This study determines effects of oxygen levels on morphology and VEGF expression of developing chicken lungs following incubation in normoxia (21% O2), hypoxia (15% O2) or hyperoxia (30% O2), until developmental days 16 or 18. Lung morphology was assessed using light microscopy, while VEGF expression was determined with ELISA. In hypoxia, the proportion of parabronchial tissue and parabronchi including lumina increased from day 16 to 18 (61 to 68% and 74.2 to 82.2%, respectively). Non-parabronchial tissue was higher in hypoxia than in hyperoxia on day 16 (26 to 20%). However, by day 18, there were no differences between groups. VEGF expression was 33% higher in hypoxia than in hyperoxia on day 16 (736 vs. 492 pg/ml). On day 18, VEGF expression was 43% higher in hyperoxia than in normoxia (673 to 381pg/ml), and remained elevated by 40% in hypoxia over normoxia (631 pg/ml). VEGF may be a mechanism by which parabronchial tissue is stimulated from day 16 to 18 following exposure to chronic hypoxia.

To fully understand the mechanisms of lower airway inflammation associated with many equine diseases such as heaves or Rhodococcus equi infection, which are age specific, we must first identify baseline “normal” structural characteristics of the horse lung. To develop a detailed understanding of the morphology of the horse lung, stereological methods were adapted and applied to the lungs from healthy adult horses (n=4) and one day (n=5) and 30 day (n=5) old foals. The left lung from each animal was fixed in situ and was then removed from the body cavity and remained in fixative overnight before beginning an unbiased sampling procedure. The tissue samples were fixed in plastic and paraffin blocks for stereological evaluation and immunohistochemistry, respectively. The lung was characterised into parenchyma and non-parenchyma, where median parenchymal density (alveolar airspace, ductal airspace and tissue) was 81.0% in one day old foals, 84.4% in 30 day old foals and 93.7% in adult lungs. The median volume density of alveolar airspace per lung was 45.9% in one day old, 55.5% in 30 day and 66.9% in adult horse lungs. Ductal airspace and alveolar tissue volume density was unchanged between the age groups. The median alveolar surface area (m^2) seemed to increase with age, from about 205.3m^2, 258.2m^2 and 629.9m^2 in one day old foals, 30 day old foals, and adults, respectively. While the median alveolar surface density decreased with age, the mean linear intercept increased with age. Alveolar surface area was consistently greater than endothelial surface area (m^2) within each lung, however the ratio between alveolar and endothelial surface density remains unchanged with age. The median endothelium surface area was 106.2m^2 in one day, 147.5m^2 in 30 day and 430m^2 in adult lungs. The data show that the foal is born with a functionally developed lung and its basic architecture changes with age. Foal lung development and remodelling postnatally is a result of alveolar expansion paralleled with angiogenesis.

Background: Intracellular glucocorticoid receptors (GRs) mediate the regulation of lung development, including alveolarization, by glucocorticoids (GCs). One potential approach to determining the role of GC-GR signalling in alveolar formation would be by pharmacologic blockade. Hypothesis: CP472555, a novel GR antagonist with negligible anti-PR activity, is a suitable tool for the study of GC-GR regulation of rat alveolarization. Design/Methods: CP472555 doses needed to block GR were estimated in vitro in fetal rat lung primary cultures. Postnatally, a variety of doses were administered intraperitoneally over a range of days. Results: During postnatal days (PN)0-PN10, when GC levels are low, CP472555 induced changes consistent with GR agonist activity. While GC levels increase after PN11, animals exposed to CP472555 from PN11-PN21 exhibit changes consistent with anti-GR antagonist activity. Conclusion: CP472555 causes a degree of GR blockade sufficient to permit further pharmacological investigation of the role of endogenous GC-GR signalling at the end of alveolarization.

