Chorioamnionitis induces enteric nervous system injury: effects of timing and inflammation in the ovine fetus.

BACKGROUND: Chorioamnionitis, inflammation of the chorion and amnion, which often results from intrauterine infection, is associated with premature birth and contributes to significant neonatal morbidity and mortality, including necrotizing enterocolitis (NEC). Recently, we have shown that chronic chorioamnionitis is associated with significant structural enteric nervous system (ENS) abnormalities that may predispose to later NEC development. Understanding time point specific effects of an intra-amniotic (IA) infection on the ENS is important for further understanding the pathophysiological processes and for finding a window for optimal therapeutic strategies for an individual patient. The aim of this study was therefore to gain insight in the longitudinal effects of intrauterine LPS exposure (ranging from 5 h to 15 days before premature delivery) on the intestinal mucosa, submucosa, and ENS in fetal lambs by use of a well-established translational ovine chorioamnionitis model. METHODS: We used an ovine chorioamnionitis model to assess outcomes of the fetal ileal mucosa, submucosa and ENS following IA exposure to one dose of 10 mg LPS for 5, 12 or 24 h or 2, 4, 8 or 15 days. RESULTS: Four days of IA LPS exposure causes a decreased PGP9.5- and S100ß-positive surface area in the myenteric plexus along with submucosal and mucosal intestinal inflammation that coincided with systemic inflammation. These changes were preceded by a glial cell reaction with early systemic and local gut inflammation. ENS changes and inflammation recovered 15 days after the IA LPS exposure. CONCLUSIONS: The pattern of mucosal and submucosal inflammation, and ENS alterations in the fetus changed over time following IA LPS exposure. Although ENS damage seemed to recover after prolonged IA LPS exposure, additional postnatal inflammatory exposure, which a premature is likely to encounter, may further harm the ENS and influence functional outcome. In this context, 4 to 8 days of IA LPS exposure may form a period of increased ENS vulnerability and a potential window for optimal therapeutic strategies.

Clinical interventions to reduce stillbirths in sub-Saharan Africa: a mathematical model to estimate the potential reduction of stillbirths associated with specific obstetric conditions.

OBJECTIVE: Stillbirths are among the most common adverse pregnancy outcomes, with 98% occurring in low-income countries. More than one-third occur in sub-Saharan Africa (SSA). However, the medical conditions causing stillbirths and interventions to reduce stillbirths from these conditions are not well documented. We estimated the reductions in stillbirths possible with combinations of interventions. DESIGN: We developed a computerised model to estimate the impact of various interventions on stillbirths caused by the most common conditions. The model considered the location of obstetric care (home, clinic or hospital) and each intervention's efficacy, penetration and utilisation. Maternal transfers were also considered. SETTING AND POPULATION: Pregnancies in SSA in 2012. METHODS: For each condition, we created a series of scenarios involving different combinations of interventions and modelled their impact on stillbirth rates. MAIN OUTCOME MEASURES: Stillbirths associated with various maternal and fetal conditions and the percentage reduction with various interventions. RESULTS: Eight to ten maternal and fetal conditions were responsible for most stillbirths, but none for more than 15%. The most common conditions causing stillbirths in SSA include obstructed labour and uterine rupture, fetal distress and umbilical cord complications, fetal growth restriction, pre-eclampsia/eclampsia, and placental abruption/placenta praevia. Syphilis and malaria contribute smaller numbers. Reducing stillbirths requires appropriate diagnosis and management of each condition, usually including hospital care for monitoring and delivery, often by caesarean section. Maternal syphilis and malaria were the only conditions for which outpatient management alone reduced stillbirth. CONCLUSIONS: Most stillbirths in low-income countries occur at term and during labour and therefore are preventable by appropriate obstetric care. Management focused on the maternal and fetal conditions that cause stillbirths is necessary to achieve stillbirth rates approaching those found in high-income countries. TWEETABLE ABSTRACT: Reducing stillbirth incidence requires appropriate management of each causative condition and often caesarean delivery.

The clinical use of corticosteroids in pregnancy.

