Antenatal maternally-administered phosphodiesterase type 5 inhibitors normalize eNOS expression in the fetal lamb model of congenital diaphragmatic hernia.

PURPOSE: Pulmonary hypertension (pHTN), a main determinant of survival in congenital diaphragmatic hernia (CDH), results from in utero vascular remodeling. Phosphodiesterase type 5 (PDE5) inhibitors have never been used antenatally to treat pHTN. The purpose of this study is to determine if antenatal PDE5 inhibitors can prevent pHTN in the fetal lamb model of CDH. METHODS: CDH was created in pregnant ewes. Postoperatively, pregnant ewes received oral placebo or tadalafil, a PDE5 inhibitor, until delivery. Near term gestation, lambs underwent resuscitations, and lung tissue was snap frozen for protein analysis. RESULTS: Mean cGMP levels were 0.53±0.11 in placebo-treated fetal lambs and 1.73±0.21 in tadalafil-treated fetal lambs (p=0.002). Normalized expression of eNOS was 82%±12% in Normal-Placebo, 61%±5% in CDH-Placebo, 116%±6% in Normal-Tadalafil, and 86%±8% in CDH-Tadalafil lambs. Normalized expression of ß-sGC was 105%±15% in Normal-Placebo, 82%±3% in CDH-Placebo, 158%±16% in Normal-Tadalafil, and 86%±8% in CDH-Tadalafil lambs. Endothelial NOS and ß-sGC were significantly decreased in CDH (p=0.0007 and 0.01 for eNOS and ß-sGC, respectively), and tadalafil significantly increased eNOS expression (p=0.0002). CONCLUSIONS: PDE5 inhibitors can cross the placental barrier. ß-sGC and eNOS are downregulated in fetal lambs with CDH. Antenatal PDE5 inhibitors normalize eNOS and may prevent in utero vascular remodeling in CDH.

Prenatal retinoic acid upregulates connexin 43 (Cx43) gene expression in pulmonary hypoplasia in the nitrofen-induced congenital diaphragmatic hernia rat model.

PURPOSE: Connexin 43 (Cx43), a major gap junction protein, is necessary for alveologenesis and plays an important role in the differentiation of type II to type I alveolar epithelial cells. Knockout mice of Cx43 display severe pulmonary hypoplasia (PH). Prenatal administration of retinoic acid (RA) is known to stimulate alveologenesis in nitrofen-induced PH. Recent studies revealed that retinoids upregulate Cx43 expression. We hypothesized that gene expression of Cx43 is downregulated during alveologenesis and that administration of RA upregulates Cx43 expression in the nitrofen-induced PH. METHODS: Pregnant rats were exposed to olive oil or nitrofen on day 9 (D9) of gestation. Retinoic acid was given intraperitoneally on D18, D19, and D20. Fetal lungs were harvested on D18 and D21 and divided into control, nitrofen, control+RA (D21), and nitrofen+RA (D21). The Cx43 expression levels were determined using reverse transcription polymerase chain reaction and immunohistochemistry. RESULTS: On D18 and D21, Cx43 relative messenger RNA expression levels were significantly downregulated in nitrofen compared with those in the control group. On D21, expression levels of Cx43 were significantly upregulated in nitrofen+RA and control+RA compared with those in nitrofen group. Immunohistochemical studies confirmed these results. CONCLUSION: Downregulation of Cx43 expression may interfere with normal alveologenesis. Upregulation of Cx43 pulmonary gene expression after RA treatment may promote lung growth by stimulating alveologenesis in nitrofen-induced PH.

Effects of estrogen on lung development in a rat model of diaphragmatic hernia.

PURPOSE: The study aimed to observe the influence of estradiol on rat models with congenital diaphragmatic hernia (CDH) and understand the potential mechanism. METHODS: Eleven pregnant female Sprague-Dawley rats were randomly divided into 3 groups on day 9.5 of gestation: group C (n = 2) was administered 2 mL of olive oil, whereas group N (n = 3) and group E (n = 6) were administered 200 mg of nitrofen. Antenatal estradiol was given subcutaneously to group E on days 18.5, 19.5, and 20.5 of gestation. Histologic evaluations, incidence of CDH, and the immunoreactivity of transforming growth factor (TGF)-ß1 in lung were observed. In addition, the mRNA levels of TGF-ß1, type I TGF-ß receptor (TßRI), and type II TGF-ß receptor (TßRII) were determined. RESULTS: Histologically, the lungs of group N fetuses were hypoplastic compared with those of group C and had thick-walled septa with poorly developed saccules. Group E showed improved mesenchymal differentiation with well-developed saccules. There was no significant difference between the incidence of CDH in group N and that in group E. The expression of TGF-ß1 in lung tissue and arterioles in group N were significantly higher than those in group C and E. Moreover, relative mRNA expression levels of TGF-ß1 and TßRI in group N were markedly higher than those in group C, whereas those in group E were significantly decreased compared with group N. CONCLUSIONS: Estradiol can promote lung development in rats with CDH. The down-regulation of TGF-ß1 and its signaling pathway may play a role in this effect.

Rapidly polymerizing hydrogel prevents balloon dislodgement in a model of fetal tracheal occlusion.

BACKGROUND/PURPOSE: This study examined whether an injectable hydrogel could buttress the balloon used in fetal tracheal occlusion, thus preventing its displacement. METHODS: Fetal lambs (n = 11) underwent tracheal occlusion through local delivery of a detachable silicone balloon and were divided in 2 groups: group I had no further manipulations, and group II received an intratracheal injection of a rapidly polymerizing hydrogel, cranially to the balloon. Near term, balloon placement was examined, the lung volume-to-body weight ratio (LV:BW) was determined, and tracheal histology was performed. Statistical analysis was by the Fisher's Exact test, with significance set at P <.05. RESULTS: Complete tracheal occlusion was achieved in all fetuses intraoperatively. The rate of balloon dislodgement was significantly higher in group I (4 of 7, or 57.1%) than in group II (0 of 4). In group II, balloons were recovered in situ with a column of residual hydrogel reinforcing their cephalad position. Animals in which balloon occlusion was maintained had significantly higher LV:BW, with no evidence of tracheal damage. CONCLUSIONS: Intratracheal delivery of a rapidly polymerizing hydrogel cephalad to detachable silicone balloons results in improved fetal tracheal occlusion, with no harmful effects to the trachea. This adjuvant principle may enhance minimally invasive balloon tracheal occlusion for treatment of severe fetal pulmonary hypoplasia.