ethylenethiourea and Disease-Models--Animal

Postoperative incontinence and constipation still remain the major complications of anorectal malformations (ARMs), despite improvements in their treatment. One of the most important factors that affect postoperative anorectal function is malformations in the lumbosacral spinal cord. However, far too little attention has been paid to the underlying mechanism that produces these malformations.. The levels of sonic hedgehog (Shh), patched homolog 1 (Ptch1), and zinc finger-containing transcription factors 1 (Gli1) expression were investigated in the lumbosacral spinal cord in ethylenethiourea-exposed rat fetus with ARMs, and Shh, Ptch1, and Gli1 expression was confirmed with immunohistochemical staining, quantitative real-time polymerase chain reaction, and western blot analyses during lumbosacral spinal cord development both in the ARMs and normal rat embryos.. Our results have shown that Shh, Ptch1, and Gli1 expression in the lumbosacral spinal cord of rat embryos with ARMs was decreased at both the messenger RNA and protein levels, when compared with their expression levels in normal tissues (P < 0.05).. This study demonstrated that the expression of Shh, Ptch1, and Gli1 in lumbosacral spinal cord was remarkably reduced during late developmental stages in fetal rats with ARMs. These findings offered some important insights into the involvement of the Shh-Ptch1-Gli1 signaling pathway in the pathogenesis of lumbosacral spinal cord maldevelopment in rat fetus with ARMs, which leads to complications after procedures for ARMs.

Topics: Animals; Anorectal Malformations; Disease Models, Animal; Embryo, Mammalian; Ethylenethiourea; Female; Gene Expression Regulation, Developmental; Hedgehog Proteins; Humans; Lumbosacral Region; Patched-1 Receptor; Rats; Rats, Wistar; Spinal Cord; Zinc Finger Protein GLI1

Embryogenesis is orchestrated by the wingless-type MMTV integration site family (WNT) signaling pathways, including Wnt3a. This study was performed to investigate the expression of Wnt3a in the terminal hindgut in ethylenethiourea (ETU)-exposed rat embryos with anorectal malformations (ARMs) and its potential association between Wnt3a and the maldevelopment of the terminal hindgut in ARMs. ARM rat embryos were induced by ethylenethiourea on embryonic day 10 (E10). The expression levels of protein and mRNA of Wnt3a were confirmed using immunohistochemistry staining, Western blotting analyses, and quantitative real-time PCR (qRT-PCR) in normal rat and ARM embryos. Immunostaining revealed a variation in the expression of Wnt3a in the developing terminal hindgut of ARM embryos. The expression of Wnt3a in the terminal hindgut of ARM rat embryos decreased at both the mRNA level and protein level (P<0.05) compared with normal tissues. This study demonstrated that the expression of Wnt3a in the ARMs of ETU-exposed rat embryos was remarkably reduced, which indicated its potential role in the pathogenesis of the terminal hindgut maldevelopment in ARMs.

Topics: Animals; Anorectal Malformations; Anus, Imperforate; Blotting, Western; Disease Models, Animal; Ethylenethiourea; Gastrointestinal Tract; Immunohistochemistry; Rats; Rats, Wistar; Real-Time Polymerase Chain Reaction; RNA, Messenger; Wnt3A Protein

To investigate the effect of folic acid (FA) in an experimental model of anorectal malformations (ARMs) ethylenethiourea (ETU) induced.. Eight female Wistar rats were divided randomly in two groups. Group A - ETU; Group B - FA+ETU; Dams from group B received daily, since two weeks before pregnancy to the end of pregnancy, FA (50mg/kg) by gavage. Dams from groups A and B, received 1% ETU (125 mk/kg) by gavage on gestational day (GD) 11. Their fetuses were harvested by cesarean section on GD21 and were examined looking for ARMs. The thickness of anal stratified squamous epithelium (ASSE) and intestinal epithelium (IE) were analyzed. p < 0.05*.. One hundred and one embryos were harvested. The number of embryos; number of ARMs; mean statistical % (± SD) were determined to be, respectively: ETU - 49 [30;65% (± 24%)] versus FA+ETU - 52 [1;02% (± 3%)] (p = 0.025). AMRs were significantly lower in FA+ETU group than in ETU group (p = 0.025). The thickness (µm) of ASSE (± SD) and IE (± SD) were measured, respectively: ETU - [27.75 (± 0.56) and 18.88 (± 0.93)] versus FA+ETU - [28.88 (± 0.61) and 21.11 (± 0.16)] (p = 0.001). The thickness of IE was significantly enlarged when FA was given (p=0.001).. Folic acid reduces the number and enlarged the IE of ARMs ETU-induced.

