Pam Farmer

The vagus and recurrent laryngeal nerves in the rodent experimental model of esophageal atresia

BACKGROUNDnAfter surgical correction of their esophageal atresia and tracheoesophageal fistula (EA-TEF), many patients exhibit evidence of esophageal dysmotility. Controversy exists as to whether the esophageal motility disorders result from denervation caused by surgery or from an inherent abnormal innervation of the esophagus.nnnMETHODSnThe present study used an Adriamycin-induced EA-TEF fetal rat model to trace the course and branching of both the vagus and recurrent laryngeal nerves. Abnormalities observed in EA-TEF rat fetuses include: (1) fewer branches from both recurrent laryngeal nerves; (2) deviation of the left vagus from its normal course below the aorta, passing behind the fistula to approach and join with the right vagus to form a single nerve trunk on the right side of the esophagus; (3) relatively few branches from the single vagal nerve trunk (composed of fibers of the left and the right vagus) on the surface of the lower esophagus.nnnCONCLUSIONSnFetuses affected by EA-TEF have inherent abnormalities in the course and branching pattern of the vagus nerves as they descend through the thorax, culminating in a deficient extrinsic nerve fiber plexus in the lower esophagus. These observations may account for the esophageal motility disorders seen in patients who have EA-TEF even before surgical intervention.

Intrinsic innervation of the oesophagus in fetal rats with oesophageal atresia

Although the aetiology of oesophageal dysmotility after repair of oesophageal atresia and tracheo-oesophageal fistula (OA-TOF) remains controversial, oesophageal dysmotility also is present in isolated TOF or OA before surgery, suggesting a congenital cause. Our previous work with a model of OA-TOF in fetal rats demonstrated an abnormality in the course and branching pattern of the vagus nerve. However, little is known about the intramural nervous components of the atretic oesophagus. The intrinsic innervation of the atretic oesophagus was examined by immunohistological staining to see if there is an abnormality that might account for dysmotility. OA-TOF was induced in fetal rats by injecting adriamycin intraperitoneally into pregnant rats. Forty-eight controls, 40 OA-TOF, and 6 treated fetuses without OA-TOF were recovered. Whole-mount preparations of each oesophagus were stained with fluorescent antibodies against neuron-specific enolase (NSE), vasoactive intestinal peptide (VIP), substance P (SP), and calcitonin gene-related peptide (CGRP). Compared with control fetuses, the density of the nerve plexus, ganglia, and number of cell bodies per ganglion immunostained by NSE, VIP, or SP was significantly reduced in OA-TOF fetuses. CGRP-immunoreactive nerve fibres in the oesophageal wall of both control and OA-TOF animals were found to be connected with extrinsic nerve bundles. No plexus-like nerve fibre network was observed. The results of the present study demonstrated significant abnormalities of the intramural nervous components of the oesophagus in OA-TOF fetal rats, involving both the excitatory (SP-labelled) and inhibitory (VIP-labelled) intramural nerves. These abnormalities may underlie the oesophageal dysmotility seen in OA-TOF patients.

In vitro fusion of human inguinal hernia with associated epithelial transformation.

The processus vaginalis (PV) is a peritoneal diverticulum which forms to allow descent of the fetal testis to the scrotum. During human development fusion and obliteration of the PV often fails to occur with the result that inguinal hernias are the most prevalent congenital abnormality requiring surgery in childhood. Androgen is proposed to regulate testicular descent via the genitofemoral nerve which releases the neuropeptide calcitonin gene-related peptide (CGRP). It is possible that subsequent fusion of the PV and tissue remodelling following descent is indirectly controlled by androgen via CGRP action. An organ culture assay was developed to assess fusion of the PV taken from inguinal herniotomy in infants. Fusion was induced in vitro by CGRP but not by CGRP 8–37, CGRP 27–37 or dihydrotestosterone in equimolar concentrations. Fusion was accompanied by transformation of the epithelium, as shown by staining of intermediate filament proteins, cytokeratin and vimentin. Localization studies for CGRP receptors on 25 specimens indicated CGRP acts on mesenchymal fibroblasts but not directly on PV epithelium suggesting an indirect pathway. Hepatocyte growth factor/scatter factor was found to induce fusion of PV and may be involved as an intermediate molecule in the fusion cascade. This study represents the first approach to understanding the humoral control and underlying mechanism by which the PV fuses.

Substance P and vasoactive intestinal peptide are reduced in right transverse colon in pediatric slow‐transit constipation

Backgroundu2002 Slow‐transit constipation (STC) is recognized in children but the etiology is unknown. Abnormalities in substance P (SP), vasoactive intestinal peptide (VIP) and nitric oxide (NO) have been implicated. The density of nerve fibers in circular muscle containing these transmitters was examined in colon from children with STC and compared to other pediatric and adult samples.

Visceral anomalies in prenatally adriamycin-exposed rat fetuses : a model for the VATER association

Abstract Adriamycin is teratogenic if given to pregnant rats. A wide range of anomalies involving the gastrointestinal, renal, and cardiovascular systems has been described, similar to the VATER association, yet it is not known if they are identical to the human pattern. The aim of this study was to document the visceral anomalies in rat fetuses exposed to adriamycin and to determine their similarities with the congenital defects in humans with the VATER association. The results revealed a spectrum of very similar anomalies. Furthermore, the characteristics of the tracheo-oesophageal anomalies had a lot of features in common with the human pattern. We conclude that the adriamycin-treated fetal rat is an excellent model for studying the VATER association.

Timing and embryology of esophageal atresia and tracheo-esophageal fistula.

The embryology of tracheo‐esophageal anomalies is controversial. The development of an adriamycin‐treated animal model has enabled improved understanding of the embryogenesis of these anomalies. Using this model, we aimed to describe the events leading to esophageal atresia and tracheo‐esophageal fistula.

