Mildred T. Stahlman

Surfactant Protein-D Regulates Surfactant Phospholipid Homeostasis in Vivo

Surfactant protein D (SP-D) is a 43-kDa member of the collectin family of collagenous lectin domain-containing proteins that is expressed in epithelial cells of the lung. The SP-D gene was targeted by homologous recombination in embryonic stem cells that were used to produce SP-D (±) and SP-D (−/−) mice. Both SP-D (−/−) and SP-D (±) mice survived normally in the perinatal and postnatal periods. Whereas no abnormalities were observed in SP-D (±) mice, alveolar and tissue phosphatidylcholine pool sizes were markedly increased in SP-D (−/−) mice. Increased numbers of large foamy alveolar macrophages and enlarged alveoli were also observed in SP-D (−/−) mice. Phospholipid composition was unaltered in SP-D (−/−) mice, but surfactant morphology was abnormal, consisting of dense phospholipid membranous arrays with decreased tubular myelin. The pulmonary lipoidosis in the SP-D (−/−) mice was not associated with accumulation of surfactant proteins B or C, or their mRNAs, distinguishing the disorder from alveolar proteinosis syndromes. Surfactant protein A mRNA was reduced and, SP-A protein appeared to be reduced in SP-D (−/−) compared with wild type mice. Targeting of the mouse SP-D gene caused accumulation of surfactant lipid and altered phospholipid structures, demonstrating a previously unsuspected role for SP-D in surfactant lipid homeostasis in vivo.

Clinical trial of vitamin A supplementation in infants susceptible to bronchopulmonary dysplasia

We conducted a randomized, double-blind, controlled trial to determine whether vitamin A supplementation from early postnatal life could reduce the morbidity associated with bronchopulmonary dysplasia in very low birth weight (VLBW) neonates. Forty VLBW neonates (700 to 1300 g birth weight, 26 to 30 weeks gestational age), who were oxygen dependent and required mechanical ventilation for at least 72 hours after birth, were given by the intramuscular route either supplemental vitamin A (retinyl palmitate 2000 IU) or 0.9% saline solution on postnatal day 4 and every other day thereafter for a total of 14 injections over 28 days. The study groups were comparable in gestational maturity, clinical characteristics, initial lung disease, and vitamin A status at entry into the trial. Vitamin A administration resulted in significantly higher mean plasma concentrations of vitamin A and retinol-binding protein in treated infants compared with controls. Bronchopulmonary dysplasia was diagnosed in nine of 20 infants given vitamin A supplement and in 17 of 20 control infants (P less than 0.008). Four of 19 infants in the vitamin A group and 11 of 20 in the control group required mechanical ventilation on study day 28 (P less than 0.029). The need for supplemental oxygen, mechanical ventilation, and intensive care was reduced in infants given vitamin A supplement compared with controls. Airway infection and retinopathy of prematurity were less frequent in the vitamin A group. We conclude that vitamin A supplementation at the dosage used in this trial in VLBW neonates not only improves their vitamin A status but also appears to promote regenerative healing from lung injury, as evidenced by a decrease in the morbidity associated with bronchopulmonary dysplasia.

Immunocytochemical Localization of Human Epidermal Growth Factor/Urogastrone in Several Human Tissues'