Topics: Anal Canal; Animals; Anorectal Malformations; Anus, Imperforate; Disease Models, Animal; Ethylenethiourea; Female; Fetus; Folic Acid; Pregnancy; Random Allocation; Rats, Wistar; Rectum; Reproducibility of Results; Vitamin B Complex

The main aim of this study was to determine Cdx2 expression patterns during anorectal development in normal and anorectal malformation (ARM) embryos with a view to establishing the possible role of Cdx2 in ARM pathogenesis. ARM was induced with ethylenethiourea on the 10th gestational day (GD10) in rat embryos, and Cesarean deliveries were performed to harvest the embryos. The temporal and spatial expression of Cdx2 was evaluated in normal rat embryos (n = 303) and ARM embryos (n = 321) from GD13 to GD16. Immunohistochemical staining revealed that, in normal embryos, Cdx2 was mainly expressed on the epithelium of the urorectal septum (URS) and the hindgut on GD13. On GD14, Cdx2-immunopositive cells were extensively detected on the URS, hindgut, and cloacal membrane. On GD15, increased immunopositive tissue staining on the anal membrane was evident. In ARM embryos, the epithelium of the cloaca, URS, and anorectum were negative or faintly immunostaining for Cdx2. Analyses by Western blot and real-time reverse transcription plus the polymerase chain reaction revealed that, in the normal group, Cdx2 protein and mRNA expression showed time-dependent changes in the developing hindgut from GD13 to GD16. Upon the URS division of the cloaca into the primitive rectum and urogenital sinus (UGS) on GD15, Cdx2 expression began to decrease. Moreover, the Cdx2 expression level in the ARM group from GD13 to GD14 was significantly lower than that in the normal group (P < 0.05). In ARM embryos, an imbalance in the spatiotemporal expression of Cdx2 was noted during anorectal morphogenesis from GD13 to GD16. Downregulation of Cdx2 at the time of cloacal separation into the primitive rectum and UGS might thus be related to the development of ARM.

Topics: Anal Canal; Animals; Anorectal Malformations; Anus, Imperforate; CDX2 Transcription Factor; Disease Models, Animal; Ethylenethiourea; Female; Homeodomain Proteins; Male; Morphogenesis; Pregnancy; Rats; Rats, Wistar; Transcription Factors

Fecal incontinence and constipation after procedures for anorectal malformations (ARMs) are closely related to the maldevelopment of the striated muscle complex (SMC). Previous studies have demonstrated that myogenic regulatory factors (MRFs) play a significant role in muscle development. Wnt signal pathway is extremely important for MRFs regulation. This study was designed to investigate the spatiotemporal expression pattern of Wnt5a in SMC in ARMs rat embryos.. Anorectal malformation embryos were induced by ethylene thiourea on embryonic day 10 (E10). Expression levels of protein and mRNA of Wnt5a were confirmed by immunohistochemistry staining, western blot and quantitative real-time PCR (qRT-PCR) between normal rat embryos and embryos with ARMs.. Immunostaining revealed that, on embryonic day 17 (E17), the Wnt5a protein was initially expressed in the SMC in normal embryos. With the growth of pregnancy, the positive staining cells gradually increased. The same time-dependent changes of Wnt5a protein were detected in ARMs embryos. Besides, immunostaining showed that Wnt5a had a significant increase in normal embryos compared with ARMs embryos. Similarly, in Western blot and qRT-PCR, the higher expression of Wnt5a protein and mRNA were remarkable in normal embryos during the SMC development, relatively.. Our study demonstrated that the downregulation of Wnt5a at the time of SMC development might partly be related to the dysplasia of SMC in ARMs.

