L K Kullama

Chronic Lung Injury in Preterm Lambs: Abnormalities of the Pulmonary Circulation and Lung Fluid Balance

Chronic lung disease of early infancy, or bronchopulmonary dysplasia, is a frequent complication of prolonged mechanical ventilation after premature birth. Pulmonary hypertension and edema are common features of this condition, which is often attributed to long-term, repetitive overinflation of incompletely developed lungs. The overall objective of this work was to examine the effects on the pulmonary circulation and lung fluid balance of different ventilation strategies using large versus small inflation volumes in an animal model of bronchopulmonary dysplasia. We studied 16 newborn lambs that were delivered prematurely (124 ± 3 d gestation, term = 147 d) by cesarean section and mechanically ventilated for 3 to 4 wk. Ten lambs were ventilated at 20 breaths/min, yielding a tidal volume of 15 ± 5 mL/kg, and six lambs were ventilated at 60 breaths/min, yielding a tidal volume of 6 ± 2 mL/kg. All lambs received surfactant at birth and had subsequent surgery for closure of the ductus arteriosus and catheter placement to allow serial measurements of pulmonary vascular resistance and lung lymph flow. Chronic lung injury, documented by serial chest radiographs and postmortem pathologic examination, developed in all lambs irrespective of the pattern of assisted ventilation. Pulmonary vascular resistance, which normally decreases during the month after birth at term, did not change significantly from the first to the last week of study. Lung lymph flow, an index of net transvascular fluid filtration, increased with time in lambs that were ventilated at 20 breaths/min, but not in lambs ventilated at 60 breaths/min. Lymph protein concentration decreased with time, indicative of increased fluid filtration pressure, without evidence of a change in lung vascular protein permeability. Postmortem studies showed interstitial lung edema, increased pulmonary arteriolar smooth muscle and elastin, decreased numbers of small pulmonary arteries and veins, and decreased capillary surface density in distal lung of chronically ventilated lambs compared with control lambs that were killed either 1 d (same postconceptional age) or 3 wk (same postnatal age) after birth at term. Thus, chronic lung injury from prolonged mechanical ventilation after premature birth inhibits the normal postnatal decrease in pulmonary vascular resistance and leads to lung edema from increased fluid filtration pressure. These abnormalities of the pulmonary circulation may contribute to the abnormal respiratory gas exchange that often exists in infants with bronchopulmonary dysplasia.

Developmental Changes in Sodium Nitroprusside and Atrial Natriuretic Factor Mediated Relaxation in the Guinea Pig Aorta

ABSTRACT: Sodium nitroprusside (SNP), a nonreceptor mediated stimulant of soluble guanylate cyclase, and atrial natriuretic factor, a receptor-dependent stimulator of particulate guanylate cyclase, mediate relaxation responses by increasing intracellular cGMP. This in vitro study was designed to compare the ontogeny of relaxation responses to SNP and atrial natriuretic factor in the guinea pig thoracic aorta. Aortic rings from fetuses at 55-60 d gestation (term = 68 d), 1- to 3-d-old newborn, and 12-wk-old adult Hartley guinea pigs were mounted in an organ bath, bathed in Krebs solution, and connected to a force-displacement transducer to measure isometric tension. Relaxation responses to SNP and atriopeptin III were studied with the vessels at optimal resting tension and after preconstriction with an EC85 concentration of norepinephrine. SNP-mediated relaxation showed a significant increase in sensitivity with development among the three age groups (p < 0.05). Methylene blue, an inhibitor of soluble guanylate cyclase, produced no inhibition of relaxation to SNP in fetal aortae, significantly decreased responses along the straight portion of the concentration-response curve in newborn aortae (p < 0.05), and significantly shifted the concentration-response curve to the right (p < 0.05) in adult aortae; but did not prevent vessels from relaxing almost 100% in any age group. However, atriopeptin III-mediated responses were similar in the three age groups and were unaffected by methylene blue. These results suggest that 1) sensitivity to SNP increases with age from fetal through adult life; 2) relaxation mediated by atriopeptin III is similar during development; 3) methylene blue does not affect SNP mediated relaxation in fetuses but progressively decreases sensitivity to SNP in newborns and adults; and4) methylene blue does not affect atriopeptin III-mediated relaxation in any age group.

