tretinoin and Hernias--Diaphragmatic--Congenital

Structural birth defects are a common cause of abnormalities in newborns. While there are cases of structural birth defects arising due to monogenic defects or environmental exposures, many birth defects are likely caused by a complex interaction between genes and the environment. A structural birth defect with complex etiology is congenital diaphragmatic hernias (CDH), a common and often lethal disruption in diaphragm development. Mutations in more than 150 genes have been implicated in CDH pathogenesis. Although there is generally less evidence for a role for environmental factors in the etiology of CDH, deficiencies in maternal vitamin A and its derivative embryonic retinoic acid are strongly associated with CDH. However, the incomplete penetrance of CDH-implicated genes and environmental factors such as vitamin A deficiency suggest that interactions between genes and environment may be necessary to cause CDH. In this review, we examine the genetic and environmental factors implicated in diaphragm and CDH development. In addition, we evaluate the potential for gene-environment interactions in CDH etiology, focusing on the potential interactions between the CDH-implicated gene, Gata4, and maternal vitamin A deficiency.

Topics: Diaphragm; Hernias, Diaphragmatic, Congenital; Humans; Infant, Newborn; Mutation; Tretinoin; Vitamin A Deficiency

Congenital diaphragmatic hernia (CDH) is a neonatal pathology in which intrathoracic herniation of abdominal viscera via diaphragmatic defect results in aberrant pulmonary and cardiovascular development. Despite decades of study and many advances in the diagnosis and treatment of CDH, morbidity and mortality remain high, largely due to pulmonary hypertension (PH), along with pulmonary hypoplasia and cardiac dysfunction. In patients with CDH, hypoplastic pulmonary vasculature and alterations in multiple molecular pathways lead to pathophysiologic pulmonary vasculopathy and, for severe CDH, sustained, elevated pulmonary arterial pressures. This review addresses the multiple anatomic and physiologic changes that underlie CDH-associated PH (CDH-PH), along with the multimodal treatment strategies that exist currently and future therapies currently under investigation.

Topics: Endothelins; Hernias, Diaphragmatic, Congenital; Humans; Hypertension, Pulmonary; Infant, Newborn; Lung; MicroRNAs; Nitric Oxide; Pulmonary Artery; Pulmonary Veins; Signal Transduction; Tretinoin; Vascular Endothelial Growth Factor A; Vascular Remodeling; Ventricular Dysfunction

Congenital diaphragmatic hernia (CDH) is the result of incomplete formation of the diaphragm that occurs during embryogenesis. The defect in the diaphragm permits the herniation of abdominal organs into the thoracic cavity contributing to the impairment of normal growth and development of the fetal lung. In addition to the hypoplastic lung, anomalies of the pulmonary arterioles worsen the pulmonary hypertension that can have detrimental effects in severe cases. Most cases of CDH can be effectively managed postnatally. Advances in neonatal and surgical care have resulted in improved outcomes over the years. When available, extracorporeal membrane oxygenation can provide temporary cardiorespiratory support for those not effectively supported by mechanical ventilation. In spite of these advances, very severe cases of CDH still carry a very high mortality and morbidity rate. Advances in imaging and evaluation now allow for early and accurate prenatal diagnosis of CDH, thereby identifying those at greatest risk who may benefit from prenatal intervention. This review article discusses some of the surgical and non-surgical prenatal interventions in the management of isolated severe congenital diaphragmatic hernia.

Topics: Female; Fetoscopy; Glucocorticoids; Hernias, Diaphragmatic, Congenital; Humans; Hypertension, Pulmonary; Pregnancy; Prenatal Diagnosis; Prognosis; Sildenafil Citrate; Trachea; Tretinoin; Ultrasonography, Prenatal; Vasodilator Agents

Congenital diaphragmatic hernia (CDH) is a congenital anomaly associated with an increased mortality and morbidity. In this article, we review the currently known etiological and pathogenic factors in CDH.

Topics: Diaphragm; Gene Expression Regulation, Developmental; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Humans; Lung; Signal Transduction; Tretinoin

Congenital diaphragmatic hernia (CDH) is a common cause of severe neonatal respiratory distress. Mortality and morbidity are determined by the amount of pulmonary hypoplasia (PH) that occurs and by the development of therapy-resistant pulmonary hypertension. The pathogenesis and aetiology of CDH and its associated anomalies are still largely unknown despite all research efforts. The pathogenesis of CDH is based on an assumption linking herniation of abdominal viscera into the thorax with compression of the developing lung. PH, however, can also result from reduced distension of the developing lung secondary to impaired fetal breathing movements. Our understanding of CDH has also been aided by basic research with the use of dietary, teratogen-induced, and knockout models of CDH. These studies indicate that lung hypoplasia may involve disturbances of mitogenic signalling pathways fundamental to embryonic lung development. Recent data reveal the role of disruption of a retinoid-signalling pathway in the pathogenesis of CDH. Although multifactorial inheritance may best explain most cases of CDH in humans, much has been learned about the genetic factors that play a role in the development of CDH by studies of patients with CDH caused by specific genetic syndromes and chromosome anomalies. More research is warranted to improve our understanding of normal and abnormal lung development in relation to CDH. Such investigations will help in the design of new treatment strategies to improve the natural course or even to prevent this anomaly.

Topics: Abnormalities, Multiple; Animals; Disease Models, Animal; Female; Genetic Association Studies; Gestational Age; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Humans; Infant, Newborn; Lung; Mice; Mice, Knockout; Persistent Fetal Circulation Syndrome; Pregnancy; Prenatal Exposure Delayed Effects; Respiratory Distress Syndrome, Newborn; Risk Factors; Tretinoin; Vitamin A

Congenital diaphragmatic hernia (CDH) is a major life-threatening cause of respiratory failure in the newborn. Although significant efforts have been undertaken to unravel the pathophysiology of CDH, our current understanding of the etiology remains spare. Here we outline recent evidence suggesting that abnormalities linked with the retinoid signaling pathway early in gestation may contribute to the etiology of CDH. These studies include 1) the effect of altering the retinoid system in vitamin A deficient and transgenic animals; 2) disruption of the retinoid system in teratogen-induced CDH in rodents, 3) the effect of co-administration of retinoids in nitrofen-induced CDH on lung and diaphragm development, and 4) clinical evidence suggesting decreased markers of vitamin A status in human CDH. Given the substantial mortality and morbidity associated with this serious developmental anomaly, advancements in this area will be critical. We feel that there is now sufficient circumstantial and direct experimental evidence to warrant further testing of the retinoid-CDH etiology hypothesis, including examination of retinoid-regulated target genes that could be candidates for involvement in CDH.

