Serum ferritin values in neonates <29 weeks' gestation are highly variable and do not correlate with reticulocyte hemoglobin content.

OBJECTIVES: To compare serum ferritin and RET-He values among extremely low gestational age neonates ELGANs with other markers of iron-deficient erythropoiesis. STUDY DESIGN: This is a secondary analysis of the NICHD Darbepoetin Trial. Study data from placebo recipients who had a serum ferritin, a RET-He, and a mean corpuscular volume (MCV) measurement within a 24-hour period were analyzed for correlation. RESULTS: Mixed linear regression models showed no association between ferritin and RET-He at both early (ß = 0.0016, p = 0.40) and late (ß = -0.0001, p = 0.96) time points. Positive associations were observed between RET-He and MCV at baseline, early, and late time points (p < 0.01, =0.01, <0.001, respectively), while ferritin was not associated with MCV at any time point. CONCLUSIONS: Our study shows that RET-He is better correlated with MCV as a marker of iron-limited erythropoiesis than ferritin. The results suggest that ferritin is limited as a marker of iron sufficiency in premature infants. STUDY IDENTIFICATION: FDA IND Number 100138; ClinicalTrials.gov number NCT03169881; NRN ID number NICHD-NRN-0058 (Darbe).

Weaning of Moderately Preterm Infants from the Incubator to the Crib: A Randomized Clinical Trial.

OBJECTIVE: To assess whether length of hospital stay is decreased among moderately preterm infants weaned from incubator to crib at a lower vs higher weight. STUDY DESIGN: This trial was conducted in the Eunice Kennedy Shriver National Institute of Child Health and Human Development Neonatal Research Network. Infants with gestational ages 29-33 weeks, birthweight <1600 g, and in an incubator were randomly assigned to a weaning weight of 1600 or 1800 g. Within 60 to 100 g of weaning weight, the incubator temperature was decreased by 1.0°C to 1.5°C every 24 hours until 28.0°C. The infants were weaned to the crib following stable temperature at 36.5°C to 37.4°C for 8 to 12 hours. Clothing and bedcoverings were standardized. The primary outcome was length of hospital stay from birth to discharge; secondary outcomes included length of stay and growth velocity from weaning to discharge. Adverse events were monitored. RESULTS: Of 1565 infants screened, 885 were eligible, and 366 enrolled-187 to the 1600-g and 179 to the 1800-g group. Maternal and neonatal characteristics did not differ among weight groups. Length of hospital stay was a median of 43 days in the lower and 41 days in the higher weight group (P = .12). Growth velocity from completion of weaning to discharge was higher in the lower weight group, 13.7 g/kg/day vs 12.8 g/kg/day (P = .005). Groups did not differ in adverse events. CONCLUSIONS: Among moderately preterm neonates, weaning from incubator to crib at a lower weight did not decrease length of stay, but was safe and was accompanied by higher weight gain after weaning. TRIAL REGISTRATION: ClinicalTrials.gov: NCT02160002.

Effects of Myo-inositol on Type 1 Retinopathy of Prematurity Among Preterm Infants <28 Weeks' Gestational Age: A Randomized Clinical Trial.

