Model Of Bronchopulmonary Dysplasia Research Articles

Searching for better animal models of BPD: a perspective.

There have been many efforts to develop good animal models of bronchopulmonary dysplasia (BPD) to better understand the pathophysiology and mechanisms underlying development of BPD as well as to test potential strategies for its prevention and treatment. This Perspectives summarizes the features of common animal models of BPD and the strengths and limitations of such models. Potential optimal approaches to development of animal models are indicated, with the underlying concepts that require emphasis.

Alteration of TGF-β-ALK-Smad signaling in hyperoxia-induced bronchopulmonary dysplasia model of newborn rats

ABSTRACTBackground: Bronchopulmonary dysplasia (BPD) is a main chronic lung disease commonly occurs in preterm infants. BPD is characterized by impaired alveolarization and vascularization of the developing lung. Transforming growth factor-β (TGF-β) signaling pathway is known to play an important role during lung vascular development. In the present study, we examined whether the regulation of TGF-β-ALK-Smad signaling pathway influence on the disruption of pulmonary vascular development in newborn rats as hyperoxia-induced BPD model. Materials and Methods: Newborn rats were continuously exposed to 21% or 85% O2 for 7 days, and subsequently kept in normoxic condition for another 14 days. Lung tissues harvested at each time point were evaluated for the expression of TGF-β1, ALK1, ALK5, phosphorylated Smad1/5, phosphorylated Smad2/3, VEGF, and endoglin, as accessed by both biochemical and immunohistological analyses. Results: Double-fluorescence immunohistochemical staining indicated these molecules were mainly expressed in pulmonary endothelial cells. The expression of TGF-β1 and ALK5 mRNA and protein were significantly increased in D5 hyperoxia group, while that of ALK1 mRNA and protein were significantly decreased. The level of phosphorylated Smad1/5 was significantly decreased in D7 hyperoxia group, whereas that of phosphorylated Smad2/3 was oppositely increased. In addition, the expression of vascular endothelial growth factor (VEGF) mRNA was increased at D1 with subsequent decrease in D7 hyperoxia group. There was no significantly difference in endoglin expression in entire experimental period. Conclusion: These results indicate that exposure to hyperoxia altered the balance between TGF-β-ALK1-Smad1/5 and TGF-β-ALK5-Smad2/3 pathways in pulmonary endothelial cells, which may ultimately lead to the development of BPD.

S-Nitrosoglutathione Attenuates Airway Hyperresponsiveness in Murine Bronchopulmonary Dysplasia

Bronchopulmonary dysplasia (BPD) is characterized by lifelong obstructive lung disease and profound, refractory bronchospasm. It is observed among survivors of premature birth who have been treated with prolonged supplemental oxygen. Therapeutic options are limited. Using a neonatal mouse model of BPD, we show that hyperoxia increases activity and expression of a mediator of endogenous bronchoconstriction, S-nitrosoglutathione (GSNO) reductase. MicroRNA-342-3p, predicted in silico and shown in this study in vitro to suppress expression of GSNO reductase, was decreased in hyperoxia-exposed pups. Both pretreatment with aerosolized GSNO and inhibition of GSNO reductase attenuated airway hyperresponsiveness in vivo among juvenile and adult mice exposed to neonatal hyperoxia. Our data suggest that neonatal hyperoxia exposure causes detrimental effects on airway hyperreactivity through microRNA-342-3p–mediated upregulation of GSNO reductase expression. Furthermore, our data demonstrate that this adverse effect can be overcome by supplementing its substrate, GSNO, or by inhibiting the enzyme itself. Rates of BPD have not improved over the past two decades; nor have new therapies been developed. GSNO-based therapies are a novel treatment of the respiratory problems that patients with BPD experience.

Alveolar formation is dysregulated by restricted nutrition but not excess sedation in preterm lambs managed by noninvasive support.

