Hyperoxia-induced Bronchopulmonary Dysplasia Research Articles

MicroRNA-mRNA interactions in a murine model of hyperoxia-induced bronchopulmonary dysplasia

BackgroundBronchopulmonary dysplasia is a chronic lung disease of premature neonates characterized by arrested pulmonary alveolar development. There is increasing evidence that microRNAs (miRNAs) regulate translation of messenger RNAs (mRNAs) during lung organogenesis. The potential role of miRNAs in the pathogenesis of BPD is unclear.ResultsFollowing exposure of neonatal mice to 80% O2 or room air (RA) for either 14 or 29 days, lungs of hyperoxic mice displayed histological changes consistent with BPD. Comprehensive miRNA and mRNA profiling was performed using lung tissue from both O2 and RA treated mice, identifying a number of dynamically regulated miRNAs and associated mRNA target genes. Gene ontology enrichment and pathway analysis revealed that hyperoxia modulated genes involved in a variety of lung developmental processes, including cell cycle, cell adhesion, mobility and taxis, inflammation, and angiogenesis. MiR-29 was prominently increased in the lungs of hyperoxic mice, and several predicted mRNA targets of miR-29 were validated with real-time PCR, western blotting and immunohistochemistry. Direct miR-29 targets were further validated in vitro using bronchoalveolar stem cells.ConclusionIn newborn mice, prolonged hyperoxia induces an arrest of alveolar development similar to that seen in human neonates with BPD. This abnormal lung development is accompanied by significant increases in the levels of multiple miRNAs and corresponding decreases in the levels of predicted mRNA targets, many of which have known or suspected roles in pathways altered in BPD. These data support the hypothesis that dynamic regulation of miRNAs plays a prominent role in the pathophysiology of BPD.

Mesenchymal Stem Cell‐Mediated Reversal of Bronchopulmonary Dysplasia and Associated Pulmonary Hypertension

Clinical trials have failed to demonstrate an effective preventative or therapeutic strategy for bronchopulmonary dysplasia (BPD), a multifactorial chronic lung disease in preterm infants frequently complicated by pulmonary hypertension (PH). Mesenchymal stem cells (MSCs) and their secreted components have been shown to prevent BPD and pulmonary fibrosis in rodent models. We hypothesized that treatment with conditioned media (CM) from cultured mouse bone marrow-derived MSCs could reverse hyperoxia-induced BPD and PH. Newborn mice were exposed to hyperoxia (FiO2=0.75) for two weeks, were then treated with one intravenous dose of CM from either MSCs or primary mouse lung fibroblasts (MLFs), and placed in room air for two to four weeks. Histological analysis of lungs harvested at four weeks of age was performed to determine the degree of alveolar injury, blood vessel number, and vascular remodeling. At age six weeks, pulmonary artery pressure (PA acceleration time) and right ventricular hypertrophy (RVH; RV wall thickness) were assessed by echocardiography, and pulmonary function tests were conducted. When compared to MLF-CM, a single dose of MSC-CM-treatment (1) reversed the hyperoxia-induced parenchymal fibrosis and peripheral PA devascularization (pruning), (2) partially reversed alveolar injury, (3) normalized lung function (airway resistance, dynamic lung compliance), (4) fully reversed the moderate PH and RVH, and (5) attenuated peripheral PA muscularization associated with hyperoxia-induced BPD. Reversal of key features of hyperoxia-induced BPD and its long-term adverse effects on lung function can be achieved by a single intravenous dose of MSC-CM, thereby pointing toward a new therapeutic intervention for chronic lung diseases.

Vasculoprotective effects of heme oxygenase-1 in a murine model of hyperoxia-induced bronchopulmonary dysplasia