Rationale/ Hypothesis: Reactive oxygen species, including lipid hydroperoxides, play a critical role as second messengers in many physiological processes in the body. Heightened reactive oxygen species production at the time of birth imposes an oxidative stress upon the lung, which may trigger postnatal alveologenesis and physiological lung cell apoptosis in the neonatal rat. Methods: Neonatal rats were subcutaneously injected daily with vehicle (corn oil) or diphenyl-phenyl diamine for the first 6 days of life to study alveologenesis and physiological lung cell apoptosis. Add-back experiments were conducted with a prototypic lipid hydroperoxide, t-butyl hydroperoxide. Main Results: Treatment with diphenyl-phenyl diamine resulted in parenchymal thickening, reduced numbers of secondary crests and enlarged air spaces, all consistent with the inhibition of alveologenesis and reduced physiological lung cell apoptosis. Conclusion: Following an oxidative stress at birth, lipid hydroperoxide formation triggers postnatal alveologenesis and physiological lung cell apoptosis in the neonatal rat.

Background: Intracellular glucocorticoid receptors (GRs) mediate the regulation of lung development, including alveolarization, by glucocorticoids (GCs). One potential approach to determining the role of GC-GR signalling in alveolar formation would be by pharmacologic blockade. Hypothesis: CP472555, a novel GR antagonist with negligible anti-PR activity, is a suitable tool for the study of GC-GR regulation of rat alveolarization. Design/Methods: CP472555 doses needed to block GR were estimated in vitro in fetal rat lung primary cultures. Postnatally, a variety of doses were administered intraperitoneally over a range of days. Results: During postnatal days (PN)0-PN10, when GC levels are low, CP472555 induced changes consistent with GR agonist activity. While GC levels increase after PN11, animals exposed to CP472555 from PN11-PN21 exhibit changes consistent with anti-GR antagonist activity. Conclusion: CP472555 causes a degree of GR blockade sufficient to permit further pharmacological investigation of the role of endogenous GC-GR signalling at the end of alveolarization.

Rationale/ Hypothesis: Reactive oxygen species, including lipid hydroperoxides, play a critical role as second messengers in many physiological processes in the body. Heightened reactive oxygen species production at the time of birth imposes an oxidative stress upon the lung, which may trigger postnatal alveologenesis and physiological lung cell apoptosis in the neonatal rat. Methods: Neonatal rats were subcutaneously injected daily with vehicle (corn oil) or diphenyl-phenyl diamine for the first 6 days of life to study alveologenesis and physiological lung cell apoptosis. Add-back experiments were conducted with a prototypic lipid hydroperoxide, t-butyl hydroperoxide. Main Results: Treatment with diphenyl-phenyl diamine resulted in parenchymal thickening, reduced numbers of secondary crests and enlarged air spaces, all consistent with the inhibition of alveologenesis and reduced physiological lung cell apoptosis. Conclusion: Following an oxidative stress at birth, lipid hydroperoxide formation triggers postnatal alveologenesis and physiological lung cell apoptosis in the neonatal rat.

Rationale: Activation of the platelet-derived growth factor receptors-α and -β (PDGF-Rα and -Rβ) is critical in the formation of secondary crests/septa during alveologenesis, and its regulation has been found to be disrupted in rat lung injury models. Objective: To determine whether the process of secondary septation can occur after transient pharmacologic inhibition of PDGF-R action during postnatal days (P)1 – 7 in rats. Hypothesis: The initial process of secondary crest formation is time-limited and, if missed, will result in a permanent loss of alveoli. Methods: Imatinib mesylate, a PDGF-R inhibitor, was injected intraperitoneally from P1 – 7. Pups were sacrificed on P2, 4, 8, 14, 28 and 65 for studies of alveolar development. Main results: The injection of imatinib inhibited PDGF-R action, resulting in a permanent decrease in alveolar number in treated rats. Conclusions: Inhibition of secondary septation during the first 7 days of life resulted in a decrease in alveolar number lasting into early adult life. This is consistent with a critical time window for secondary septation, which, if disrupted, results in long-term adverse effects on lung development.