Preterm birth and respiratory support with invasive mechanical ventilation frequently leads to bronchopulmonary dysplasia (BPD). A hallmark feature of BPD is alveolar simplification. For our preterm lamb model of BPD, invasive mechanical ventilation is associated with postnatal feeding intolerance (reduced nutrition) and sedation. In contrast, preterm lambs managed by noninvasive support (NIS) have normal alveolar formation, appropriate postnatal nutrition, and require little sedation. We used the latter, positive-outcome group to discriminate the contribution of reduced nutrition vs. sedation on alveolar simplification. We hypothesized that, restricted nutrition, but not sedation with pentobarbital, contributes to impaired indices of alveolar formation in preterm lambs managed by NIS. Preterm lambs managed by NIS for 21d were randomized into three groups: NIS control, NIS plus restricted nutrition, and NIS plus excess sedation with pentobarbital. We quantified morphological and biochemical indices of alveolar formation, as well as mesenchymal cell apoptosis and proliferation. Restricted nutrition impaired morphological and biochemical indices of alveolar formation, and reduced mesenchymal cell apoptosis and proliferation. Excess sedation with pentobarbital did not alter these indices, although mesenchymal cell apoptosis was less. Our results demonstrate that restricted nutrition, but not excess sedation, contributes to impaired alveolar formation during the evolution of BPD in chronically ventilated preterm lambs.

Effects of prolonged hyperoxia on pulmonary microvascular development and expression of Ehprin-B2 in neonatal mouse lungs

Objective To investigate the effects of long-term oxygen exposure on the pulmonary microvascular development and the expression of Ephrin-B2 of lungt issue in neonatal mice. Methods Forty-eight 2-day-old Kunming mice were randomly divided into hyperoxia group and air group, with 24 mice in each group. Mice in hyperoxia group were exposed to 70% oxygen to establish a model of bronchopulmonary dysplasia (BPD). Six mice from each group were sacrificed at 3, 7, 14 and 21 days of age, and lung tissue was collected for further test. The lung sections were stained with hematoxylin and eosin for histological evaluation, radial alveolar counts (RAC) and microvessel density (MVD) measurement by CD34 immunohistochemistry. Location and expression of Ephrin-B2 in lung tissue were measured by immunohistochemistry. Ephrin-B2 mRNA and protein levels were detected by fluorescent quantitative reverse transcriptase-polymerase chain reaction and Western blot, respectively. Two independent samples t-test was used for statistical analysis. Results (1) Pathological changes: The pathology of lung tissue in hyperoxia group showed typical BPD-like changes with advancing postnatal age, presenting mainly with simplified alveolar development and decreased microvessel number. Compared with the air group, RAC and MVD were significantly decreased in 14-day-old mice (6.067±0.432 vs 6.950±0.243, t=4.365, P<0.05; 4.133±0.476 vs 4.867±0.472, t=2.680, P<0.01) and 21-day-old mice in the hyperoxia group (8.050±0.362 vs 9.817±0.487, t=7.127, P<0.05; 4.333±0.532 vs 6.017±0.937, t=3.828, P<0.01). (2) Location and expression of Ephrin-B2: Ephrin-B2 was mainly expressed in alveolar epithelial cells, and weakly expressed in alveolar septum. Compared with the air group, the average optical density of Ephrin-B2 was significantly decreased in 7-day-old (0.146±0.013 vs 0.153±0.009), 14-day-old (0.140±0.007 vs 0.161±0.006) and 21-day-old mice in the hyperoxia group (0.138±0.008 vs 0.166±0.009)(t=-2.049, -9.442 and -10.087, all P<0.05). (3) Ephrin-B2 mRNA and protein levels: The Ephrin-B2 mRNA levels in 14-day-old (0.65±0.14 vs 1.05±0.16, t=4.609, P<0.01) and 21-day-old mice (0.57±0.09 vs 1.13±0.18, t=6.816, P<0.01) were significantly lower in hyperoxia group than in the air group. The Ephrin-B2 protein levels were also significantly lower in hyperoxia group than in the air group in 21-day-old mice (0.13±0.03 vs 0.29±0.08, t=4.587, P=0.000). Conclusions Oxygen-induced BPD model mice have simplified alveolar development, reduced MVD and decreased expression of Ephrin-B2 in lung tissue, which may play an important role in the pathogenesis of BPD. Key words: Bronchopulmonary dysplasia; Microvessels; Ephrin-B2; Disease models, animal

Phenotypic assessment of pulmonary hypertension using high-resolution echocardiography is feasible in neonatal mice with experimental bronchopulmonary dysplasia and pulmonary hypertension: a step toward preventing chronic obstructive pulmonary disease.

