantly responsible for increased fetal mortality and growth retardation.
The Role of Medicine for Children with Developmental Disabilities*
Authors: TAKAMATSU Tsurukichi
Published: May 2008

Abstract The trend of the delivery of health care services for developmentally disabled children, especially those with congenital anomalies, is described. The importance of how the organization of community health services and the role of medicine and medical facilities mesh together in Japan is emphasized.
A Clinical Case of Persistent Stapedial Artery
Authors: MASUDA Yu , WATANABE Shuichi , OMICHI Takuya , OGURA Yoshio
Published: May 2008

Abstract A case of persistent stapedial artery is presented.

The patient was a 49‐year‐old man who had suffered from tinnitus in the left ear and bilateral hearing impairment for 20 years. The preoperative audiological examinations showed considerable combined hearing loss, suggesting middle ear anomalies or otosclerosis. An exploratory tympanotomy on the left ear was performed, and complete fixation of the stapedial footplate and a small vessel running vertically on the footplate were observed. By reconstructive surgery using a prosthesis, postoperative hearing improved. The histological examination of the removed portion of the stapes showed an otosclerotic lesion. Cases of persistent stapedial artery found in the literature were discussed from the clinical and embryological viewpoint.
Results of the NCTR Collaborative Behavioral Teratology Study (CBTS)*
Authors: BUELKE‐SAM Judy
Published: May 2008

Abstract Behavioral measures used in the CBTS were negative geotaxis, olfactory discrimination, auditory startle habituation, 1‐hr and 23‐hr activity, an operant discrete trial discrimination task, and activity following amphetamine challenge. Body weights and certain physical landmarks of development also were monitored in two separate studies, one using prenatal treatment with 0, 0.5 or 2.0 mg/kg d‐amphetamine sulfate sc on gestation days 12‐15, the other using 0, 2.0 or 6.0 mg/kg methylmercuric chloride by gavage on gestational days 6‐9. An untreated control group also was included in each study. The CBTS design allowed evaluation of reproducibility and detection sensitivity of these methods, as well as the impact of early testing experience on later assessments, offspring sex differences in response levels and variability, and the contribution of litter‐to‐litter variation to behavioral measures used in this standardized protocol. Results obtained at the six participating laboratories are discussed in relation to each of these factors and to the degree of overt toxicity obtained with each compound. Behavioral data were reproducible, and detection sensitivities of the tests were very good, requiring no more than 5‐25% change from control values for either sex. Early testing experience was not found to be a major factor in determining later behavioral levels, but litter was found to contribute considerable variability to all physical and behavioral data.
Developmental Abnormalities of Dendrites of the Cerebral Cortical Neurons in Mental Retardation*
Authors: TAKASHIMA Sachio , MITO Takashi , HASHIMOTO Kimio
Published: May 2008

Abstract This paper of Golgi studies describes abnormal dendritic structure and development assessed by camera lucida representation, quantitation of dendritic spines, and dendritic branch measurement in metabolic and chromosomal diseases with mental retardation, reviewing the literatures of Golgi study.

Neuronal lipid storage diseases with progressive mental and motor deterioration show characteristic meganeurites between the soma and origin of the axon, and all dendrites have fewer number of branches, shorter total basal dendritic length and marked reduction of spines. Phenylketonuria and Lesch‐Nyhan syndrome also present decreased number of spines.

On the other hand, children and adults with Down's syndrome show poor dendritic spine density and dendritic branchings as well as structural irregularity of spines. This poor' development of dendrites suggesting postnatal poor synaptogenesis may be related to mental retardation in children. These postnatal dendritic abnormalities may be based on genetic factors such as diminished number of cortical granular cells and extrinsic factors which include poor perception of external stimuli as noted in sensory deprivation.