Topics: Animals; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Humans; Infant, Newborn; Signal Transduction; Tretinoin

The mortality and morbidity of patients with congenital diaphragmatic hernia (CDH) is primarily caused by treatment-resistant, persistent pulmonary hypertension. Structural vascular changes, exemplified by extensive muscularization, are already present early in gestation, but the origin of these abnormalities is unknown. Understanding the origin of the vascular defects is important to improve treatment modalities. Here, we show that the distribution of pericytes is different and may thereby potentially initiate the vascular pathology in CDH. Transient inhibition of retinoic acid (RA) signaling early during pregnancy, the basis of the CDH mouse model, led to an increase in the number of pericytes, thereby affecting the angiogenic potential of pericytes in the fetuses. Pericytes of CDH lungs showed reduced proliferation and an increased ACTA2 expression, which indicates that these pericytes are more contractile than in control lung pericytes. This resulted in increased pericyte coverage of pulmonary vessels and reduced expansion of the capillary bed, the earliest pathological sign of the structural changes in CDH. Furthermore, the pericytes had reduced and altered collagen IV deposition in CDH, pointing to a loss of basal membrane integrity between pericytes and endothelial cells. Inhibition of RA signaling in vitro resulted in reduced migration of pericytes, reduced angiogenesis, and loss of collagen IV expression. Importantly, we confirmed our findings in lungs of human CDH patient samples. In summary, inhibition of RA signaling affects the lung pericyte population, leading to increased contractility, reduced pulmonary angiogenesis, and aberrant lung development, as observed in CDH.

Topics: Animals; Cell Differentiation; Disease Models, Animal; Endothelial Cells; Hernias, Diaphragmatic, Congenital; Humans; Hypertension, Pulmonary; Lung; Mice; Pericytes; Signal Transduction; Tretinoin

Abnormal retinoic acid (RA) signalling is considered a major cause of congenital diaphragmatic hernia (CDH). Pulmonary hypoplasia and pulmonary hypertension are the major causes of morbidity and mortality in infants born with CDH. Experimental studies in animals have found that RA signalling is involved in lung and liver development, but animal models of CDH do not directly correlate with CDH in human fetuses. This study investigated if RA status is also linked to lung and liver growth in human fetuses with CDH.. Hepatic stellate cells (HSC) in autopsy human fetal liver tissue were identified using cRBP-1 immunohistochemistry and the numbers of HSC manually counted. In mammals, RA is principally stored in HSC complexed to cRBP-1 and therefore cRBP-1. Fetal RA stores, reflected in the number of cRBP-1

Topics: Autopsy; Case-Control Studies; Female; Gestational Age; Glial Fibrillary Acidic Protein; Hepatic Stellate Cells; Hernias, Diaphragmatic, Congenital; Humans; Liver; Lung; Male; Organ Size; Pregnancy; Retinol-Binding Proteins, Cellular; Tretinoin

Topics: Fetus; Hernias, Diaphragmatic, Congenital; Humans; Liver; Lung; Tretinoin; Ultrasonography, Prenatal

Lung and pulmonary vascular maldevelopment in congenital diaphragmatic hernia (CDH) results in significant morbidity and mortality. Retinoic acid (RA) and imatinib have been shown to improve pulmonary morphology following prenatal administration in the rat nitrofen-induced CDH model. It remains unclear if these changes translate into improved function. We evaluated the effect of prenatal RA and imatinib on postnatal lung function, structure, and pulmonary artery (PA) blood flow in the rat CDH model.. Olive oil or nitrofen was administered alone or in combination with RA or imatinib to pregnant rats. Pups were assessed for PA blood flow by ultrasound and pulmonary function/morphology following delivery, intubation, and short-term ventilation.. Neither RA nor imatinib had a negative effect on lung and body growth. RA accelerated lung maturation indicated by increased alveoli number and thinner interalveolar septa and was associated with decreased PA resistance and improved oxygenation. With the exception of a decreased PA pulsatility index, no significant changes in morphology and pulmonary function were noted following imatinib.. Prenatal treatment with RA but not imatinib was associated with improved pulmonary morphology and function, and decreased pulmonary vascular resistance. This study highlights the potential of prenatal pharmacologic therapies, such as RA, for management of CDH.

Topics: Animals; Drug Administration Schedule; Female; Hernias, Diaphragmatic, Congenital; Imatinib Mesylate; Lung; Phenyl Ethers; Pregnancy; Prenatal Care; Protein Kinase Inhibitors; Pulmonary Alveoli; Pulmonary Artery; Random Allocation; Rats; Rats, Sprague-Dawley; Treatment Outcome; Tretinoin

Erythropoietin (EPO), an essential stimulator of erythropoiesis produced by the fetal liver, is important both in vascular remodeling and modulation of the endothelial response in the pulmonary vasculature. In addition, EPO guides alveolar development, along with retinoic acid (RA). EPO is a direct target of RA, and the retinoid pathway is altered in the nitrofen-induced congenital diaphragmatic hernia (CDH) model. In the present study, we tested the hypothesis that the synthesis of EPO is suppressed in a rat model of CDH.. Pregnant rats were treated with either nitrofen or vehicle on gestational day 9 (D9). Fetuses were sacrificed on D19 and D21 and divided into control and CDH groups. Immunohistochemistry and quantitative real-time polymerase chain reaction (RT-PCR) were performed to determine the expression of EPO in the fetal liver and kidney. We also estimated the expression of EPO receptor in the fetal lung.. The relative EPO mRNA expression in the liver on D19 and in the kidney on D21 were significantly lower in the CDH group than in the controls (P = 0.0008 and P = 0.0064, respectively). In addition, the results of immunohistochemistry supported the findings from the RT-PCR analysis. No significant changes were noted in the expression pattern or EPO receptor levels in the fetal lungs of the CDH group compared to the controls.. Our results reveal the suppressed EPO synthesis in the CDH fetus, which may contribute to the pathogenesis of lung hypoplasia and modification of pulmonary vasculature in the CDH rat model. Pediatr Pulmonol. 2017;52:606-615. © 2016 Wiley Periodicals, Inc.