Bronchopulmonary dysplasia (BPD) and chronic obstructive pulmonary disease (COPD) are chronic lung diseases of human infants and adults, respectively, that are characterized by alveolar simplification. One-third of the infants with severe BPD develop pulmonary hypertension (PH). More importantly, PH increases morbidity and mortality in BPD patients. Additionally, COPD is a common respiratory morbidity in former BPD patients. The lack of an appropriate small animal model wherein echocardiography (Echo) can demonstrate PH is one of the major barriers to understand the molecular mechanisms of the disease and, thereby, develop rational therapies to prevent and/or treat PH in BPD patients. Thus, the goal of this study was to establish a model of experimental BPD and PH and investigate the feasibility of Echo to diagnose PH in neonatal mice. Since hyperoxia-induced oxidative stress and inflammation contributes to the development of BPD with PH, we tested the hypothesis that exposure of newborn C57BL/6J mice to 70% O2 (hyperoxia) for 14 days leads to lung oxidative stress, inflammation, alveolar and pulmonary vascular simplification, pulmonary vascular remodeling, and Echo evidence of PH. Hyperoxia exposure caused lung oxidative stress and inflammation as evident by increased malondialdehyde adducts and inducible nitric oxide synthase, respectively. Additionally, hyperoxia exposure caused growth restriction, alveolar and pulmonary vascular simplification, and pulmonary vascular remodeling. At 14 days of age, Echo of these mice demonstrated that hyperoxia exposure decreased pulmonary acceleration time (PAT) and PAT/ejection time ratio and increased right ventricular free wall thickness, which are indicators of significant PH. Thus, we have demonstrated the feasibility of Echo to phenotype PH in neonatal mice with experimental BPD with PH, which can aid in discovery of therapies to prevent and/or treat BPD with PH and its sequelae such as COPD in humans.

Protective effects of BMSCs in combination with erythropoietin in bronchopulmonary dysplasia-induced lung injury.

Bronchopulmonary dysplasia (BPD) is the most common type of chronic lung disease in infancy, for which no effective therapy is currently available. The aim of the present study was to investigate the effect of treatment with bone marrow mesenchymal stem cells (BMSCs) in combination with recombinant human erythropoietin (rHuEPO) on BPD‑induced mouse lung injury, and discuss the underlying mechanism. The BPD model was established by the exposure of neonatal mice to continuous high oxygen exposure for 14days, following which 1x106 BMSCs and 5,000U/kgrHuEPO were injected into the mice 1h prior to and 7days following exposure to hyperoxia. The animals received four treatments in total (n=10 in each group). After 14days, the body weights, airway structure, and levels of matrix metalloproteinase‑9 (MMP‑9) and vascular endothelial growth factor (VEGF) were detected using histological and immunohistochemical analyses. The effect on cell differentiation was observed by examining the presence of platelet endothelial cell adhesion molecule (PECAM) and VEGF using immunofluorescence. Compared with the administration of BMSCs alone, the body weight, airway structure, and the levels of MMP‑9 and VEGF were significantly improved in the BMSCs/rHuEPO group. The results of the present study demonstrated that the intravenous injection of BMSCs significantly improved lung damage in the hyperoxia‑exposed neonatal mouse model. Furthermore, the injection of BMSCs in combination with intraperitoneal injection of rHuEPO had a more marked effect, compared with BMSCs alone, and the mechanism may be mediated by the promoting effects of BMSCs and EPO. The results of the present study provided information, which may assist in future clinical trials.

MicroRNA in late lung development and bronchopulmonary dysplasia: the need to demonstrate causality.

MicroRNA are emerging as powerful regulators of cell differentiation and tissue and organ development. Several microRNA have been described to play a role in branching morphogenesis, a key step in early lung development. However, considerably less attention has been paid to microRNA as regulators of the process of secondary septation, which drives lung alveolarization during late lung development. Secondary septation is severely perturbed in bronchopulmonary dysplasia (BPD), a common complication of preterm birth characterized by blunted alveolarization. A number of studies to date have reported microRNA microarray screens in animal models of BPD; however, only two studies have attempted to demonstrate causality. Although the expression of miR-150 was altered in experimental BPD, a miR-150−/− knockout mouse did not exhibit appreciable protection in a BPD animal model. Similarly, while the expression of miR-489 in the lung was reduced in clinical and experimental BPD, antagomiR and over-expression approaches could not validate a role for miR-489 in the impaired alveolarization associated with experimental BPD. This mini-review aims to highlight microRNA that have been revealed by multiple microarray studies to be potential causal players in normal and pathological alveolarization. Additionally, the challenges faced in attempting to demonstrate a causal role for microRNA in lung alveolarization are discussed. These include the tremendous variability in the animal models employed, and the limitations and advantages offered by the available tools, including antagomiRs and approaches for the validation of a specific microRNA-mRNA interaction during lung alveolarization.

