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Little is known about the effects of fetal ethanol exposure on lung development. Our aim was to determine the effects of repeated ethanol exposure during late gestation on fetal lung growth, maturation, and inflammatory status. Pregnant ewes were chronically catheterized at 91 days of gestational age (DGA; term ~147 days). From 95-133 DGA, ewes were given a 1-h daily infusion of either 0.75 g ethanol/kg (n = 9) or saline (n = 8), with tissue collection at 134 DGA. Fetal lungs were examined for changes in tissue growth, structure, maturation, inflammation, and oxidative stress. Between treatment groups, there were no differences in lung weight, DNA and protein contents, percent proliferating and apoptotic cells, tissue and air-space fractions, alveolar number and mean linear intercept, septal thickness, type-II cell number and elastin content. Ethanol exposure caused a 75% increase in pulmonary collagen I [alpha]1 mRNA levels (P < 0.05) and a significant increase in collagen deposition. Surfactant protein (SP)-A and SP-B mRNA levels were approximately one third of control levels following ethanol exposure (P < 0.05). The mRNA levels of the proinflammatory cytokines interleukin (IL)-1[beta] and IL-8 were also lower (P < 0.05) in ethanol-exposed fetuses compared with controls. Pulmonary malondialdehyde levels tended to be increased (P = 0.07) in ethanol-exposed fetuses. Daily exposure of the fetus to ethanol during the last third of gestation alters extracellular matrix deposition and surfactant protein gene expression, which could increase the risk of respiratory distress syndrome after birth. Changes to the innate immune status of the fetus could increase the susceptibility of the neonatal lungs to infection. lung growth; lung morphometry; surfactant proteins; proinflammatory cytokines

Maternal alcohol consumption during pregnancy can affect fetal development, but little is known about the effects on the developing kidney. Our objectives were to determine the effects of repeated ethanol exposure during the latter half of gestation on glomerular (nephron) number and expression of key genes involved in renal development or function in the ovine fetal kidney. Pregnant ewes received daily intravenous infusion of ethanol (0.75 g/kg, n = 5) or saline (control, n = 5) over 1 h from 95 to 133 days of gestational age (DGA; term is ~147 DGA). Maternal and fetal arterial blood samples were taken before and after the start of the daily ethanol infusions for determination of blood ethanol concentration (BEC). Necropsy was performed at 134 DGA, and fetal kidneys were collected for determination of total glomerular number using the physical disector/fractionator technique; at this gestational age nephrogenesis is completed in sheep. Maximal maternal and fetal BECs of 0.12 [+ or -] 0.01 g/dl (mean [+ or -] SE) and 0.11 [+ or -] 0.01 g/dl, respectively, were reached 1 h after starting maternal ethanol infusions. Ethanol exposure had no effect on fetal body weight, kidney weight, or the gene expression of members of the renin-angiotensin system, insulin-like growth factors, and sodium channels. However, fetal glomerular number was lower after ethanol exposure (377,585 [+ or -] 8,325) than in controls (423,177 [+ or -] 17,178, P < 0.001). The data demonstrate that our regimen of fetal ethanol exposure during the latter half of gestation results in an 11% reduction in nephron endowment without affecting the overall growth of the kidney or fetus or the expression of key genes involved in renal development or function. A reduced nepliron endowment of this magnitude could have important implications for the cardiovascular health of offspring during postnatal life. nephron number; kidney; fetus; ethanol exposure; sheep

Experimentally, a typical ≈2 min cord clamp-to-ventilation interval in preterm lambs is accompanied by increased hemodynamic lability of the birth transition. However, whether this lability is related to development of asphyxia after cord clamping, or can be avoided with a shorter clamp-to-ventilation interval, is unknown. To address these questions, anesthetized preterm fetal lambs (gestation 127±2 days) were instrumented with ductus arteriosus and left pulmonary artery flow probes to obtain right ventricular (RV) output, brachiocephalic trunk and aortic isthmus flow probes to measure left ventricular (LV) output, and aortic trunk catheters for pressure measurement and blood gas analysis. With hemodynamics recorded continuously, fetuses were delivered onto the ewe's abdomen and the cord clamped for 1.5 min before ventilation (n=8), with aortic sampling at 15, 30, 45 & 60 sec, or for 0.5 min, with sampling at 15 sec (n=4). With 1.5 mins' cord clamping, an asphyxial state (Po2<10mmHg) was evident at ≥45 sec, with bradycardia and marked falls in LV and RV outputs (by 60% & 50%, P<0.001), followed after ventilation onset by tachycardia and LV and RV output surges (4- & 3-fold, P<0.001). By contrast, heart rate and outputs remained stable after 0.5 mins' cord clamping, with no post-ventilation change in heart rate or RV output, and a lesser rise in LV output (22%, P<0.005). In preterm lambs, rapid development of an asphyxial state within 45 sec in the cord clamp-to-ventilation interval increased hemodynamic lability of the birth transition, which was reduced with a shorter (≈0.5 min) cord clamp-to-ventilation interval. [ABSTRACT FROM AUTHOR]