Topics: Animals; Disease Models, Animal; Erythropoietin; Female; Gene Expression Regulation, Developmental; Hernias, Diaphragmatic, Congenital; Lung; Phenyl Ethers; Pregnancy; Rats; Rats, Sprague-Dawley; Tretinoin

Low pulmonary retinol levels and disrupted retinoid signaling pathway (RSP) have been implicated in the pathogenesis of congenital diaphragmatic hernia (CDH) and associated pulmonary hypoplasia (PH). It has been demonstrated that nitrofen disturbs the main retinol-binding protein (RBP)-dependent trophoblastic retinol transport. Several studies have demonstrated that prenatal treatment with retinoic acid (RA) can reverse PH in the nitrofen-induced CDH model. We hypothesized that maternal administration of RA can increase trophoblastic RBP-dependent retinol transport in a nitrofen model of CDH.. Pregnant rats were treated with nitrofen or vehicle on gestational day 9 (D9) and sacrificed on D21. RA was given i.p. on D18, D19, and D20. Retinol and RA levels were measured using high-performance liquid chromatography. Immunohistochemistry was performed to evaluate trophoblastic expression of RBP. Expression levels of the primary RSP genes were determined using quantitative real-time PCR and immunohistochemistry.. Markedly increased trophoblastic RBP immunoreactivity was observed in CDH+RA compared to CDH. Significantly increased serum and pulmonary retinol and RA levels were detected in CDH+RA compared to CDH. Pulmonary expression of RSP genes and proteins were increased in CDH+RA compared to CDH.. Increased trophoblastic RBP expression and retinol transport after antenatal administration of RA suggest that retinol-triggered RSP activation may attenuate CDH-associated PH by elevating serum and pulmonary retinol levels.

Topics: Animals; Biological Transport; Disease Models, Animal; Female; Fetal Weight; Hernias, Diaphragmatic, Congenital; Lung; Phenyl Ethers; Pregnancy; Rats; Retinol-Binding Proteins; Tretinoin; Trophoblasts; Vitamin A

To investigate the effect of combined prenatal treatment with retinoic acid (RA) and tracheal occlusion (TO) on the pulmonary vascular morphology and expression of vascular endothelial growth factors (VEGF) and its receptors in a rat model of congenital diaphragmatic hernia (CDH).. Rats were given nitrofen at 9 days of gestation followed by no treatment (CDH), RA (CDH + RA), TO (CDH + TO), or both (CDH + RA + TO) (n = 16). We measured the median wall thickness of the pulmonary arterioles (MWT) and analyzed the expression of VEGF and its receptors (VEGFR1 and VEGFR2).. Compared to control animals, CDH had increased MWT (44 ± 15 vs. 58 ± 7; p < 0.05) and decreased expression of VEGF, VEGFR1, and VEGFR2 (p < 0.05). Treatment with RA or TO alone, and RA + TO reduced the MWT (46 ± 9, 42 ± 11, 46 ± 8, respectively) and improved the expression of VEGF, VEGFR1, and VEGFR2 compared to CDH (p < 0.05). However, the combination of RA + TO did not confer additional benefit in the reduction of the MWT or in increasing the VEGF and its receptors compared to either treatment alone.. Antenatal treatment with either RA or TO improved the MWT and expression of VEGF and its receptors in a CDH rat model. However, combined treatment with RA + TO was not superior to either treatment alone.

Topics: Animals; Antineoplastic Agents; Balloon Occlusion; Disease Models, Animal; Female; Hernias, Diaphragmatic, Congenital; Pregnancy; Pregnancy, Animal; Rats; Rats, Sprague-Dawley; Trachea; Tretinoin

The high mortality rate in congenital diaphragmatic hernia (CDH) is attributed to pulmonary hypoplasia (PH). Insulin-like growth factor 2 (IGF2) is an important regulator of fetal growth. The highest levels of IGF2 expression are found in the placenta, which are negatively regulated by decidual retinoid acid receptor alpha (RARα). It has been demonstrated that prenatal administration of retinoic acid (RA) suppresses decidual RARα expression. Previous studies have further shown that prenatal administration of RA can reverse PH in nitrofen-induced CDH model. In IGF2 knockout animals, low levels of IGF2 are associated with decreased placental growth and PH. We therefore hypothesized that nitrofen decreases trophoblastic IGF2 expression and prenatal administration of RA increases it through decidual RARα in the nitrofen-induced CDH model.. Pregnant rats were exposed to either olive oil or nitrofen on day 9 of gestation (D9). RA was given intraperitoneally on D18, D19 and D20. Fetuses were harvested on D21 and divided into three groups: control, CDH and nitrofen+RA. Immunohistochemistry was performed to evaluate decidual RARα and trophoblastic IGF2 expression. Protein levels of IGF2 in serum, intra-amniotic fluid and left lungs were measured by enzyme-linked immunosorbent assay.. Significant growth retardation of placenta and left lungs was observed in the CDH group compared to control and nitrofen+RA group. Markedly increased decidual RARα and decreased IGF2 immunoreactivity were found in the CDH group compared to control and nitrofen+RA group. Significantly decreased IGF2 protein levels were detected in serum, intra-amniotic fluid and left lungs in the CDH group compared to control and nitrofen+RA group.. Our findings suggest that nitrofen may disturb trophoblastic IGF2 expression through decidual RARα resulting in retarded placental growth and PH in the nitrofen-induced CDH. Prenatal administration of RA may promote lung and placental growth by increasing trophoblastic IGF2 expression.

Topics: Animals; Antineoplastic Agents; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Female; Gene Expression Regulation, Developmental; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Insulin-Like Growth Factor II; Lung; Phenyl Ethers; Pregnancy; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction; Tretinoin; Trophoblasts; Up-Regulation

Prenatal administration of all-trans retinoic acid (ATRA) has been shown to stimulate alveolarization in nitrofen-induced pulmonary hypoplasia (PH) associated with congenital diaphragmatic hernia (CDH). Lipid-containing interstitial lipofibroblasts (LIFs), characterized by adipocyte differentiation-related protein (ADRP), play a critical role in alveolar development by coordinating lipid homeostasis. Previous studies have demonstrated that ATRA positively affects LIF expression in developing lungs. We hypothesized that pulmonary LIF expression is increased after prenatal ATRA treatment in the nitrofen model of CDH-associated PH.. Timed-pregnant rats were treated with nitrofen or vehicle on E9.5, followed by injection of ATRA or placebo on E18.5, E19.5, and E20.5. Fetal lungs were dissected on E21.5 and divided into Control+Placebo, Control+ATRA, Nitrofen+Placebo, and Nitrofen+ATRA. Pulmonary gene expression levels of ADRP were analyzed by quantitative real-time polymerase chain reaction, and LIF expression was investigated by ADRP immunohistochemistry, oil-red-O-, and immunofluorescence-double-staining.. Relative mRNA expression of pulmonary ADRP was significantly increased in Nitrofen+ATRA compared to Nitrofen+Placebo (0.31±0.02 vs. 0.08±0.01; P<0.0001). ADRP immunoreactivity and oil-red-O-staining were markedly increased in alveolar interstitium of Nitrofen+ATRA compared to Nitrofen+Placebo. Immunofluorescence-double-staining confirmed markedly increased LIF expression in alveolar walls of Nitrofen+ATRA compared to Nitrofen+Placebo.. Increased LIF expression after prenatal treatment with ATRA in nitrofen-induced PH suggests that ATRA may have a therapeutic potential in attenuating CDH-associated PH by stimulating alveolar development.