Importance: Previous studies of myo-inositol in preterm infants with respiratory distress found reduced severity of retinopathy of prematurity (ROP) and less frequent ROP, death, and intraventricular hemorrhage. However, no large trials have tested its efficacy or safety. Objective: To test the adverse events and efficacy of myo-inositol to reduce type 1 ROP among infants younger than 28 weeks' gestational age. Design, Setting, and Participants: Randomized clinical trial included 638 infants younger than 28 weeks' gestational age enrolled from 18 neonatal intensive care centers throughout the United States from April 17, 2014, to September 4, 2015; final date of follow-up was February 12, 2016. The planned enrollment of 1760 participants would permit detection of an absolute reduction in death or type 1 ROP of 7% with 90% power. The trial was terminated early due to a statistically significantly higher mortality rate in the myo-inositol group. Interventions: A 40-mg/kg dose of myo-inositol was given every 12 hours (initially intravenously, then enterally when feeding; n = 317) or placebo (n = 321) for up to 10 weeks. Main Outcomes and Measures: Type 1 ROP or death before determination of ROP outcome was designated as unfavorable. The designated favorable outcome was survival without type 1 ROP. Results: Among 638 infants (mean, 26 weeks' gestational age; 50% male), 632 (99%) received the trial drug or placebo and 589 (92%) had a study outcome. Death or type 1 ROP occurred more often in the myo-inositol group vs the placebo group (29% vs 21%, respectively; adjusted risk difference, 7% [95% CI, 0%-13%]; adjusted relative risk, 1.41 [95% CI, 1.08-1.83], P = .01). All-cause death before 55 weeks' postmenstrual age occurred in 18% of the myo-inositol group and in 11% of the placebo group (adjusted risk difference, 6% [95% CI, 0%-11%]; adjusted relative risk, 1.66 [95% CI, 1.14-2.43], P = .007). The most common serious adverse events up to 7 days of receiving the ending dose were necrotizing enterocolitis (6% for myo-inositol vs 4% for placebo), poor perfusion or hypotension (7% vs 4%, respectively), intraventricular hemorrhage (10% vs 9%), systemic infection (16% vs 11%), and respiratory distress (15% vs 13%). Conclusions and Relevance: Among premature infants younger than 28 weeks' gestational age, treatment with myo-inositol for up to 10 weeks did not reduce the risk of type 1 ROP or death vs placebo. These findings do not support the use of myo-inositol among premature infants; however, the early termination of the trial limits definitive conclusions.

Prolonged respiratory support of any type impacts outcomes of extremely low birth weight infants.

OBJECTIVES: This study tested the hypothesis that longer duration of any type of respiratory support is associated with an increased rate of death or neurodevelopmental impairment (NDI) at 18-22 months. METHODS: Retrospective cohort study using the Generic Database of NICHD Neonatal Research Network from 2006 to 2010. Infants were born at <27 weeks gestational age with birth weights of 401-1000 g. Respiratory support received during initial hospitalization from birth was characterized as follows: no support, only invasive support, only non-invasive support or mixed invasive, and non-invasive support. The primary outcome was death after 24 h of life or NDI at 18-22 months corrected age. RESULTS: In a cohort of 3651 infants, 1494 (40.9%) died or had NDI. Cumulative respiratory support of any type beyond 60 days was associated with the likelihood of death or NDI. Infants who only received invasive support had the highest rate (89.1%), followed by those received mixed support (26.1%). Infants who received only non-invasive support had the lowest rate (7.7%). When compared to the only non-invasive support group, both invasive [OR 62.7 (95%CI 25.7, 152.6)] and mixed [OR 6.1 (95%CI 2.6, 14.4)] support groups were significantly more likely to die or have NDI. CONCLUSION: Prolonged respiratory support, whether invasive or non-invasive, is associated with increased odds of a poor outcome. The proportion of infants with a poor outcome increased in a dose dependent manner by two factors: the cumulative duration of respiratory support beyond 60 days, and the extent to which invasive support is provided.

Effect of Therapeutic Hypothermia Initiated After 6 Hours of Age on Death or Disability Among Newborns With Hypoxic-Ischemic Encephalopathy: A Randomized Clinical Trial.

Importance: Hypothermia initiated at less than 6 hours after birth reduces death or disability for infants with hypoxic-ischemic encephalopathy at 36 weeks' or later gestation. To our knowledge, hypothermia trials have not been performed in infants presenting after 6 hours. Objective: To estimate the probability that hypothermia initiated at 6 to 24 hours after birth reduces the risk of death or disability at 18 months among infants with hypoxic-ischemic encephalopathy. Design, Setting, and Participants: A randomized clinical trial was conducted between April 2008 and June 2016 among infants at 36 weeks' or later gestation with moderate or severe hypoxic-ischemic encephalopathy enrolled at 6 to 24 hours after birth. Twenty-one US Neonatal Research Network centers participated. Bayesian analyses were prespecified given the anticipated limited sample size. Interventions: Targeted esophageal temperature was used in 168 infants. Eighty-three hypothermic infants were maintained at 33.5°C (acceptable range, 33°C-34°C) for 96 hours and then rewarmed. Eighty-five noncooled infants were maintained at 37.0°C (acceptable range, 36.5°C-37.3°C). Main Outcomes and Measures: The composite of death or disability (moderate or severe) at 18 to 22 months adjusted for level of encephalopathy and age at randomization. Results: Hypothermic and noncooled infants were term (mean [SD], 39 [2] and 39 [1] weeks' gestation, respectively), and 47 of 83 (57%) and 55 of 85 (65%) were male, respectively. Both groups were acidemic at birth, predominantly transferred to the treating center with moderate encephalopathy, and were randomized at a mean (SD) of 16 (5) and 15 (5) hours for hypothermic and noncooled groups, respectively. The primary outcome occurred in 19 of 78 hypothermic infants (24.4%) and 22 of 79 noncooled infants (27.9%) (absolute difference, 3.5%; 95% CI, -1% to 17%). Bayesian analysis using a neutral prior indicated a 76% posterior probability of reduced death or disability with hypothermia relative to the noncooled group (adjusted posterior risk ratio, 0.86; 95% credible interval, 0.58-1.29). The probability that death or disability in cooled infants was at least 1%, 2%, or 3% less than noncooled infants was 71%, 64%, and 56%, respectively. Conclusions and Relevance: Among term infants with hypoxic-ischemic encephalopathy, hypothermia initiated at 6 to 24 hours after birth compared with noncooling resulted in a 76% probability of any reduction in death or disability, and a 64% probability of at least 2% less death or disability at 18 to 22 months. Hypothermia initiated at 6 to 24 hours after birth may have benefit but there is uncertainty in its effectiveness. Trial Registration: clinicaltrials.gov Identifier: NCT00614744.

