Developing Mouse Lung Research Articles

Gestational vitamin D deficiency impairs fetal lung development through suppressing type II pneumocyte differentiation

Gestational vitamin D deficiency is associated with pulmonary diseases. This study aimed to investigate the effect of gestational vitamin D deficiency on fetal lung development in mice. Absolute and relative weights of fetal lungs were reduced in vitamin D deficient (VDD) group. Incrassate mesenchyme, measured by septal wall thickness, accompanied by lessened saccular space, was shown in VDD group. Numerous immature type II pneumocytes, as determined by PAS staining, were observed in VDD group. Moreover, increased Ki67-positive cells, a marker of cell proliferation, was detected in VDD group. The additional experiments showed that Sftpa, Sftpb, Sftpc and Sftpd, four surfactant genes, were downregulated and pro–surfactant protein B was reduced in VDD group. FoxA1, FoxA2 and TTF-1, three transcription factors that regulate surfactant genes, and VEGF, a key regulator for pulmonary maturation, were downregulated in VDD group. These results suggest that gestational vitamin D deficiency impairs fetal lung development partially through suppressing type II pneumocyte differentiation.

Lung developmental arrest caused by PDGF-A deletion: consequences for the adult mouse lung.

PDGF-A is a key contributor to lung development in mice. Its expression is needed for secondary septation of the alveoli and deletion of the gene leads to abnormally enlarged alveolar air spaces in mice. In humans, the same phenotype is the hallmark of bronchopulmonary dysplasia (BPD), a disease that affects premature babies and may have long lasting consequences in adulthood. So far, the knowledge regarding adult effects of developmental arrest in the lung is limited. This is attributable to few follow-up studies of BPD survivors and lack of good experimental models that could help predict the outcomes of this early age disease for the adult individual. In this study, we used the constitutive lung-specific Pdgfa deletion mouse model to analyze the consequences of developmental lung defects in adult mice. We assessed lung morphology, physiology, cellular content, ECM composition and proteomics data in mature mice, that perinatally exhibited lungs with a BPD-like morphology. Histological and physiological analyses both revealed that enlarged alveolar air spaces remained until adulthood, resulting in higher lung compliance and higher respiratory volume in knockout mice. Still, no or only small differences were seen in cellular, ECM and protein content when comparing knockout and control mice. Taken together, our results indicate that Pdgfa deletion-induced lung developmental arrest has consequences for the adult lung at the morphological and functional level. In addition, these mice can reach adulthood with a BPD-like phenotype, which makes them a robust model to further investigate the pathophysiological progression of the disease and test putative regenerative therapies.

Circular RNAs are abundant and dynamically expressed during the embryonic lung development of C57BL/6 mice

Circular RNAs (circRNAs), a novel type of endogenous RNAs, can function as microRNA (miRNA) sponges capable of regulating gene transcription, binding to RNA-associated proteins, and even encoding proteins. CircRNAs are involved in various cell behaviors, such as proliferation and apoptosis. The mouse model has also been demonstrated to be similar to that of humans in many studies. To explore the profile of circRNAs during embryonic lung development and their potential functions in lung development-related diseases, mouse embryos at the pseudoglandular phase, canalicular phase, saccular phase, and alveolar phase were collected. High-throughput sequencing was then used to identify a total of 1,735 circRNAs (junction reads ≥5 and p < 0.05). It is well known that the functions of circRNAs are related to host genes. In our study, bioinformatics analysis indicated that the screened host genes were closely associated with lung development and included the Hippo signaling pathway, PI3K-Akt signaling pathways, and TGF-β signaling pathways. Moreover, miRNA sponges are another mechanism involved in lung development. Therefore, we predicted many miRNAs binding to circRNAs, such as miR-17 and miR-20, using the TargetScan and miRanda databases. Previously, miRNAs were proven to be necessary for lung development. The peak expression of circRNAs is distributed at different time points, suggesting their involvement in different stages of embryonic mouse lung development.

In utero exposures to electronic-cigarette aerosols impair the Wnt signaling during mouse lung development.

