Down-regulation of thyroid transcription factor-1 gene expression in fetal lung hypoplasia is restored by glucocorticoids.

The thyroid transcription factor (TTF)-1 has an essential role in lung morphogenesis and development. It is involved in the transcription of surfactant proteins (SP), which are critical in respiratory function. Neonates with congenital diaphragmatic hernia die of respiratory failure caused by pulmonary hypoplasia with associated biochemical immaturity. To gain new insights into the causes of this disorder and the effect of prenatal hormonal treatment on reducing mortality in these infants, we evaluated the expression of TTF-1 as marker of lung morphogenesis and SP-B as marker of lung maturity. Using a rat model of lung immaturity, we show that TTF-1 and SP-B messenger RNA (mRNA) levels are drastically reduced in congenital lung hypoplasia. Interestingly, prenatal dexamethasone (Dex) treatment increased both TTF-1 and SP-B mRNAs over control levels when administered to rats with lung hypoplasia, but it had no effect on TTF-1 or a moderate effect on SP-B mRNA when administered to control rats. TRH alone also increases TTF-1 and SP-B mRNA levels but to a lesser extent than Dex. When administered together with Dex, TRH counteracts the induction observed with the glucocorticoid. The decrease in TTF-1 mRNA levels in lung hypoplasia is paralleled by a down-regulation of TTF-1 protein levels, as well as by a decrease in the TTF-1/DNA complex when the TTF-1-binding site of the SP-B promoter was used as a probe. Both parameters were reestablished after glucocorticoid treatment. Moreover, the regulation of TTF-1 gene expression described in this report is accompanied by the same regulation in its promoter activity, as demonstrated in transfection experiments performed in H-441 human lung-derived adenocarcinoma cells. In conclusion, our data demonstrate, for the first time, that lung hypoplasia and the associated respiratory dysfunction caused by SP-B deficiency are caused, in part, by down-regulation of TTF-1 gene expression. The observations that prenatal glucocorticoid treatment induces the expression of TTF-1 supports routine in utero glucocorticoid treatment of patients expected to have lung hypoplasia.

Histologic classification of undescended testes.

On the basis of testicular biopsy study in 203 patients and study of a second biopsy specimen from 27 of these patients, prepubertal undescended testes were classified into four categories according to the mean tubular diameter, the tubular fertility index, and the Sertoli cell index. Type I cases (testes with minimal lesions) were characterized by a normal mean tubular diameter and normal tubular fertility and Sertoli cell indexes or slight tubular hypoplasia. This group represented 26 per cent of the undescended testes. The corresponding lesions can be observed from two years of age onward and are probably acquired. After puberty normal spermatogenesis occurs. Type II cases (24 per cent of the undescended testes) included testes with marked germinal hypoplasia as well as slight or marked tubular hypoplasia and a normal Sertoli cell index. After puberty these testes develop a degree of marked hypospermatogenesis, maturation arrest, or Sertoli cells with only isolated spermatogonia and primary spermatocytes. In type III cases (testes with diffuse tubular hypoplasia) the mean tubular diameter and the tubular fertility and Sertoli cell index values were severely reduced. This group represented 33 per cent of the undescended testes, and after puberty most of them showed seminiferous tubules with exclusively adult Sertoli cells. Type IV testes (diffuse Sertoli cell hyperplasia) were associated with a nearly normal mean tubular diameter and variable tubular fertility index values and represented 17 per cent of all the undescended testes. After puberty Sertoli cells do not mature completely, and therefore in spite of the earlier tubular fertility index, the germinal cell line does not reach adult development. Although early orchiopexy prevents tubular fertility index and mean tubular diameter deterioration due to the noxious effects of temperature in type I testes, we believe that there is no such benefit in the other types. These patients may present only slight modifications in these indexes.

Electrophysiologic changes during acute rejection of heterotopically transplanted rat hearts.

Noninvasive monitoring of heart allograft rejection was performed by electrophysiologic techniques using an implanted electrode located on the epicardial surface of the heterotopic transplanted heart of rats. R wave and slew rate determinations were performed daily in 30 syngeneic and 66 allogeneic transplants. These determinations were later compared with histopathologic studies of the transplanted hearts. R wave and slew rate values of allogeneic or rejecting hearts were found to decrease significantly compared with syngeneic or nonrejecting hearts on the days studied. This noninvasive electrophysiologic method may be a promising method for monitoring heart allograft rejection.