Early Caffeine Prophylaxis and Risk of Failure of Initial Continuous Positive Airway Pressure in Very Low Birth Weight Infants.

OBJECTIVE: To test the hypothesis that early caffeine treatment on the day of birth, compared with later treatment in very low birth weight (VLBW, <1500 g) infants receiving continuous positive airway pressure (CPAP) therapy, is associated with a decreased risk of CPAP failure in the first week of life. STUDY DESIGN: Multicenter, observational cohort study in 366 US neonatal intensive care units. We evaluated inborn, VLBW infants discharged from 2000 to 2014, who received only CPAP therapy without surfactant treatment on day of life (DOL) 0, had a 5-minute Apgar ≥3, and received caffeine in the first week of life. We used multivariable conditional logistic regression to compare the risk of CPAP failure, defined as invasive mechanical ventilation or surfactant therapy on DOL 1-6, by timing of caffeine treatment as either early (initiation on DOL 0) or routine (initiation on DOL 1-6). RESULTS: We identified 11 133 infants; 4528 (41%) received early caffeine and 6605 (59%) received routine caffeine. Median gestational age was lower in the early caffeine group, 29 weeks (25th, 75th percentiles; 28, 30) vs the routine caffeine group, 30 weeks (29, 31); P < 0.001. The incidence of CPAP failure on DOL 1-6 was similar between the early and routine caffeine groups: 22% vs 21%; adjusted OR = 1.05 (95% CI: 0.93, 1.18). CONCLUSIONS: Early caffeine treatment on the day of birth was not associated with a decreased risk of CPAP failure in the first week of life for VLBW infants initially treated with CPAP.

Early characteristics of infants with pulmonary hypertension in a referral neonatal intensive care unit.

BACKGROUND: Approximately 8-23% of premature infants develop pulmonary hypertension (PH), and this diagnosis confers a higher possibility of mortality. As a result, professional societies recommend PH screening in premature infants. However, the risk factors for and the outcomes of PH may differ depending on the timing of its diagnosis, and little evidence is available to determine at-risk infants in the referral neonatal population. ï»¿The objective of this study was to define clinical and echocardiographic characteristics of infants with pulmonary hypertension during the neonatal hospital course and at or near-term. METHODS: Infants who had the following billing codes: < 32 weeks, birth weight < 1500 g, neonatal unit, and echocardiograph had records abstracted from a data warehouse at Children's Healthcare of Atlanta. The outcome was defined as late PH on the final echocardiogram for all patients, and, separately, for patients with multiple studies. Descriptive statistics, univariable, and multivariable models were evaluated, and odds ratios and 95% confidence intervals are expressed below as (OR, CI). RESULTS: 556 infants were included in the overall study, 59 had PH on their final echocardiogram (11%). In multivariable analyses, atrial septal defect (2.9, 1.4-6.1), and intrauterine growth restriction (2.7, 1.2-6.3) increased the odds of late PH, whereas caffeine therapy decreased PH (0.4, 0.2-0.8). When the analyses were restricted to 32 infants who had multiple echocardiograms during their hospitalization, the association between atrial septal defect (5.9, 2.0-16.5) and growth restriction (3.7, 1.3-10.7) and late PH was strengthened, but the effect of caffeine therapy was no longer significant. In this smaller subgroup, infants with late PH had their final echocardiogram at a median of 116 days of life, and 42-74% of them had right ventricular pathology. CONCLUSIONS: Early clinical variables are associated with PH persistence in a referral neonatal population. Identification of early clinical factors may help guide the ascertainment of infant risk for late PH, and may aid in targeting sub-groups that are most likely to benefit from PH screening.