Topics: Animals; Cell Differentiation; Female; Fibroblasts; Gene Expression Regulation, Developmental; Hernias, Diaphragmatic, Congenital; Immunohistochemistry; Lung; Membrane Proteins; Organogenesis; Perilipin-2; Phenyl Ethers; Pregnancy; Pregnancy, Animal; Rats; Rats, Sprague-Dawley; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tretinoin; Up-Regulation

Pulmonary hypoplasia (PH), characterized by alveolar immaturity, is one of the leading causes of respiratory insufficiency in newborns with congenital diaphragmatic hernia (CDH). Leptin (Lep) and its receptor (Lep-R) play an important role in fetal lung growth by stimulating alveolar differentiation and maturation. Lep and Lep-R are strongly expressed by alveolar cells during the saccular stage of fetal lung development. Lep-deficient mice exhibit decreased alveolarization with reduced pulmonary surfactant phospholipid synthesis, similar to human and nitrofen-induced PH. Prenatal administration of all-trans retinoic acid (ATRA) has been shown to stimulate alveolarization in nitrofen-induced PH. Recent studies have demonstrated that Lep and Lep-R expression in developing lungs is regulated by ATRA. We hypothesized that prenatal treatment with ATRA increases pulmonary Lep and Lep-R expression in the nitrofen model of CDH-associated PH.. Time-mated rats received either 100 mg nitrofen or vehicle via oral-gastric lavage on embryonic day 9.5 (E9.5). Control and nitrofen-exposed dams were randomly assigned to either intraperitoneal ATRA (5 mg/kg/d) or placebo administration on E18.5, E19.5 and E20.5. Fetal lungs were harvested on E21.5, and divided into Control+Placebo, Control+ATRA, Nitrofen+Placebo and Nitrofen+ATRA. Alveolarization was assessed using stereo- and morphometric analysis techniques. Surfactant phospholipid synthesis was analyzed by labeling for surfactant protein B (SP-B). Pulmonary gene expression levels of Lep and Lep-R were determined using quantitative real-time polymerase chain reaction. Immunohistochemical staining for Lep and Lep-R was performed to evaluate alveolar protein expression and localization.. In vivo administration of ATRA resulted in significantly increased lung-to-body weight ratio with enhanced radial alveolar count and decreased mean linear intercept compared to placebo treatment. Immunofluorescence analysis demonstrated markedly increased pulmonary SP-B expression in Nitrofen+ATRA compared to Nitrofen+Placebo. Relative mRNA expression of Lep and Lep-R was significantly increased in Nitrofen+ATRA compared to Nitrofen+Placebo. Lep and Lep-R immunoreactivity was markedly increased in interstitial and alveolar epithelial cells of Nitrofen+ATRA compared to Nitrofen+Placebo.. Increased Lep and Lep-R expression after prenatal administration of ATRA in nitrofen-induced PH suggests that ATRA may have therapeutic potential in attenuating CDH-associated PH by stimulating alveolarization and de novo surfactant production.

Topics: Animals; Disease Models, Animal; Female; Hernias, Diaphragmatic, Congenital; Immunohistochemistry; Leptin; Lung; Organogenesis; Pregnancy; Pregnancy, Animal; Rats; Rats, Sprague-Dawley; Real-Time Polymerase Chain Reaction; RNA, Messenger; Signal Transduction; Tretinoin; Up-Regulation

Congenital diaphragmatic hernia (CDH) is a common birth defect that results in a high degree of neonatal morbidity and mortality, but its pathological mechanisms are largely unknown. Therefore, we performed a forward genetic screen in mice to identify unique genes, models, and mechanisms of abnormal diaphragm development. We identified a mutant allele of kinesin family member 7 (Kif7), the disorganized diaphragm (dd). Embryos homozygous for the dd allele possess communicating diaphragmatic hernias, central tendon patterning defects, and increased cell proliferation with diaphragmatic tissue hyperplasia. Because the patterning of the central tendon is undescribed, we analyzed the expression of genes regulating tendonogenesis in dd/dd mutant embryos, and we determined that retinoic acid (RA) signaling was misregulautted. To further investigate the role of Kif7 and RA signaling in the development of the embryonic diaphragm, we established primary mesenchymal cultures of WT embryonic day 13.5 diaphragmatic cells. We determined that RA signaling is necessary for the expression of tendon markers as well as the expression of other CDH-associated genes. Knockdown of Kif7, and retinoic acid receptors alpha (Rara), beta (Rarb), and gamma (Rarg) indicated that RA signaling is dependent on these genes to promote tendonogenesis within the embryonic diaphragm. Taken together, our results provide evidence for a model in which inhibition of RA receptor signaling promotes CDH pathogenesis through a complex gene network.

Topics: Alleles; Animals; Body Patterning; Cell Differentiation; Cell Line; Diaphragm; Gene Expression Regulation, Developmental; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Kinesins; Mice; Mice, Mutant Strains; Muscle Proteins; Receptors, Retinoic Acid; Retinoic Acid Receptor alpha; Retinoic Acid Receptor gamma; Signal Transduction; Tendons; Tretinoin

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.. 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.. 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.. 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.

Topics: Animals; Cell Differentiation; Connexin 43; Disease Models, Animal; Drug Evaluation, Preclinical; Epithelial Cells; Female; Fetal Therapies; Gene Expression Regulation, Developmental; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Injections, Intraperitoneal; Lung; Phenyl Ethers; Pregnancy; Random Allocation; Rats; Rats, Sprague-Dawley; Real-Time Polymerase Chain Reaction; Respiratory Mucosa; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tretinoin; Up-Regulation

We sought to investigate the effects of antenatal retinoic acid on the pulmonary vasculature and vascular endothelial growth factor (VEGF) and VEGF receptors (VEGFR) expression in a nitrofen-induced congenital diaphragmatic hernia (CDH) model.. Rat fetuses were exposed to nitrofen at gestational day 9.5 and/or all-trans retinoic acid (ATRA) at gestational days 18.5-20.5. We assessed lung growth, airway, and vascular morphometry. VEGF, VEGFR1, and VEGFR2 expression was analyzed by Western blotting and immunohistochemistry. Continuous data were analyzed by analysis of variance and Kruskal-Wallis test.. CDH decreased lung to body weight ratio, increased mean linear intercept and mean transection length/airspace, and decreased mean airspace cord length. ATRA did not affect lung growth or morphometry. CDH increased proportional medial wall thickness of arterioles while ATRA reduced it. ATRA recovered expression of VEGF and receptors, which were reduced in CDH.. Retinoic acid and VEGF may provide pathways for preventing pulmonary hypertension in CDH.