Nitric Oxide Inhalation Decreases Pulmonary Vascular Resistance in Preterm Lambs With Evolving Chronic Lung Disease. • 1470

Nitric Oxide Inhalation Decreases Pulmonary Vascular Resistance in Preterm Lambs With Evolving Chronic Lung Disease. • 1470

SURFACTANT ALTERS LUNG LIQUID PRODUCTION AND EPITHELIAL ION TRANSPORT IN FETAL SHEEP. |[dagger]| 1949

Surfactant content progressively increases in the lung lumen around the time of birth. During this same period, net production of lung liquid slows and reabsorption of liquid across the lung epithelium begins. To see if surfactant participates in the regulation of net production of lung liquid(Jv), we placed tracheal catheters in 9 fetal lambs (studied at 132 ± 2 d gestation; term 147 d) to measure Jv before and after surfactant (Infasurf, 350 mg) instillation into the lung lumen. We measured Jv by calculating changes in lung liquid volume over time as determined by the concentration of125 I-albumin placed into the lung lumen before each experiment. When we found that surfactant significantly decreased Jv (n=8, baseline: 12 ± 7; surfactant: 2 ± 9 ml/h; mean ± SD), we performed studies in which we measured Jv following the instillation of surfactant together with amiloride (≈10-4M), an inhibitor of transepithelial Na movement. Jv was significantly greater following the combination of surfactant and amiloride than after surfactant alone (n=8, baseline: 14 ± 7; amiloride/surfactant: 9 ± 7 ml/h), suggesting that enhanced Na reabsorption was in part responsible for the decrease in Jv after surfactant instillation. Because changes in lung liquid movement around the time of birth occur in association with increased Na-K-ATPase activity in distal lung epithelium, we studied fetal lamb distal lung epithelial cells in culture following acute surfactant exposure and assessed Na-K-ATPase activity by calculating the fraction of 86Rb (a K substitute) uptake that was inhibited by ouabain (10-4M). In 6 studies (passages 5-9), surfactant(160 μg/ml) increased ouabain-sensitive 86Rb uptake on average by 88% (control: 82 ± 29; surfactant: 132 ± 37 nmol/106 cells/h). These results suggest that surfactant slows net production of lung liquid by a mechanism involving Na transport and Na-K-ATPase. Thus, in addition to promoting alveolar stability, surfactant may help regulate water balance in the perinatal lung through an effect on lung epithelial ion transport. (Supported by NIH HL49098).

PROLONGED MECHANICAL VENTILATION OF PRETERM LAMBS LEADS TO SMOOTH MUSCLE EXTENSION AND ELASTIN ACCUMULATION ALONG PULMONARY BLOOD VESSELS.|[dagger]| 1923

Chronic lung injury from prolonged mechanical ventilation after premature birth is characterized in part by pulmonary edema and sustained pulmonary vascular resistance (PVR) (Bland et al; FASEB J 9:A275, 1995). These observations suggest that chronic lung injury after premature birth may be associated with abnormal structural development of the pulmonary circulation. We tested this hypothesis by determining smooth muscle extension and elastin accumulation along pulmonary arteries (PAs) and veins (PVs) in 3 groups of lambs. 5 preterm lambs each were mechanically ventilated at 60 breaths/min(tidal volume 5 ± 2 ml/kg) or 20 breaths/min (tidal volume 15 ± 5 ml/kg) for 3-4 wks postnatally. Inspired O2 was adjusted to keep PaO2 ≈60-90 mmHg. 5 term lambs <24 h old served as gestation-matched controls. Lung lobes were fixed by clamping them at the prevailing inflation pressure. Paraffin-embedded tissue sections were analyzed for vascular smooth muscle extension (trichrome stain) and elastin accumulation (Harts elastic fiber stain) along PAs and PVs by quantitative histology (mean ± SD; *:p < 0.05 vs control). Muscularization of PAs and elastin accumulation in PAs and PVs may contribute to the histologic and extravascular lung water content evidence of pulmonary edema, and the sustained PVR by increasing the filtration pressure and resistance along the pulmonary microcirculation. (Supported by March of Dimes #6 FY95 0981)Table

CHRONIC LUNG INJURY BLUNTS THE PULMONARY VASOCONSTRICTOR RESPONSE TO HYPOXIA IN PRETERM LAMBS. • 1944

CHRONIC LUNG INJURY BLUNTS THE PULMONARY VASOCONSTRICTOR RESPONSE TO HYPOXIA IN PRETERM LAMBS. • 1944

Sustained Hyperoxia Inhibits the Postnatal Increase in Na,K-ATPase Activity in Distal Lung Epithelial Cells From Chronically Ventilated Preterm Baboons|[bull]| 1649