Topics: Animals; Anorectal Malformations; Anus, Imperforate; Blotting, Western; Disease Models, Animal; Ethylenethiourea; Female; Gene Expression Regulation, Developmental; Gestational Age; Immunohistochemistry; Male; Maternal Exposure; Muscle Development; Muscle, Striated; Pregnancy; Rats, Wistar; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Time Factors; Wnt Proteins; Wnt-5a Protein

The pathophysiology of abnormalities associated with myenteric plexus lesions remains imperfectly understood. Such abnormalities have been correlated with subocclusive intestinal conditions in children with Hirschsprung's disease, cases of chronic constipation and, postoperatively, in cases of anorectal anomalies. This study evaluated abnormalities of the myenteric plexus in fetus from female rats that received ethylenethiourea.. Female rats were exposed to ethylenethiourea on the 11(th) day of pregnancy (experimental group) or to 0.9% physiological solution (control group). Abnormalities were only found in the experimental group. The digestive tract muscle layer was analyzed morphometrically and changes to the frequencies of nerve plexus cells and interstitial cells of Cajal were evaluated, using hematoxylin-eosin, S-100 protein, neuron-specific enolase and C-Kit, respectively.. Muscle and skeletal abnormalities were observed in 100%, anorectal anomalies in 86%, absent tail in 71%, short tail in 29%, duodenal atresia in 5%, esophageal atresia in 5% and persistent omphalomesenteric duct in 5%. Histopathological analysis showed a thinner muscle layer associated with lower frequencies of ganglion cells and interstitial cells of Cajal, in all gastrointestinal tract.. Severe nerve plexus abnormalities associated with muscle layer atrophy were observed throughout the gastrointestinal tract in newborn rats exposed to ethylenethiourea.

Topics: Abdominal Muscles; Abnormalities, Drug-Induced; Animals; Animals, Newborn; Digestive System Abnormalities; Disease Models, Animal; Ethylenethiourea; Female; Fetus; Ganglia; Interstitial Cells of Cajal; Muscular Atrophy; Myenteric Plexus; Pregnancy; Prenatal Exposure Delayed Effects; Random Allocation; Rats; Rats, Wistar; Staining and Labeling; Statistics, Nonparametric

Wnt5a is involved in the initiating and patterning morphological adaptations of gut. However, its role remained unknown during terminal hindgut development in the normal and anorectal malformation (ARM) rat embryos. This study was designed to investigate the expression pattern of Wnt5a in the terminal hindgut in ARM rat embryos.. Ethylenethiourea-induced ARM model was introduced to investigate the expression pattern of Wnt5a during terminal hindgut development using immunohistochemical staining, reverse transcriptase polymerase chain reaction (RT-PCR), and Western blot analysis.. Immunostaining revealed that Wnt5a expression showed space-dependent changes in the developing terminal hindgut. On embryonic day 17 (E17) in normal embryos, the Wnt5a protein was initially expressed in the mesenchyme of the terminal hindgut. From E18 to 19, the positive staining cells gradually increased. The expression was detected mainly in the circular muscle and myenteric plexus of hindgut. In the ARM embryos, on E17, the Wnt5a protein was also expressed in the hindgut. However, from E18 to 19, the positive staining cells in the middle hindgut gradually increased but in the terminal hindgut decreased. In Western blot and RT-PCR, time-dependent changes of Wnt5a protein and mRNA expression were remarkable during the terminal hindgut development in normal and ARM embryos.. These data implied that the downregulation of Wnt5a at the time of hindgut neuromuscular development might partly be related to the maldevelopment of terminal hindgut in ARM.

Topics: Anal Canal; Animals; Blotting, Western; Disease Models, Animal; Ethylenethiourea; Fetus; Gene Expression Regulation, Developmental; Immunohistochemistry; Rats; Rats, Wistar; Rectum; Reverse Transcriptase Polymerase Chain Reaction; Wnt Proteins; Wnt-5a Protein

Despite evidence for a conserved role of thyroid-stimulating hormone (TSH) in regulating vertebrate thyroid function, molecular data on thyroid responses to TSH are mainly limited to mammalian species. In this study, we examined histological and molecular changes in the thyroid of Xenopus laevis tadpoles during a 12-day treatment with 20mg/l perchlorate (PER) and 50mg/l ethylenethiourea (ETU). Inhibition of thyroid hormone (TH) synthesis by PER and ETU was evident from developmental retardation, reduced expression of TH-regulated genes and up-regulation of tshb-A mRNA. Thyroid histopathology revealed goiters with strikingly different follicular morphologies following PER and ETU treatment. Using real-time PCR, we analyzed thyroids sampled on day 12 for differential expression of 60 candidate genes. Further temporal analyses were performed for a subset of 14 genes. Relative to the control, PER and ETU treatment modulated the expression of 51 and 49 transcripts, respectively. Particularly genes related to TH synthesis and protein metabolism were similarly affected by PER and ETU. However, several genes were differentially expressed in PER- and ETU-treated tadpoles. Specifically, goiter formation in the PER treatment was associated with low expression of genes related to DNA replication but high expression of negative growth regulators. Results from this work provide for the first time a characterization of gene expression profiles during goitrogenesis in a non-mammalian vertebrate model. Overall, our data suggest that, in addition to TSH over-stimulation, further mechanisms related to the mode of goitrogen action contribute to the regulation of thyroid gene expression.