Safety and pharmacokinetics of multiple dose myo-inositol in preterm infants.

BACKGROUND: Preterm infants with respiratory distress syndrome (RDS) given inositol had reduced bronchopulmonary dysplasia (BPD), death and severe retinopathy of prematurity (ROP). We assessed the safety and pharmacokinetics of daily inositol to select a dose providing serum levels previously associated with benefit, and to learn if accumulation occurred when administered throughout the normal period of retinal vascularization. METHODS: Infants ≤ 29 wk GA (n = 122, 14 centers) were randomized and treated with placebo or inositol at 10, 40, or 80 mg/kg/d. Intravenous administration converted to enteral when feedings were established, and continued to the first of 10 wk, 34 wk postmenstrual age (PMA) or discharge. Serum collection employed a sparse sampling population pharmacokinetics design. Inositol urine losses and feeding intakes were measured. Safety was prospectively monitored. RESULTS: At 80 mg/kg/d mean serum levels reached 140 mg/l, similar to Hallman's findings. Levels declined after 2 wk, converging in all groups by 6 wk. Analyses showed a mean volume of distribution 0.657 l/kg, clearance 0.058 l/kg/h, and half-life 7.90 h. Adverse events and comorbidities were fewer in the inositol groups, but not significantly so. CONCLUSION: Multiple dose inositol at 80 mg/kg/d was not associated with increased adverse events, achieves previously effective serum levels, and is appropriate for investigation in a phase III trial.

Hyperoxia induces alveolar epithelial-to-mesenchymal cell transition.

Myofibroblast accumulation is a pathological feature of lung diseases requiring oxygen therapy. One possible source for myofibroblasts is through the epithelial-to-mesenchymal transition (EMT) of alveolar epithelial cells (AEC). To study the effects of oxygen on alveolar EMT, we used RLE-6TN and ex vivo lung slices and found that hyperoxia (85% O2, H85) decreased epithelial proteins, presurfactant protein B (pre-SpB), pro-SpC, and lamellar protein by 50% and increased myofibroblast proteins, α-smooth muscle actin (α-SMA), and vimentin by over 200% (P < 0.05). In AEC freshly isolated from H85-treated rats, mRNA for pre-SpB and pro-SpC was diminished by ∼50% and α-SMA was increased by 100% (P < 0.05). Additionally, H85 increased H2O2 content, and H2O2 (25-50 µM) activated endogenous transforming growth factor-ß1 (TGF-ß1), as evident by H2DCFDA immunofluorescence and ELISA (P < 0.05). Both hyperoxia and H2O2 increased SMAD3 phosphorylation (260% of control, P < 0.05). Treating cultured cells with TGF-ß1 inhibitors did not prevent H85-induced H2O2 production but did prevent H85-mediated α-SMA increases and E-cadherin downregulation. Finally, to determine the role of TGF-ß1 in hyperoxia-induced EMT in vivo, we evaluated AEC from H85-treated rats and found that vimentin increased ∼10-fold (P < 0.05) and that this effect was prevented by intraperitoneal TGF-ß1 inhibitor SB-431542. Additionally, SB-431542 treatment attenuated changes in alveolar histology caused by hyperoxia. Our studies indicate that hyperoxia promotes alveolar EMT through a mechanism that is dependent on activation of TGF-ß1 signaling.

Serum tocopherol levels in very preterm infants after a single dose of vitamin E at birth.

