Association of rare non-coding SNVs in the lung-specific FOXF1 enhancer with a mitigation of the lethal ACDMPV phenotype.

Haploinsufficiency of FOXF1 causes alveolar capillary dysplasia with misalignment of pulmonary veins (ACDMPV), a lethal neonatal lung developmental disorder. We describe two similar heterozygous CNV deletions involving the FOXF1 enhancer and re-analyze FOXF1 missense mutation, all associated with an unexpectedly mitigated disease phenotype. In one case, the deletion of the maternal allele of the FOXF1 enhancer caused pulmonary hypertension and histopathologically diagnosed MPV without the typical ACD features. In the second case, the deletion of the paternal enhancer resulted in ACDMPV rather than the expected neonatal lethality. In both cases, FOXF1 expression in lung tissue was higher than usually seen or expected in patients with similar deletions, suggesting an increased activity of the remaining allele of the enhancer. Sequencing of these alleles revealed two rare SNVs, rs150502618-A and rs79301423-T, mapping to the partially overlapping binding sites for TFAP2s and CTCF in the core region of the enhancer. Moreover, in a family with three histopathologically-diagnosed ACDMPV siblings whose missense FOXF1 mutation was inherited from the healthy non-mosaic carrier mother, we have identified a rare SNV rs28571077-A within 2-kb of the above-mentioned non-coding SNVs in the FOXF1 enhancer in the mother, that was absent in the affected newborns and 13 unrelated ACDMPV patients with CNV deletions of this genomic region. Based on the low population frequencies of these three variants, their absence in ACDMPV patients, the results of reporter assay, RNAi and EMSA experiments, and in silico predictions, we propose that the described SNVs might have acted on FOXF1 enhancer as hypermorphs.

Ibuprofen and indomethacin for the closure of the patent ductus arteriosus / Ibuprofen, indometacina e fechamento do canal arterial

The ductus arteriosus connects the pulmonary artery with the aorta and allows right ventricular blood to bypass the unexpanded lungs. In mature infants, the ductus arteriosus closes after birth. Patent ductus arteriosus occurs in 70% of preterm infants with a birth weight < 1,000 grams. Failure of the ductus arteriosus to close has been associated with intraventricular hemorrhage, necrotizing enterocolitis, bronchopulmonary dysplasia, periventricular leukomalacia, renal failure, and persistent pulmonary hypertension. The drugs used to treat the patent ductus arteriosus are ibuprofen and indomethacin which are potent non-selective inhibitors of cyclo-oxygenase (COX) and therefore inhibit prostaglandin E2 synthesis. Prostaglandin E2 relaxes smooth muscle and tends to inhibit the closure of the patent ductus arteriosus. Intravenous ibuprofen and indomethacin inhibit prostaglandin E2 synthesis and thereby close the patent ductus arteriosus with similar efficacy. Indomethacin reduces the blood flow velocity in kidneys, intestine and brain. Ibuprofen has less effect on blood flow velocity in these organs. There is a significant increase in serum creatinine after indomethacin administration but not after ibuprofen and infants treated with ibuprofen have higher creatinine clearance. Oliguria (urine output < 1 ml/kg/h) occurs more frequently with indomethacin than with ibuprofen. Indomethacin requires furosemide for urine output more often than ibuprofen. Ibuprofen reduces the risk of necrotizing enterocolitis and transient renal insufficiency and it is the drug of choice for closing the patent ductus arteriosus. Ibuprofen and indomethacin may be administered orally. In conclusion, intravenous ibuprofen and indomethacin close the patent ductus arteriosus at the same rate, but indomethacin is more toxic than ibuprofen.