Topics: Animals; Antithyroid Agents; Brain; Disease Models, Animal; Endocrine Disruptors; Ethylenethiourea; Gene Expression Profiling; Gene Expression Regulation, Developmental; Goiter; Life Cycle Stages; Perchlorates; Thyroid Gland; Thyrotropin; Vertebrates; Xenopus laevis

The receptor tyrosine kinase of the Eph family is a large group of highly conserved molecules that function in diverse intercellular recognition events. It has been reported that EphB2 is related to caudal remodeling events. The aim of this study is to investigate EphB2 expression in anorectal development in normal and rat embryos with anorectal malformations (ARMs) and attempt to define its role in anorectal morphogenesis.. The ethylenethiourea (ETU) rat model of the ARMs was used in this study. Immunohistochemical analyses and real time quantitative polymerase chain reaction (PCR) were carried out to investigate EphB2 protein localizations and messenger RNA (mRNA) expression levels. (1) Rat embryos with ARMs were obtained by treating pregnant rats (n = 24) with administration of ETU on gestation day (Gd) 10. Normal rat embryos (n = 111) and embryos treated by ETU without ARMs (n = 90) were the control groups, and embryos with ARMs (n = 108) from Gd13 to Gd16 were divided according to the sections taken from specimens. (2) Embryos were sequentially sectioned in the sagittal and transversal planes before staining with a specific antibody to EphB2. Spatiotemporal study was carried out on EphB2 expression. (3) Individual frozen sections were used to manually microdissect the cloaca and anorectal specimens for total RNA extraction. EphB2 expression was evaluated by real time quantitative PCR.. On the immunologic labeling study, EphB2 expression was confined to the cloaca in control groups, whereas EphB2 expression was mainly located at the urorectal septum (URS) and cloacal membrane on Gd13 and Gd14. The increased positive expression was observed in the fused tissue of the URS and cloacal membrane on Gd15. On Gd16, the anal membrane broke down, and the rectum was able to be in contact with the anus, and EphB2 expression was then noted in mucous membrane of rectum. EphB2 expression was seen in the cloacal and anorectal tissues of embryos with ARMs. By integrated optical density (IOD) measurement, IOD value of EphB2 protein was significantly lower in the ARM group than that in the control groups on Gd13 to Gd16 (P < .05), respectively. As shown by real time quantitative PCR, EphB2 expression was detected in 3 groups. EphB2 mRNA level increased on Gd13 to Gd16 but gradually decreased after Gd16. The expression level of EphB2 mRNA in the ARM embryos was lower on Gd13 to Gd16 than that in control groups (P < .05).. EphB2 expression decreased in the ARM embryos and was confined to URS and cloaca, whereas it was higher in control group. Our data thus indicated that EphB2 molecules possibly contributed to the anorectal morphogenesis and the decreased expression of EphB2 might be related to the development of ARMs.

Topics: Anal Canal; Animals; Cloaca; Disease Models, Animal; Ethylenethiourea; Immunohistochemistry; Intestinal Mucosa; Polymerase Chain Reaction; Rats; Rats, Wistar; Receptor, EphB2; Rectum; RNA, Messenger

Intestinal constipation is one of the most commonly occurring complaints in the postoperative period after correction of anorectal malformation (ARM). An abnormal density of interstitial cells of Cajal (ICC) is one potential cause. The objective of this study was to analyze the density of ICC in the terminal intestine of fetuses of rats with anorectal anomaly induced by ethylenethiourea (ETU).. The fetuses were distributed into three groups: Group A--normal fetuses obtained from pregnant female rats that did not receive ETU; Group B--fetuses with no ARM, obtained from pregnant rats that received ETU, and Group C--fetuses with ARM, obtained from pregnant rats that received ETU. ETU was administered on the 11th day of pregnancy at a dose of 125 mg/kg. The fetuses were extracted by means of laparotomy on the 21st day of pregnancy. The terminal intestine of the fetuses was removed and analyzed by immunohistochemistry to evaluate ICC.. Statistically significant differences were found between groups A, B and C regarding the density of ICC. Group A presented with the highest density, followed by groups B and C.. There is a lower density of ICC in the terminal intestine of rats with ARM.