A new rodent experimental model of esophageal atresia and tracheoesophageal fistula: preliminary report.

In spite of the interest paid by pediatric surgeons to esophageal atresia (EA) with tracheoesophageal fistula (TEF), no animal experimental model has been available for investigation. This preliminary report describes a reproducible fetal model of these malformations. Time-mated pregnant rats were given 1.5, 1.75, or 2 mg/kg of Adriamycin intraperitoneally on days 6 to 9 of gestation, and the litters were recovered on day 21 (near full-term). The amount of amniotic fluid was measured, and the fetuses were dissected and studied histologically. The findings were compared with those of suitable control fetuses. Adriamycin-exposed fetuses weighed less than controls. EA with TEF (Gross' type C) was found in 28%, 45%, and 41% of animals in the three dose groups (respectively). The malformation was anatomically identical to that of the human neonate, and the amount of amniotic fluid in affected fetuses increased significantly. In one instance, an H-type fistula was observed. In addition to esophageal interruption, many other malformations fitting within the human VATER association were found: duodenal atresia (41%, 50%, and 47%, respectively), anorectal (28%, 50%, and 41%), renal (81%, 100%, and 100%), and limb malformations (0%, 2.3%, and 13.8%). This new, easily reproducible and relatively inexpensive experimental model of one of the most interesting pediatric surgical malformations permits new research into both its embryogenesis and the biology of the malformed fetus.

Transdiaphragmatic pressure gradients and the lower esophageal sphincter after tight abdominal wall plication in the rat.

BACKGROUND: Gastroesophageal reflux (GER) is increasingly recognized as a complication of surgical closure of gastroschisis and omphalocele. AIM: This study tests the hypothesis that forceful abdominal wall closure reinforces the transdiaphragmatic pressure gradients that constitute the main GER-driving force and challenges the antireflux barrier. MATERIALS AND METHODS: Abdominal and esophageal pressures as well as lower esophageal sphincter pressures (LESP) and length (LESL) were measured in 17 adult rats before tight abdominal wall plication, after it, and 1 week later. RESULTS: This maneuver increased the transdiaphragmatic expiratory gradient from 0.67 +/- 1.31 to 6.97 +/- 2.68 mm Hg (P < .01) and the inspiratory gradient from 4.36 +/- 1.13 to 10.79 +/- 2.31 mm Hg (P < .01) by markedly increasing both the expiratory (from 1.47 +/- 0.74 to 9.44 +/- 1.85 mm Hg; P < .01) and inspiratory (from 0.98 +/- 0.69 to 6.83 +/- 1.55 mm Hg; P < .01) intraabdominal pressures. These changes were transient, and all pressures became normal after 1 week. The antireflux barrier functioned properly under these new conditions because both LESP and the diaphragmatic pinch-cock pressure (DPP) increased, from 20.3 +/- 3.63 to 26.5 +/- 4.31 mm Hg (P < .01) and from 16.4 +/- 7.25 to 22.5 +/- 4.36 mm Hg (P < .01), respectively, while LESL remained unchanged. CONCLUSION: Tight abdominal wall plication in the rat generates high intraabdominal pressures and thus reinforces the transdiaphragmatic pressure gradients, but these conditions elicit a healthy barrier response with sphincteric reinforcement. In addition, these changes are transient and fade out some time after operation. These facts should be taken into account for understanding the pathogenesis of GER after repair of abdominal wall defects in human babies.

Intestinal rotation in experimental congenital diaphragmatic hernia.

This study examines whether experimental congenital diaphragmatic hernia (CDH) induced by nitrofen in rats is accompanied by intestinal malrotation similar to that observed in the human condition. Time-dated pregnant rats were fed 100 mg of nitrofen on day 9.5 gestation, and fetuses were examined on days 17, 19, and 21. Body weight, lung weight, grade of bowel herniation into the umbilical cord and grade of intestinal malrotation were compared with those of age-matched controls. Body and lung weights were significantly lower in nitrofen-exposed on days 17, 19, and 21. The umbilical hernia persisted on day 17 in 100% of experimental animals and 66% of controls (P < .01). Intestinal malrotation was more severe in experimental rats than in controls on days 19 (63% v 17% grade 2; P < .01) and 21 (27% v 0% grade 1; P < .01). Finally, 52% of nitrofen-fed fetuses with CDH had malrotation at term, whereas only 18.2% of those without it did (P < .05). There was a significant (P < .001) negative correlation between the lung weight/body weight ratio and the degree of malrotation in nitrofen-treated fetuses. In conclusion, maternal nitrofen exposure on gestational day 9.5 induces intestinal malrotation in fetal rats by (1) delaying fetal growth and maturation; (2) causing CDH, which permits displacement of the liver and the gut into the thorax during the critical period of reintegration and fixation; and (3) inducing lung hypoplasia and reducing thoracic volume during this period.