Thus, dendritic spine abnormalities may be correlated with mental retardation. However, we need better staining method and more sytemic study.
Appearance of a Stage‐specific Prominence during Mouse Forelimb Morphogenesis
Authors: KAWAMURA Satoshi , NAKAMURA Harukazu , YASUDA Mineo
Published: May 2008

Abstract A prominence on the preaxial border at the proximal end of the forelimb was found to appear in a limited period of limb development in mouse embryos. This prominence is observable around day 11 (vaginal plug = day 0). When compared with the number of tail somites, this prominence first appears in embryos with 10–12 tail somites, becomes prominent in embryos with 14–18 tail somites, and disappears in embryos with 20–22 tail somites. This prominence may be utilized as a simple morphological indicator for staging developing mouse forelimb.
Bayesian Estimation of the Incidence Rate in Birth Defects Monitoring
Authors: HEMMI Isao
Published: May 2008

Abstract Although non‐Bayesian statistical methods have been used to detect a significant increase of birth defect incidence in monitoring, they require a large number of data. In this paper Bayesian statistics is applied to estimate the incidence rate particularly in a small population. The Bayesian method is as follows. Assuming that the incidence follows a binomial distribution and the prior distribution of the incidence rate is a beta distribution, the posterior distribution of the incidence rate is a beta distribution. The hyperparameters of the prior distribution are determined objectively by the method of maximum marginal likelihood or minimum ABIC of the posterior distribution based on past observations. The Bayesian method is compared with the non‐Bayesian method when applied to the monthly data of anencephaly from the birth defects monitoring program in the Tokyo Metropolitan hospitals between 1979 and 1984. The variation of the Bayesian point estimates is 25% that of the non‐Bayesian ones. The average confidence interval by the Bayesian method is 29% that by the non‐Bayesian method. The results indicate that the Bayesian estimation is more appropriate than the non‐Bayesian estimation in this type of monitoring analysis.
The Use of Animal Models in Understanding Human Teratogens
Authors: WEBSTER William S.
Published: May 2008

Abstract The testing of drugs and other chemicals in pregnant animals is required by legislation in a number of countries as a screening procedure for teratogenic potential in the human. The testing procedure involves methodology designed in the 1960s which was based on regimens established in the 1940s for toxicity testing. The requirement that animals are dosed to maternally toxic levels, frequently means that the embryos are exposed to inappropriately high concentrations of the test substance. Positive results in this type of experiment may have no relevence to the human situation where the exposure profile is often quite different, with the human embryo being exposed for prolonged periods to much lower drug concentrations. One way of duplicating the anticipated human exposure is to grow rat embryos in serum containing the drug and/or its metabolites at concentrations determined in the human during early clinical testing. It is proposed that mammalian embryos will respond in a similar manner to a particular concentration of a test substance.

In vitro experiments using isotretinoin and its main metabolite 4‐oxo‐isotretinoin showed that the metabolite was teratogenic at concentrations which occurred in the human during normal repetitive dosing and hence the metabolite was the likely human teratogen. Similarly, rat embryo culture studies showed that the anticonvulsant drug, valproic acid, was teratogenic at blood concentrations which occurred during normal dosing in the human. Other in vitro studies showed that cadmium is unlikely to be a human teratogen, despite the fact that it is well established as a teratogen in experimental animals in vivo.

It is proposed that embryo culture should be used as an adjunct procedure during teratology testing making use of metabolic and pharmacokinetic data obtained from the human during clinical testing.
Developmental Disturbance of the Cerebral Cortex of Mouse Offspring from Dams Treated with Ochratoxin A during Pregnancy
Authors: HOSHINO Kiyoshi , FUKUI Yoshihiro , HAYASAKA Ikuo , KAMEYAMA Yoshiro
Published: May 2008

ABSTRACT  This study was designed to elucidate long‐term pathological effects of ochratoxin A, one of the food contaminating mycotoxins, on the developing brain in mice. A single intraperitoneal injection of ochratoxin A at a dose of 2 or 3 mg/kg was given to pregnant mice on day 10 of gestation. They were allowed to give birth and the offspring were examined 6 weeks after birth. Their body weight, brain weight, brain size, thickness and cell packing density of dorsal neo‐cortex in histological sections were examined. Growth of basal dendrites of pyramidal neurons in the layer V of dorsal neocortex was also examined with Golgi‐Cox specimens in terms of number of dendrites directly arising from the perikaryon, number of endings of dendrites, branching index (No. of endings/No. of arisings), mean distance from the perikaryon to the tips of dendrites, and number of intersections of dendrites with the zonal boundaries.