Topics: Analysis of Variance; Angiogenesis Inducing Agents; Animals; Arterioles; Biomarkers; Blotting, Western; Disease Models, Animal; Female; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Lung; Phenyl Ethers; Pregnancy; Rats; Rats, Sprague-Dawley; Teratogens; Treatment Outcome; Tretinoin; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factor Receptor-1; Vascular Endothelial Growth Factor Receptor-2

Pulmonary hypoplasia (PH), the leading cause of mortality in congenital diaphragmatic hernia (CDH), is associated with arrested alveolarization. Late gestation lung protein 1 (LGL1) plays a crucial role in the regulation of alveolarization. Inhibition of LGL1 impairs alveolar maturation in fetal rat lungs. LGL1 heterozygotus knockout mice display delayed lung maturation. It is well known that prenatal administration of retinoic acid (RA) stimulates alveologenesis in nitrofen-induced PH. In vitro studies have reported that RA is a key modulator of LGL1 during alveologenesis. We hypothesized, that pulmonary gene expression of LGL1 is downregulated in the late stage of lung development, and that prenatal administration of RA upregulates pulmonary LGL1 expression in the nitrofen CDH model.. Pregnant rats were exposed to nitrofen on day 9 (D9) of gestation. RA was given intraperitoneally on D18, D19 and D20. Fetal lungs were dissected on D21 and divided into control, control + RA, CDH and CDH + RA group. Expression levels of LGL1 were determined using RT-PCR and immunohistochemistry.. On D21, LGL1 relative mRNA expression levels were significantly downregulated in CDH group compared to controls. After RA treatment, gene expression levels of LGL1 were significantly upregulated in CDH + RA and control + RA compared to CDH group. Immunohistochemical studies confirmed these results.. Downregulation of pulmonary LGL1 gene expression in the late stage of lung development may interfere with normal alveologenesis. Upregulation of LGL1 pulmonary gene expression after RA treatment may promote lung growth by stimulating alveologenesis in the nitrofen CDH model.

Topics: Animals; Female; Gene Expression Regulation, Developmental; Gestational Age; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Immunohistochemistry; Lung; Lung Diseases; Phenyl Ethers; Pregnancy; Pregnancy, Animal; Proteins; Rats; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tretinoin

Nitrofen-induced congenital diaphragmatic hernia (CDH) model has been widely used to investigate the pathogenesis of pulmonary hypoplasia (PH) in CDH. Recent studies have suggested that retinoids may be involved in the molecular mechanisms of PH in CDH. Prenatal treatment with retinoic acid (RA) has been reported to improve the growth of hypoplastic lung in the nitrofen CDH model. Midkine (MK), a RA-responsive growth factor, plays key roles in various organogenesis including lung development. In fetal lung, MK mRNA expression has its peak at E13.5-E16.5 and is markedly decreased during mid-to-late gestation, indicating its important role in early lung morphogenesis. We designed this study to investigate the hypothesis that the pulmonary MK gene expression is downregulated in the early lung morphogenesis in the nitrofen-induced PH, and to evaluate the effect of prenatal RA treatment on pulmonary MK gene expression in the nitrofen-induced CDH model.. Pregnant rats were exposed to either olive oil or nitrofen on day 9 of gestation (D9). Fetal lungs were harvested on D15, D18, and D21 and divided into control, nitrofen with or without CDH [CDH(+) or CDH(-)]. In addition, RA was given on days D18, D19, and D20 and fetal lungs were harvested on D21, and then divided into control + RA and nitrofen + RA. The pulmonary gene expression levels of MK were evaluated by real-time RT-PCR and statistically analyzed. Immunohistochemistry was also performed to examine protein expression/distribution of MK in fetal lung.. The relative mRNA expression levels of MK were significantly downregulated in nitrofen group compared to controls at D15 ((§)p < 0.01), whereas there were no significant differences at D18 and D21. MK gene expression levels were significantly upregulated in nitrofen + RA (0.71 ± 0.17) compared to the control (0.35 ± 0.16), CDH(-) (0.24 ± 0.15), CDH(+) (0.39 ± 0.19) and control + RA (0.47 ± 0.13) (*p < 0.05). Immunoreactivity of MK was also markedly decreased in nitrofen lungs compared to controls on D15, and increased in nitrofen + RA lungs compared to the other lungs on D21.. Downregulation of MK gene on D15 may contribute to primary PH in the nitrofen CDH model by disrupting early lung morphogenesis. Upregulation of MK gene after RA treatment in the nitrofen-induced hypoplastic lung suggests that RA may have a therapeutic potential to rescue PH in CDH through RA-responsive growth factor signaling.

Topics: Animals; Cytokines; Disease Models, Animal; Down-Regulation; Female; Gene Expression Regulation, Developmental; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Immunohistochemistry; Lung; Midkine; Pregnancy; Pregnancy, Animal; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tretinoin

Recent studies have suggested that retinoids may be involved in the molecular mechanisms of pulmonary hypoplasia (PH) in congenital diaphragmatic hernia (CDH). Connective tissue growth factor (CTGF) plays a key role in foetal lung development and remodelling during later gestation. CTGF knockout mice exhibit PH with similar characteristics to the human and nitrofen-induced PH. Prenatal administration of retinoic acid (RA) has been shown to stimulate alveologenesis in nitrofen-induced PH. In vitro studies have revealed that RA can induce CTGF gene expression. We hypothesized that pulmonary gene expression of CTGF is downregulated during the later stages of lung development, and that prenatal administration of RA upregulates CTGF in the nitrofen CDH model.. Pregnant rats were exposed to either olive oil or nitrofen on day 9 (D9) of gestation. RA was given intraperitoneally on D18, D19 and D20. Foetuses were harvested on D21 and divided into control, CDH, control + RA and CDH + RA group. Pulmonary CTGF gene and protein expression levels were determined using RT-PCR and immunohistochemistry.. On D21, CTGF relative mRNA expression levels were significantly downregulated in CDH group compared to controls. After RA treatment, expression levels of CTGF were significantly upregulated in CDH + RA and control + RA compared to the CDH group. Immunohistochemical studies confirmed these results.. Downregulation of pulmonary CTGF gene and protein expression during later stages of lung development may interfere with normal alveologenesis in the nitrofen CDH model. Upregulation of CTGF pulmonary gene expression after prenatal RA treatment may promote lung growth by promoting alveologenesis in the nitrofen-induced CDH model.