OBJECTIVE: Our aim was to examine the impact of a single enteral dose of vitamin E on serum tocopherol levels. The study was undertaken to see whether a single dose of vitamin E soon after birth can rapidly increase the low α-tocopherol levels seen in very preterm infants. If so, this intervention could be tested as a means of reducing the risk of intracranial hemorrhage. METHODS: Ninety-three infants <27 weeks' gestation and <1000 g were randomly assigned to receive a single dose of vitamin E or placebo by gastric tube within 4 hours of birth. The vitamin E group received 50 IU/kg of vitamin E as dl-α-tocopheryl acetate (Aquasol E). The placebo group received sterile water. Blood samples were taken for measurement of serum tocopherol levels by high-performance liquid chromatography before dosing and 24 hours and 7 days after dosing. RESULTS: Eighty-eight infants received the study drug and were included in the analyses. The α-tocopherol levels were similar between the groups at baseline but higher in the vitamin E group at 24 hours (median 0.63 mg/dL vs. 0.42 mg/dL, P = .003) and 7 days (2.21 mg/dL vs 1.86 mg/dL, P = .04). There were no differences between groups in γ-tocopherol levels. At 24 hours, 30% of vitamin E infants and 62% of placebo infants had α-tocopherol levels <0.5 mg/dL. CONCLUSIONS: A 50-IU/kg dose of vitamin E raised serum α-tocopherol levels, but to consistently achieve α-tocopherol levels >0.5 mg/dL, a higher dose or several doses of vitamin E may be needed.

Chronic lung disease in preterm lambs: effect of daily vitamin A treatment on alveolarization.

Neonatal chronic lung disease is characterized by failed formation of alveoli and capillaries, and excessive deposition of matrix elastin, which are linked to lengthy mechanical ventilation (MV) with O(2)-rich gas. Vitamin A supplementation has improved respiratory outcome of premature infants, but there is little information about the structural and molecular manifestations in the lung that occur with vitamin A treatment. We hypothesized that vitamin A supplementation during prolonged MV, without confounding by antenatal steroid treatment, would improve alveolar secondary septation, decrease thickness of the mesenchymal tissue cores between distal air space walls, and increase alveolar capillary growth. We further hypothesized that these structural advancements would be associated with modulated expression of tropoelastin and deposition of matrix elastin, phosphorylated Smad2 (pSmad2), cleaved caspase 3, proliferating cell nuclear antigen (PCNA), VEGF, VEGF-R2, and midkine in the parenchyma of the immature lung. Eight preterm lambs (125 days' gestation, term approximately 150 days) were managed by MV for 3 wk: four were treated with daily intramuscular Aquasol A (vitamin A), 5,000 IU/kg, starting at birth; four received vehicle alone. Postmortem lung assays included quantitative RT-PCR and in situ hybridization, immunoblot and immunohistochemistry, and morphometry and stereology. Daily vitamin A supplementation increased alveolar secondary septation, decreased thickness of the mesenchymal tissue cores between the distal air space walls, and increased alveolar capillary growth. Associated molecular changes were less tropoelastin mRNA expression, matrix elastin deposition, pSmad2, and PCNA protein localization in the mesenchymal tissue core of the distal air space walls. On the other hand, mRNA expression and protein abundance of VEGF, VEGF-R2, midkine, and cleaved caspase 3 were increased. We conclude that vitamin A treatment partially improves lung development in chronically ventilated preterm neonates by modulating expression of tropoelastin, deposition of elastin, and expression of vascular growth factors.

Antenatal betamethasone depresses maternal and fetal aldosterone levels.

Antenatal glucocorticoids are used to mature lung function in fetuses at risk for preterm delivery, but they also suppress cortisol synthesis in both pregnant women and their fetuses. We recently discovered in pregnant rabbits that even though exogenous betamethasone is not a mineralocorticoid, it also suppresses production of aldosterone. Lower aldosterone levels were linked to reduced P450 side chain cleavage(P450scc) messenger RNA levels in the rabbit maternal and fetal adrenal cortex. To establish whether this occurs in humans, we assayed aldosterone levels in women and newborns treated with antenatal betamethasone for preterm labor. In mothers treated with betamethasone, maternal cortisol depression after 48 hours was accompanied by aldosterone depression. Both pregnant women and their newborns treated with betamethasone showed depressed aldosterone levels in a 1- to 3-day period after the first betamethasone dose. We conclude that suppression of aldosterone biosynthesis is a side effect of antenatal steroids that has been largely overlooked, but may be clinically relevant at a time when the newborn is learning to control plasma electrolytes and blood volume.

Permissive hypercapnia and risk for brain injury and developmental impairment.