Significant changes were found in brain weight, anteroposterior length of cerebrum, mean distance from the perikaryon to the tips of dendrites and number of intersections of dendrites with the zonal boundaries. These parameters can be recommended as sensitive ones in order to estimate rather weak effects of cyto‐toxic chemical agents exposed in utero on the development of cerebral cortex.
Congenital Anomalies of Central Nervous System and Neuronal Plasticity*
Authors: SHIMADA Morimi
Published: May 2008

Abstract Pathological studies on autopsied brains of mentally and physically handicapped children disclose not infrequently various abnormalities at the histological and cytological levels, such as the abnormal cytoarchitecture, heterotopic neuronal nests, abnormal orientation of neurons and tortuous dendrites. These abnormalities have generally been considered to result from the disorders of neuron production and/or mechanical obstruction of neuronal migration including abnormal radial fibers.

Our experimental studies on the microcephalic mouse, which was induced either by prenatal or by postnatal administration of cytosine arabinoside, indicated that the following factors play important roles for above mentioned histological and cytological changes: (1) Disturbance of the neuron production following the destruction of the matrix layer. (2) Regional and incomplete regeneration of the matrix layer and delayed neuron production in the regenerated matrix layer. (3) Unbalanced and plastic synaptogenesis between the remaining neurons which have been produced prior to the destruction of the matrix layer, migrating young neurons produced in the regenerated matrix layer and the afferent fiber terminals which originate from subcortical neurons.
The Use of Flexible Hysterofiberscope (FUJINON) in Transcervical Chorionic Villi Sampling, and Its Evaluation
Authors: HAYASHI Kazuhiko , NOSAKA Keisuke , IWATA Michie , ISHIJIMA Koh , HAMADA Hiroshi , NAGAE Manabu , KANAMORI Norie , KUROKI Yoshikazu
Published: May 2008

Abstract Under real‐time ultrasonographic guidance, chorionic villi sampling (CVS) was performed either by direct punch biopsy with FUJINON therapeutic flexible hysterofiberscope (HYS‐FT), or by catheter aspiration biopsy after observation with FUJINON hysterofiberscope (HYS‐F). The former method was performed in 2 pregnant women, the latter in 11. The chorion frondosum was observed three dimensionally in all cases. Except for one unsuccessful sampling in our first cases in which CVSs were performed, all attempts were successful, and diagnoses duly completed (11 cases of chromosome analyses and one case of DNA analysis). No serious complications appeared in any case immediately after sampling or on the 1st and 7th days. Ultrasonography showed no adverse effects on fetal heart rate, no hematoma at the site of CVS and no injury to the gestational sac. Uterine bleeding was rarely witnessed. Except for one spontaneous abortion and one intra‐uterine fetal death (IUFD) due to placenta previa, all other newborns were healthy. With some improvements, this CVS is believed to be an effective method.
Avoidance Learning Deficits in Prenatally Gamma‐Ray Irradiated Rats
Authors: TAMAKI Yoshitaka , HOSHINO Kiyoshi , KAMEYAMA Yoshiro
Published: May 2008

Abstract Pregnant female rats of the Fischer344 (F344/Du Crj) strain were exposed to gamma‐ray radiation of 1.46Gy on day 15 of gestation (vaginal plug = day 0). When male offspring of control and irradiated dams matured, they were trained with various avoidance conditioning paradigms. In the avoidance acquisition paradigm in which one of the tone‐on and lights‐off signals was presented with predetermined random sequences, the irradiated group had lower overall rates of active avoidance responding than controls. In the go/no‐go (active‐passive) discrimination avoidance paradigm after rats learned only active avoidance responses to these two different signals, irradiated rats established stable active avoidance rates under the go (tone‐on) signal, while they decreased gradually the rate of passive avoidance responding to the no‐go (lights‐off) signal as the amount of discrimination training increased. In the standard avoidance acquisition paradigm in which only the lights‐off signal was used, the irradiated group also was inferior in active avoidance learning to controls. However, the irradiated group showed a high level of anticipatory responding, which was maintained in avoidance conditioning paradigms even though no shock‐avoidance contingency was operating. These results indicated that poor associative ability for the warning signal and the shock might result in the active avoidance learning deficits in irradiated rats, and that active responding was more persistent and passive responding declined due to response excitation within a shock paradigm over the subsequent further training.
Epidemiological Approaches to Identify Risk Factors for Human Congenital Malformations: Descriptive and Analytical Epidemiology of Cleft Lip and/or Palate in Japan*
Authors: OHNO Yoshiyuki , TOHNAI Iwai
Published: May 2008