O canal arterial conecta a artéria pulmonar com a aorta e permite que o sangue oriundo do ventrículo direito evite passar pelos pulmões fetais não expandidos. Em recém-nascidos maduros, o canal arterial se fecha após o nascimento. A persistência do canal arterial ocorre em 70% dos recém-nascidos prematuros com peso de nascimento < 1.000 gramas. O não fechamento do canal arterial associa-se a hemorragia intraventricular, enterocolite necrosante, displasia bronco-pulmonar, leucomalacia periventricular, insuficiência renal e hipertensão pulmonar persistente. Os medicamentos utilizados para tratar a persistência do canal arterial são o ibuprofeno e a indometacina. Ambos são potentes inibidores não seletivos da ciclo-oxigenase e inibem a síntese de prostaglandina E2. Esta relaxa a musculature vascular lisa e tende a inibir o fechamento do canal arterial. O ibuprofeno e a indometacina inibem a síntese de prostaglandina E2 e favorecem o fechamento do canal arterial. A indometacina reduz a velocidade do fluxo sanguíneo renal, intestinal e cerebral. O Ibuprofeno tem efeito menor sobre a velocidade do fluxo de sangue nesses órgãos. Há um aumento significativo da creatinina sérica após a administração de indometacina, mas não após o ibuprofeno; por isso, recém-nascidos tratados com ibuprofeno têm maior depuração da creatinina. A oligúria ocorre mais frequentemente com a indometacina vs. ibuprofeno. A indometacina requer furosemida para a produção de urina mais frequentemente do que o ibuprofeno. O ibuprofeno reduz o risco de enterocolite necrotizante e de insuficiência renal transitória e é a droga de escolha para o fechamento do canal arterial patente. O ibuprofeno e a indometacina podem ser ministrados por via oral. Em conclusão, o ibuprofeno e a indometacina fecham o canal arterial patente com a mesma velocidade, mas a indometacina é mais tóxica.

Ghrelin ameliorates hypoxia-induced pulmonary hypertension via phospho-GSK3 ß/ß-catenin signaling in neonatal rats.

Effective treatment and/or prevention strategies for neonatal persistent pulmonary hypertension of the newborn (PPHN) have been an important topic in neonatal medicine. However, mechanisms of impaired pulmonary vascular structure in hypoxia-induced PPHN are poorly understood and consequently limit the development of effective treatment. In this study, we aimed to explore the molecular signaling cascades in the lungs of a PPHN animal model and used primary cultured rat pulmonary microvascular endothelial cells to analyze the physiological benefits of ghrelin during the pathogenesis of PPHN. Randomly selected newborn rats were exposed to hypoxia (10-12%) or room air and received daily s.c. injections of ghrelin (150 µg/kg) or saline. After 2 weeks, pulmonary hemodynamics and morphometry were assessed in the rats. Compared with the control, hypoxia increased pulmonary arterial pressure, right ventricle (RV) hypertrophy, and arteriolar wall thickness. Ghrelin treatment reduced both the magnitude of PH and the RV/(left ventricle+septum (Sep)) weight ratio. Ghrelin protected neonatal rats from hypoxia-induced PH via the upregulation of phosphorylation of glycogen synthase kinase 3ß (p-GSK3ß)/ß-catenin signaling and associated with ß-catenin translocation to the nucleus in the presence of growth hormone secretagogue receptor-1a. Our findings suggest that s.c. administration of ghrelin improved PH and attenuated pulmonary vascular remodeling after PPHN. These beneficial effects may be mediated by the regulation of p-GSK3ß/ß-catenin expression. We propose ghrelin as a novel potential therapeutic agent for PPHN.

Pentoxifylline in preterm neonates: a systematic review.

Sepsis, necrotizing enterocolitis (NEC), and chronic lung disease (CLD) in preterm neonates are associated with significant mortality and morbidity, including long-term neurodevelopmental impairment and socioeconomic burden. Safe and effective drugs for the prevention and treatment of these conditions are urgently needed. Pentoxifylline, a synthetic theobromine derivative, is a non-steroidal immunomodulating agent with unique hemorrheologic effects which has been used in a range of infectious, vascular, and inflammatory conditions in adults and children. The unique properties of pentoxifylline explain its potential benefits in preterm neonates with sepsis, NEC, and CLD, conditions characterized by activation of the inflammatory cytokine cascade, free radical toxicity, and impaired microcirculation. Pentoxifylline has anti-inflammatory properties resulting from inhibition of erythrocyte phosphodiesterase. It lowers blood viscosity and improves microcirculation and tissue perfusion. As a phosphodiesterase inhibitor, pentoxifylline downregulates pro-inflammatory cytokines such as tumor necrosis factor-alpha, interleukin-6, and interferon-gamma. Methylxanthines, including caffeine, theophylline, and theobromine are relatively non-toxic drugs; of these, theobromine is the least toxic. Pentoxifylline-related significant adverse events are thus very rare. Unlike other methylxanthines, pentoxifylline does not have significant cardiac and bronchodilating effects at therapeutic doses. Although it is contraindicated in adults with recent cerebral hemorrhage due to its effect on platelets, red blood cells, and plasma fibrinogen levels, no significant adverse effects including thrombocytopenia and bleeding have been reported in critically ill preterm neonates with sepsis or NEC after treatment with pentoxifylline. Based on data from pilot randomized trials and observational studies, our systematic review suggests that pentoxifylline may reduce mortality and/or morbidity in preterm neonates with sepsis, NEC, and CLD. Results of experimental studies also indicate that pentoxifylline may potentially be beneficial in meconium aspiration syndrome and hypoxic ischemic encephalopathy. Given the substantial burden of sepsis, NEC, and CLD in high-risk preterm neonates, and the findings of this systematic review, pentoxifylline needs to be evaluated urgently as a preventative and therapeutic agent for these conditions in randomized controlled trials that can detect minimal clinically significant effect sizes. Further clinical and experimental studies are also necessary to evaluate whether pentoxifylline is safe and effective in meconium aspiration syndrome and hypoxic ischemic encephalopathy.