Bronchopulmonary dysplasia (BPD) is characterized by simplified alveolarization and arrested vascular development of the lung with associated evidence of endothelial dysfunction, inflammation, increased oxidative damage, and iron deposition. Heme oxygenase-1 (HO-1) has been reported to be protective in the pathogenesis of diseases of inflammatory and oxidative etiology. Because HO-1 is involved in the response to oxidative stress produced by hyperoxia and is critical for cellular heme and iron homeostasis, it could play a protective role in BPD. Therefore, we investigated the effect of HO-1 in hyperoxia-induced lung injury using a neonatal transgenic mouse model with constitutive lung-specific HO-1 overexpression. Hyperoxia triggered an increase in pulmonary inflammation, arterial remodeling, and right ventricular hypertrophy that was attenuated by HO-1 overexpression. In addition, hyperoxia led to pulmonary edema, hemosiderosis, and a decrease in blood vessel number, all of which were markedly improved in HO-1 overexpressing mice. The protective vascular response may be mediated at least in part by carbon monoxide, due to its anti-inflammatory, antiproliferative, and antiapoptotic properties. HO-1 overexpression, however, did not prevent alveolar simplification nor altered the levels of ferritin and lactoferrin, proteins involved in iron binding and transport. Thus the protective mechanisms elicited by HO-1 overexpression primarily preserve vascular growth and barrier function through iron-independent, antioxidant, and anti-inflammatory pathways.

Activation of Akt Protects Alveoli from Neonatal Oxygen-Induced Lung Injury

Bronchopulmonary dysplasia (BPD) is the main complication of extreme prematurity, resulting in part from mechanical ventilation and oxygen therapy. Currently, no specific treatment exists for BPD. BPD is characterized by an arrest in alveolar development and increased apoptosis of alveolar epithelial cells (AECs). Type 2 AECs are putative distal lung progenitor cells, capable of regenerating alveolar homeostasis after injury. We hypothesized that the protection of AEC2 death via the activation of the prosurvival Akt pathway prevents arrested alveolar development in experimental BPD. We show that the pharmacologic inhibition of the prosurvival factor Akt pathway with wortmannin during the critical period of alveolar development impairs alveolar development in newborn rats, resulting in larger and fewer alveoli, reminiscent of BPD. Conversely, in an experimental model of BPD induced by oxygen exposure of newborn rats, alveolar simplification is associated with a decreased activation of lung Akt. In vitro studies with rat lung epithelial (RLE) cells cultured in hyperoxia (95% O(2)) showed decreased apoptosis and improved cell survival after the forced expression of active Akt by adenovirus-mediated gene transfer. In vivo, adenovirus-mediated Akt gene transfer preserves alveolar architecture in the newborn rat model of hyperoxia-induced BPD. We conclude that inhibition of the prosurvival factor Akt disrupts normal lung development, whereas the expression of active Akt in experimental BPD preserves alveolar development. We speculate that the modulation of apoptosis may have therapeutic potential in lung diseases characterized by alveolar damage.

From the authors

We would like to thank C. Mehats and colleagues for their comments regarding our paper on phosphodiesterase (PDE)4 inhibition in hyperoxia-induced bronchopulmonary dysplasia (BPD) 1. In our article, we stated that inhibition of PDE4 with cilomilast leads to an improved alveolarisation in hyperoxia-induced lung injury in mice 1. C. Mehats and colleagues raised concerns about our study design and the obtained results. We would like to respond to the issues in question. The first point of criticism addresses differences regarding the therapeutic impact of the PDE4 inhibitor cilomilast in our study of mice, as compared to rolipram in the study of MEhats et al. 2, which was conducted on rats. MEhats et al. 2 demonstrated that treatment of hyperoxia-exposed rats with 0.5 mg·kg−1·day−1 rolipram prolonged the survival of the treated rats …

Inhibition of phosphodiesterase 4 enhances lung alveolarisation in neonatal mice exposed to hyperoxia

Bronchopulmonary dysplasia (BPD) is characterised by impaired alveolarisation, inflammation and aberrant vascular development. Phosphodiesterase (PDE) inhibitors can influence cell proliferation, antagonise inflammation and restore vascular development and homeostasis, suggesting a therapeutic potential in BPD. The aim of the present study was to investigate PDE expression in the lung of hyperoxia-exposed mice, and to assess the viability of PDE4 as a therapeutic target in BPD. Newborn C57BL/6N mice were exposed to normoxia or 85% oxygen for 28 days. Animal growth and dynamic respiratory compliance were reduced in animals exposed to hyperoxia, paralleled by decreased septation, airspace enlargement and increased septal wall thickness. Changes were evident after 14 days and were more pronounced after 28 days of hyperoxic exposure. At the mRNA level, PDE1A and PDE4A were upregulated while PDE5A was downregulated under hyperoxia. Immunoblotting confirmed these trends in PDE4A and PDE5A at the protein expression level. Treatment with cilomilast (PDE4 inhibitor, 5 mg.kg(-1).day(-1)) between days 14 and 28 significantly decreased the mean intra-alveolar distance, septal wall thickness and total airspace area and improved dynamic lung compliance. Pharmacological inhibition of phosphodiesterase improved lung alveolarisation in hyperoxia-induced bronchopulmonary dysplasia, and thus may offer a new therapeutic modality in the clinical management of bronchopulmonary dysplasia.