Abstract This paper described three major epidemiological approaches to explore etiological clues to and risk factors for human congenital malformations, taking cleft lip and/or palate as an example. (1) An analysis of mortality statistics demonstrated an apparently downward trend of infant mortality from cleft lip and/or palate over the recent three decades in Japan. This particular finding was found to be plausibly ascribable to changing clinical assignment of causes of death from “cleft lip and/or palate” to “congenital heart diseases” or “multiple anomalies, syndromes or chromosome abnormalities”, when previous reports on incidence rate and autopsy series were examined. (2) A case‐control study of 194 infants with cleft lip and/or palate (cases) and 194 normal infants (controls), who were matched to cases for sex, maternal age to within one year, birth order and residential area, yielded the following major findings, (a) A significantly increased risk was associated with parental highest educational attainment of less than 19 years, parental occupation of unskilled/service workers, positive family history of cleft lip and/or palate, positive past history of artificial abortion, maternal smoking habits at first trimester, and maternal episodes at first trimester of suffering from any diseases and of ingesting any drugs, (b) A significantly decreased risk was linked with frequent maternal intake at first trimester of such animal proteins as meats, fishes and shells, eggs and milk, (c) Maternal drinking habits and radiation exposures, and frequent maternal ingestion of Japanese/black tea were not associated. (d) Maternal coffee drinking at first trimester significantly elevated the risk, but turned to be unrelated when maternal smoking habits were statistically corrected. (3) A cohort study of approximately 10,000 pregnants, which is ongoing from April 1989 in Nagoya and aims to disclose the associations of parental life‐style habits with general pregnancy outcomes including major congenital malformations, was briefly discussed.
Pathogenesis of an Affected Organ Can Help Us to Understand Normal Organ Development*
Authors: YASUDA Yoshiko
Published: May 2008

Abstract This review discusses the pathogenesis of spina bifida and describes the microscopic appearance of the initial pathological features and their further development in the spinal cords. The embryos and fetuses were derived from pregnant mice given orally 60, 40 or 0 mg/kg of retinoic acid in olive oil on day 8 of gestation and killed one, two or 10 days later. Separation of the primary neural fold from the secondary neural tube was seen in 9‐ and 10‐day‐old embryos in which the caudal end of the neuroepithelium was disorganized and the posterior neuropore was open; underneath it, the secondary neural tissue was seen. At term, fetuses showed spina bifida, including myeloschisis, myelocele and diplo‐myelia. The cord lesions were located between the third lumbar and second coccygeal levels. Thus, the differentiation and development of the spinal cord proceeds as in chick and human embryos, and the spinal cord caudal to the third lumbar level originates from both neuroepithelium and mesenchyme‐like cells while that caudal to the third sacral level originates from mesenchyme‐like cells only.
Congenital Amputation of All the Limbs Related with Amniotic Rupture
Authors: SINGH Jay Datta , SINGH Gajendra
Published: May 2008

A female infant born with all the four limbs amputated close to the proximal ends is reported. The parents both Indians, having six normal children with poor socio‐economic background denied family history of any congenital anomalies. The mother neither suffered any illness nor had exposure to any known teratogen to the best of their knowledge during this pregnancy. However the mother had attempted to abort this baby in the week 7 of gestation with help of an indigenous mechanical device, a herbal stick, per vaginam which resulted in gushing out of some fluid (amniotic fluid) only. This unsuccessfully attempted abortion neither affected the mother nor the pregnancy apparently and on full term this baby was born. When examined on postnatal day 33, the baby looked very healthy, alert and responsive weighing 3500 g, with CR length 38 cm and occipito‐frontal circumference 35 cm. No abnormalities were observed except that all the limbs were amputated just distal to their proximal end. The baby could move all the stumps of the amputated limbs actively. This case seems to be first of its kind being reported from India.

The etio‐pathogenesis of amputation of limbs was reviewed and evaluated in the light of the available literature. Of the possible etiological factors suggested in the literature the most convincing mode of amputation of all the limbs simultaneously in the present case seems to be the multiple amniotic band formation.
Morphological analysis of neural tube defects: Chlorambucil‐induced exencephaly in mice
Authors: TANAKA Osamu , YOSHIOKA Natsumi , YOSHIOKA Takafumi , NAORA Hiroyuki , HATTA Toshihisa
Published: May 2008