Morbid musculoskeletal sequelae of thoracotomy for tracheoesophageal fistula.

The standard surgical approach for tracheoesophageal fistula (TEF) is right dorso-lateral thoracotomy. The late musculoskeletal consequences of the operation have been evaluated only rarely. Two hundred and seventy-seven patients with TEF were operated upon during the past 16 years, 117 of whom were available for long term (3 to 16 year) study. Twenty-nine of the patients had significant musculoskeletal deformities: (1) Twenty-one patients (23.8%) had prominent elevation of the right shoulder or "winged" scapula secondary to partial paralysis of the latissimus dorsi muscle; (2) Eighteen (20%) had marked asymmetry of the thoracic wall from atrophy of the serratus anterior muscle; (3) Nine (10%) had fusion of the ribs, in one of whom major respiratory dysfunction was a consequence; (4) Seven (7.8%) had severe thoracic scoliosis. The deformity was not of sufficient severity to warrant surgical correction but all patients required physiotherapy; (5) In two children (2.2%), fixation of the skin cicatrix to the bony thorax limited the mobility of the ipsilateral shoulder; (6) And finally, in three girls (3.3%), the thoracotomy scar disfigured the right breast leading to mammary maldevelopment in one adolescent. The latter child required plastic release of the entrapped breast. The dorso-lateral thoracic incision for tracheoesophageal atresia may lead to significant musculoskeletal complications and, since other alternatives are available, should be reevaluated as the recommended surgical approach.

An experimental study on the pathogenesis of gastroesophageal reflux after repair of diaphragmatic hernia.

BACKGROUND/PURPOSE: Gastroesophageal reflux (GER) is increasingly reported after surgical repair of congenital diaphragmatic hernia (CDH) and eventration. The aim of this study was to test the hypothesis that transdiaphragmatic pressure gradients are increased and that the antireflux barrier is weakened after plication of a previously paralyzed diaphragm. METHODS: Abdominal and esophageal pressures as well as lower esophageal sphincter pressures (LESP) and diaphragmatic pinchcock pressure (DPP) were measured before and after diaphragmatic plication in 16 rats in which the diaphragm had been previously eventrated by phrenic nerve section. RESULTS: This maneuver increased the transdiaphragmatic inspiratory pressure gradient from 2.75 +/- 0.54 to 4.51 +/- 0.86 mm Hg (P < .05) by rising both the inspiratory (-2.02 +/- 0.39 v -3.11 +/- 0.92 mm Hg, P < .05) and the expiratory (1.47 +/- 0.87 v 0.51 +/- 0.41 mm Hg, P < .05) intrathoracic pressures. At the same time, the antireflux barrier was weakened because LESP decreased from 17.5 +/- 5.59 to 10.59 +/- 5.74 mm Hg (P < .05) and DPP tended to decrease from 13.57 +/- 8.67 to 6.07 +/- 1.72 mm Hg (ns). CONCLUSIONS: Plication of the previously paralyzed diaphragm in the rat reinforces the GER driving forces while weakening the antireflux barrier. This may explain why reflux is frequent in children surviving repair of diaphragmatic hernia and eventration.

Skeletal malformations associated with congenital diaphragmatic hernia: experimental and human studies.