Mesenchymal stem cells in combination with erythropoietin repair hyperoxia-induced alveoli dysplasia injury in neonatal mice via inhibition of TGF-β1 signaling.

The aim of the present study is to investigate the protection effects of bone marrow mesenchymal stem cells (MSCs) in combination with EPO against hyperoxia-induced bronchopulmonary dysplasia (BPD) injury in neonatal mice. BPD model was prepared by continuous high oxygen exposure, 1×106 bone marrow MSCs and 5000U/kg recombinant human erythropoietin (EPO) were injected respectively. Results showed that administration of MSCs, EPO especially MSCs+EPO significant attenuated hyperoxia-induced lung damage with a decrease of fibrosis, radical alveolar counts and inhibition of the occurrence of epithelial-mesenchymal transition (EMT). Furthermore, MSCs+EPO co-treatment more significantly suppressed the levels of transforming growth factor-β1(TGF-β1) than MSCs or EPO alone. Collectively, these results suggested that MSCs, EPO in particular MSCs+EPO co-treatment could promote lung repair in hyperoxia-induced alveoli dysplasia injury via inhibition of TGF-β1 signaling pathway to further suppress EMT process and may be a promising therapeutic strategy.

Gene Expression Profiling Identifies Cell Proliferation and Inflammation as the Predominant Pathways Regulated by Aryl Hydrocarbon Receptor in Primary Human Fetal Lung Cells Exposed to Hyperoxia.

Exposure to hyperoxia contributes to the development of bronchopulmonary dysplasia (BPD) in premature infants. We observed that aryl hydrocarbon receptor (AhR) signaling protects newborn mice and primary fetal human pulmonary microvascular endothelial cells (HPMECs) against hyperoxic injury. Additionally, a recent genome-wide transcriptome study in a newborn mouse model of BPD identified AhR as a key regulator of hyperoxia-induced gene dysregulation. Whether the AhR similarly deregulates genes in HPMEC is unknown. Therefore, the objective of this study was to characterize transcriptome level gene expression profile in AhR-sufficient and -deficient HPMEC exposed to normoxic and hyperoxic conditions. Global gene expression profiling was performed using Illumina microarray platform and selected genes were validated by real-time RT-PCR. AhR gene expression and hyperoxia independently affected the expression of 540 and 593 genes, respectively. Two-way ANOVA further identified 85 genes that were affected by an interaction between AhR expression and exposure to hyperoxia. Kyoto Encyclopedia of Genes and Genomes (KEGG), Gene Ontology, and Reactome pathway analysis identified cell proliferation, immune function, cytokine signaling, and organ development as the major pathways affected in AhR-deficient cells. The biological processes that were significantly enriched by hyperoxia included metabolic process, stress response, signal transduction, cell cycle, and immune regulation. Cell cycle was the predominant pathway affected by the combined effect of AhR knockdown and hyperoxia. Functional analysis of cell cycle showed that AhR-deficient cells had decreased proliferation compared with AhR-sufficient cells. These findings suggest that AhR modulates hyperoxic lung injury by regulating the genes that are necessary for cell proliferation and inflammation.

Bronchopulmonary dysplasia impairs L-type amino acid transporter-1 expression in human and baboon lung.