Topics: Animals; Connective Tissue Growth Factor; Disease Models, Animal; Female; Gene Expression Regulation, Developmental; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Immunohistochemistry; Phenyl Ethers; Pregnancy; Pregnancy, Animal; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tretinoin; Up-Regulation

Pulmonary hypoplasia (PH) is the main cause of mortality in newborns with congenital diaphragmatic hernia (CDH). Prenatal administration of retinoic acid (RA) stimulates alveologenesis in the nitrofen-induced pulmonary hypoplasia. Insulin-like growth factor receptors (IGFRs) play a crucial role in alveologenesis during lung development. We recently demonstrated that IGFRs were downregulated in later stages of lung development in the nitrofen CDH model. Several studies suggest the ability of RA to regulate insulin-like growth factor signaling. We hypothesized that IGFRs pulmonary gene expression is upregulated after the administration of RA in the nitrofen-induced CDH model.. Pregnant rats were exposed to either olive oil or nitrofen on day 9 (D9) of gestation. RA was given intraperitoneally on days D18, D19, and D20. Fetal lungs were dissected on D21 and divided into control, control + RA, CDH, and CDH + RA group. IGFRs gene and protein expression were determined using RT-PCR and immunohistochemistry.. mRNA expression levels of IGFRs were significantly increased in control + RA and CDH + RA compared with CDH group. Immunoreactivity of IGFRs was markedly increased in control + RA and CDH + RA compared with CDH lungs.. Upregulation of pulmonary gene and protein expression of IGFRs after prenatal RA treatment in the nitrofen model suggests that RA may promote lung growth by stimulating IGFRs mediated alveologenesis.

Topics: Animals; Animals, Newborn; Disease Models, Animal; Down-Regulation; Female; Gene Expression Regulation, Developmental; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Immunohistochemistry; Lung; Lung Diseases; Phenyl Ethers; Pregnancy; Prenatal Exposure Delayed Effects; Rats; Rats, Sprague-Dawley; Receptor, IGF Type 1; Receptor, IGF Type 2; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tretinoin; Up-Regulation

Congenital diaphragmatic hernia (CDH) is a frequently occurring source of severe neonatal respiratory distress. It has been hypothesized that abnormal retinoid signaling contributes to the etiology of this developmental anomaly. Here, we use rodent models toward specifically understanding the role of retinoid signaling in the developing diaphragm and how its perturbation is a common mechanism in drug-induced CDH. This includes monitoring of retinoic acid (RA) response element (RARE) activation with RARE-lacZ mice, RA supplementation studies, systematic analyses of the expression profile of key elements in the RA signaling pathway within the developing diaphragm, and the in utero delivery of a RA receptor (RAR) antagonist. These data demonstrate the timing of RARE perturbation by CDH-inducing teratogens and the efficacy of RA supplementation. Furthermore, a detailed profile of retinoid binding proteins, synthetic enzymes, and retinoid receptors within primordial diaphragm cells was obtained. The expression profile of RAR-alpha was particularly striking in regard to its overlap with the regions of primordial diaphragm affected in multiple CDH models. Blocking of RAR signaling with the pan-RAR antagonist BMS493 induced a very high degree of CDH, with a marked left-right sidedness that depended on the timing of drug delivery. Collectively, these data demonstrate that retinoid signaling is essential for normal diaphragm development, providing further support to the hypothesis that abnormalities related to the retinoid signaling pathway cause diaphragmatic defects. This study also yielded a novel experimental model that should prove particularly useful for further studies of CDH.

Topics: Animals; beta-Galactosidase; Diaphragm; Dietary Supplements; Enzyme Activation; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Mice; Rats; Receptors, Retinoic Acid; Response Elements; Retinal Dehydrogenase; Retinoids; Signal Transduction; Stilbenes; Teratogens; Tretinoin

Although there is strong evidence that genetic factors play a pathogenic role in congenital diaphragmatic hernia (CDH), few causal genes have been identified in humans. A number of studies, essentially in animal models, have suggested that disruption of the retinoid signaling pathway plays a major role in the pathogenesis of CDH. Our hypothesis is that human fetal skin fibroblasts express some metabolic and molecular actors of the retinoid pathway and that they offer convenient cellular material for investigating the molecular retinoid pathway defects associated with CDH.. We first established the expression of receptors, enzymes and binding proteins involved in the retinoic acid (RA) pathway in non-CDH fetal skin fibroblasts using RT-PCR and immunocytochemistry approaches. We then studied the expression of these genes in skin fibroblasts from seven fetuses with isolated and nonisolated CDH.. Fetal skin fibroblasts expressed enzymes involved in RA metabolism as well as nuclear receptors for signal transduction. Basal levels of retinoic acid receptor, retinaldehyde dehydrogenase 2, and CYP26 (cytochrome P450 RAI) expression were altered in two of seven fetuses. Interestingly, these genes were previously described as abnormally expressed in CDH physiopathology.. Our results suggest that human fetal skin fibroblasts could be useful for studying retinoid signaling pathway disruption in the context of CDH. Our proposal is strengthened by the fact that we identified CDH fetuses for which molecular and metabolic actors of the retinoid pathway were not detected.

Topics: Cells, Cultured; Fetus; Fibroblasts; Gene Expression Regulation, Developmental; Gestational Age; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Humans; Retinoid X Receptors; RNA, Messenger; Signal Transduction; Skin; Tretinoin

Retinoids play an important role in lung development. A recent study has demonstrated that prenatal treatment with retinoic acid (RA) stimulates alveologenesis in hypoplastic lungs in the nitrofen model of congenital diaphragmatic hernia (CDH). Furthermore, it has also been demonstrated that the differentiation from alveolar epithelial cells type II (AECs-II) into alveolar epithelial cells type I (AECs-I), which is the key process in lung development, is disturbed in this model. We hypothesized that retinoids promote alveologenesis by stimulating differentiation of AECs-II to AECs-I at the end of gestation; and therefore, we investigated the effect of RA on the pulmonary expression of intercellular adhesion molecule 1 (ICAM-1), a marker for AECs-I, and thyroid transcription factor 1 (Ttf-1), a marker for AECs-II, in nitrofen-induced hypoplastic lungs.. Pregnant rats were exposed to either olive oil or 100 mg nitrofen on day of gestation (D) 9. Five milligrams per kilogram of RA was given intraperitoneally on D18, D19, and D20; and fetuses were recovered on D21. We had 4 study groups: control (n = 7), control + RA (n = 7), CDH (n = 6), and CDH + RA (n = 6). The expression of ICAM-1 and Ttf-1 was analysed in each lung by real-time reverse transcription polymerase chain reaction and immunohistochemistry. One-way analysis of variance test was used for statistical analysis.. Expression levels of ICAM-1 were significantly reduced in CDH lungs compared with normal controls, whereas levels increased significantly in CDH group after the addition of RA (P < .05). Expression levels of Ttf-1 were significantly decreased in lungs from RA-treated CDH animals compared with CDH without RA (P < .05). The ICAM-1 and Ttf-1 immunoreactivity demonstrated similar pattern of expression in various groups.. Our results demonstrate that prenatal treatment with RA accelerates AEC-I proliferation in the hypoplastic lung in CDH.