Cardiovascular malformations in rat fetuses with oesophageal atresia and tracheo-oesophageal fistula induced by adriamycin

Associated congenital anomalies have emerged as the most significant prognostic factor in babies born with oesophageal atresia and/or tracheo-oesophageal fistula (OA-TOF). The most frequently encountered groups of anomalies are cardiovascular (CV) and gastrointestinal, the former being more significant from a prognostic point of view. Some, such as a right-sided aortic arch (RAA), vascular ring, or major heart defects, may alter the timing and surgical approach for the repair of OA-TOF or adversely affect the prognosis. The rat fetal OA model induced by adriamycin (Adr) has been described previously. In the present experiments, information was sought regarding the incidence and type of CV abnormalities in fetal rats obtained from Adr-treated dams. OA-TOF was induced in 24 of 36 fetal rats from Adr-treated dams. DV abnormalities were found in 18 (75%) OA-TOF fetuses and 10 (83%) Adr-treated fetuses without OA-TOF. The difference was not significant (P > 0.05). The most frequently found anomalies were ventricular and atrial septal defects. A RAA was present in 8/36 fetuses and a double aortic arch in 2/36. A patent ductus arteriosus was present in all treated fetuses compared with two-thirds of controls. The findings in the present study emphasise the importance of CV anomalies in association with OA, and reinforce the value of the Adr-induced rat fetal OA model by adding to our knowledge of the basic embryogenesis of both OA and CV anomalies.

Tracheomalacia with esophageal atresia and tracheoesophageal fistula in fetal rats

BACKGROUNDnMany patients who have esophageal atresia and tracheoesophageal fistula (EA-TEF) have associated tracheomalacia, which is thought to be one of the reasons for respiratory complications after surgical correction of the abnormality.nnnMETHODSnIn this study, tracheas from Adriamycin-induced EA-TEF fetal rats were examined histologically and relevant cross-sectional parameters of the tracheas were measured.nnnRESULTSnThe tracheal lumen in tracheomalacia was small and irregular, losing its normal D shape. In most rats, the cartilaginous ring was broken into two to four segments, making the trachea lose its rigid support. The submucosa was thickened with prominent bulging of its membranous part into the tracheal lumen. The ratio of the inner luminal cross-sectional area to the outer tracheal cross-sectional area in EA-TEF rats was 15.7%, compared with a control ratio of 47.2%. In EA-TEF rats, the length of the cartilaginous ring was significantly shortened (P < .001), but not the length of membranous trachea, thus resulting in a cartilaginous/membranous (C/M) ratio of 1.55:1, markedly lower than that of normal rats (4.34:1, P < .001). The reduction of anterior-posterior diameter of the tracheal lumen was more marked than that of the transverse diameter.nnnCONCLUSIONSnThese observations suggest that the trachea in EA-TEF rats has a smaller lumen and is more flaccid than normal, making it prone to airway obstruction. The fact that tracheomalacia developed only in fetuses who had EA-TEF indicates that the factors that result in EA-TEF also cause tracheomalacia.

Calcitonin gene-related peptide is a survival factor, inhibiting apoptosis in neonatal rat gubernaculum in vitro

BACKGROUNDnTesticular descent is proposed to occur in 2 stages. During the second stage, calcitonin gene-related peptide (CGRP) released from the genitofemoral nerve (GFN) causes maximal mitosis in the gubernacular bulb. As normal development requires a balance between cell proliferation and apoptosis, this study explored the effect of CGRP on apoptosis in the rat gubernacular bulb.nnnMETHODSnGubernacula were collected from male Sprague-Dawley rats at birth (D0) or 2 days post birth (D2), and placed in organ culture for 24 hours with or without CGRP (0.001 mol/L). The D2 rats were pretreated with capsaicin (sensory nerve toxin) or flutamide (antiandrogen) or untreated. D0 rats were untreated (n = 64). Sections of the bulb were stained using the TUNEL method to identify apoptotic cells. Apoptosis was calculated as the percentage of positive cells per hundred cells.nnnRESULTSnNormal Sprague-Dawley rat gubernacula showed reduced apoptosis when cultured with CGRP, in D0 (7.0% vs 4.8%, P < .05) and D2 (4.9% vs 2.3%, P < .001). Greater apoptosis occurred at D0 compared to D2, without CGRP added (7.0% vs 4.9%, P < .05) and with CGRP (4.8% vs 2.3%, P < .001). For D2 gubernacula, capsaicin treatment increased apoptosis compared to controls, without CGRP added (4.9% vs 7.3%, P < .05) and with CGRP (2.3% vs 6.7%, P < .001). There was no difference in apoptosis when cultured with or without CGRP (7.3% vs 6.7%, nonsignificant) after capsaicin treatment. Flutamide treatment increased apoptosis compared to controls, but only with CGRP (2.3% vs 7.3%, P < .001). There was no difference in apoptosis when cultured with or without CGRP (7.1% vs 7.3%, nonsignificant) after flutamide.nnnCONCLUSIONSnCalcitonin gene-related peptide (CGRP) acts as a survival factor in the rat gubernaculum, possibly to steer cells away from a defined apoptotic pathway. Greater apoptosis occurs earlier in development. However, in vivo CGRP released from the genitofemoral nerve may be required to prevent apoptosis, as shown by pretreatment with the sensory nerve toxin capsaicin. Androgen is also involved in the pathway controlling apoptosis, as androgen blockade with flutamide inhibited the action of CGRP.

Embryogenesis of Tracheal Atresia

A spectrum of tracheo‐esophageal anomalies has been described in an adriamycin‐treated model with common features to the human pattern. Tracheal agenesis was part of this spectrum. It is a rare congenital anomaly that has not been described in embryos.