BACKGROUND/PURPOSE: Skeletal malformations are seen occasionally in infants with congenital diaphragmatic hernia (CDH). This study examines whether nitrofen, able to produce CDH in fetal rats, also induces skeletal anomalies and, if so, whether these are similar to those seen in CDH patients. METHODS: Pregnant rats received either nitrofen (100 mg, n = 7) or no treatment (n = 2) on gestational day 9.5. Skeletal anatomy was studied in fetuses recovered on day 21 after alcian blue-alizarin red staining. The charts and postmortem records of 117 stillborns or newborns who died of CDH were investigated retrospectively for skeletal defects. The proportions of anomalies found in the different groups were compared. RESULTS: The 15 control rat fetuses were normal, whereas 57 of 90 nitrofen-exposed animals (63%) had CDH accompanied by other malformations. Skeletal defects limited to vertebral segmentation or identity anomalies (split vertebra or absent, hypoplastic, or fused ribs) were seen at low thoracic and high lumbar levels in 68% of animals with CDH and in 57% of those without. Delayed ossification of limbs was seen in treated animals. There were skeletal malformations in 31.6% of the 117 human patients with CDH. Costovertebral defects (malformed, extra or defective vertebral bodies or ribs and spina bifida) were comparably frequent in infants with syndromes and in those without them (31.2% v 17.8%, not significant), whereas limb defects were significantly more frequent in those with syndromes (56.2% v 13.9%, P<.05). CONCLUSION: The nature and location of costovertebral malformations found in both CDH patients and nitrofen-exposed rats suggest that the diaphragmatic defect and the associated organ malformations might be caused by the same early embryonal disturbance involving axial and para-axial mesoderm.

Ambulatory 24-hour manometric and pH metric evidence of permanent impairment of clearance capacity in patients with esophageal atresia.

Dysphagia and gastroesophageal reflux (GER) probably caused by structural disorganization of the esophagus occur frequently after repair of tracheoesophageal fistula (TEF), and the extent to which they may improve beyond childhood is not known. The aim of the present study is to assess by combined ambulatory 24-hour manometry and pH-metry the esophageal peristaltic activity and acid clearing capacity in adolescents and adults who had been operated on for TEF at birth. Twenty-two patients, aged 17.1 +/- 4.5 years (mean +/- SD), were examined with combined three-channel manometry and two-channel pH-metry followed by endoscopy and biopsy. Although they considered themselves healthy, on careful interrogation, 16 (72%) were found to have dysphagia, 13 (59%) had heartburn, 10 (45%) had foreign body impaction, and 7 (31%) had chronic respiratory tract disease. GER was detected in 12 (54%) patients (5 with histological esophagitis), 10 of whom had a pattern of prolonged nocturnal episodes with very slow clearance. All patients had diminished contractile activity with low-amplitude and short-duration waves that decreased from 0.53 +/- 0.35 waves per minute to 0.28 +/- 0.2 waves per minute during sleep. Propulsive activity was uniformly disorganized, with peristaltic sequences being few (less than 50% overall) and incomplete (above 80%). Finally, the acid-clearing capacity was nil; the proportions of ineffective sequences were above 90% for all periods considered, including sleep and mealtimes. The motor behavior of nonrefluxing and refluxing patients was identical despite the differences in esophageal acid exposure.(ABSTRACT TRUNCATED AT 250 WORDS)

Cardiovascular malformations in congenital diaphragmatic hernia: human and experimental studies.

BACKGROUND/PURPOSE: Cardiovascular malformations (CVM) associated with congenital diaphragmatic hernia (CDH) account in part for the high mortality caused by this defect. The aim of this study is to examine the nature of these malformations in a large series of autopsies and to assess if similar defects are also present in rat fetuses with experimental CDH. METHODS: The incidence of CVM and their nature were examined in the autopsy records of 136 stillborns and neonates with CDH admitted to our institution in the last 30 years. Experimental CDH was induced in rat fetuses by giving 100 mg of nitrofen to their mothers on gestational day 9.5, and the fetuses were harvested on day 21 (near full term). The presence of CDH and the anatomy of the heart and great vessels were studied under dissecting microscope after formalin fixation. Unexposed fetuses were used as controls. RESULTS: Thirty-three newborns with CDH (24%) had CVM, either isolated or associated with other defects, and 7 had heart hypoplasia. Most CVM (ventricular septal defect, tetralogy of Fallot, transposition of the great vessels, double-outlet right ventricle) involved the outflow tract. In our animal experiments, no malformations were found in 21 control pups. Conversely, 80 of 130 nitrofen-exposed fetuses (61%) had CDH, and 59 of them (74%) had CVM. A significant association (Fisher's Exact test, P<.01) was found between CDH and CVM because only 25 of the 50 exposed animals without CDH (50%) had CVM. Again, most defects involved the outflow tract and were similar to those seen in human CDH (tetralogy of Fallot, persistent truncus, ventricular septal defect, double-outlet right ventricle, aberrant right subclavian artery, agenetic ductus, and interrupted aortic arch). Animals with CDH had significantly decreased heart weight to fetal weight ratio in comparison with controls and with those without CDH. CONCLUSIONS: The similar nature of the cardiovascular defects found in babies succumbing to CDH and in nitrofen-exposed rats suggests that a similar disturbance of the regional organogenesis related to the neural crest might be involved in both settings, and further validates the use of this animal model for clarifying the cellular and molecular pathogenetic mechanisms.