Lung epithelial cell Na,K-ATPase activity increases at birth in association with clearance of fluid from the potential airspaces. We hypothesized that the pulmonary edema that occurs in preterm baboons after sustained hyperoxia and mechanical ventilation might be associated with inhibition of the perinatal increase in Na,K-ATPase activity. To test this hypothesis, we measured Na,K-ATPase activity in distal lung epithelial cells, and we assessed Na,K-ATPase immunoreactivity and α1 and β1subunit mRNA content in whole lung tissue harvested from neonatal baboons that were delivered prematurely (140d gestation, term=180d) and mechanically ventilated for 6-10d with either 100% O2 (hyperoxia) or sufficient O2 to maintain normal PaO2 (normoxia). Results were compared with those from gestation-matched control fetuses. Na,K-ATPase activity, determined by measurement of ouabain-sensitive 86Rb uptake (nmol/106 cells/h), averaged 14 ± 3 in cells from 8 control fetuses, 24 ± 3(significantly different from control, p<0.05) in cells from 10 newborns with normoxia, and 14 ± 2 in lung epithelial cells from 14 newborns with hyperoxia. These results demonstrate that ventilation with 100% O2 is associated with inhibition of the normal postnatal increase in lung epithelial cell Na,K-ATPase activity that occurs after premature birth without subsequent hyperoxia. Immunohistochemistry, performed with antisera to Na,K-ATPase holoenzyme, showed localization to the epithelial membrane in all groups, but there was decreased signal intensity in lung sections from the hyperoxia group compared to the normoxia group and to fetal controls. mRNA content for the α1 and β1 Na,K-ATPase subunits was determined in whole lung tissue by northern blot analysis and quantified by phosphorimagery. A 6-fold increase (p<0.05, n=3) in α1 subunit mRNA content was observed in the hyperoxic group relative to normoxic newborns and control fetuses. No significant difference was observed between the groups in mRNA content for the β1subunit. Thus, sustained postnatal hyperoxia enhanced α1 subunit mRNA abundance, but inhibited Na,K-ATPase activity in the lungs of preterm baboons. We speculate that the failure of epithelial cell Na,K-ATPase activity to increase postnatally in non-human primates that are mechanically ventilated with 100% O2 probably contributes to the postnatal lung edema and associated respiratory distress that occurs in these animals.

ENDOTHELIAL NITRIC OXIDE SYNTHASE EXPRESSION IS DECREASED IN CHRONIC LUNG INJURY IN PRETERM LAMBS. † 2016

ENDOTHELIAL NITRIC OXIDE SYNTHASE EXPRESSION IS DECREASED IN CHRONIC LUNG INJURY IN PRETERM LAMBS. † 2016

ADRENAL HORMONE EFFECTS ON LUNG LIQUID PRODUCTION IN FETAL SHEEP. ▴ 358

Aldosterone is a mineralocorticoid hormone that increases Na,K-ATPase activity in both airway and distal lung epithelium of fetal sheep (Kullama et al, FASEB J 10: in press, 1996). We previously reported (Kullama et al, FASEB J 9:A568, 1995) that intravenous infusion of aldosterone for 24 h increased net production of lung liquid (Jv) in immature fetal sheep (125 d gestation; term = 147 d), but decreased Jv in more mature fetal sheep (≥136 d gestation). These different effects of aldosterone on Jv at different stages of development may reflect maturational changes in the respiratory tract epithelium, which is predominantly a Cl- secreting membrane early in gestation, switching to a Na+ absorbing membrane around the time of birth. To see if glucocorticoid treatment might modify the response of the immature lung epithelium to aldosterone, we measured Jv in 5 preterm (125± 1 d gestation) fetal sheep before and at the end of a 24-h iv infusion of aldosterone (5 μg/h), and again at the end of a 24-h simultaneous iv infusion of aldosterone and cortisol (10 mg/24 h). Each fetus had a chronically implanted tracheal loop cannula that enabled us to assess Jv by measuring serial changes in the concentration of an impermeant tracer,125 I-albumin, that was added to lung liquid and well mixed within the lung lumen at the start of each study. Jv (mean ± SD) averaged 11.6± 4.4 ml/h before aldosterone, 14.4 ± 8.5 ml/h after 24 h of iv aldosterone, and 8.7 ± 5.8 ml/h after 24 h of iv aldosterone plus cortisol. This decrease in Jv occurred in all 5 studies. Thus, glucocorticoid administration to immature fetal sheep modifies the respiratory epithelial response to aldosterone, with a resultant decrease in net production of lung luminal liquid. It is therefore possible that birth-related increases in the plasma concentration of adrenal hormones, including aldosterone and cortisol, contribute to the decrease in lung liquid that occurs near birth. (Supported by NIH #HL 49098)