Bronchopulmonary dysplasia (BPD) is an inflammatory lung disorder common in premature infants who undergo mechanical ventilation with supplemental oxygen. Inhaled nitric oxide (iNO) has been used to prevent experimental and clinical BPD. Earlier studies showed that NO effects in alveolar epithelial cells (AEC) are mediated by S-nitrosothiol uptake via L-type amino acid transporter-1 (LAT1). Because LAT1 expression could influence the efficacy of iNO therapy, we sought to determine whether pulmonary LAT1 expression is altered in preterm baboons with experimental BPD and in human newborns susceptible to developing BPD. Using fixed lung obtained from 125 d to 140 d gestation baboon models of BPD, LAT1 immunostaining was measured in control and BPD animals. In adult controls and in 140 d gestational controls (GC), LAT1 was expressed in both type I and type II AECs. In 140 d BPD lungs, LAT1 expression density in alveolar tissue was decreased. In 125 d GC baboons, LAT1 immunostaining was largely confined to cuboidal AECs, whereas animals given 14 d of mechanical ventilation exhibited diminished alveolar septal LAT1 Labeling. The pattern in adult human donor lung was comparable to that observed in adult baboons. LAT1 was expressed in lungs obtained from some but not all very premature newborns at autopsy. In human and baboon lung, adult and newborn, pulmonary vascular cells expressed LAT1. In summary, LAT1 is expressed in AECs and pulmonary vascular cells in baboons and humans. Experimental BPD in premature baboons decreases pulmonary LAT1 expression and alters its spatial localization. Heterogeneity of functional LAT1 could affect S-nitrosothiol importation, which could impair iNO therapy. Pediatr Pulmonol. 2016;51:1048-1056. © 2016 Wiley Periodicals, Inc.

Caffeine Prevents Hyperoxia-Induced Functional and Structural Lung Damage in Preterm Rabbits

Background: Caffeine is a commonly used drug for apnea of prematurity. It may, however, also have a beneficial effect on bronchopulmonary dysplasia (BPD), which is the most common complication of extreme preterm birth. Objectives: To study the inflammatory, structural and functional effects of caffeine in an animal model of BPD. Methods: Preterm New Zealand-Dendermonde rabbits (gestational day 28; term 31) were randomized to three groups: normoxia-placebo (N-P), hyperoxia-placebo (H-P) and hyperoxia-caffeine (H-C). Lung function was assessed on postnatal day 5, along with airway morphometry, vascular morphometry and a score observing airway inflammation. Results: Caffeine improved lung function by increasing lung volume [mean displaced volume N-P: 40.1 ± 6 ml/kg, H-P: 27.8 ± 8 ml/kg and H-C: 34.4 ± 7 ml/kg (p < 0.05); total lung capacity: N-P: 1.17 ± 0.1 ml, H-P: 0.67 ± 0.1 ml and H-C: 1.1 ± 0.1 ml (p < 0.05)], decreasing tissue damping [N-P: 2.7 ± 0.3 cm H₂O/ml, H-P: 4.6 ± 0.6 cm H₂O/ml and H-C: 3.2 ± 0.4 cm H₂O/ml (p < 0.05)], elastance [N-P: 9.3 ± 2.4 cm H₂O/ml, H-P: 19.2 ± 7.4 cm H₂O/ml and H-C: 10.7 ± 2 cm H₂O/ml (p < 0.05)] and compliance [N-P: 0.06 ± 0.01 cm H₂O/ml, H-P: 0.054 ± 0.01 cm H₂O/ml and H-C: 0.07 ± 0.013 cm H₂O/ml (p < 0.05)]. Caffeine also improved histology by decreasing alveolar size [linear intercepts; N-P: 83.6 ± 1.7, H-P: 82.9 ± 1.6 and H-C: 67.3 ± 1.4 (p < 0.05)], increasing radial alveolar count (N-P: 6.6 ± 0.5, H-P: 5.7 ± 0.6 and H-C: 7.05 ± 0.5) and decreasing the acute inflammation score [N-P: 0.3 ± 0.1, H-P: 0.5 ± 0.1 and H-C: 0.4 ± 0.1 (p < 0.05)]. Conclusion: In preterm rabbits, caffeine reduces the functional, architectural and inflammatory pulmonary changes induced by hyperoxia in the lung.

Fractal analysis of alveolarization in hyperoxia-induced rat models of bronchopulmonary dysplasia.