Topics: Anal Canal; Animals; Anus, Imperforate; Cell Count; Disease Models, Animal; Ethylenethiourea; Female; Muscle, Smooth; Myenteric Plexus; Pregnancy; Rats; Rats, Wistar; Rectum

This study was designed to determine the expression of Sonic hedgehog (Shh) and its downstream genes during development of the enteric nervous system (ENS) in ethylenethiourea (ETU)-exposed fetal rats with anorectal malformations (ARMs).. Anorectal malformations were induced by 1% ETU (125 mg/kg) given on gestational day 10, and the litter was harvested at term. The fetal anorectum and rectosigmoid region, including any communication with the urinary tract, were collected for gene expression studies and immunofluorescence study of the ENS. Gene expression of Shh cascade was performed using reverse transcription and real-time polymerase chain reaction (PCR). The myenteric plexuses of the ENS in normal and ARM rats were visualized with fluorescent antibodies.. Reverse transcription-PCR confirmed expression of Shh and its target genes in all parts of the ARMs. Quantitative PCR demonstrated that levels of expression of the genes of the Shh cascade were low in the ARMs. The immunoreactivity of neuromarkers was markedly reduced in high ARMs and slightly reduced in low ARMs.. This study demonstrates reduced expression of Shh and its target genes in ARMs in ETU-exposed fetal rats. Neurons in the myenteric plexus were decreased in high and low types of ARMs. Our results support a role for the Shh cascade during development of the ENS during hindgut development.

Topics: Anal Canal; Animals; Base Sequence; Digestive System Abnormalities; Disease Models, Animal; Enteric Nervous System; Ethylenethiourea; Female; Fetal Diseases; Fluorescent Antibody Technique; Gene Expression Regulation, Developmental; Hedgehog Proteins; Molecular Sequence Data; Pregnancy; Random Allocation; Rats; Rats, Sprague-Dawley; Rectum; Reference Values; Reverse Transcriptase Polymerase Chain Reaction; Sensitivity and Specificity

shh signaling pathway has been shown to be involved in the morphogenesis of many organ systems. In this study, we investigated the expression of shh and its targets, BMP4 and Hox genes, in the development of anorectal malformations in Ethylenethiourea (ETU)-exposed embryos.. We used ETU murine model of the vertebral, anal, cardiac, tracheoesophageal, renal, and limb association. Ethylenethiourea 1% (125 mg/kg) was given to the pregnant females via gavage feeding on gestational day (gD) 10 and saline to control animals. Embryos were collected at gD12 to gD16 and gD21; hindguts were dissected and snap frozen. Highly purified RNA was isolated, and expression of shh, BMP4, Hoxa13, and Hoxd13 genes was confirmed with RT-PCR. Relative quantitative expression of shh and target genes at each time point was done with SYBR Green I qPCR. Normalized gene of interest expression was calculated by geNorm, and data analysis was done with 2-tail Student t test.. shh, BMP4, Hoxa13, and Hoxd13 transcripts were detected in all samples, confirming that shh cascade is active during the process of hindgut development in fetal rats. Relative quantitation demonstrated that shh cascade expression shows time-dependent changes in the developing hindgut.. This study shows that ETU disturbs the expression of shh signaling pathway during the development of hindgut. We provide evidence that shh plays a pivotal role in the hindgut morphogenesis, and its misexpression affect the expression of targets, BMP4 and Hox genes.