OBJECTIVE: Permissive hypercapnia is a respiratory-care strategy that is used to reduce the risk for lung injury. The goal of this study was to evaluate whether permissive hypercapnia is associated with higher risk for intraventricular hemorrhage and early childhood behavioral and functional problems than normocapnia among very low birth weight infants. METHODS: Very low birth weight infants from a statewide cohort were eligible for this study when they were born at <32 weeks' gestational age and survived at least 24 hours. Infants were classified as receiving a permissive hypercapnia, normocapnia, or unclassifiable respiratory strategy during the first 24 hours after birth according to an algorithm based on Pco(2) values and respiratory-treatment decisions that were abstracted from medical charts. Intraventricular hemorrhage diagnosis was also abstracted from the medical chart. Behavioral and functional outcomes were assessed by parent interview at 2 to 3 years. Logistic regression was used to evaluate the relationship between intraventricular hemorrhage and respiratory strategy; ordinary linear regression was used to evaluate differences in behavior and function scores between children by respiratory strategy. RESULTS: Infants who received a permissive hypercapnia strategy were not more likely to have intraventricular hemorrhage than those with normocapnia. There were no differences in any of the behavioral or functional scores among children according to respiratory strategy. There was a significant interaction between care strategy and 1-minute Apgar score, indicating that infants with lower Apgar scores may be at higher risk for intraventricular hemorrhage with permissive hypercapnia. CONCLUSIONS: This study suggests that permissive hypercapnia does not increase risk for brain injury and impairment among very low birth weight children. The interaction between respiratory strategy and Apgar score is a potential worrisome exception to this conclusion. Future research should further evaluate the effect of elevated Pco(2) levels among those who are sickest at birth.

Blunted hypoxic pulmonary vasoconstriction in experimental neonatal chronic lung disease.

RATIONALE: Neonatal chronic lung disease (CLD), caused by prolonged mechanical ventilation (MV) with O(2)-rich gas, is the most common cause of long-term hospitalization and recurrent respiratory illness in extremely premature infants. Recurrent episodes of hypoxemia and associated ventilator adjustments often lead to worsening CLD. The mechanism that causes these hypoxemic episodes is unknown. Hypoxic pulmonary vasoconstriction (HPV), which is partially controlled by O(2)-sensitive voltage-gated potassium (K(v)) channels, is an important adaptive response to local hypoxia that helps to match perfusion and ventilation in the lung. OBJECTIVES: To test the hypothesis that chronic lung injury (CLI) impairs HPV. METHODS: We studied preterm lambs that had MV with O(2)-rich gas for 3 weeks and newborn rats that breathed 95%-O(2) for 2 weeks, both of which resulted in airspace enlargement and pulmonary vascular changes consistent with CLD. MEASUREMENTS AND MAIN RESULTS: HPV was attenuated in preterm lambs with CLI after 2 weeks of MV and in newborn rats with CLI after 2 weeks of hyperoxia. HPV and constriction to the K(v)1.x-specific inhibitor, correolide, were preferentially blunted in excised distal pulmonary arteries (dPAs) from hyperoxic rats, whose dPAs exhibited decreased K(v)1.5 and K(v)2.1 mRNA and K(+) current. Intrapulmonary gene transfer of K(v)1.5, encoding the ion channel that is thought to trigger HPV, increased O(2)-sensitive K(+) current in cultured smooth muscle cells from rat dPAs, and restored HPV in hyperoxic rats. CONCLUSIONS: Reduced expression/activity of O(2)-sensitive K(v) channels in dPAs contributes to blunted HPV observed in neonatal CLD.

Reticulocyte enrichment of zinc protoporphyrin/heme discriminates impaired iron supply during early development.

In infants and children, elevated whole blood zinc protoporphyrin/heme (ZnPP/H) measures iron-deficient (ID) erythropoiesis. Because immature erythrocytes are less dense than mature erythrocytes, we hypothesized that the sensitivity of ZnPP/H is improved if measured in the least dense cells. Blood was collected from control suckling, mildly and severely ID suckling rats. Cord blood was collected after uncomplicated pregnancies (control), diabetic pregnancies (severe ID) and after pregnancies at-risk for iron deficiency (mild ID). ZnPP/H was measured before and after a two-step density centrifugation to obtain the lightest 6.25% of erythrocyte (top fraction). The difference between whole blood and top fraction was defined as DeltaZnPP/H. In rats, although the whole or top ZnPP/H differed by postnatal age, DeltaZnPP/H was greatest after the interval with least body iron accrual. In either rats or humans with mild ID, whole blood ZnPP/H was similar to, but DeltaZnPP/H was greater than controls. In rats and newborn humans, DeltaZnPP/H is more sensitive than whole blood ZnPP/H in identifying conditions associated with impaired erythrocyte iron delivery and may become a useful tool in measuring erythrocyte iron incorporation in early development.