Preterm neonates are born while nephrogenesis is ongoing and are commonly exposed to factors in the extrauterine environment that may impair renal development. Supplemental oxygen therapy exposes the preterm infant to a hyperoxic environment that may induce oxidative stress. Our aim was to determine the immediate and long-term effects of exposure to hyperoxia, during the period of postnatal nephrogenesis, on renal development. Newborn mice (C57BL/6J) were kept in a normoxic (room air, 21% oxygen) or a controlled hyperoxic (65% oxygen) environment from birth to postnatal day 7 (P7d). From P7d, animals were maintained in room air until early adulthood at postnatal day 56 (P56d) or middle age (10 mo; P10mo). Pups were assessed for glomerular maturity and renal corpuscle cross-sectional area at P7d (control n = 14; hyperoxic n = 14). Nephron number and renal corpuscle size were determined stereologically at P56d (control n = 14; hyperoxic n = 14) and P10mo (control n = 10; hyperoxic n = 10). At P7d, there was no effect of hyperoxia on glomerular size or maturity. In early adulthood (P56d), body weights, relative kidney weights and volumes, and nephron number were not different between groups, but the renal corpuscles were significantly enlarged. This was no longer evident at P10mo, with relative kidney weights and volumes, nephron number, and renal corpuscle size not different between groups. Furthermore, hyperoxia exposure did not significantly accelerate glomerulosclerosis in middle age. Hence, our findings show no overt long-term deleterious effects of early life hyperoxia on glomerular structure. [ABSTRACT FROM AUTHOR]

Gas flow is fundamental for driving tidal ventilation and, thus, the speed of lung motion, but current bias flow settings to support the preterm lung after birth do not have an evidence base. We aimed to determine the role of gas bias flow rates to generate positive pressure ventilation in initiating early lung injury pathways in the preterm lamb. Using slower speeds to inflate the lung during tidal ventilation (gas flow rates 4-6 L/min) did not affect lung mechanics, mechanical power, or gas exchange compared with those currently used in clinical practice (8-10 L/min). Speed of pressure and volume change during inflation were faster with higher flow rates. Lower flow rates resulted in less bronchoalveolar fluid protein, better lung morphology, and fewer detached epithelial cells. Overall, relative to unventilated fetal controls, there was greater protein change using 8-10 L/min, which was associated with enrichment of acute inflammatory and innate responses. Slowing the speed of lung motion by supporting the preterm lung from birth with lower flow rates than in current clinical use resulted in less lung injury without compromising tidal ventilation or gas exchange.

Background: The incidence of chronic lung disease is increasing, suggesting a need to explore novel ways to understand ventilator induced lung injury (VILI) in preterm infants. Mechanical power (MP) is a unifying measure of energy transferred to the respiratory system and a proposed determinant of VILI. The gold-standard method for calculating MP (geometric method) is not feasible in the clinical setting. This has prompted the derivation of simplified equations for calculating MP., Objective: To validate the agreement between a simplified calculation of MP (MP Simple ) and the true MP calculated using the geometric method (MP Ref )., Methods: MP Simple and MP Ref was calculated in mechanically ventilated preterm lambs (n = 71) and the agreement between both measures was determined using intraclass correlation coefficients (ICC), linear regression, and Bland-Altman analysis., Results: A strong linear relationship (adjusted R 2  = 0.98), and excellent agreement (ICC = 0.99, 95% CI = 0.98-0.99) between MP Simple and MP Ref was demonstrated. Bland-Altman analysis demonstrated a negligible positive bias (mean difference = 0.131 J/min·kg). The 95% limits of agreement were -0.06 to 0.32 J/min·kg., Conclusions: In a controlled setting, there was excellent agreement between MP Simple and gold-standard calculations. MP Simple should be validated and explored in preterm neonates to assess the cause-effect relationship with VILI and neonatal outcomes., Impact Statement: Mechanical power (MP) unifies the individual components of ventilator induced lung injury (VILI) and provides an estimate of total energy transferred to the respiratory system during mechanical ventilation. As gold-standard calculations of mechanical power at the bedside are not feasible, alternative simplified equations have been proposed. In this study, MP calculated using a simplified equation had excellent agreement with true MP in mechanically ventilated preterm lambs. These results lay foundations to explore the role of MP in neonatal VILI and determine its relationship with short and long term respiratory outcomes., (© 2024. Crown.)