Vascular Endothelial Growth Factor Gene Therapy Increases Survival, Promotes Lung Angiogenesis, and Prevents Alveolar Damage in Hyperoxia-Induced Lung Injury

Bronchopulmonary dysplasia (BPD) and pulmonary emphysema, both significant global health problems, are characterized by a loss of alveoli. Vascular endothelial growth factor (VEGF) is a trophic factor required for endothelial cell survival and is abundantly expressed in the lung. We report that VEGF blockade decreases lung VEGF and VEGF receptor 2 (VEGFR-2) expression in newborn rats and impairs alveolar development, leading to alveolar simplification and loss of lung capillaries, mimicking BPD. In hyperoxia-induced BPD in newborn rats, air space enlargement and loss of lung capillaries are associated with decreased lung VEGF and VEGFR-2 expression. Postnatal intratracheal adenovirus-mediated VEGF gene therapy improves survival, promotes lung capillary formation, and preserves alveolar development in this model of irreversible lung injury. Combined VEGF and angiopoietin-1 gene transfer matures the new vasculature, reducing the vascular leakage seen in VEGF-induced capillaries. These findings underscore the importance of the vasculature in what is traditionally thought of as an airway disease and open new therapeutic avenues for lung diseases characterized by irreversible loss of alveoli through the modulation of angiogenic growth factors.

Lung inflammation in hyperoxia can be prevented by antichemokine treatment in newborn rats.

Hyperoxia may contribute to lung disease in newborns through effects on alveolar neutrophils which predominate in respiratory distress syndrome and other acute lung injuries. Neutrophil chemokines such as interleukin-8 (IL-8) regulate chemoattraction, and are elevated in tracheal aspirates of newborns who develop bronchopulmonary dysplasia (BPD). Blockade of neutrophil chemokines may reduce hyperoxia-induced inflammatory lung injury and BPD. We therefore tested the hypothesis that hyperoxia contributes to elevations of rat neutrophil chemokines, cytokine-induced neutrophil chemoattractant-1 (CINC-1), and macrophage inflammatory protein-2 (MIP-2) in newborn rat lung. Newborn rats were exposed to air or 95% O(2) for 8 d. CINC-1 and MIP-2 were measured in whole lung homogenates by ELISA. Newborn 95% O(2)-exposed animals were given anti-CINC-1 or anti-MIP-2, 1, 5, or 10 microg on Days 3 and 4 of 95% O(2) exposure. Bronchoalveolar lavage (BAL) was performed after perfusion on day 6 to evaluate airway neutrophils, and myeloperoxidase (MPO) was measured in perfused whole lung. Lungs were examined histologically and immunohistochemically for effects of 95% O(2) +/- antichemokine. CINC-1 and MIP-2 increased nearly tenfold by Day 8 95% O(2) treatment versus air control. CINC-1 and MIP-2 immunolabeling was increased in alveolar macrophages and alveolar epithelium in 95% O(2). Anti-CINC-1 and anti-MIP-2 treatment at every dose reduced neutrophil number > 90% in BAL. Anti-CINC-1 10 microg reduced tissue MPO by 50%. Antichemokine treatment on days 3 and 4 prevented alveolar septal thickening and reduced chemokine immunolabeling on Day 6. Hyperoxia-induced neutrophil influx is mediated in part by CINC-1 and MIP-2 in newborn rats and can be partially prevented by treatment with anti-CINC-1 and anti-MIP-2.

Early high dose antioxidant vitamins do not prevent bronchopulmonary dysplasia in premature baboons exposed to prolonged hyperoxia: a pilot study.