Abstract: Exencephaly has been induced in mouse embryos by chlorambucil (CA), a cytotoxic agent. To understand the course of development of this malformation, open neural tube defects in CA‐treated mice were examined using light and scanning electron microscopes (SEM). CA was given to the pregnant mice on day 7.4 of gestation. Embryos were removed and fixed on gestational days 9.3–9.4, 9.7–9.8 and 10.4, and compared to control embryos from untreated mice. By gestational day 9.4, all control embryos had closed neural tubes, except for the posterior neuropores, and well developed brain vesicles. By contrast, in the experimental embryos the frequencies of open neural tube were 26/33 (78.8%), 32/ 40 (80.0%) and 23/66 (34.8%) on each day examined, respectively. Open neural tube defects were classified into six patterns according to the location and magnitude of the open area. The patterns of open neural tube on day 9.7–9.8 were diverse; however, in almost all cases on day 10.4 the open neural tube appeared in a region from the caudal forebrain to the rostral hindbrain. It was evident that an unusual pattern of closure of the neural tube was involved in forming the cranial neural tube. The present study shows that failure of closure of the cranial neural tube in the CA‐treated mouse embryos can be defined as a primary neural tube defect (NTD), which can be, in part, repaired by unusual closure of the neural tube.

At high magnifications of SEM, the neuroectodermal surfaces of the 9.0‐day affected embryos often had a number of slender processes projecting from the neuroepithelial cells. “Ruffles” and “blebs” at the lateral edges of the neural folds were observed in both control and affected embryos.
Growth Retardation during Organogenesis in Genetically Diabetic Mice, Yellow KK
Authors: OOSHIMA Yojiro
Published: May 2008

Yellow KK mice with non‐insulin dependent diabetes were mated at 7 (group A) or 13 (group B) weeks of age. On day 13, 16, or 18 of pregnancy, the plasma glucose concentration in dams was determined, and the embryos/fetuses and placentae were weighed and examined.

The rate of early embryonic death was high and invariable on all three days. The rate of late death increased with advancing pregnancy in both groups and the incidence in group B was higher than in group A on each of the three days. On day 13 of pregnancy, maternal plasma glucose concentration in group B was significantly higher than in group A (465mg/dl vs 230 mg/dl). Embryonic and placental weights in group B were less than those in group A (84 vs 113 mg and 59 vs 68 mg). In group B, there was an inverse correlation between the maternal plasma glucose concentration and the embryonic weight. A similar inverse correlation existed between the maternal plasma glucose concentration and the placental weight. On days 16 and 18 of pregnancy, fetal weights in group B were less than those in group A, but placental weights did not differ significantly between the two groups.

From these results, it appears that in the yellow KK mouse fetal growth retardation has already begun in the organogenesis period and is closely related to maternal hyperglycemia.
An Autopsy Case of Klippel‐Feil Syndrome with Cardiovascular Anomalies and Upper Limb Anomaly
Authors: KAJIWARA Yoshifumi , IKEDA Takayoshi , MATSUO Takeshi , IWASAKI Keisuke , SHIMOKAWA Isao , IKENO Yuji , UEHARA Yutaka
Published: May 2008

Abstract An autopsy case of Klippel‐Feil syndrome with the classical triad of short neck, low posterior hairline and limitation of neck movement is reported. The patient died of cardiac failure resulting from complicated serious cardiovascular anomalies such as double outlet right ventricle, congenital mitral valve stenosis and coarctation of aorta. The condition was also complicated by radial ray deficiency of the left arm and asymmetry of the thyroid gland. Narrowing of the left subclavian and axillary arteries was detected by angiography while the patient was alive, a finding which may suggest the etiology of this congenital anomaly complex.
Effects of Chlorambucil on the Brain Development in Mice during Post‐neurulation Period
Authors: TANAKA Osamu , YOSHIOKA Natsumi , YOSHIOKA Takafumi , OTANI Hiroki , HATTA Toshihisa , HASHIMOTO Ryuju , ABU‐MUSA Antoine
Published: May 2008

Abstract The teratogenic effect of chlorambucil (CA), given during proliferation of matrix cells of the brain following the closure of the neural tube, on the development of central nervous system in mouse embryos was investigated histologically. CA (4, 8 or 15 mg/kg) was given orally to pregnant mice on day 9, 10, 11 or 12 of gestation by single shot, and the fetuses were examined on day 18 of gestation.