Tidal ventilation is essential in supporting the transition to air-breathing at birth, but excessive tidal volume (V T ) is an important factor in preterm lung injury. Few studies have assessed the impact of specific V T levels on injury development. Here, we used a lamb model of preterm birth to investigate the role of different levels of V T during positive pressure ventilation (PPV) in promoting aeration and initiating early lung injury pathways. V T was delivered as 1 ) 7 mL/kg throughout (V Tstatic ), 2 ) begun at 3 mL/kg and increased to a final V T of 7 mL/kg over 3 min (V Tinc ), or 3 ) commenced at 7 mL/kg, decreased to 3 mL/kg, and then returned to 7 mL/kg (V Talt ). V T , inflating pressure, lung compliance, and aeration were similar in all groups from 4 min, as was postmortem histology and lung lavage protein concentration. However, transient decrease in V T in the V Talt group caused increased ventilation heterogeneity. Following TMT-based quantitative mass spectrometry proteomics, 1,610 proteins were identified in the lung. Threefold more proteins were significantly altered with V Talt compared with V Tstatic or V Tinc strategies. Gene set enrichment analysis identified V Talt specific enrichment of immune and angiogenesis pathways and V Tstatic enrichment of metabolic processes. Our finding of comparable lung physiology and volutrauma across V T groups challenges the paradigm that there is a need to rapidly aerate the preterm lung at birth. Increased lung injury and ventilation heterogeneity were identified when initial V T was suddenly decreased during respiratory support at birth, further supporting the benefit of a gentle V T approach. NEW & NOTEWORTHY There is little evidence to guide the best tidal volume (V T ) strategy at birth. In this study, comparable aeration, lung mechanics, and lung morphology were observed using static, incremental, and alternating V T strategies. However, transient reduction in V T was associated with ventilation heterogeneity and inflammation. Our results suggest that rapidly aerating the preterm lung may not be as clinically critical as previously thought, providing clinicians with reassurance that gently supporting the preterm lung maybe permissible at birth.

The glucocorticosteroid betamethasone is routinely administered via maternal intramuscular injection to enhance fetal lung maturation before anticipated preterm birth. Although antenatal betamethasone increases fetal pulmonary arterial (PA) blood flow, whether this agent alters the contribution of 1 ) right ventricular (RV) output or 2 ) left-to-right shunting across the ductus arteriosus to rises in PA blood flow after preterm birth is unknown. To address this question, anesthetized control ( n = 7) and betamethasone-treated ( n = 7) preterm fetal lambs (gestation 127 ± 1 days, means ± SD) were instrumented with aortic, pulmonary, and left atrial catheters as well as ductus arteriosus and left PA flow probes to calculate RV output, with hemodynamics measured for 30 min after cord clamping and mechanical ventilation. Mean PA blood flow was higher in betamethasone-treated than in control lambs over the initial 10 min after birth ( P < 0.05). This higher PA flow was accompanied by 1) a greater pulmonary vascular conductance ( P ≤ 0.025), 2) a larger proportion of RV output passing to lungs ( P ≤ 0.01), despite a fall in this output, and 3) earlier reversal and a greater magnitude ( P ≤ 0.025) of net ductal shunting, due to the combination of higher left-to-right ( P ≤ 0.025) and lesser right-to-left phasic shunting ( P ≤ 0.025). These results suggest that antenatal betamethasone augments the initial rise in PA blood flow after birth in preterm lambs, with this augmented rise supported by the combination of 1 ) a greater redistribution of RV output toward the lungs and 2 ) a faster and larger reversal in net ductal shunting underpinned not only by greater left-to-right, but also by lesser right-to-left phasic shunting.