No papers are available about potentiality of fractal analysis in quantitative assessment of alveolarization in bronchopulmonary dysplasia (BPD). Thus, we here performed a comparative analysis between fractal [fractal dimension (D) and lacunarity] and stereological [mean linear intercept (Lm), total volume of alveolar air spaces, total number of alveoli, mean alveolar volume, total volume and surface area of alveolar septa, and mean alveolar septal thickness] parameters in experimental hyperoxia-induced models of BPD. At birth, rats were distributed between the following groups: 1) rats raised in ambient air for 2 wk; 2) rats exposed to 60% oxygen for 2 wk; 3) rats raised in normoxia for 6 wk; and 4) rats exposed to 60% hyperoxia for 2 wk and to room air for further 4 wk. Normoxic 6-wk rats showed increased D and decreased lacunarity with respect to normoxic 2-wk rats, together with changes in all stereological parameters except for mean alveolar volume. Hyperoxia-exposed 2-wk rats showed significant changes only in total number of alveoli, mean alveolar volume, and lacunarity with respect to equal-in-age normoxic rats. In the comparison between 6-wk rats, the hyperoxia-exposed group showed decreased D and increased lacunarity, together with changes in all stereological parameters except for septal thickness. Analysis of receiver operating characteristic curves showed a comparable discriminatory power of D, lacunarity, and total number of alveoli; Lm and mean alveolar volume were less discriminative. D and lacunarity did not show significant changes when different segmentation thresholds were applied, suggesting that the fractal approach may be fit to automatic image analysis.

Hydrogen-rich water ameliorates bronchopulmonary dysplasia (BPD) in newborn rats.

Bronchopulmonary dysplasia (BPD) is characterized by developmental arrest of the alveolar tissue. Oxidative stress is causally associated with development of BPD. The effects of hydrogen have been reported in a wide range of disease models and human diseases especially caused by oxidative stress. We made a rat model of BPD by injecting lipopolysaccharide (LPS) into the amniotic fluid at E16.5. The mother started drinking hydrogen-rich water from E9.5 and also while feeding milk. Hydrogen normalized LPS-induced abnormal enlargement of alveoli at P7 and P14. LPS increased staining for nitrotyrosine and 8-OHdG of the lungs, and hydrogen attenuated the staining. At P1, LPS treatment decreased expressions of genes for FGFR4, VEGFR2, and HO-1 in the lungs, and hydrogen increased expressions of these genes. In contrast, LPS treatment and hydrogen treatment had no essential effect on the expression of SOD1. Inflammatory marker proteins of TNFα and IL-6 were increased by LPS treatment, and hydrogen suppressed them. Treatment of A549 human lung adenocarcinoma epithelial cells with 10% hydrogen gas for 24 hr decreased production of reactive oxygen species in both LPS-treated and untreated cells. Lack of any known adverse effects of hydrogen makes hydrogen a promising therapeutic modality for BPD. Pediatr Pulmonol. 2016; 51:928-935. © 2016 Wiley Periodicals, Inc.

Standardisation of oxygen exposure in the development of mouse models for bronchopulmonary dysplasia.

ABSTRACTProgress in developing new therapies for bronchopulmonary dysplasia (BPD) is sometimes complicated by the lack of a standardised animal model. Our objective was to develop a robust hyperoxia-based mouse model of BPD that recapitulated the pathological perturbations to lung structure noted in infants with BPD. Newborn mouse pups were exposed to a varying fraction of oxygen in the inspired air (FiO2) and a varying window of hyperoxia exposure, after which lung structure was assessed by design-based stereology with systemic uniform random sampling. The efficacy of a candidate therapeutic intervention using parenteral nutrition was evaluated to demonstrate the utility of the standardised BPD model for drug discovery. An FiO2 of 0.85 for the first 14 days of life decreased total alveoli number and concomitantly increased alveolar septal wall thickness, which are two key histopathological characteristics of BPD. A reduction in FiO2 to 0.60 or 0.40 also caused a decrease in the total alveoli number, but the septal wall thickness was not impacted. Neither a decreasing oxygen gradient (from FiO2 0.85 to 0.21 over the first 14 days of life) nor an oscillation in FiO2 (between 0.85 and 0.40 on a 24 h:24 h cycle) had an appreciable impact on lung development. The risk of missing beneficial effects of therapeutic interventions at FiO2 0.85, using parenteral nutrition as an intervention in the model, was also noted, highlighting the utility of lower FiO2 in selected studies, and underscoring the need to tailor the model employed to the experimental intervention. Thus, a state-of-the-art BPD animal model that recapitulates the two histopathological hallmark perturbations to lung architecture associated with BPD is described. The model presented here, where injurious stimuli have been systematically evaluated, provides a most promising approach for the development of new strategies to drive postnatal lung maturation in affected infants.