Epidermal growth factor (EGF) stimulates the growth of many tissues and inhibits stimulated gastric acid secretion. Its primary tissue of origin in man is still unknown. We used polyclonal anti-human EGF sera in the peroxidase-antiperoxidase immunocytochemical staining technique to identify immunoreactive human EGF (ihEGF) in tissue sections from 29 subjects ranging from fetuses to 63 years in age. In addition to acinar cells in the submandibular salivary glands and cells of Brunners duodenal glands, previously reported to contain ihEGF, we found ihEGF in most anterior pituitary glycopeptide hormone-secreting cells, in gastric and pyloric gland cells of the stomach, and in bone marrow cells that resembled mononuclear phagocytes in subjects of all ages. The eccrine sweat glands in the skin of adults also contained ihEGF. Cells containing ihEGF were found singly or in clusters in the trachea of the fetus only. No fetal pancreatic islet cells stained, but occasional cells in neonates and a majority of islet cells in older subjects contained ihEGF; there was no constant association with insulin, glucagon, or somatostatin. Only the lactating breast contained ihEGF. In adults, outer adrenomedullary cells contained ihEGF. Intense immunostaining was observed in the renal medulla, apparently limited to the extracellular area between the renal tubules, and increased with age; the cortex was devoid of ihEGF. No ihEGF was detected in posterior pituitary gland, thyroid gland, heart, lung, or liver at any age. An adult prostate contained ihEGF only in an area of local injury, and some primordial follicles from the ovary of a newborn appeared to contain ihEGF. Thus, many tissues appear to synthesize hEGF, which may exert exocrine, endocrine, or paracrine functions in different tissues and at different ages.

Foxa2 regulates alveolarization and goblet cell hyperplasia

The airways are lined by several distinct epithelial cells that play unique roles in pulmonary homeostasis; however, the mechanisms controlling their differentiation in health and disease are poorly understood. The winged helix transcription factor, FOXA2, is expressed in the foregut endoderm and in subsets of respiratory epithelial cells in the fetal and adult lung. Because targeted mutagenesis of the Foxa2 gene in mice is lethal before formation of the lung, its potential role in lung morphogenesis and homeostasis has not been determined. We selectively deleted Foxa2 in respiratory epithelial cells in the developing mouse lung. Airspace enlargement, goblet cell hyperplasia, increased mucin and neutrophilic infiltration were observed in lungs of the Foxa2-deleted mice. Experimental goblet cell hyperplasia caused by ovalbumin sensitization, interleukin 4 (IL4), IL13 and targeted deletion of the gene encoding surfactant protein C (SP-C), was associated with either absent or decreased expression of Foxa2 in airway epithelial cells. Analysis of lung tissue from patients with a variety of pulmonary diseases revealed a strong inverse correlation between FOXA2 and goblet cell hyperplasia. FOXA2 is required for alveolarization and regulates airway epithelial cell differentiation in the postnatal lung.

Vitamin A Status of Neonates with Bronchopulmonary Dysplasia

ABSTRACT: We prospectively assessed and compared the vitamin A status of two groups of preterm neonates (<1500 g birth weight, <32 wk gestation), one who developed clinical and radiographic evidence of bronchopulmonary dysplasia (BPD) (n = 10), and the other (control) who developed no significant lung disease (n = 8). The infants with BPD in this study required prolonged mechanical ventilation and supplemental O2 therapy, and had a higher incidence of cardiorespiratory complications when compared to controls. Their mean plasma vitamin A concentrations were significantly lower than those of controls at four sampling times in the 1st postnatal month. In contrast to the controls, infants with BPD showed a substantial decline in their plasma vitamin A concentrations from the initial values, and a high percentage of individual values of plasma vitamin A concentration in these infants were <10 μg/dl during the 8-wk postnatal period of observation. Delayed establishment of gastrointestinal feeding and a lower vitamin A intake in these infants relative to controls may have accounted for this decline. Our data show that preterm neonates who develop BPD have suboptimal plasma vitamin A concentrations for extended periods of time postnatally. We speculate that the necrotizing bronchiolitis and squamous metaplasia of conducting airways associated with vitamin A deficiency could influence the orderly repair of lung injury in susceptible neonates who are mechanically ventilated and could contribute to the pathophysiology of BPD in these infants. Further studies are needed to determine whether more efficient management of these neonates with regard to their vitamin A nutrition and early achievement of vitamin A sufficiency could result in a decrease in the incidence and severity of BPD.