BACKGROUND: The use of antenatal steroid therapy is common in pregnancy. In early pregnancy, steroids may be used in women for the treatment of recurrent miscarriage or fetal abnormalities such as congenital adrenal hyperplasia. In mid-late pregnancy, the antenatal administration of corticosteroids to expectant mothers in anticipation of preterm birth is one of the most important advances in perinatal medicine; antenatal corticosteroids are now standard care for pregnancies at risk of premature delivery in high- and middle-income countries. The widespread uptake of this therapy is due to a compelling body of evidence demonstrating improved neonatal outcomes following antenatal corticosteroid exposure, stemming most notably from corticosteroid-driven maturation of fetal pulmonary function. As we approach the 50th anniversary of landmark work in this area by Liggins and Howie, it is apparent that much remains to be understood with regards to how we might best apply antenatal corticosteroid therapy to improve pregnancy outcomes at both early and mid to late gestation. METHODS: Drawing on advances in laboratory science, pre-clinical and clinical studies, we performed a narrative review of the scientific literature to provide a timely update on the benefits, risks and uncertainties regarding antenatal corticosteroid use in pregnancy. Three, well-established therapeutic uses of antenatal steroids, namely recurrent miscarriage, congenital adrenal hyperplasia and preterm birth, were selected to frame the review. RESULTS: Even the most well-established antenatal steroid therapies lack the comprehensive pharmacokinetic and dose-response data necessary to optimize dosing regimens. New insights into complex, tissue-specific corticosteroid signalling by genomic-dependent and independent mechanisms have not been used to inform corticosteroid treatment strategies. There is growing evidence that some fetal corticosteroid treatments are either ineffective, or may result in adverse outcomes, in addition to lasting epigenetic changes in a variety of homeostatic mechanisms. Nowhere is the need to better understand the intricacies of corticosteroid therapy better conveyed than in the findings of Althabe and colleagues who recently reported an increase in overall neonatal mortality and maternal morbidity in association with antenatal corticosteroid administration in low-resource settings. CONCLUSIONS: New research to clarify the benefits and potential risks of antenatal corticosteroid therapy is urgently needed, especially with regard to corticosteroid use in low-resource environments. We conclude that there is both significant scope and an urgent need for further research-informed refinement to the use of antenatal corticosteroids in pregnancy.

Antenatal ureaplasma infection impairs development of the fetal ovine gut in an IL-1-dependent manner.

Ureaplasma infection of the amniotic cavity is associated with adverse postnatal intestinal outcomes. We tested whether interleukin-1 (IL-1) signaling underlies intestinal pathology following ureaplasma exposure in fetal sheep. Pregnant ewes received intra-amniotic injections of ureaplasma or culture media for controls at 3, 7, and 14 d before preterm delivery at 124 d gestation (term 150 d). Intra-amniotic injections of recombinant human interleukin IL-1 receptor antagonist (rhIL-1ra) or saline for controls were given 3 h before and every 2 d after Ureaplasma injection. Ureaplasma exposure caused fetal gut inflammation within 7 d with damaged villus epithelium and gut barrier loss. Proliferation, differentiation, and maturation of enterocytes were significantly reduced after 7 d of ureaplasma exposure, leading to severe villus atrophy at 14 d. Inflammation, impaired development and villus atrophy of the fetal gut was largely prevented by intra-uterine rhIL-1ra treatment. These data form the basis for a clinical understanding of the role of ureaplasma in postnatal intestinal pathologies.

Antenatal exposure to chorioamnionitis affects lipid metabolism in 7-week-old sheep.

Antenatal exposure of the fetus to inflammation may alter postnatal organ development. In our previous work, we demonstrated that the fetal liver is involved in the systemic inflammation associated with chorioamnionitis, leading to metabolic changes. On the basis of these findings, we hypothesized that chorioamnionitis can lead to postnatal inflammation-related liver injury and disturbed lipid metabolism. Chorioamnionitis was induced in sheep by intra-amniotic injection of lipopolysaccharide (LPS) or saline at 90, 100 and 110 days of gestation. Liver homeostasis and lipid metabolism were analyzed at term and at 7 weeks of age. At term, hepatic T-lymphocytes and apoptotic hepatocytes were increased. In addition, hepatic cholesterol and triglyceride levels were decreased in LPS-exposed animals compared with controls. At 7 weeks of age, no hepatic inflammation could be detected. However, liver triglycerides and plasma cholesterol levels were increased in LPS-exposed animals relative to controls. The changes in lipid levels at 7 weeks of age were associated with increased leptin receptor mRNA levels, increased lipid peroxidation, increased expression of cytochrome c oxidase subunit 4 as a marker for mitochondrial function and increased circulating ceramide levels. These findings demonstrate that chorioamnionitis-mediated antenatal inflammation-related liver disturbances have long-lasting postnatal effects on lipid metabolism.

All-trans retinoic acid and intra-amniotic endotoxin-mediated effects on fetal sheep lung.