Executive summary of the workshop on oxygen in neonatal therapies: controversies and opportunities for research.

One of the most complex areas in perinatal/neonatal medicine is the use of oxygen in neonatal therapies. To address the knowledge gaps that preclude optimal, evidence-based care in this critical field of perinatal medicine, the National Institute of Child Health and Human Development organized a workshop, Oxygen in Neonatal Therapies: Controversies and Opportunities for Research, in August 2005. The information presented at the workshop included basic and translational oxygen research; a review of completed, ongoing, and planned clinical trials; oxygen administration for neonatal resuscitation; and a review of the collaborative home infant monitoring evaluation study. This article provides a summary of the discussions, focusing on major knowledge gaps, with prioritized suggestions for studies in this area.

Update on neonatal resuscitation.

Pediatricians and other practitioners who attend deliveries maintain resuscitation skills throughout their careers. NRP certification and continuing education programs (PCEP) have done much to standardize the care a newborn receives in the first minutes after birth. Neonates have benefited tremendously from strong national programs (NRP and PCEP) with great local execution of training modules.

Involvement of thromboxane A2 and prostacyclin in the early pulmonary hypertension after porcine meconium aspiration.

Severe perinatal aspiration of meconium is frequently complicated by unsuccessful neonatal adaptation with associated pulmonary hypertension. This vascular complication is supposedly related to pulmonary release of vasoconstrictory agents, including metabolites of arachidonic acid. Thus, to investigate the role of prostanoids on these meconium-induced circulatory changes in the lungs, the hemodynamic response to meconium instillation was studied in acetylsalicylic acid-pretreated juvenile pigs. Twelve 10-wk-old pigs with adapted lung circulation received 3 mL/kg of 65 mg/mL human meconium via the endotracheal tube. Six of them were medicated with 10 mg/kg acetylsalicylic acid 30 min before meconium insufflation. Hemodynamic parameters and urinary excretion of stable metabolites of thromboxane A2 and prostacyclin were measured serially for 6 h after the insult. Meconium administration induced a biphasic increase in mean pulmonary artery pressure and pulmonary vascular resistance, and a rapid rise in urinary levels of prostanoid metabolites. Acetylsalicylic acid pretreatment prevented the initial (0-1 h) pulmonary hypertensive response and increase in prostanoid excretion. During the second phase (1-6 h), acetylsalicylic acid did not attenuate the progressive increase in mean pulmonary artery pressure and pulmonary vascular resistance nor did it affect the longitudinal distribution of the pulmonary resistances. Our results thus show that in adapted porcine lungs, arachidonic acid metabolites contribute to the early hypertensive response, but have only minor effects during the second phase vascular hypertension.

Antenatal glucocorticoid therapy suppresses angiotensin-converting enzyme activity in rats with nitrofen-induced congenital diaphragmatic hernia.