Skeletal malformations associated with esophageal atresia: clinical and experimental studies.

BACKGROUND/PURPOSE: Patients with esophageal atresia (EA) often have skeletal malformations. The purpose of this study is to examine if similar defects occur in rat fetuses prenatally exposed to Adriamycin, a chemical capable of causing EA in these animals. METHODS: The charts of 443 babies with EA were reviewed to assess the incidence and nature of these defects in them. Time-mated female rats were given either 2 mg/kg intraperitoneal Adriamycin (experimental group, n = 16) or no treatment (control group, n = 4) on gestational days 8 and 9, and the fetuses were removed near term. Skeletal anatomy was studied after alcian blue and alizarin red staining. RESULTS: A total of 528 skeletal malformations, mainly abnormal segmentation and vertebral identity (extra or defective bodies or ribs), mishaped vertebral bodies, and limb malformations like radial aplasia or hypoplasia were found in 245 babies (55%). Costal fusion and sternal anomalies were present in 17 and 4 babies, respectively. In the animal study, all control fetuses were normal, whereas 83 of 134 experimental fetuses (62%) had EA accompanied by other malformations. No segmentation or vertebral identity anomalies were seen, but butterfly, wedged, and asymmetric vertebral bodies were found at various levels in all animals with EA and in about half of those without it. Three fetuses had rib anomalies, and 3 more had sternal malformations. Ossification of limbs was delayed in treated fetuses and short, thick, and crooked bones were seen in 4 of 31 fetuses with EA and in none of the Adriamycin-exposed ones without EA. CONCLUSIONS: Adriamycin exposure induces in fetal rats, in addition to esophageal, duodenal, and anorectal atresias, high proportions of vertebral malformations and some limb defects of nature not identical but quite similar to that of babies with EA. This further validates this model for investigating the nature of the processes leading to EA and its associated malformations.

Effects of esophageal shortening on the gastroesophageal barrier: an experimental study on the causes of reflux in esophageal atresia.

BACKGROUND/PURPOSE: Gastroesophageal reflux (GER) is frequently recognized after surgical repair of esophageal atresia. The aim of this study was to test the hypothesis that one or more components of the gastroesophageal pressure barrier are weakened by esophageal anastomosis under tension. METHODS: Lower esophageal sphincter pressure (LESP), crural sling pressure (CSP), and the length of the intraabdominal segment of the esophagus (LIAE) were measured by pull-through perfusion manometry in 20 rats before and after resection of 15 mm of the cervical esophagus, and in eight rats before and after esophageal transection (control group). RESULTS: This manouver decreased the LESP from 44.9+/-17.4 to 30.9+/-12.3 mm Hg and the LIAE from 17.9+/-2.8 to 15.8+/-2.4 mm (P < .05) in experimental animals, whereas they did not significantly change in controls. CSP did not change significantly. CONCLUSIONS: Anastomosis of the esophagus under tension in this model decreases significantly the lower esophageal sphincter tone and length of the intraabdominal esophagus, but it does not change the crural sling pressure. Postoperative reflux in patients operated on for esophageal atresia might be in part, caused by this mechanism.

Embryology of esophageal atresia in the adriamycin rat model.

