Bao Quan Qi

Evidence of a common pathogenesis for foregut duplications and esophageal atresia with tracheo‐esophageal fistula

The pathogenesis of the alimentary tract duplications, including foregut duplications (FgD) remains speculative. The accidental finding of FgD in fetal rats with esophageal atresia and tracheoesophageal fistula (EA‐TEF) induced by Adriamycin provided an animal model to investigate a possible relationship between these two entities. Timed‐pregnant rats were intraperitoneally injected with Adriamycin (1.75 mg/kg) on gestational Days 6 to 9. Their embryos were harvested by Caesarean section from gestational Days 14 to 21. Forty‐six of embryos were processed and serially sectioned in the transverse or sagittal planes. EA‐TEF occurred in 43/46 (93%) embryos of which 11 (24%) were found to have an associated FgD located at the level where the esophagus was absent. Six FgDs communicated with the foregut or the trachea. Five noncommunicating FgDs were located between the foregut and the vertebral column. In the control embryo, the notochord was located in the centre of the vertebral column from Day 11 of the gestation. In Day 14, 15 and 16, however, embryos exposed to Adriamycin, an abnormal notochord or branch frequently was located within the mesenchyme of the maldeveloped foregut or attached to the duplication cyst. In some, it appeared that the notochord was drawing the cyst‐like structure away from the foregut. The present study confirms that duplications adjacent to the esophagus arise from the foregut and that failure of the foregut to detach from the notochord at the normal time may contribute to the development of foregut duplications. Anat Rec 264:93–100, 2001.

The contribution of the adriamycin-induced rat model of the VATER association to our understanding of congenital abnormalities and their embryogenesis

Abstract The adriamycin-induced rat model of the VATER association has provided a means of studying the morphogenesis of a variety of major congenital structural abnormalities similar to those seen in humans with the VATER association. Most interest has been centered on the foregut, where the model has clarified some aspects of the development of esophageal atresia (EA), tracheal agenesis, and other communicating bronchopulmonary foregut malformations. It has demonstrated aberrations in the nerve supply to the esophagus in EA and allowed the study of tracheomalacia. A relationship between an abnormal notochord, foregut abnormalities, and vertebral defects has been shown, and the model has reignited interest in the role of the notochord as a regional organizer of axial development. The normal temporospatial characteristics of apoptosis during fore- and hindgut development is disturbed in this model, resulting in abnormal morphology. The indications are that this model will continue to clarify the processes that lead to many of the structural congenital abnormalities that are seen in infants born with the VATER association.

Relationship of the notochord to foregut development in the fetal rat model of esophageal atresia

BACKGROUND/PURPOSE The notochord (Nt) is thought to act as a primary organizer for adjacent axial embryonic organs. The current study used the Adriamycin-induced fetal rat model of esophageal atresia and tracheoesophageal fistula (EA-TEF) to determine whether anomalies of the foregut (FG) were associated with an abnormal Nt. METHODS Eight experimental female Sprague-Dawley rats received intraperitoneal injection of Adriamycin (1.75 mg/kg) on gestational days 6 to 9 inclusive, and 4 control rats received saline injection only. Their embryos were harvested on gestational days 11, 12, 13, and 14. Embryos from each age subgroup were serially sectioned and stained with H&E. The FG and Nt were traced from the primitive pharynx to the level of the stomach. RESULTS By day 11, the Nt of control embryos had completely separated from the FG and was located immediately ventral to the neural tube. On gestational day 12, the Nt detached from the neural tube, and the trachea and esophagus were separating. On day 11, in the Adriamycin-treated embryos, the Nt was still attached to an FG that was narrowed or occluded. On day 12, the Nt remained adherent to the FG from the primitive pharynx to the level above the primitive respiratory buds, at which point it became thicker and branched sagittally, with the anterior branch contacting or merging with the FG. The FG usually loses its lumen or continuity when in contact with the Nt. CONCLUSIONS Exposure of rat embryos to Adriamycin leads to abnormal development of the Nt, including prolonged attachment to or fusion with the FG, and abnormal branching. Traction on the FG by the Nt produces occlusion of its lumen and may result in its complete interruption. Separation of the Nt from the FG would appear to be a prerequisite for the normal development of the FG into its derivatives: the esophagus and trachea.

Hedgehog in the human: A possible explanation for the VATER association

Abstract: Foregut malformations are relatively common anomalies, occurring in 1 in 2000–5000 live births. The adriamycin‐induced rat model of the VATER association has provided a means of studying the morphogenesis of a variety of major congenital structural abnormalities similar to those seen in humans with VATER association. The secreted glycoprotein, Sonic hedgehog (Shh), may act as an endodermal signal that controls gut and lung patterning. Mice with targeted deletion of Shh have foregut defects that are consistent with those produced by administration of adriamycin. It is possible that mutations induced by adriamycin may result from the breakdown of the Shh signalling pathway.

Stages of normal tracheo-bronchial development in rat embryos: Resolution of a controversy

The embryonic events surrounding tracheo‐esophageal separation remain controversial. The present study was undertaken to clarify early tracheo‐bronchial development in the rat embryo at a critical period of organogenesis. Twenty‐seven timed‐mated Sprague‐Dawley rats were divided into nine groups of three rats. Their embryos were harvested on gestational days 11–15 at intervals of 8 h, processed and sectioned transversely. The sections were stained with hematoxylin and eosin and examined serially. The foregut is a single tube on gestational day 11. During the following 16 h, there is localized and rapid growth of the respiratory epithelium and a latero‐ caudal expansion to form the bronchial buds and a protuberance on the ventral wall of the foregut (future tracheal carina). From gestational days 12–12 + 8, cellular debris and apoptotic epithelial cells are specifically located in the tracheo‐esophageal groove, resulting in collapse and fusion of the lateral walls of the foregut, effectively separating the trachea and esophagus. Afterwards, the epithelial proliferation dominates the process of tracheo‐esophageal separation until it reaches the caudal end of the laryngeal epithelial lamina on gestational day 15. The present study shows that separation of the trachea from the esophagus involves three consecutive stages: (i) epithelial proliferation resulting in the formation of bronchial buds and the tracheal carina; (ii) epithelial apoptosis leading to separation of the trachea and esophagus; and (iii) epithelial proliferation to complete the separation process.