All-trans retinoic acid (RA) is a potent modulator of lung development. Chorioamnionitis, which is frequently associated with preterm birth, causes fetal lung inflammation and improves lung function but also results in alveolar simplification and microvascular injury. Endotoxin-mediated chorioamnionitis reduces RA concentration in the fetal lung to 16% of control values. We hypothesized that administration of RA to the fetus before induction of chorioamnionitis would preserve septation of the distal airspaces. Time-mated ewes with singletons were assigned to receive a fetal intramuscular treatment with 20,000 IU of RA in olive oil (or olive oil only) 3 hr prior to intra-amniotic injection of endotoxin (20 mg, E. coli 055:B5) or saline, at 124-day gestational age and 7 days after the fetal treatment. The right cranial lung lobe was processed for morphometric analysis. RA treatment did not affect chorioamnionitis-induced fetal and systemic inflammation or interleukin-8 concentrations in lung tissue. RA administration alone did not alter lung structure. Relative to control lungs (5 +/- 3 mL/kg), lung volume increased similarly with endotoxin (22 +/- 4 mL/kg) or RA plus endotoxin (20 +/- 3 mL/kg; P < 0.05). Alveolar wall thickness was 4.2 +/- 0.3 mum after endotoxin-induced chorioamnionitis, 6.0 +/- 0.4 mum in controls (P < 0.05 versus endotoxin) and 5.5 +/- 0.2 mum after RA and endotoxin (P < 0.05 versus control, n.s. versus endotoxin). The ratio of airspace versus tissue was 4.6 +/- 0.3 in endotoxin-induced chorioamnionitis, 2.1 +/- 0.3 in controls and 4.1 +/- 0.5 after RA and endotoxin. We conclude that fetal treatment with RA did not prevent inflammation-induced alveolar simplification.

The cardiopulmonary system: research and training opportunities.

The new biology has the potential to provide mechanistic insights into the causes and progression of complex cardiopulmonary diseases such as congenital heart disease and bronchopulmonary dysplasia. Such research requires collaborative investigation supported by sophisticated infrastructures and core facilities. Translating basic observations to clinical outcomes will require networks for collaborative translational research. The research initiatives require excellently trained and motivated clinician-scientists, but there are numerous barriers to the training and support of clinician-scientists in cardiology and neonatology.

Contribution of inflammation to lung injury and development.

Inflammation interferes with lung development in model systems and is present chronically in the lungs of preterm infants who develop bronchopulmonary dysplasia (BPD). Antenatal inflammation is very commonly associated with preterm deliveries, but there is generally minimal information about the duration, intensity, or organisms associated with chorioamnionitis. In preterm lamb models, chorioamnionitis causes a lung injury similar to BPD and also causes clinical lung maturation. Continuous exposure of the developing lung before and after delivery to inflammation may be central to the development of BPD.

Antenatal infection/inflammation and postnatal lung maturation and injury.

Chorioamnionitis is frequently associated with preterm deliveries before 30 weeks gestation. Chorioamnionitis correlates both with an increased risk of bronchopulmonary dysplasia and with a decreased risk of respiratory distress syndrome. Both interleukin-1alpha and endotoxin can induce inflammation in the fetal lungs and lung maturation after preterm birth when given by intra-amniotic injection. Inflammation can also result in an arrest of alveolarization, and this lung developmental abnormality is prominent in the lungs of preterm infants that die of bronchopulmonary dysplasia. The mechanisms by which infection/inflammation can have both beneficial and injurious effects on the preterm lung remain to be characterized.

Dose and time response after intraamniotic endotoxin in preterm lambs.

Intraamniotic endotoxin causes chorioamnionitis, which is followed by improved fetal lung function after 4 d in fetal sheep. We evaluated 0.1 mg, 1 mg, 4 mg, and 10 mg endotoxin for inflammation and lung maturation effects after 7 d. Four and 10 mg endotoxin caused similar lung maturation and inflammation in the lung and chorioamnion. The number of neutrophils in cord blood and the inflammatory cells in alveolar lavage and fetal lung tissue increased in a dose-dependent manner. Lower endotoxin doses induced indicators of chorioamnionitis, lung and systemic inflammation without inducing lung maturation. Therefore, some degree of inflammation can occur without subsequent lung maturation. The inflammatory changes caused by 4 mg endotoxin were assessed after 5 h, 24 h, 72 h, and 7 d to discern local versus systemic inflammation after intraamniotic endotoxin. At 5 h active inflammatory cells were in the airways producing hydrogen peroxide, and interleukin-6 and -8 were increased in the cord blood indicating both lung and systemic responses. Cells recruited into the amniotic fluid produced proinflammatory cytokine mRNA for 7 d with no cytokine mRNA in chorioamnion, lung, or spleen after 72 h. The cells in the amniotic fluid may be a source of prolonged fetal exposure to proinflammatory cytokines.

Biology of surfactant.

Surfactant is a metabolically active assembly of phospholipids and surfactant-specific proteins that is essential for normal lung mechanics. The surfactant proteins SP-A and SP-D also have important innate host defense functions. Surfactant metabolism in the developing lung differs from the adult lung by having slower kinetics of secretion with a longer half-life and more efficient recycling. Ventilation styles that injure the lung also result in altered surfactant function.

Effects of ventilation with different positive end-expiratory pressures on cytokine expression in the preterm lamb lung.