BACKGROUND/PURPOSE: Neonates with congenital diaphragmatic hernia (CDH) have a high morbidity and mortality rate caused by pulmonary hypoplasia associated with pulmonary hypertension (PH). In experimental CDH, antenatal glucocorticoid therapy improves surfactant biochemical immaturity, enhances lung compliance, and induces morphological maturation in CDH rats. The effects of steroid therapy on preventing or treating PH in this condition have not been studied. Angiotensin converting enzyme (ACE), which is produced by the vascular endothelium, is implicated in the pathogenesis of pulmonary hypertension. The aim of this study was to evaluate the effect of antenatal glucocorticoid therapy on ACE activity and expression in CDH rat lungs. METHODS: CDH was induced in fetal rats by the maternal administration of 100 mg nitrofen on day 9.5 of gestation (term, day 22). Dexamethasone (Dex) (0.25 mg/kg) was given by intraperitoneal injection on days 18.5 and 19.5 before delivery of the fetuses by cesarean section on day 21.5. Control animals received olive oil (OO) by gavage and normal saline (NS) as vehicle injection. ACE activity was measured spectrophotometrically in the lungs of rats from four treatment groups: CDH-NS, non-CDH-NS, CDH-Dex, and OO-NS controls. Total lung ACE activity (mU per lung) was significantly lower in CDH-NS (P = .002) and CDH-Dex (P = .004) rats compared with non-CDH-NS and OO-NS controls (9.1 +/- 1.0 and 10.7 +/- 1.3 v 16.2 +/- 1.6 and 15.4 +/- 1.7). When specific ACE activity (mU/mg protein) was derived by expressing ACE activity per milligram of lung protein, CDH-NS animals showed elevated specific ACE activity (P = .05) compared with OO-NS controls (6.31 +/- 1.1 v 4.4 +/- 0.4). CDH-Dex animals had a significantly lower specific ACE activity (P = .01) compared with CDH-NS and Non-CDH-NS rats (4.0 +/- 0.4 v 6.31 +/- 1.1 and 5.83 +/- 0.54). The specific ACE activity levels of CDH-Dex rats were equivalent to those seen in the lungs of OO-NS controls (P = .24). CONCLUSION: Antenatal steroid therapy, by suppressing pulmonary ACE activity, may reduce the risk of pulmonary hypertension developing in human newborns with antenatally diagnosed CDH.

Effect of prenatal glucocorticoids on pulmonary vascular muscularisation in nitrofen-induced congenital diaphragmatic hernia.

BACKGROUND/PURPOSE: Pulmonary hypertension (PH) contributes significantly to the mortality of congenital diaphragmatic hernia (CDH). Pulmonary vascular changes in CDH include a reduced vascular bed with increased arterial medial wall thickness and peripheral extension of muscle into intraacinar vessels. Antenatal steroids improve biochemical immaturity, lung compliance, and morphology in experimental CDH animals. The aim of this study was to examine the effects of prenatal glucocorticoid therapy on pulmonary artery muscularisation in CDH rats. METHODS: CDH was induced in fetal rats by the maternal administration of 100 mg of nitrofen by gavage on day 9.5 of gestation (term, day 22). Control animals received olive oil (OO). Dexamethasone (Dex, 0.25 mg/kg) or normal saline (NS) was given by intraperitoneal injection on days 18.5 and 19.5, and fetuses were delivered by caesarean section on day 21.5. Lung sections from five fetuses in each of four experimental groups were studied by a blinded investigator- OO-NS controls, CDH-NS, CDH-Dex, and non-CDH-NS. The external diameter (ED), medial wall thickness (MT), percent of medial wall thickness, and wall structure were evaluated from preacinar arteries accompanying conducting airways, and the intraacinar arterioles associated with the respiratory bronchi and saccules. RESULTS: In the preacinar arteries, CDH-NS animals had a significantly increased MT percentage compared with OO-NS controls (21.2+/-8.8 v 17.8+/-10.3, P = .0001). CDH-Dex rats had a lower MT percentage than CDH-NS rats (15.5+/-6.7 v 21.2+/-8.8, P = .0001). In the intraacinar region, CDH-Dex fetuses had a reduced percentage of muscularised intraacinar blood vessels compared with CDH-NS and OO-NS controls (10% v 24% and 28%, respectively, P = .01). Dexamethasone-treated CDH pups also displayed a significantly lower MT percentage of the intraacinar arteries compared with CDH-NS and OO-NS animals (15.7+/-13 v 23.4+/-9 and 25.4+/-12, P = .003). CONCLUSIONS: Medial hypertrophy is present in the preacinar but not the intraacinar blood vessels of CDH rats before birth. Dexamethasone inhibits medial hypertrophy and reduces the number of muscularised intraacinar vessels. Antenatal glucocorticoids may reduce the risk of PH developing in human newborns with antenatally diagnosed CDH.

Reversible tracheal obstruction in the fetal sheep: effects on tracheal fluid pressure and lung growth.