Inhaled nitric oxide effects on lung structure and function in chronically ventilated preterm lambs.

RATIONALE: Inhaled nitric oxide (iNO) can reverse neonatal pulmonary hypertension and bronchoconstriction and reduce proliferation of cultured arterial and airway smooth muscle cells. OBJECTIVES: To see if continuous iNO from birth might reduce pulmonary vascular and respiratory tract resistance (PVR, RE) and attenuate growth of arterial and airway smooth muscle in preterm lambs with chronic lung disease. METHODS: Eight premature lambs received mechanical ventilation for 3 weeks, four with and four without iNO (5-15 ppm). Four term lambs, mechanically ventilated without iNO for 3 weeks, served as additional control animals. MEASUREMENTS: PVR and RE were measured weekly. After 3 weeks, lung tissue was processed for quantitative image analysis of smooth muscle abundance around small arteries (SMart) and terminal bronchioles (SMtb). Radial alveolar counts were done to assess alveolar number. Endothelial NO synthase (eNOS) protein in arteries and airways was measured by immunoblot analysis. MAIN RESULTS: At study's end, PVR was similar in iNO-treated and untreated preterm lambs; PVR was less in iNO-treated preterm lambs compared with term control animals. RE in iNO-treated lambs was less than 40% of RE measured in preterm control animals. SMart was similar in iNO-treated and both groups of control lambs; SMtb in lambs given iNO was significantly less (approximately 50%) than in preterm control animals. Radial alveolar counts of iNO-treated lambs were more than twice that of preterm control animals. eNOS was similar in arteries and airways of iNO-treated preterm lambs compared with control term lambs. CONCLUSIONS: iNO preserves structure and function of airway smooth muscle and enhances alveolar development in preterm lambs with chronic lung disease.

Pulmonary vascular dysfunction in preterm lambs with chronic lung disease.

Chronic lung injury from prolonged mechanical ventilation after premature birth inhibits the normal postnatal decrease in pulmonary vascular resistance (PVR) and leads to structural abnormalities of the lung circulation in newborn sheep. Compared with normal lambs born at term, chronically ventilated preterm lambs have increased pulmonary arterial smooth muscle and elastin, fewer lung microvessels, and reduced abundance of endothelial nitric oxide synthase. These abnormalities may contribute to impaired respiratory gas exchange that often exists in infants with chronic lung disease (CLD). Nitric oxide inhalation (iNO) reduces PVR in human infants and lambs with persistent pulmonary hypertension. We wondered whether iNO might have a similar effect in lambs with CLD. We therefore studied the effect of iNO on PVR in lambs that were delivered prematurely at approximately 125 days of gestation (term = 147 days) and mechanically ventilated for 3 wk. All of the lambs had chronically implanted catheters for measurement of pulmonary vascular pressures and blood flow. During week 2 of mechanical ventilation, iNO at 15 parts/million for 1 h decreased PVR by approximately 20% in 12 lambs with evolving CLD. When the same study was repeated in eight lambs at the end of week 3, iNO had no significant effect on PVR. To see whether this loss of iNO effect on PVR might reflect dysfunction of lung vascular smooth muscle, we infused 8-bromo-guanosine 3',5'-cyclic monophosphate (cGMP; 150 microg. kg(-1). min(-1) iv) for 15-30 min in four of these lambs at the end of week 3. PVR consistently decreased by 30-35%. Lung immunohistochemistry and immunoblot analysis of excised pulmonary arteries from lambs with CLD, compared with control term lambs, showed decreased soluble guanylate cyclase (sGC). These results suggest that loss of pulmonary vascular responsiveness to iNO in preterm lambs with CLD results from impaired signaling, possibly related to deficient or defective activation of sGC, the intermediary enzyme through which iNO induces increased vascular smooth muscle cell cGMP and resultant vasodilation.