Rationale: Activation of the platelet-derived growth factor receptors-α and -β (PDGF-Rα and -Rβ) is critical in the formation of secondary crests/septa during alveologenesis, and its regulation has been found to be disrupted in rat lung injury models. Objective: To determine whether the process of secondary septation can occur after transient pharmacologic inhibition of PDGF-R action during postnatal days (P)1 – 7 in rats. Hypothesis: The initial process of secondary crest formation is time-limited and, if missed, will result in a permanent loss of alveoli. Methods: Imatinib mesylate, a PDGF-R inhibitor, was injected intraperitoneally from P1 – 7. Pups were sacrificed on P2, 4, 8, 14, 28 and 65 for studies of alveolar development. Main results: The injection of imatinib inhibited PDGF-R action, resulting in a permanent decrease in alveolar number in treated rats. Conclusions: Inhibition of secondary septation during the first 7 days of life resulted in a decrease in alveolar number lasting into early adult life. This is consistent with a critical time window for secondary septation, which, if disrupted, results in long-term adverse effects on lung development.

Magister Scientiae (Medical Bioscience) - MSc(MBS) / Several studies conducted in laboratories at the University of the Western Cape has demonstrated an interference with the parenchymal lung tissue of the offspring when exposed to nicotine (smoking cigarettes and/or Nicotine Replacement Therapy [NRT]), maternally i.e. during gestation and lactation. This in turn, decreases the amount of air sacs (alveolar number) resulting in a reduced surface area available for efficient gas exchange in the offspring. Since the foetus and offspring are only exposed to nicotine during gestation and lactation, emphysema- like lesions appear to develop after nicotine withdrawal in the foetus. It has been proposed that during lung development in utero, a change in the "program" that controls the maintenance of lung integrity will occur in the long term due to the initial maternal nicotine exposure. Therefore, animals that were exposed to maternal nicotine resemble lungs that have undergone rapid, premature aging caused by cellular senescence. Furthermore, energy metabolism and structural changes in the glycolytic pathways appear irreversibly slower compared to animals that were not exposed to nicotine via the mother during gestation and lactation, resulting in a reduction in the anti-oxidant capacity of lung development. Previous studies have also shown that strong anti-oxidants supplemented by smoking mothers during gestation and lactation could possibly resist change in the "program" which controls lung development and integrity of the offspring in the long term. Lycopene – as a strong anti-oxidant supplementation have shown to decrease the alveolar volume and increase the alveolar surface area for better gas exchange after the offspring has been exposed to maternal nicotine. In this study I have treated pregnant wistar rats with nicotine, tomato juice (containing lycopene among other phytonutrients), and a combination of nicotine and tomato juice during gestation, to determine various changes in the lung structure and signs of premature aging in the lungs of the offspring. I have also performed various staining techniques such as H&E, connective tissue and β- galactosidase staining which indicated whether maternal nicotine exposure indeed induced premature cellular senescence in the lungs of the offspring. / National Research Foundation

We tested the hypothesis that oxytocin-induced labor augmented IL-1β-induced/-stimulated lung fluid absorption in preterm guinea pig fetuses. IL-1β was administered subcutaneously daily to timed-pregnant guinea pigs for 3 days with and without simultaneous cortisol synthesis inhibition by metyrapone. At day 3, oxytocin was administered, and fetuses were delivered by abdominal hysterotomy at 61 and by oxytocin-induced birth at 68 days gestation. Delivered fetuses were instilled with isosmolar 5% albumin into the lungs, and lung fluid movement was measured over 1 h by mass balance. Lung fluid absorption was induced in 61-day and stimulated in 68-day gestation lungs by IL-1β. Labor induction by oxytocin augmented IL-1β-induced/- stimulated lung fluid absorption. Metyrapone pretreatment did not affect oxytocin-induced/-stimulated lung fluid absorption, while completely blocking IL-1β-induced/-stimulated fluid absorption. Fetal lung fluid absorption, when present, was always propranolol and amiloride sensitive, suggesting that β-adrenoceptor stimulation and amiloride-sensitive sodium channels were critical for fluid absorption. Epithelial sodium channel and Na-K-ATPase subunit expressions were both increased by IL-1β, but not further by oxytocin. Our results indicate that IL-1β release into the maternal blood circulation positively affects lung maturation due to the IL-1β-induced release of cortisol and thus prepares the lungs for the epinephrine surge associated with labor.