Significant decrease in the brain size and microcephaly occurred in fetuses treated with CA regardless of the gestational age studied. The microcephalic brains were characterized by the severe reduction of the cerebral hemispheres in a rostrocaudal direction rather than in the lateral directions. The fetal brains from mice given a low dosage of 4 mg/kg CA maintained normal cerebral cortex, but higher dosages (8 or 15 mg/kg) showed severe dysplasia of all the cerebral histological structures. In the groups treated with 8 or 15 mg/kg CA on day 10 or 11 of gestation, severe cranial malformations, identified as cranioschisis or encephalocele, were often observed. The malformed brains with cranioschisis showed an extensive necrosis of the brain tissue around the lesions. The present study shows that the microcephaly induced by cytotoxic teratogens such as CA, given during the proliferative period of matrix cells after closure of the neural tube, is different from that induced during the migratory period of the neuroblasts, and suggests that the degeneration of the brain tissue by the cytotoxic agent may induce the partial anencephaly in mice.
Behavioral Testing in the Context of Reproductive and Developmental Toxicity Screening in the West*
Authors: SCHARDEIN J. L. , YORK R. G. , WEISENBURGER W. P.
Published: May 2008

Abstract Methods for assessing animal neurobehavior following prenatal/perinatal exposures have been in use for almost 20 years and screening tests incorporating them are regulated in three settings: in Japanese Segment II and III studies on drugs, European (EEC/UK) Segment I studies on drugs, and American neurotoxicity studies on chemicals.

The methods used in the West for assessing behavior in these contexts at present are described in detail. The evolution of the methods over time, the concepts of apical tests and functional domains, the use of positive control agents, what the U.S. Collaborative Study has shown us, and justification for assessing animal behavior in predicting outcome from human exposures are discussed.
Mouse Embryo Culture for Chromosome Analysis*
Authors: TATEWAKI Reiko , TANAKA Osamu
Published: May 2008

Abstract Mouse embryo culture has been established and applied in the study of the causal mechanisms of abnormal development. Mouse embryo culture can be used as an experimental system to examine the influence of teratogen on early development, together with investigations using developmental engineering such as chimera, transgenic mice and nuclear transfer. We combined mouse embryo culture with embryo transfer and chromosome analysis, and studied the relationship between environmental factors (including maternal factors) and genetic predisposition in developmental abnormalities in the embryos of diabetic mice.

In mouse embryo culture from one‐cell to the blastocyst stage, fertilized eggs were incubated in modified Whitten's medium and mixed gas (5% O2, 5% CO2 and 90% N2) at 37°C for three days. β‐Mercaptoethanol 20 μM and EDTA‐2Na 100 μM were added in order to overcome “two‐cell block’ in which two‐cell stage embryos could not develop further. Under theses conditions, from 55 to 100% of the cultured one‐cell stage embryos developed into blastocysts depending on the strain.

For chromosome preparation, Mikamo and Hamaguchi's method (1975) was modified. This method can be applied from one‐cell to pre‐implantation stage embryos, and allows a higher rate of success (63% of prepared embryos) in preparing chromosome slides than that reported in the literature. Using improved methods of embryo culture, embryo transfer and chromosome analysis, we have obtained evidence that both environmental and genetic factors are the causes of malformation during diabetic pregnancy in mice. Mouse embryo culture associated with chromosome analysis is a useful procedure, and offers important data for the clarification of the causal mechanisms in developmental abnormalities.
Teratogenic Effect of Tedral (Theophylline, Ephedrine, and Phenobarbital) on Cardiac Development in Chick Embryos
Authors: MATSUOKA Rumiko , NISHIKAWA Toshio , Jr. Harold J. BRUYERE , GILBERT Enid F.
Published: May 2008