The Extracellular Matrix in Bronchopulmonary Dysplasia: Target and Source.

Bronchopulmonary dysplasia (BPD) is a common complication of preterm birth that contributes significantly to morbidity and mortality in neonatal intensive care units. BPD results from life-saving interventions, such as mechanical ventilation and oxygen supplementation used to manage preterm infants with acute respiratory failure, which may be complicated by pulmonary infection. The pathogenic pathways driving BPD are not well-delineated but include disturbances to the coordinated action of gene expression, cell–cell communication, physical forces, and cell interactions with the extracellular matrix (ECM), which together guide normal lung development. Efforts to further delineate these pathways have been assisted by the use of animal models of BPD, which rely on infection, injurious mechanical ventilation, or oxygen supplementation, where histopathological features of BPD can be mimicked. Notable among these are perturbations to ECM structures, namely, the organization of the elastin and collagen networks in the developing lung. Dysregulated collagen deposition and disturbed elastin fiber organization are pathological hallmarks of clinical and experimental BPD. Strides have been made in understanding the disturbances to ECM production in the developing lung, but much still remains to be discovered about how ECM maturation and turnover are dysregulated in aberrantly developing lungs. This review aims to inform the reader about the state-of-the-art concerning the ECM in BPD, to highlight the gaps in our knowledge and current controversies, and to suggest directions for future work in this exciting and complex area of lung development (patho)biology.

Efficacy of Leukadherin-1 in the Prevention of Hyperoxia-Induced Lung Injury in Neonatal Rats.

Lung inflammation plays a key role in the pathogenesis of bronchopulmonary dysplasia (BPD), a chronic lung disease of premature infants. The challenge in BPD management is the lack of effective and safe antiinflammatory agents. Leukadherin-1 (LA1) is a novel agonist of the leukocyte surface integrin CD11b/CD18 that enhances leukocyte adhesion to ligands and vascular endothelium and thus reduces leukocyte transendothelial migration and influx to the injury sites. Its functional significance in preventing hyperoxia-induced neonatal lung injury is unknown. We tested the hypothesis that administration of LA1 is beneficial in preventing hyperoxia-induced neonatal lung injury, an experimental model of BPD. Newborn rats were exposed to normoxia (21% O2) or hyperoxia (85% O2) and received twice-daily intraperitoneal injection of LA1 or placebo for 14 days. Hyperoxia exposure in the presence of the placebo resulted in a drastic increase in the influx of neutrophils and macrophages into the alveolar airspaces. This increased leukocyte influx was accompanied by decreased alveolarization and angiogenesis and increased pulmonary vascular remodeling and pulmonary hypertension (PH), the pathological hallmarks of BPD. However, administration of LA1 decreased macrophage infiltration in the lungs during hyperoxia. Furthermore, treatment with LA1 improved alveolarization and angiogenesis and decreased pulmonary vascular remodeling and PH. These data indicate that leukocyte recruitment plays an important role in the experimental model of BPD induced by hyperoxia. Targeting leukocyte trafficking using LA1, an integrin agonist, is beneficial in preventing lung inflammation and protecting alveolar and vascular structures during hyperoxia. Thus, targeting integrin-mediated leukocyte recruitment and inflammation may provide a novel strategy in preventing and treating BPD in preterm infants.