Topics: Anal Canal; Animals; Bone Morphogenetic Protein 4; Bone Morphogenetic Proteins; Disease Models, Animal; Ethylenethiourea; Female; Genes, Homeobox; Hedgehog Proteins; Pregnancy; Prenatal Exposure Delayed Effects; Rats; Rats, Sprague-Dawley; Rectal Diseases; Rectum; Signal Transduction

Lumbosacral vertebral abnormalities are a common association of anorectal malformations (ARMs) and are one of the determinants of the eventual level of fecal continence that can be achieved. This study used a fetal rat model to investigate the spectrum of axial skeletal maldevelopment that may occur with ARMs. Time-mated pregnant rats received 125 mg/kg of 1% ethylenethiourea (ETU) (experimental group) or vehicle only (control). Their fetuses were examined for external malformations and prepared for staining of their skeletons using Alcian blue and Alizarin red S. ARMs developed in 67/68 (98%) of ETU-exposed fetuses, of which 28 (42%) also developed rachischisis, mainly involving the lumbosacral vertebrae. No skeletal abnormality was found in control fetuses. ETU-exposed fetuses with ARMs and rachischisis had abnormal ossification of the vertebral centrum, abnormal fusion between the neural arches of vertebrae, localized narrow or interrupted thoracic vertebral canal, a widely open vertebral canal in the lumbosacral area (rachischisis), and absence of the lower two sacral and coccygeal vertebrae. Rib abnormalities included absence of two to three floating ribs, abnormal fusion of adjacent proximal segments, and abnormal ramification, irregularity, and angulation of their distal segments. The vertebral and rib abnormalities found in ETU-exposed fetuses with ARMs but no rachischisis were much less severe. In addition to the lumbosacral anomalies that are common with ARMs, severe abnormalities of the thoracic vertebrae and their corresponding ribs may occur also. Fetuses with both ARM and rachischisis tend to have more extensive and severe vertebral and rib anomalies. These observations imply a possible common aetiology for ARMs and vertebral anomalies and are consistent with our understanding of the perceived role of the notochord in axial development.

Topics: Abnormalities, Drug-Induced; Anal Canal; Animals; Disease Models, Animal; Ethylenethiourea; Female; Fetus; Lumbar Vertebrae; Notochord; Osteogenesis; Pregnancy; Rats; Rats, Sprague-Dawley; Rectum; Ribs; Sacrum; Spinal Canal; Spinal Dysraphism; Thoracic Vertebrae

The notochord is known to organize normal development of central axial structures, such as the spinal cord, vertebral column, and anorectum, but its role in abnormal development of these organs has not been well documented. The current study has used Ethylenethiourea to induce anorectal malformations in fetal rats, allowing investigation of abnormalities of the notochord and their relationship to the axial structural abnormalities that occur.. Timed-mated pregnant rats were fed Ethylenethiourea by gavage on gestational day 10. Their embryos were harvested on gestational days 13 to 16 and sectioned in either the transverse or sagittal plane. Sections were stained with H and E and examined serially.. Anorectal malformations were identified in 29 of 34 embryos and neural tube defects in 24, ranging from an accessory neural tube to lumbo-sacral rachischisis. There was no tail or only a rudimentary tail in the majority of embryos. Abnormalities of the notochord in the lumbo-sacral area included ventro-dorsal branching, ventral deviation, and ectopic notochordal tissue. Most abnormal notochord branches and ectopic notochordal tissue were abnormally close to or in contact with the wall of the cloaca or neural tube.. Given the known role of the notochord in controlling normal development, this study would suggest that abnormal notochord development may be pivotal in producing neural tube defects and anorectal malformations, possibly by altering sonic hedgehog signalling.

Topics: Anal Canal; Animals; Digestive System Abnormalities; Disease Models, Animal; Ethylenethiourea; Neural Tube Defects; Notochord; Organogenesis; Rats; Spinal Cord; Spine

Deficiency of motoneuron innervation to the sphincter mechanism has been described in patients with anorectal malformation. Whether this event is primary or secondary remains unclear.. The authors quantified the motoneuron innervation of the sphincter mechanism by Fluorogold (FG) retrograde tracing experiment in fetal rats with anorectal malformation. Anorectal malformation was induced in rat fetuses by ethylenethiourea (ETU). Serial longitudinal sections encompassing the whole width of lumbosacral spinal cord were examined. The number of FG-labelled motoneurons were scored and compared between male fetuses with or without malformation in the ETU-fed group and normal controls.. The number of FG-labelled motoneurons in the fetuses without defect, with imperforate anus (IA), with neural tube anomalies (NTA), with combined IA and NTA, and normal controls were determined to be (mean +/- SEM) 109.13 +/- 37.88, 55.05 +/- 25.85, 48.20 +/- 30.34, 54.43 +/- 28.55, and 135.22 +/- 28.78, respectively. FG-labelled motoneurons in the fetuses with IA, NTA, and combined IA and NTA are significantly fewer than that in fetuses without defects (P <.05) and in normal controls (P <.005).. These findings suggest that defective motoneuron innervation to the sphincter mechanism is a primary anomaly that coexists with the alimentary tract anomaly in anorectal malformation during fetal development. The intrinsic neural deficiency is an important factor likely to contribute to poor postoperative anorectal function despite surgical correction of anorectal malformation.