Abstract We employed the chick embryo in a comparative study of the effect of Tedral and theophylline on anatomical development. Tedral, which contains theophylline in combination with ephedrine and phenobarbital in the relative dose ratio of theophylline 33: ephedrine 6: phenobarbital 2, or saline for control was dropped onto the surface of the chorioallantoic membrane of 2, 3, 4 and 5‐day‐old chick embryos. In groups A (theophylline 0.015 M, ephedrine 0.0027 M, phenobarbital 0.0009 M) and B (theophylline 0.011 M, ephedrine 0.0020 M, phenobarbital 0.0007 M), 100% of embryos treated on the 5th day of incubation developed aortic aneurysms with associated cardiovascular malformations. In group C (theophylline 0.005 M, ephedrine 0.0009 M, phenobarbital 0.0003 M), 36.8% of embryos treated on the 5th day of incubation developed aortic aneurysms, and 73.7% of embryos had cardiovascular malformations. These data were compared with those obtained by using theophylline, ephedrine or phenobarbital alone. The commonest cardiovascular anomalies induced by Tedral were double‐outlet right ventricle with ventricular septal defect. Aneurysms were larger in size and cardiovascular malformations were more complex in embryos exposed to Tedral than in those treated with theophylline alone. Complex cardiac defects produced by Tedral included transposition of the great arteries, double‐outlet right ventricle with double‐inlet left ventricle, and truncus arteriosus communis. A slit‐like small ventricular septal defect was found in 9.5% of the ephedrine‐treated (0.51 mg) chick embryo group and 9.1% in the control embryos. These data show: (1) theophylline, a major component of Tedral, produces cardiovascular anomalies in embryonic chick hearts; (2) there is synergy of all three drugs in Tedral; (3) the specific type of conotruncal defect depends on the timing of administration, e.g., truncus arteriosus communis was produced on day 4 of Tedral administration in chick embryos.
Prenatal Diagnosis Reveals the Relative Frequencies of Lysosomal Storage Diseases in Japan
Authors: MORIYA Momoko , OWADA Misao , KITAGAWA Teruo
Published: May 2008

Abstract Data collected in our institution, in which approximately one‐fourth of the prenatal diagnoses for inborn errors of metabolism in Japan are performed, suggest somewhat higher prevalences of Gaucher disease and mucolipidoses in Japan than in the United States, European countries and Australia. Hunter disease seems to be more frequent than Hurler disease in Japan while the reverse is true in Europe and Australia. In contrast, family histories of Pompe disease or cystinosis were much less frequent indications for prenatal diagnosis in Japanese than in Caucasian populations. The results of a recent multicenter survey conducted in Japan on the indications for and reliability of prenatal diagnoses for inherited metabolic disease support the conclusions drown from our data. The relative frequencies of each of the lysosomal storage diseases, with the exception of the mucopolysaccharidoses, obtained from our prenatal diagnosis surveys were quite similar to those derived from an epidemiological study of lysosomal storage diseases in Japan. We conclude that prenatal diagnosis can be used an indicator of the prevalences of lysosomal storage diseases in Japan.
Stage Differences in Developmental Disorders in ICR Mouse Embryos Irradiated with Gamma‐Rays
Authors: KUSAMA Tomoko , HASEGAWA Yasunari
Published: May 2008

This study was designed to determine precisely the radiosensitive period in the development of ICR mouse embryos during which external malformations and growth retardation tend to occur. Female and male mice were placed together for only three hours to allow fairly precise identification of the time of conception. The pregnant mice were divided into 31 groups, which were irradiated in turn with 1.5 Gy gamma radiation at 6‐hour intervals during the period of organogenesis. They were then observed on day 18 of gestation. Items recorded were intrauterine death, external malformations, sex ratio and fetal body weight. Death of the embryo/fetus, especially death in the early period of organogenesis, was most frequent in mice irradiated between days 6.75 and 8.25 of gestation, but there was no statistically significant difference in the frequency of early‐ and late‐period deaths between irradiated and control groups. The types and frequencies of external malformations observed differed according to the exposure period. The most highly sensitive period for each malformation lasted no more than 12 hours. Reduction of fetal body weight was a good indicator of radiation effects, and was observed mostly in the groups irradiated between days 9.75 and 11.00 of gestation. The sex ratio was not affected by the period in which irradiation was performed.
The Pathogenesis of Anorectal Malformation Induced by All‐Trans Retinoic Acid in Mice
Authors: HASHIMOTO Ryozo , ODA Sen‐ichi , INOUYE Minoru , YAMAMURA Hideki
Published: May 2008

The pathogenesis of anorectal malformations has been discussed for more than 100 years, however, much remains to be elucidated. We studied the pathogenesis of high‐type anorectal malformations induced by all‐trans retinoic acid (all‐trans RA) in mice. Pregnant females were injected intraperitoneally once with all‐trans RA, suspended in corn oil (5 mg/ml), at a dose of 100 mg/kg on day 9 of pregnancy. The all‐trans RA‐treated females were killed on one of days 10 to 18 of pregnancy. All fetuses examined on day 18 of pregnancy following in utero exposure to all‐trans RA had anorectal malformations. In the affected male fetuses, the rectum was positioned away from the retroperitoneum toward the ventral side leading to the opening of the urethra just underneath the urinary bladder. Deficiency of the cloacal plate at the dorsal part was also observed in the affected embryos during days 10–11 of pregnancy. The cloacal plate is considered to bring the cloacal cavity to the anus, and thus the deficiency of the dorsal part may be the major cause of the hightype anorectal malformations.
Craniofacial Dysmorphia of Frontal Proboscis in Holoprosencephaly
Authors: LEE Suk Keun , SUH Yeon‐Lim , CHI Je G.
Published: May 2008