Ventilator-induced lung injury increases proinflammatory cytokines in the adult lung. We asked if positive end-expiratory pressure (PEEP) affects proinflammatory cytokine mRNA expression in the preterm lung. Preterm lambs at 129 +/- 3 d gestation were treated with 100 mg/kg recombinant human surfactant protein-C surfactant and ventilated for 2 or 7 h with 0, 4, or 7 cm H(2)O of PEEP. Unventilated fetal lambs were used as controls. Within 2 h of ventilation, alveolar total protein and activated neutrophils were increased and expression of mRNAs for the proinflammatory cytokines interleukin (IL)-1beta, IL-6, IL-8, and tumor necrosis factor-alpha (TNF-alpha) was increased in lung tissue of all ventilated animals relative to unventilated controls. Alveolar protein and neutrophils were higher for 0 and 7 PEEP animals than 4 PEEP animals. IL-1beta, IL-6, and IL-8 mRNAs were significantly elevated in animals ventilated with 0 PEEP compared with 4 PEEP. The percentage fractional area of collapsed alveoli was significantly higher for 0 PEEP compared with 4 and 7 PEEP groups. Mechanical ventilation increased the expression of proinflammatory mediators in surfactant-treated preterm lungs and the use of 4 PEEP minimized this response.

Macrophage and type II cell catabolism of SP-A and saturated phosphatidylcholine in mouse lungs.

Type II cells and macrophages are the major cells involved in the alveolar clearance and catabolism of surfactant. We measured type II cell and macrophage contributions to the catabolism of saturated phosphatidylcholine and surfactant protein A (SP-A) in mice. We used intratracheally administered SP-A labeled with residualizing (125)I-dilactitol-tyramine, radiolabeled dipalmitoylphosphatidylcholine ([(3)H]DPPC), and its degradation-resistant analog [(14)C]DPPC-ether. At 15 min and 7, 19, 29, and 48 h after intratracheal injection, the mice were killed; alveolar lavage was then performed to recover macrophages and surfactant. Type II cells and macrophages not recovered by the lavage were subsequently isolated by enzymatic digestion of the lung. Radioactivity was measured in total lung, lavage fluid macrophages, alveolar washes, type II cells, and lung digest macrophages. Approximately equal amounts of (125)I-dilactitol-tyramine-SP-A and [(14)C]DPPC-ether associated with the macrophages (lavage fluid plus lung digest) and type II cells when corrected for the efficiency of type II cell isolation. Eighty percent of the macrophage-associated radiolabel was recovered from lung digest macrophages. We conclude that macrophages and type II cells contribute equally to saturated phosphatidylcholine and SP-A catabolism in mice.

Lung morphometry after repetitive antenatal glucocorticoid treatment in preterm sheep.

Antenatal glucocorticoids are thought to be less effective when delivery occurs more than 7 d after initiation of treatment; therefore, repeat courses are often administered. We examined lung structure after single or repetitive antenatal glucocorticoid injections in fetal sheep. Pregnant ewes received single or repetitive doses of 0.5 mg/kg betamethasone at 7-d intervals by maternal or fetal injection, beginning at D104 or D114 with delivery at D125, D135, or D146 gestation (term = 150 d). Changes in lung structure were more pronounced after repetitive versus single injections. Repetitive fetal or maternal injections beginning at D104 (delivery at D125) resulted in comparable structural changes: alveolar volume increased by 50 to 80%, alveolar numerical density decreased by 30 to 40%, and pleural and interlobular septal volumes decreased by as much as 70%. Similar changes were seen in animals delivered at D135 after repetitive maternal injections beginning at D114. There were no structural differences between control and repetitive betamethasone animals when delivery was delayed until D146, indicating that betamethasone induced structural changes were reversible.

Effects of gestation and antenatal steroid on airway and tissue mechanics in newborn lambs.

The aim of this study was to partition airway and parenchymal mechanics in newborn lambs at different gestations and following variable exposure to antenatal maternal betamethasone using the forced oscillation technique (FOT). Pulmonary impedance data were collected in 37 sedated and intubated apneic lambs with the FOT between 0.5 and 20 Hz and fitted by a model to estimate airway resistance (Raw) and inertance (Iaw) and the coefficients of tissue resistance (GL) and elastance (HL). Total respiratory resistance (Rrs) was also determined during tidal ventilation by using the multiple linear regression technique. Advancing gestation or increasing antenatal steroid exposure had no clinically significant effect on the values of Raw and Iaw, whereas Rrs and both GL and HL decreased markedly. There was a decrease in tissue hysteresivity (GL/HL) with repeated antenatal steroid exposure. Partitioning of lung mechanics highlights the dominant contribution of the tissues to the total respiratory resistance in the immature ovine lung. Clinically relevant changes in lung mechanics associated with structural and functional maturation of the immature ovine lung are primarily confined to the tissue compartment.