PURPOSE: The aim of this study was to describe the dysmorphogenetic process leading to esophageal atresia and tracheoesophageal fistula (EA + TEF) in the recently developed Adriamycin model of the malformation. METHODS: Time-mated pregnant rats were given either Adriamycin (1.75 mg/kg intraperitoneally) or saline on days 6 to 9 of gestation, and their embryos recovered on days 12, 12.5, and 13 were serially sectioned in the transversal plane and studied microscopically after H&E and PAS staining. The findings were compared with those of age-matched untreated embryos. RESULTS: All untreated and saline embryos were normal, whereas 49% of Adriamycin embryos had foregut malformations. Tracheoesophageal separation was complete on day 12 in control embryos, whereas 9 of 10 Adriamycin-exposed embryos had a common esophagotrachea with low emergence of the bronchi at that stage. This pattern had evolved into that of a regular EA + TEF in all nine malformed embryos by day 13. On day 12.5, esophagotrachea was found in 6 of 13 and EA + TEF in 5 of 13 embryos. Two had less well-defined malformations. CONCLUSIONS: Esophagotrachea equivalent to complete tracheoesophageal cleft is the first step leading to EA + TEF in this model. The full-blown malformation is finally acquired by partial loss of the posterior wall of the foregut, which tapers-off in the mediastinal mesenchyme and respiratory differentiation of the anterior wall down to the level of bronchial bifurcation, where it constitutes the fistula and the distal esophagus.

Experimental ureterohydronephrosis in fetal rats.

This study tests the hypothesis that prenatal exposure to 2,4-dichlorophenyl-p-nitrophenyl ether (nitrofen), an herbicide known to induce pulmonary hypoplasia and other malformations in fetal rats, also may induce ureterohydronephrosis (UHN) and oligohydramnios. Time-dated pregnant Wistar rats were given 100 mg of the chemical on day 9 or 11 of gestation, and the findings in their fetuses were compared with those of suitable controls. Marked bilateral UHN was found in the majority of exposed fetuses, but without evidence of either mechanical obstruction or dysplastic parenchymal lesions. These animals had various degrees of lung hypoplasia. The amount of fluid in their amniotic sacs was increased rather than decreased and it was independent of lung weight but correlated to some extent with UHN grade. Urinary tract dilatation and polyhydramnios in this model most likely are attributable to polyuria caused by nitrofen-induced impairment of renal concentrating capacity. This relatively simple animal model might facilitate research into some aspects of the physiology of nonobstructive, prenatally dilated urinary tracts.

Tracheobronchial malformations in experimental esophageal atresia.

BACKGROUND: Malformations of the tracheobronchial tree may account for postoperative respiratory symptoms in patients with esophageal atresia (EA). This study examines the respiratory tract in fetal rats with EA induced by Adriamycin. METHODS: Time-mated female rats were given either 2 mg/kg intraperitoneal Adriamycin on gestational days 8 and 9 (adria group, n = 6) or no treatment (control group, n = 2), and the fetuses were recovered on day 21. Laryngo-tracheo bronchial tree was studied after transparentation and alcian blue-alizarin red staining that depicts the cartilage in blue and make the surrounding tissues transparent. RESULTS: There were no malformations in any of the 1 1 control animals studied. Conversely, 31 of 46 (67%) Adriamycin fetuses had EA with distal TEF. These had more tracheal rings than controls (32+/-2 v 26+/-1.5, P < .05) at the expense of those of the mainstem bronchi (3.2+/-1 v 6.6+/-1.1 in the right, P< .05 and 6.2+/-2.1 v 11+/-1.1 in the left, P < .05). There were tracheal stenoses in 16 pups with EA (some severe and five double), and all these had fragmented rings in the trachea or bronchi. In six cases there was an ectopic upper right bronchus, and 1 had a grossly abnormal larynx. The malformations in the 15 Adriamycin-exposed fetuses without EA were limited to some fragmented or mishaped rings. CONCLUSIONS: Laryngo-tracheobronchial malformations entailing the whole length of the tract are very constant and severe in rats with EA and tracheoesophageal fistula and correspond to an abnormal development of the tracheobronchial anlage from the ventral foregut. Their nature and extent invite a careful investigation of the respiratory tracts in EA babies in whom they could be underscored.

Gastric heterotopy of the biliary tract.

The case of a 12-year-old girl having cholestatic syndrome, due to the presence of gastric tissue in the gallbladder and extrahepatic biliary tract, is reported here.

Heart hypoplasia in experimental congenital diaphragmatic hernia.