A three-dimensional study of alveologenesis in mouse lung

Alveologenesis is the final step of lung maturation, which subdivides the alveolar region of the lung into smaller units called alveoli. Each of the nascent dividers serves as a new gas-exchange surface, and collectively they drastically increase the surface area for breathing. Disruption of alveologenesis results in simplification of alveoli, as is seen in premature infants diagnosed with bronchopulmonary dysplasia (BPD), a prevalent lung disease that is often associated with lifelong breathing deficiencies. To date, a majority of studies of alveologenesis rely on two-dimensional (2D) analysis of tissue sections. Given that an overarching theme of alveologenesis is thinning and extension of the epithelium and mesenchyme to facilitate gas exchange, often only a small portion of a cell or a cellular structure is represented in a single 2D plane. Here, we use a three-dimensional (3D) approach to examine the structural architecture and cellular composition of myofibroblasts, alveolar type 2 cells, elastin and lipid droplets in normal as well as BPD-like mouse lung. We found that 2D finger-like septal crests, commonly used to depict growing alveolar septae, are often artifacts of sectioning through fully established alveolar walls. Instead, a more accurate representation of growing septae are 3D ridges that are lined by platelet-derived growth factor receptor alpha (PDGFRA) and alpha smooth muscle actin (α-SMA)-expressing myofibroblasts, as well as the elastin fibers that they produce. Accordingly in 3D, both α-SMA and elastin were each found in connected networks underlying the 3D septal ridges rather than as isolated dots at the tip of 2D septal crests. Analysis through representative stages of alveologenesis revealed unappreciated dynamic changes in these patterns. PDGFRA-expressing cells are only α-SMA-positive during the first phase of alveologenesis, but not in the second phase, suggesting that the two phases of septae formation may be driven by distinct mechanisms. Thin elastin fibers are already present in the alveolar region prior to alveologenesis, suggesting that during alveologenesis, there is not only new elastin deposition, but also extensive remodeling to transform thin and uniformly distributed fibers into thick cables that rim the nascent septae. Analysis of several genetic as well as hyperoxia-induced models of BPD revealed that the myofibroblast organization is perturbed in all, regardless of whether the origin of defect is epithelial, mesenchymal, endothelial or environmental. Finally, analysis of relative position of PDGFRA-positive cells and alveolar type 2 cells reveal that during alveologenesis, these two cell types are not always adjacent to one another. This result suggests that the niche and progenitor relationship afforded by their close juxtaposition in the adult lung may be a later acquired property. These insights revealed by 3D reconstruction of the septae set the foundation for future investigations of the mechanisms driving normal alveologenesis, as well as causes of alveolar simplification in BPD.

Neonatal bronchopulmonary dysplasia increases neuronal apoptosis in the hippocampus through the HIF-1α and p53 pathways

Neonatal bronchopulmonary dysplasia (BPD) might lead to an increased risk for brain injury. The present study aims to investigate the effects of neonatal BPD on neuronal apoptosis in the hippocampus and cognitive function and to explore the underlying mechanisms. The results revealed that BPD model rat pups exhibited more apoptotic cells in the hippocampus and longer escape latencies in the Morris maze test. Both the caspase-dependent and caspase-nondependent signal pathways were activated. Further examinations showed an elevated p53 level by BPD via HIF-1α induction, while the caspase-3 in the hippocampus was suppressed by both HIF-1α and p53 inhibitor. These findings suggested that neonatal BPD caused impaired cognitive function and neuron apoptosis in hippocampus via p53 and HIF-1α. Although the precise mechanism requires further investigation, this study provided new evidence for and an explanation of the impaired CNS developmental outcomes of BPD.

The Effect of Continuous Positive Airway Pressure in a Mouse Model of Hyperoxic Neonatal Lung Injury

Background: Continuous positive airway pressure (CPAP) and supplemental oxygen have become the mainstay of neonatal respiratory support in preterm infants. Although oxygen therapy is associated with respiratory morbidities including bronchopulmonary dysplasia (BPD), the long-term effects of CPAP on lung function are largely unknown. We used a hyperoxia-induced mouse model of BPD to explore the effects of daily CPAP in the first week of life on later respiratory system mechanics. Objective: We wanted to test the hypothesis that daily CPAP in a newborn-mouse model of BPD improves longer-term respiratory mechanics. Methods: Mouse pups from C57BL/6 pregnant dams were exposed to room air (RA) or hyperoxia (50% O₂, 24 h/day) for the first postnatal week with or without exposure to daily CPAP (6 cm H₂O, 3 h/day). Respiratory system resistance (Rrs) and compliance (Crs) were measured following a subsequent 2-week period of RA recovery. Additional measurements included radial alveolar and macrophage counts. Results: Mice exposed to hyperoxia had significantly elevated Rrs, decreased Crs, reduced alveolarization and increased macrophage counts at 3 weeks when compared to RA-treated mice. Daily CPAP treatment significantly improved Rrs, Crs and alveolarization and decreased lung macrophage infiltration in the hyperoxia-exposed pups. Conclusions: We have demonstrated that daily CPAP had a longer-term benefit on baseline respiratory system mechanics in a neonatal mouse model of BPD. We speculate that this beneficial effect of CPAP was the consequence of a decrease in the inflammatory response and resultant alveolar injury associated with hyperoxic lung injury in newborns.