Demonstration of abnormal notochord development by three-dimensional reconstructive imaging in the rat model of esophageal atresia.

Abstract The notochord (Nt) is believed to have a role in the development of axial organs. This study was undertaken to reconstruct in three dimensions (3D) the relationship of the Nt to abnormal development of the foregut (Fg) in the adriamycin-induced rat model of esophageal atresia (EA). Pregnant Sprague-Dawley rats were given 1.75 mg/kg adriamycin intraperitoneally on gestational days 6–9 inclusive; control rats received i.p. saline of equal volume, or no injection. Rats were killed between days 11 and 14 and their embryos harvested, histologically sectioned serially, and stained with hematoxylin and eosin. Digitized photographs were taken of serial transverse sections; these photos were traced and used as the basis for 3D reconstruction. From day 11 the normal Nt is no longer in contact with the respiratory or Fg mesenchyme. In adriamycin-treated embryos the Nt branches abnormally as it enters the Fg mesenchyme. Adherence of the Nt to the mesenchyme of the Fg exerts mechanical traction pulling the upper Fg dorsally. The severity of the Fg abnormalities correlates with the length of the ventral extension of the Nt within the Fg mesenchyme: the embryo develops atresia of the esophagus or trachea when the Nt is grossly abnormal. The Nt undergoes reactive thickening in the absence of Fg structures ventral to it. Thus, structural lesions of the Fg (e.g., atresias) are associated with abnormalities of the Nt. The relationship of the Nt to the Fg mesenchyme determines the severity of the abnormality induced by adriamycin: extensive adherence produces tracheal agenesis and EA.

Evidence that the notochord may be pivotal in the development of sacral and anorectal malformations

BACKGROUND/PURPOSE 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. METHODS 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. RESULTS 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. CONCLUSIONS 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.

Does the urorectal septum fuse with the cloacal membrane

PURPOSE Traditional theories of cloacal embryogenesis assume that the urorectal septum fuses with the cloacal membrane before the anal membrane disintegrates. However, recent observations in humans and other species raise doubt about this assumption. We determined whether urorectal septum fusion occurs in rats. MATERIALS AND METHODS Rat embryos were harvested at specific times between days 11 and 16 of gestation. We evaluated the morphology, growth and relationship of the urorectal septum to the cloacal membrane on serial histological sections. RESULTS The urorectal septum consistently fused with the cloacal membrane on day 15 of gestation before the cloacal membrane began to disintegrate. CONCLUSIONS In rats the urorectal septum fuses with the cloacal membrane, after which the urogenital membrane and anal membrane disintegrate by a process of apoptosis.

PATTERNS OF APOPTOSIS DURING DEGENERATION OF THE PRONEPHROS AND MESONEPHROS

PURPOSE The adult mammalian kidney is preceded developmentally by 2 primitive kidneys. We determine whether apoptosis is involved in the regression of these primitive organs and document its temporospatial characteristics. MATERIALS AND METHODS Timed pregnant rats were sacrificed at 11 to 16 days of gestation inclusively. The fetuses were histologically examined to determine the timing and pattern of apoptosis in the primitive kidneys, including pronephros and mesonephros. RESULTS Apoptosis of the pronephros occurred primarily at 12 days of gestation. Apoptosis of the distal mesonephros occurred primarily at 13 days and was completed by 14, which left the proximal mesonephros a functioning kidney during the early period of development. The location and timing of apoptosis were consistent and specific. CONCLUSIONS Apoptosis appears to be an important mechanism of the normal regression of the primitive kidney and follows a strict temporospatial pattern.

Abnormalities of the tracheal cartilage in the rat fetus with tracheo-oesophageal fistula or tracheal agenesis.

Abstract Many infants with oesophageal atresia and tracheo-oesophageal fistula (OA/TOF) have associated tracheomalacia (TM), which is one of the reasons for respiratory complications after surgical correction of the atresia. OA/TOF was induced in the offspring of pregnant rats by intraperitoneal injection of adriamycin. Fetuses were harvested by caesarean section. The trachea, oesophagus, lungs, and stomach were removed en bloc and stained for cartilage using Alcian blue. The tracheas were examined, photographed, and relevant parameters pertaining to the tracheal cartilage were measured. Exposure to adriamycin resulted in a range of anatomical defects including OA/TOF (47%) and tracheal agenesis (TA) (41%). Adriamycin-treated fetuses were smaller (P < 0.01), yet had longer tracheas (P < 0.001) than control fetuses. The OA/TOF fetuses had more tracheal cartilage rings than controls (P < 0.01), whereas TA fetuses had fewer (P < 0.001). Both OA/TOF and TA fetuses had more malformed tracheal cartilage rings than controls (P < 0.001 and P < 0.05, respectively). Cartilage in the proximal part of the trachea was most frequently and severely affected (P < 0.05). These observations clarify the structural abnormalities of tracheal cartilage that occur in rat fetuses with OA/TOF or TA induced by adriamycin, and may explain the functional disturbances of TM seen in OA/TOF.