Congenital diaphragmatic hernia (CDH) continues to carry high morbidity and mortality. A number of treatment modalities including extracorporeal membrane oxygenation and in utero repair have improved the mortality rate only minimally. With this condition, there is often insufficient lung mass at birth and persistent pulmonary hypertension postnatally. Experimental fetal tracheal ligation has been shown to increase lung growth in utero and to reduce the hernial contents in CDH. The purpose of this study was to determine the effect of reversible fetal tracheal occlusion on tracheal pressure and lung development. Nine fetal sheep were divided into two groups. Group 1 had intratracheal balloons placed, and the balloons were left inflated for 21 to 28 days. Group 2 consisted of littermates that served as controls. They either had uninflated balloons placed or were left unoperated. Tracheal pressure measurements were recorded periodically, and the amniotic fluid pressure served as a reference. The animals were killed near term, and the lungs, heart, and liver were weighed and corrected for body weight. Standard morphometry was used to compare the lungs further, and the lung DNA and protein content were measured. Tracheal damage from the balloon catheter also was assessed. The tracheal pressure was 3.85 (+/- .49 SE) mm Hg in experimental animals, and it averaged -0.27 (+/- .27 SE) mm Hg in controls (P < .0001). Tracheal occlusion increased lung weight and volume by two to three times (P < .0001 and P < .0006, respectively) while heart and liver weights remained similar to those of controls.(ABSTRACT TRUNCATED AT 250 WORDS)

A new protocol for the perinatal management of patients with congenital diaphragmatic hernia with severe hypoplastic lungs and its clinical application.

It is prerequisite for the pre-operative management of patients with congenital diaphragmatic hernia (CDH) to avoid the factors that increase pulmonary vascular resistance, because such patients easily fall into the state of persistent fetal circulation (PFC). In this paper, a new protocol is proposed for the perinatal management of CDH patients to prevent the PFC which is facilitated by the enlargement of hernia and the deviation of mediastinum caused by spontaneous breathing just after birth. We also describe the usefulness of this protocol based on our experience in which we successfully treated a patient with CDH with severe hypoplastic lungs that were diagnosed antenatally by ultrasonographic examination. In treating the CDH patient with severe hypoplastic lungs, it is effective for the respiratory control and the prevention of PFC to administer morphine and pancuronium to the neonate through the umbilical vein before the resection of the umbilical cord. In the case of Cesarean section, in addition to the direct administration of morphine to the patient, administration of morphine to the patient's mother just before the delivery is more effective to prevent PFC, which can be easily induced by the initial resuscitation at birth.

Association of elective repeat cesarean delivery and persistent pulmonary hypertension of the newborn.

Seventy-one cases of persistent pulmonary hypertension of the newborn have been reviewed in an attempt to identify possibly preventable causes. Three groups of infants were identified. The first group consisted of 36 infants with evidence of perinatal asphyxia. The second group was made up of 23 infants who exhibited a variety of associated factors including pneumonia, septic shock, and congenital diaphragmatic hernia. A third group included 12 infants delivered by elective repeat cesarean section. Infants in the third group did not have evidence of perinatal asphyxia, meconium aspiration, or infection. Chest roentgenograms revealed amniotic fluid aspiration in seven cases, retained lung fluid in three cases, and normal findings in two cases. All 12 infants in the third group developed respiratory distress which eventually progressed to respiratory failure and persistent pulmonary hypertension of the newborn. These data suggest that infants of elective repeat cesarean deliveries are at risk for developing persistent pulmonary hypertension of the newborn and constitute a group of patients with a potentially preventable course of events.

Reversion to fetal circulation in congenital diaphragmatic hernia: a preventable postoperative complication.

A review of 26 patients with congenital diaphragmatic hernia, diagnosed in the first 24 hours of life, supports the hypothesis that the postoperative fetal circulation syndrome is an iatrogenic complication, due to the rapid expansion of both lungs, when they are severely hypoplastic. This complication is preventable, when no aspiration of air from the chest cavity is done, and when no tube attached to an underwater seal is inserted. When assisted ventilation is necessary, small volumes at a rapid rate allows satisfactory gaseous exchanges, without pulmonary overinflation.

Persistent pulmonary hypertension of the neonate (persistent fetal circulation syndrome).

This 15-year-old disease has been clearly described anatomically. Some understanding of possible in utero predisposing conditions has emerged from clinical and animal studies. However, we have very little understanding of the cellular processes that trigger and/or prolong the abnormal medial smooth muscle hypertrophy underlying the condition. Empiric observation has resulted in the development of hyperventilation as a fairly successful treatment modality, although the underlying mechanism of this salubrious effect is unknown. Drugs, a major focus of clinical and laboratory investigations, sometimes are marginally successful (and sometimes are utter failures). Translated into the neonatal intensive care unit, the disease is more frequently accurately diagnosed than in the past, but it remains frustratingly difficult to manage, and thus far is impossible to prevent. The research foundations laid in the past decade provide impetus for accelerated search into the fundamental cellular and biochemical derangements that cause persistent pulmonary hypertension. It is to be hoped that the next decade will yield major advances in both mechanistic understanding and in treatment.