BACKGROUND/PURPOSE: Heart hypoplasia is associated with congenital diaphragmatic hernia (CDH) and decisively influences survival rate. This study examines whether nitrofen-exposed fetal rats have heart hypoplasia. METHODS: Pregnant rats received either 100 mg nitrofen or vehicle on gestational day 9.5. The hearts recovered near full term were either formalin fixed for anatomic studies or snap-frozen for biochemical studies. Heart weight, ventricular chamber diameters and aortic-to-pulmonary root diameter ratios were measured in fixed hearts. Protein and DNA were determined in frozen hearts. Analysis of variance (ANOVA) and correlation-regression studies were used for statistical assessment. RESULTS: All control fetuses were normal, whereas 61% of those exposed to nitrofen had CDH. Cardiovascular malformations were found in 73% of CDH and in 50% of non-CDH animals. Wet and fixed heart weights in percent of fetal weight, left-to-right ventricular diameter ratio, and aortic-to-pulmonary root diameter ratio were significantly decreased in fetuses with CDH in comparison with controls. Only wet heart was significantly decreased in nitrofen-treated fetuses without CDH, although all other variables showed a trend in the same direction. Protein to DNA ratios were similar in the three groups. The structure of the myocytes was histologically similar in all groups. CONCLUSIONS: The spectrum of lesions in the nitrofen model of CDH encompasses heart hypoplasia, further validating its use for research on this condition. Heart hypoplasia is related to cardiopulmonary compression, but its presence in treated animals without CDH demonstrates that the teratogen itself participate directly in its pathogenesis, and this finding invites further research on this line.

Thyroid hormones in the pathogenesis of lung hypoplasia and immaturity induced in fetal rats by prenatal exposure to nitrofen.

BACKGROUND/PURPOSE: Nitrofen is believed to act on prenatally exposed fetuses by changing maternal or fetal thyroid hormone physiology. The aim of this study was to determine whether the amounts of circulating and lung tissue T3 and T4 are decreased in rat fetuses with nitrofen-induced pulmonary hypoplasia and diaphragmatic hernia. METHODS: Timed-pregnant rats were given 100 mg of nitrofen in oil on gestational day 9.5, and their fetuses were recovered on the 21st day. Lung weight to body weight ratio was determined. Hormonal studies consisted in measurement of plasma T3, T4, and TSH, and of T3, T4, and DNA in lung tissue. Suitable groups of control fetuses prenatally exposed to oil were used for comparison. RESULTS: The lungs of nitrofen-treated fetuses were hypoplastic and those who had congenital diaphagmatic hernia were even more so. Nitrofen treatment led to decreased plasma T3 and T4 levels without TSH changes. T3 and T4 in lung tissue were apparently decreased in treated fetuses when expressed by weight, but these differences disappeared when expressed by DNA (cell content). CONCLUSIONS: Lung hypoplasia and immaturity induced by nitrofen treatment are not related to decreased levels of thyroid hormones in tissue near term. This should be kept in mind when proposing hormonal treatment for prenatal induction of lung maturation.

Prenatal dexamethasone rescues heart hypoplasia in fetal rats with congenital diaphragmatic hernia.

BACKGROUND/PURPOSE: Patients and rats with congenital diaphragmatic hernia (CDH) have lung and heart hypoplasia. Prenatal steroids improve lung hypoplasia in CDH rats. The current study tests the hypothesis that prenatal dexamethasone could rescue heart hypoplasia in rats with CDH. METHODS: Timed pregnant rats received intragastrically either 100 mg nitrofen or oil on day 9.5, and other animals had the same treatment with, in addition, either 0.25 mg/kg dexamethasone intraperitoneally or no treatment on days 19 and 20. Fetuses were recovered on day 21, and heart weight to body weight ratios, heart DNA, protein, and glycogen were measured in fresh specimens. Left-to-right ventricular diameter and aortic-to-pulmonary diameter ratios were measured after formalin fixation. RESULTS: Wet heart weight to body weight, left-to-right ventricular diameter, and aortic-to-pulmonary root diameter ratios, which were lower in fetuses exposed only to nitrofen than in their oil controls, were similar in those exposed to nitrofen plus dexamethasone than in their corresponding oil plus dexamethasone controls. Total heart DNA, which was decreased in fetuses exposed to nitrofen with CDH in comparison with their controls, was increased in those receiving nitrofen and dexamethasone in comparison with theirs. Protein to DNA ratio was decreased in all rats with CDH irrespective of their exposure or not to dexamethasone. Glycogen to DNA ratio was higher in all dexamethasone-treated fetuses than in those without this treatment. No gross histologic differences were seen among groups. CONCLUSIONS: Heart hypoplasia in rats with CDH is in part rescued by prenatal dexamethasone treatment as expressed by increased number of smaller myocytes with higher glycogen content. Prenatal steroids could modify heart involvement in human fetuses with CDH as well.