Compensatory Roles of Foxa1 and Foxa2 during Lung Morphogenesis

Foxa1 and Foxa2 are closely related family members of the Foxa group of transcription factors that are coexpressed in subsets of respiratory epithelial cells throughout lung morphogenesis. Shared patterns of expression, conservation of DNA binding, and transcriptional activation domains indicate that they may serve complementary functions in the regulation of gene expression during lung morphogenesis. Whereas branching morphogenesis of the fetal lung occurs normally in the Foxa2Δ/Δ and Foxa1–/– mice, deletion of both Foxa1 and Foxa2 (in Foxa2Δ/Δ, Foxa1–/– mice) inhibited cell proliferation, epithelial cell differentiation, and branching. Dilation of terminal lung tubules and decreased branching were observed as early as embryonic day 12.5. Foxa1 and Foxa2 regulated Shh (sonic hedgehog) and Shh-dependent genes in the respiratory epithelial cells that influenced the expression of genes in the pulmonary mesenchyme that are required for branching morphogenesis. Epithelial cell differentiation, as indicated by lack of expression of surfactant protein B, surfactant protein C, the Clara cell secretory protein, and Foxj1, was inhibited. Foxa family members regulate signaling and transcriptional programs required for morphogenesis and cell differentiation during formation of the lung.

Randomized trial of early closure of symptomatic patent ductus arteriosus in small preterm infants.

As a result of randomized assignment, 15 preterm infants weighing 1,500 gm or less at birth and who had a symptomatic PDA were treated according to a medical management protocol, and ten according to an early surgical closure protocol. All infants required mechanical ventilation at the time of study entry, which was one week after birth. Birth weight, gestational age, age at onset of congestive failure, age at study entry, and the initial morbidity of members of the two groups were similar. The nine surviving infants managed according to the surgical closure protocol were weaned from mechanical ventilation sooner, had a decreased need for digoxin and furosemide, achieved gastrointestinal function sooner, and had a smaller hospital bill than the 12 survivors of the medical management group. These results indicate that infants with a symptomatic PDA still requiring mechanical ventilation at one week after birth will benefit from surgical closure of the ductus at that time.

Effect of Epidermal Growth Factor on Lung Maturation in Fetal Rabbits

Summary: Injection of epidermal growth factor (EGF) into 24-day rabbit fetuses (5 μg, im or ip) induced accelerated maturation of the lung. On sacrifice at day 27, there was greater distensibility and stability on deflation associated with the appearance of a complement of type II cells approaching that of the rabbit at term. EGF treatment had no demonstrable effect on body weight or lung weight in this group of animals. Saline-injected control fetuses were not affected significantly.Speculation: EGF is capable not only of promoting epithelial cell growth but also differentiation in the fetal rabbit lung. It may be an important hormone in the maturation of the lung and capable of protecting the prematurely delivered fetus against the development of hyaline membrane disease.

Biogenesis of lamellar bodies, lysosome-related organelles involved in storage and secretion of pulmonary surfactant.

Lamellar bodies are members of a subclass of lysosome-related organelles referred to as secretory lysosomes. The principal constituents of the lamellar body, surfactant phospholipids, are organized into tightly packed, bilayer membranes in a process that is strongly influenced by the lung-specific, hydrophobic peptide SP-B. Newly synthesized SP-B is transported from the Golgi to the lamellar body via multivesicular bodies; in contrast, preliminary evidence suggests that newly synthesized surfactant phospholipids are transported from the ER and incorporated into the internal membranes of the lamellar body via a distinct pathway.

Medical management of small preterm infants with symptomatic patient ductus arteriosus

During 1975, 38 of 44 infants with a birth weight of less than or equal to 1,500 gm who developed pulmonary edema and congestive heart failure due to a patent ductus arteriosus were managed medically until the ductus closed spontaneously days or weeks later. Overall survival was 71%, and there were no deaths among 11 infants weighing more than 1,250 gm. Pulmonary complications were prevalent and were attributed to the extensive use of mechanical ventilation required to control pulmonary edema. The results of this study document the results to be expected when small preterm infants with a symptomatic patent ductus arteriosus are managed without surgical or pharmacologic intervention and provide a basis for the rational design of clinical trials evaluating other management approaches.