We have examined eleven cases of holoprosencephaly with special emphasis on the frontal proboscis. Although the nostril appears blind ended grossly, the proboscis has a single slender midline nostril lined with squamous mucosa containing numerous sebaceous glands. It is actually connected to a poorly formed but definite nasal cavity microscopically. The inner portion of the nostril tunnel is lined by olfactory mucosa containing numerous subjacent secretary glands. Despite the absence of olfactory bulbs and tracts the cartilaginous nasal capsule is identified along with abnormal nasal cavity. However, the nasal capsule is hypoplastic, and has an aberrant nasal septum and no concha. In severe form of holoprosencephaly, i.e., cyclopia, the nasal capsule is noted above the level of the ethmoid plate, in which case the nasal cavity dose not communicate with nasopharynx because it is too distant from the hypoplastic maxilla. Therefore, choanal atresia is resulted. Although unique maxillary bone comprises the whole upper jaw, the failure of fusion between vomeronasal septum and palatal processes in holoprosencephaly results in abnormal maxillary bone which is characterized by a narrow palate without palatal arch form. Nevertheless, through serial microscopic sections we could find a space between the posterior maxilla and the sphenoid bone, which is composed of adipose tissue and is similar in shape to the nasal capsule. This space is thought to be made for the nasal capsule during early developmental period. The present study suggests that the frontal proboscis in holoprosencephaly is an undescended nose. In most cases it is connected to the aberrant nasal cavity which is located beneath the proboscis. The level of proboscis descends gradually toward normal nose position as the severity of holoprosencephaly decreased from cyclopia, ethmocephaly, cebocephaly, to premaxillary agenesis. Key words: frontal proboscis, holoprosencephaly, craniofacial dysmorphia
Consultation System for Congenital Malformation Syndrome*
Authors: SUGIYAMA Kohachiro
Published: May 2008

ABSTRACT Accurate diagnosis of a specific birth defect is a necessary prerequisite to a plan for management of an affected infant, as well as genetic counselling for the parents. But it requires much knowledge on the part of the physician about birth defects from textbooks or journals. Remembering each diagnostic model of a particular birth defect is no easy task, because birth defects by whatever definition exhibit great variety or complexity, and often overlap. With these problems in mind, we have been developing a consultation system for congenital malformation syndromes, which is composed of a knowledge data base and an inference‘engine.’Here our consultation system will be introduced, and several points to put the system into practical use will be mentioned.
Potentiated Embryotoxicity of Pyrimethamine by Folic Acid in Mice
Authors: KUDO Gen , TAHARA Atsushi , HO‐CHUL Shin , KOKUE Ei‐ichi
Published: May 2008

ABSTRACT Effects of folic acid (FA) and folinic acid (FNA) on the embryotoxicity of pyrimethamine (PYR), an antifolate drug, were examined in mice. PYR and FA were administered orally, and FNA was injected intraperitonially for 7 days from day 9 to 15 of pregnancy. The incidence of embryotoxicity including intrauterine deaths and malformations, was 33.5 % in the PYR 50 mg/kg/day group. However, all the fetuses were resorbed in the PYR 50 + FA 50 mg/kg/day group, and FA potentiated the embryotoxicity of PYR. On the contrary, FNA reduced the embryotoxicity of PYR (PYR 50 + FNA 10mg/kg/day; 8.5%). Plasma concentrations of PYR and 5‐methyltetrahydrofolic acid (5MF), a principal form of folate in plasma, were determined after single oral administration of PYR 50 mg/kg with or without FA 50 mg/kg to non‐pregnant mice. Plasma 5MF concentration decreased drastically in the group receiving PYR with FA, compared with the PYR alone group. The pharmacokinetics of PYR were not affected by co‐administration of FA. Therefore, we consider that the potentiated embryotoxicity of PYR by oral FA results from a decrease of plasma 5MF concentration in dams.