Topics: Anal Canal; Animals; Anus, Imperforate; Disease Models, Animal; Ethylenethiourea; Female; Fluorescent Dyes; Male; Neural Tube Defects; Pelvic Floor; Rats; Rats, Wistar; Spinal Cord; Stilbamidines

Ethylenethiourea (ETU) administered to timed-pregnant rats can induce anorectal malformations (ARMs) in about 80% of rat fetuses, thus providing an ideal animal model to study the embryogenesis of ARMs. The current study was undertaken to investigate the embryogenetic events that may be responsible for the development of ARMs in rats.. Time-mated pregnant rats were divided randomly into control and experimental groups. The experimental rats received 1% ETU (125 mg/kg) by gavage on gestational day 10, and control rats received only the vehicle. Their embryos were harvested by cesarean section on gestational days 13, 14, 15, and 16. They were fixed and embedded in paraffin and serially sectioned in either the sagittal or transverse plane. The sections were stained with H&E, examined, and photographed. The comparative morphogenesis of the hindgut, cloaca, and tailgut of age-matched embryos was studied.. The key abnormalities in the experimental embryos were: (1) maldevelopment of cloaca and urorectal septum with no sign of the fusion between the urorectal septum and the cloacal membrane, (2) delay of tailgut regression, (3) abnormal and massive apoptotic cell death involving the posterior cloacal wall, and (4) underdevelopment of the dorsal aspect of the cloaca and its membrane. The type of ARM that was developing was discernible by gestational day 15 and 16.. ARMs induced by ETU in rat embryos seem to be caused by the cumulative effect of aberrations in the development of several components of the hindgut and cloaca. Variation in the extent of maldevelopment of these structures may result in a spectrum of ARMs.

Topics: Anal Canal; Animals; Anus, Imperforate; Cloaca; Disease Models, Animal; Ethylenethiourea; Female; Pregnancy; Rats; Rats, Sprague-Dawley; Rectum

Overgrowth of the myeloschisis, namely the excessive amount of the neural plate tissue, has been reported in the human myeloschisis. However, it is still debatable how the overgrowth develops and whether the overgrowth is the cause, or the secondary effect of spinal dysraphism. The author induced myeloschisis in the fetuses of Long-Evans rats by the administration of ethylenethiourea (ETU) to pregnant rats on day 10 of gestation. The fetuses were removed 1 hour after the treatment with bromodeoxyuridine (BrdU) to the dams on day 14 and 21. The fetuses were fixed in alcohol and embedded in paraffin. H-E staining and the immunohistologic examination were performed on the staining patterns to anti-neurofilament (NFP), anti-glial fibrillary acidic protein (GFAP) and anti-BrdU antibody by ABC method. On day 14, the lateral portion of everted neural plate showed a loose arrangement of cells and there was rosette formation in the mesoderm. On day 21, cell necrosis was observed at the dorsolateral portion of myeloschisis, although the ventral portion showed almost normal cytoarchitecture and was positive to NFP and GFAP. The cause of myeloschisis in this model is supposed to be the local and direct cytotoxic effect of ETU to neuro-ectodermal junction. On day 14, control animals contained few BrdU-incorporated cells at the basal plate of neural tube. In contrast, everted neural plate showed an active uptake of BrdU diffusely in the subependymal matrix layer cells. Overgrowth was not yet identified. On day 21, overgrowth of myeloschisis was found in spite of a few positive cells to BrdU which was identical to the control animals. These findings seem to suggest that cells in the myeloschisis retain their ability of DNA synthesis for longer periods of development and overgrowth found on day 21 is possibly a secondary effect of spinal dysraphism in this model.

Topics: Animals; Bromodeoxyuridine; Disease Models, Animal; DNA; Ethylenethiourea; Female; Male; Pregnancy; Rats; Spinal Cord; Spinal Dysraphism