This study tested the hypothesis that varying degrees of hemodynamic fluctuations seen after birth following immediate cord clamping were related to development of asphyxia with longer cord clamp-to-ventilation intervals, resulting in higher perinatal circulating levels of the catecholamines norepinephrine (NE) and epinephrine (Epi), and thus increased heart rate, blood pressures, and cardiac contractility after birth. Anesthetized preterm fetal lambs were instrumented with 1 ) aortic (AoT) and pulmonary trunk (PT) micromanometers to obtain pressures and the maximal rate of pressure rise (dP/d t max ) as a surrogate measure of ventricular contractility, and 2 ) an AoT catheter to obtain samples for blood gas and catecholamine analyses. After delivery, immediate cord clamping was followed by ventilation ∼40 s ( n = 7), ∼60 s ( n = 8), ∼90 s ( n = 9), or ∼120 s later ( n = 8), with frequent blood sampling performed before and after ventilation. AoT O 2 content fell rapidly after immediate cord clamping ( P < 0.001), with an asphyxial state evident at ≥60 s. Plasma NE and Epi levels increased progressively with longer cord clamp-to-ventilation intervals, with an exponential relation between falling AoT O 2 content and rising catecholamines ( R 2  = 0.64-0.67). Elevated circulating catecholamines persisted for some minutes after ventilation onset, with postbirth surges in heart rate, AoT and PT pressures, and AoT and PT dP/d t max linearly related to log e of catecholamine levels ( R 2  = 0.41-0.54, all P < 0.001). These findings suggest that 1 ) a greater degree of asphyxia-induced sympathoadrenal activation (reflected in elevated circulating catecholamine levels) occurs with longer intervals between immediate cord clamping and subsequent ventilation, and 2 ) this activation is a major determinant of hemodynamic fluctuations evident with birth., (Copyright © 2017 the American Physiological Society.)

High levels of alcohol (ethanol) exposure during fetal life can affect liver development and can increase susceptibility to infection after birth. Our aim was to determine the effects of a moderate level of ethanol exposure in late gestation on the morphology, iron status, and inflammatory status of the ovine fetal liver. Pregnant ewes were chronically catheterized at 91 days of gestation (DG; term ~145 DG) for daily intravenous infusion of ethanol (0.75 g/kg maternal body wt; n = 8) or saline (n = 7) over 1 h from 95 to 133 DG. At necropsy (134 DG), fetal livers were collected for analysis. Liver weight, general liver morphology, hepatic cell proliferation and apoptosis, perivascular collagen deposition, and interleukin (IL)-1β, IL-6, or IL-8 mRNA levels were not different between groups. However, ethanol exposure led to significant decreases in hepatic content of ferric iron and gene expression of the iron-regulating hormone hepcidin and tumor necrosis factor (TNF)-α (all P < 0.05). In the placenta, there was no difference in transferrin receptor, divalent metal transporter 1, and ferritin mRNA levels; however, ferroportin mRNA levels were increased in ethanol-exposed animals (P < 0.05), and ferroportin protein tended to be increased (P = 0.054). Plasma iron concentration was not different between control and ethanol-exposed groups; control fetuses had significantly higher iron concentrations than their mothers, whereas maternal and fetal iron concentrations were similar in ethanol-exposed animals. We conclude that daily ethanol exposure during the third-trimester-equivalent in sheep does not alter fetal liver morphology; however, decreased fetal liver ferric iron content and altered hepcidin and ferroportin gene expression indicate that iron homeostasis is altered.