The antioxidant vitamins ascorbic acid (AA) and alpha-tocopherol (alpha-TP) effectively inhibit oxygen free radical-induced lipid peroxidation. Using a premature baboon model of hyperoxia-induced bronchopulmonary dysplasia (BPD), we measured concentrations of AA, alpha-TP, and conjugated dienes (CD, marker of lipid peroxidation) in four animals (hyperoxic antioxidant group) receiving high dose antioxidant vitamin supplementation (AA, 100 mg x kg x(-1) x d(-1); alpha-TP; 20 mg x kg x(-1) x d(-1)) and one animal receiving standard dose antioxidant vitamin supplementation (AA, 10 mg x kg x(-1) x d(-1); alpha-TP, 1 mg x kg x(-1) x d(-1)). Respiratory and histopathologic data were compared with data from 10 historical animals exposed to hyperoxia (hyperoxic control group) and 11 historical animals treated as required with oxygen (normoxic control group) who had received standard dose antioxidant vitamin supplementation. Compared with standard dose antioxidant vitamin supplementation, high dose antioxidant vitamin supplementation effectively raised AA concentrations in plasma (37 +/- 22 micromol/L and 395 +/- 216 micromol/L, respectively) and tracheal aspirates (62 +/- 35 micromol/L and 286 +/- 205 micromol/L, respectively), and alpha-TP concentrations in plasma (10.1 +/- 2.5 micromol/L and 24.6 +/- 17.5 micromol/L, respectively). However, there was no apparent effect on tracheal aspirate CD concentrations (482 +/- 333 micromol/L and 1050 +/- 1111 micromol/L, respectively), and respiratory parameters in the hyperoxic antioxidant group were comparable to those of the hyperoxic control group but significantly worse than in the normoxic control group. Finally, no protective effect of high dose antioxidant vitamin supplementation was noted at the histopathologic level.

Lung manganese superoxide dismutase protein expression increases in the baboon model of bronchopulmonary dysplasia and is regulated at a posttranscriptional level.

The expression of lung manganese superoxide dismutase (MnSOD) mRNA and protein were examined in a premature baboon model of hyperoxia-induced bronchopulmonary dysplasia (BPD) and BPD superimposed with bacterial infection. When 140-d gestation baboons were delivered by hysterotomy and treated for 16 d with appropriate ventilatory and oxygen support (pro re nada controls), there was an increase in both MnSOD mRNA and protein compared with 140-d or 156-d gestation, nonventilated controls. The concentration of MnSOD protein was also elevated when the prematurely delivered baboons were ventilated with a high fraction of inspired O2 to produce a primate homolog of BPD, but there was a significant decrease in the concentration of MnSOD mRNA in BPD animals compared with pro re nada controls. In the lungs of premature baboons in which Escherichia coli infection was superimposed on hyperoxia-induced BPD, MnSOD mRNA was diminished to approximately the same extent as in BPD alone, but MnSOD protein was significantly increased compared with all other groups. Taken together these data indicate that the premature baboon is capable of mounting an antioxidant response and that increased MnSOD protein expression in BPD and BPD-infected premature baboons is regulated, at least in part, at a posttranscriptional level.

Surfactant protein-A deficiency in a primate model of bronchopulmonary dysplasia.

Pathophysiologic and biochemical (surfactant protein and phospholipid) features were studied in a baboon model of hyperoxia-induced bronchopulmonary dysplasia (BPD) and superimposed infection. A total of 20 baboons were delivered by hysterotomy at 76% of gestation (140 d of gestational age) and were randomized into four groups, consisting of two control and two injury groups. Animals constituting a group that was managed on a pro re nata (PRN) basis were ventilated with clinically appropriate oxygen for the 16-d experimental period and served as ventilatory controls. They underwent an initial period of 42 h during which they demonstrated evidence of hyaline membrane disease (HMD), but began recovery at 42 h and by Day 6 appeared to have maximally recovered. At the time of these animals' killing, concentrations of surfactant proteins, messenger ribonucleic acids (mRNAs), and phospholipids were similar to those of normal adult baboons. Gestational control animals were delivered and killed without ventilation at 156 d gestational age. Surfactant protein-A (SP-A) and phospholipid concentrations in these animals' lavage fluids were about 10% of those in the PRN animals. Animals with BPD were subjected to positive-pressure ventilation and an FIO2 of 1.0 for 11 d, followed by 5 d of an FIO2 sufficient to maintain PaO2 at 40 to 50 mm Hg. The animals with BPD and infection were treated in the same way as the BPD group, except that 10(8) Escherichia coli were instilled intratracheally on Day 11, concomitantly with the reduction in FIO2.(ABSTRACT TRUNCATED AT 250 WORDS)