Topics: Animals; Cell Proliferation; Disease Models, Animal; Drug Administration Schedule; Epithelial Cells; Female; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Immunohistochemistry; Lung; Phenyl Ethers; Pregnancy; Pregnancy, Animal; Prenatal Care; Probability; Pulmonary Alveoli; Random Allocation; Rats; Rats, Sprague-Dawley; Reference Values; Respiratory System Abnormalities; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sensitivity and Specificity; Tretinoin

Severe pulmonary hypoplasia remains the main cause of the high mortality in newborn infants with congenital diaphragmatic hernia (CDH). Retinoids are a family of molecules derived from vitamin A, which play an important role in lung development. We hypothesized that retinoids promote alveologenesis at the end of gestation and therefore designed this study to investigate the effects of retinoid acid on nitrofen-induced hypoplastic lungs in CDH.. Pregnant rats were exposed to either olive oil or 100 mg nitrofen on day 9 of gestation. Retinoic acid 5 mg/kg was given intraperitoneally on days 18, 19, and 20 of gestation and fetuses were recovered on day 21. We had 4 study groups: control (n = 24), control + retinoic acid (n = 22), CDH (n = 24), and CDH + retinoic acid (n = 19). Lungs from the 4 study groups were fixed, and the following stereological measurements were performed on vertical random sections: total lung volume, volume density of airspaces, volume density of air walls, gas exchange surface area, alveolar volume, and total number of alveoli per lung. Total DNA content of each lung was measured using a spectrophotometer.. Total lung volume increased in CDH lungs after the addition of retinoic acid but remained the same in the control group. Gas exchange surface area was larger in CDH lungs after the addition of retinoic acid but remained unchanged in the control group. The total number of alveoli per lung was higher after the addition of retinoic acid. Total DNA content as well as total DNA content-lung weight ratio of the left lung increased significantly in the CDH group after the addition of retinoic acid compared with CDH without retinoic acid.. Our results demonstrate that prenatal treatment with retinoic acid stimulates alveologenesis in hypoplastic lungs in CDH.

Topics: Animals; Disease Models, Animal; Female; Fetal Organ Maturity; Hernias, Diaphragmatic, Congenital; Injections, Intraperitoneal; Lung; Phenyl Ethers; Pregnancy; Pregnancy, Animal; Prenatal Care; Probability; Pulmonary Alveoli; Random Allocation; Rats; Rats, Sprague-Dawley; Reference Values; Respiratory System Abnormalities; Sensitivity and Specificity; Tretinoin

Maternal retinoid administration has beneficial effects on lung development in the nitrofen rodent toxic model of congenital diaphragmatic hernia (DH). We wanted to investigate the effects in a surgical model, where the retinoid signaling pathway is not primarily disrupted by the toxic agent. We created DH in fetal rabbits at day 23 of gestation, administrated to the does all trans-retinoic acid (ATRA) or vehicle (VHC) intramuscularly for 8 consecutive days and harvested normal and operated (DH) fetuses at 31 d (n = 7 in each group). Normal lungs exposed to ATRA had increased surfactant protein mRNA levels without change in type II pneumocyte density. There was no measurable effect on lung-to-body weight ratio and airway morphometry by ATRA. In DH lungs (DH/VHC) surfactant protein mRNA levels were increased, as well as the density of type II pneumocytes. When supplemented with ATRA (DH/ATRA) these parameters returned to normal (VHC). Cell proliferation or apoptosis were not influenced by ATRA supplementation. In conclusion, maternal ATRA supplementation does not affect gross anatomic, morphologic or proliferation indices in hypoplastic lungs related to surgically induced DH in rabbit. However, ATRA lowers surfactant protein expression and normalizes type I/II pneumocyte ratio to what is observed in normal lungs.

Topics: Animals; Blotting, Western; Caveolin 1; Cell Death; Female; Fetal Organ Maturity; Fetus; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Lung; Models, Animal; Pregnancy; Prenatal Exposure Delayed Effects; Proliferating Cell Nuclear Antigen; Pulmonary Surfactant-Associated Protein A; Pulmonary Surfactant-Associated Protein B; Pulmonary Surfactant-Associated Protein C; Rabbits; Reverse Transcriptase Polymerase Chain Reaction; Tretinoin; Vitamins

Retinoids are a group of molecules derived from vitamin A, which play an important role in lung development. Within the cell, retinol can either be oxidized to retinal or esterified to retinyl esters by lecithin : retinol acyltransferase (LRAT) for storage. Retinal is then oxidized to an active metabolite of vitamin A, retinoic acid (RA) by retinal dehydrogenase (RALDH). RA is the active metabolite of vitamin A. Cyp26 (a1,b1, and c1), which is a member of the cytochrome P450 family, acts by reducing the activity of RA. Cyp26 type b1 is the predominant subtype expressed in the murine lung. Several studies have suggested that nitrofen may interfere with the retinoid pathway resulting in congenital diaphragmatic hernia (CDH) and pulmonary hypoplasia. Recently, it was reported that nitrofen may act by inhibiting RALDH2. The aim of this study was to examine the pulmonary expression of Cyp26b1, LRAT, and RALDH2, the key enzymes involved in the synthesis of RA, in order to understand the mechanisms underlying pulmonary hypoplasia in the nitrofen CDH model. Pregnant rats were exposed to either olive oil or 100 mg of nitrofen on day 9 of gestation (D9). Fetal lungs were harvested at D15, D17, D19, and D21. D17, D19, and D21 lungs were divided into three groups: control, nitrofen without CDH and nitrofen with CDH, whereas D15 lungs were divided into only two groups; control and nitrofen as the diaphragm is not fully formed yet at this stage. Real- time PCR was performed to evaluate the relative level of Cyp26b1, LRAT, and RALDH2 expression in the lung. Relative levels of Cyp26b1 mRNA were significantly decreased in the lungs of nitrofen with CDH (D17;0.19 +/- 0.09, D19;0.70 +/- 0.20, D21;0.40 +/- 0.36) and nitrofen without CDH (D17;0.14 +/- 0.06, D19;0.54 +/- 0.42, D21;0.51 +/- 0.56) compared to controls (D17;0.35 +/- 0.16, D19;1.15 +/- 0.48, D21;1.28 +/- 0.78) (P < 0.05). LRAT expression was also significantly decreased in nitrofen with CDH (D17; 19.3 +/- 7.8, D19; 4.3 +/- 1.1, D21; 3.3 +/- 1.6) and nitrofen without CDH (D17; 21.2 +/- 11.1, D19; 4.5 +/- 3.6, D21; 4.1 +/- 1.6) compared to controls (D17; 153.7 +/- 29.8, D19; 26.8 +/- 16.8 D21; 10.1 +/- 3.8) (P < 0.05). There was no significant difference in the relative levels of Cyp26b1 and LRAT between nitrofen with CDH and nitrofen without CDH. There were no significant differences in RALDH2 expression among the groups at any stages. Down-regulation of Cryp26b1 and LRAT demonstrates that RA content is decreased i