High levels of ethanol (EtOH) consumption during pregnancy adversely affect fetal development; however, the effects of lower levels of exposure are less clear. Our objectives were to assess the effects of daily EtOH exposure (3.8 USA standard drinks) on fetal-maternal physiological variables and the fetal brain, particularly white matter. Pregnant ewes received daily intravenous infusions of EtOH (0.75 g/kg maternal body wt over 1 h, 8 fetuses) or saline (8 fetuses) from 95 to 133 days of gestational age (DGA; term ∼145 DGA). Maternal and fetal arterial blood was sampled at 131-133 DGA. At necropsy (134 DGA) fetal brains were collected for analysis. Maternal and fetal plasma EtOH concentrations reached similar maximal concentration (∼0.11 g/dl) and declined at the same rate. EtOH infusions produced mild reductions in fetal arterial oxygenation but there were no changes in maternal oxygenation, maternal and fetal Pa(CO(2)), or in fetal mean arterial pressure or heart rate. Following EtOH infusions, plasma lactate levels were elevated in ewes and fetuses, but arterial pH fell only in ewes. Fetal body and brain weights were similar between groups. In three of eight EtOH-exposed fetuses there were small subarachnoid hemorrhages in the cerebrum and cerebellum associated with focal cortical neuronal death and gliosis. Overall, there was no evidence of cystic lesions, inflammation, increased apoptosis, or white matter injury. We conclude that daily EtOH exposure during the third trimester-equivalent of ovine pregnancy has modest physiological effects on the fetus and no gross effects on fetal white matter development.

Prenatal exposure to high levels of ethanol is associated with cardiac malformations, but the effects of lower levels of exposure on the heart are unclear. Our aim was to investigate the effects of daily exposure to ethanol during late gestation, when cardiomyocytes are undergoing maturation, on the developing myocardium. Pregnant ewes were infused with either ethanol (0.75 g/kg) or saline for 1 h each day from gestational days 95 to 133 (term ∼145 days); tissues were collected at 134 days. In sheep, cardiomyocytes mature during late gestation as in humans. Within the left ventricle (LV), cardiomyocyte number was determined using unbiased stereology and cardiomyocyte size and nuclearity determined using confocal microscopy. Collagen deposition was quantified using image analysis. Genes relating to cardiomyocyte proliferation and apoptosis were examined using quantitative real-time PCR. Fetal plasma ethanol concentration reached 0.11 g/dL after EtOH infusions. Ethanol exposure induced significant increases in relative heart weight, relative LV wall volume, and cardiomyocyte cross-sectional area. Ethanol exposure advanced LV maturation in that the proportion of binucleated cardiomyocytes increased by 12%, and the number of mononucleated cardiomyocytes was decreased by a similar amount. Apoptotic gene expression increased in the ethanol-exposed hearts, although there were no significant differences between groups in total cardiomyocyte number or interstitial collagen. Daily exposure to a moderate dose of ethanol in late gestation accelerates the maturation of cardiomyocytes and increases cardiomyocyte and LV tissue volume in the fetal heart. These effects on cardiomyocyte growth may program for long-term cardiac vulnerability.

Prenatal ethanol exposure increases collagen deposition and alters surfactant protein (SP) expression and immune status in lungs of near-term fetal sheep. Our objectives were to determine 1) whether these prenatal effects of repeated gestational ethanol exposure persist after birth and 2) whether surfactant phospholipid composition is altered following prenatal ethanol exposure. Pregnant ewes were chronically catheterized at 90 days of gestational age (DGA) and given a 1-h daily infusion of ethanol (0.75 g/kg, n = 9) or saline (n = 7) from 95 to 135 DGA; ethanol administration ceased after 135 DGA. Lambs were born naturally at full term (146 ± 0.5 DGA). Lung tissue was examined at 9 wk postnatal age for alterations in structure, SP expression, and inflammation; bronchoalveolar lavage fluid was examined for alterations in surfactant phospholipid composition. At 134 DGA, surfactant phospholipid concentration in amniotic fluid was significantly reduced (P < 0.05) by ethanol exposure, and the composition was altered. In postnatal lambs, there were no significant differences between treatment groups in birth weight, postnatal growth, blood gas parameters, and lung weight, volume, tissue fraction, mean linear intercept, collagen content, proinflammatory cytokine gene expression, and bronchoalveolar lavage fluid surfactant phospholipid composition. Although SP-A, SP-B, and SP-C mRNA levels were not significantly different between treatment groups, SP-D mRNA levels were significantly greater (P < 0.05) in ethanol-treated animals; as SP-D has immunomodulatory roles, innate immunity may be altered. The adverse effects of daily ethanol exposure during late gestation on the fetal lung do not persist to 2 mo after birth, indicating that the developing lung is capable of repair.