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To characterize surfactant kinetics in vivo in two groups of premature infants on different levels of mechanical ventilation and at different risk of developing bronchopulmonary dysplasia.Controlled observational study in two independent groups of infants.Neonatal intensive care unit.Thirteen preterm infants (26 ± 0.5 wks, birth weight 801 ± 64 g) on high ventilatory setting and who finally all developed bronchopulmonary dysplasia (MechVentBPD), and eight (26 ± 0.5 wks, birth weight 887 ± 103 g) who had minimal or no lung disease and of whom none developed bronchopulmonary dysplasia (MechVentNoBPD).Endotracheal 13C-labeled dipalmitoyl-phosphatidylcholine was administered and subsequent measurements of the 13C enrichment of surfactant-disaturated phosphatidylcholine (DSPC) from serial tracheal aspirates were made by gas chromatography-mass spectrometry. We calculated disaturated phosphatidylcholine pharmacokinetic variables in terms of half-life and apparent pool size from the enrichment decay curves over time. DSPC concentration from tracheal aspirates was expressed as milligrams/milliliter epithelial lining fluid (ELF-DSPC). Data are presented as mean ± se. In MechVentBPD infants vs. MechVentNoBPD, ELF-DSPC was much reduced, 2.9 ± 0.6 vs. 9.4 ± 3.0 mg/mL ELF (p = .03), half-life was shorter, 19.4 ± 2.8 vs. 42.5 ± 6.3 hrs (p = .002), and apparent pool size larger, 136 ± 21 vs. 65.8 ± 16.0 mg/kg (p = .057). In MechVentBPD, apparent DSPC pool size positively correlated with mean airway pressure × Fio2 and inversely correlated with ELF-DSPC. ELF-DSPC was inversely correlated with mean airway pressure × Fio2. No significant correlations were found in the MechVentNoBPD group.MechVentBPD infants showed profound alteration of surfactant kinetics compared with preterm infants with minimal lung disease, and these alterations were correlated with severity of ventilatory support.