Topics: Acyltransferases; Animals; Cytochrome P-450 Enzyme System; Disease Models, Animal; Down-Regulation; Female; Gene Expression Regulation, Developmental; Herbicides; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Lung; Lung Diseases; Olive Oil; Phenyl Ethers; Plant Oils; Pregnancy; Prenatal Exposure Delayed Effects; Rats; Rats, Sprague-Dawley; Retinal Dehydrogenase; Retinoic Acid 4-Hydroxylase; Reverse Transcriptase Polymerase Chain Reaction; Tretinoin

This review highlights the relevance of the neural crest (NC) as a developmental control mechanism involved in several pediatric surgical conditions and the investigative interest of following some of its known signaling pathways.. The participation of the NC in facial clefts, ear defects, branchial fistulae and cysts, heart outflow tract and aortic arch anomalies, pigmentary disorders, abnormal enteric innervation, neural tumors, hemangiomas, and vascular anomalies is briefly reviewed. Then, the literature on clinical and experimental esophageal atresia-tracheoesophageal fistula (EA-TEF) and congenital diaphragmatic hernia (CDH) is reviewed for the presence of associated NC defects. Finally, some of the molecular signaling pathways involved in both conditions (sonic hedgehog, Hox genes, and retinoids) are summarized.. The association of facial, cardiovascular, thymic, parathyroid, and C-cell defects together with anomalies of extrinsic and intrinsic esophageal innervation in babies and/or animals with both EA-TEF and CDH strongly supports the hypothesis that NC is involved in the pathogenesis of these malformative clusters. On the other hand, both EA-TEF and CDH are observed in mice mutant for genes involved in the previously mentioned signaling pathways.. The investigation of NC-related molecular pathogenic pathways involved in malformative associations like EA-TEF and CDH that are induced by chromosomal anomalies, chemical teratogens, and engineered mutations is a promising way of clarifying why and how some pediatric surgical conditions occur. Pediatric surgeons should be actively involved in these investigations.

Topics: Abnormalities, Multiple; Blood Vessels; Branchial Region; Cardiovascular Abnormalities; Cell Lineage; Cell Movement; Child; Child, Preschool; Enteric Nervous System; Esophageal Atresia; Face; Genes, Homeobox; Hedgehog Proteins; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Homeodomain Proteins; Humans; Infant; Infant, Newborn; Neoplasms; Neural Crest; Patched Receptors; Pigmentation Disorders; Receptors, Cell Surface; Receptors, G-Protein-Coupled; Receptors, Retinoic Acid; Signal Transduction; Smoothened Receptor; Syndrome; Transcription Factors; Tretinoin; Zinc Finger Protein GLI1

There is increasing evidence to suggest that the retinoid pathway is involved in the pathogenesis of congenital diaphragmatic hernia (CDH). We hypothesised that retinoids are involved in the pathogenesis of associated pulmonary hypoplasia in CDH and therefore designed this study to investigate the effects of retinoid acid on nitrofen-induced hypoplastic lungs. Pregnant rats were exposed to either olive oil or 100 mg nitrofen on day 9.5 of gestation. Foetal lungs were harvested on embryonic day 13.5 and were cultured for 96 h with or without exogenous retinoic acid (RA) (1 muM) added daily to the culture medium. Lungs were divided into four study groups: control (n=31); control + RA (n=19); nitrofen (n=19); and nitrofen + RA (n=12). Lung growth was assessed in each group by measuring branching morphogenesis, total DNA content and the proportion of proliferating cells stained by immunohistochemistry. One-way ANOVA test was used for statistical analysis. Retinoic acid significantly increased the growth of nitrofen-induced hypoplastic lungs, whilst growth of control lungs did not change. The number of lung buds and lung area of nitrofen-exposed hypoplastic lungs after 96 h of culture significantly increased after the addition of RA compared to the non-treated hypoplastic lungs (25.75+/-6.47 vs 15.11+/-3.29 and 0.98+/-0.18 mm(2) vs 0.65+/-0.13 mm(2), respectively; P<0.0001). Lung perimeter was also higher when RA was added to hypoplastic lungs compared to the non-treated ones, although it did not reach significance (12.51+/-2.53 mm vs 11.19+/-2.56 mm; P=0.17). Conversely, the addition of RA to control lungs did not affect the number of lung buds, lung area or lung perimeter after 96 h in culture compared to the non-treated ones (31.28+/-4.66 vs 31.81+/-6.67; 1.29+/-0.18(2) vs 1.29+/-0.23 mm(2) and 18.47+/-3.47 mm vs 17.89+/-2.94 mm, respectively; P=NS). Retinoic acid also increased the total DNA content and the proportion of proliferating cells in hypoplastic lungs compared to the non-treated ones (2.59+/-0.58 mug vs 1.96+/-0.31 mug and 57.89+/-9.46% vs 36.76+/-8.15%, respectively; P<0.001). The addition of RA did not affect either total DNA content or the proportion of proliferating cells in control lungs compared to the non-treated ones (4.04+/-0.64 mug vs 3.79+/-0.85 mug and 58.67+/-11.23% vs 56.03+/-10.36%, respectively; P=NS). This study demonstrates for the first time that RA rescues lung hypoplasia in nitrofen-induced hypoplastic lungs. These results su

Topics: Analysis of Variance; Animals; Disease Models, Animal; Female; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Lung; Pregnancy; Random Allocation; Rats; Rats, Sprague-Dawley; Retinoids; Signal Transduction; Tretinoin

Defects very similar to those seen in infants born with congenital diaphragmatic hernias can be induced in rodents by the administration of the teratogen nitrofen. There is an interest in understanding the biochemical mechanisms of nitrofen's actions in hopes of gaining insights into the etiology of congenital diaphragmatic hernia. In this study, we test the hypothesis that nitrofen is acting to perturb the retinoid signaling pathway by utilizing genetically engineered mice that have the lacZ reporter gene linked to a retinoic acid response element (RARE). We demonstrate a pronounced suppression of RARE-lacZ expression by nitrofen in vitro (by approximately 64%) and in vivo (by approximately 43%).

Topics: Animals; beta-Galactosidase; Disease Models, Animal; Drug Interactions; Gene Expression; Herbicides; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; In Vitro Techniques; Lac Operon; Mice; Phenyl Ethers; Response Elements; Teratogens; Tretinoin