Currently, more than 9 million American adults, including women of childbearing age, use electronic-cigarettes (e-cigs). Further, the prevalence of maternal vaping now approaching 10% is similar to that of maternal smoking. Little, however, is known about the effects of fetal exposures to nicotine-rich e-cig aerosols on lung development. In this study, we assessed whether in utero exposures to e-cig aerosols compromised lung development in mice. A third-generation e-cig device was used to expose pregnant BALB/c mice by inhalation to 36 mg/mL of nicotine cinnamon-flavored e-cig aerosols for 14-31 days. This included exposures for either 12 days before mating plus during gestation (preconception groups) or only during gestation (prenatal groups). Respective control mice were exposed to filtered air. Subgroups of offspring were euthanized at birth or at 4 wk of age. Compared with respective air-exposed controls, both preconception and prenatal exposures to e-cig aerosols significantly decreased the offspring birth weight and body length. In the preconception group, 7 inflammation-related genes were downregulated, including 4 genes common to both dams and fetuses, denoting an e-cig immunosuppressive effect. Lung morphometry assessments of preconception e-cig-exposed offspring showed a significantly increased tissue fraction at birth. This result was supported by the downregulation of 75 lung genes involved in the Wnt signaling, which is essential to lung organogenesis. Thus, our data indicate that maternal vaping impairs pregnancy outcomes, alters fetal lung structure, and dysregulates the Wnt signaling. This study provides experimental evidence for future regulations of e-cig products for pregnant women and developmentally vulnerable populations.

Simultaneous isolation of endothelial and alveolar epithelial type I and type II cells during mouse lung development in the absence of a transgenic reporter.

Mouse lung developmental maturation and final alveolarization phase begin at birth. During this dynamic process, alveolar cells modify their morphology and anchorage to the extracellular matrix. In particular, alveolar epithelial cell (AEC) type I undergo cytoplasmic flattening and folding to ensure alveoli lining. We developed FACS conditions for simultaneous isolation of alveolar epithelial and endothelial cells in the absence of specific reporters during the early and middle alveolar phase. We evidenced for the first time a pool of extractable epithelial cell populations expressing high levels of podoplanin at postnatal day (pnd)2, and we confirmed by RT-qPCR that these cells are already differentiated but still immature AEC type I. Maturation causes a decrease in isolation yields, reflecting the morphological changes that these cell populations are undergoing. Moreover, we find that major histocompatibility complex II (MHCII), reported as a good marker of AEC type II, is poorly expressed at pnd2 but highly present at pnd8. Combined experiments using LysoTracker and MHCII demonstrate the de novo acquisition of MCHII in AEC type II during lung alveolarization. The lung endothelial populations exhibit FACS signatures from vascular and lymphatic compartments. They can be concomitantly followed throughout alveolar development and were obtained with a noticeable increased yield at the last studied time point (pnd16). Our results provide new insights into early lung alveolar cell isolation feasibility and represent a valuable tool for pure AEC type I preparation as well as further in vitro two- and three-dimensional studies.

Effect of Intranasal Instillation of Lipopolysaccharide on Lung Development and Its Related Mechanism in Newborn Mice.

Premature infants are prone to repeated lung infections after birth, which can disrupt the development of lung structure and function. However, the effects of postnatal pulmonary inflammation on lung development in newborn mice have not been reported and may play an important role in the development of bronchopulmonary dysplasia (BPD). This study aimed to establish a BPD model of postnatal pulmonary inflammation in premature infants and to explore its role and possible mechanisms in the pathogenesis of BPD. We exposed postnatal day 1 mice to lipopolysaccharide (LPS) and normal saline for 14 days. Pulmonary inflammation and alveolar microvascular development were assessed by histology. In addition, we also examined the expression of vascular endothelial growth factor (VEGF), VEGFR2, nuclear factor-kappa-B (NF-κB) and related inflammatory mediators [interleukin-1β (IL-1β), tumor necrosis factor-alpha (TNF-α), macrophage inflammatory protein-1α (MIP-1α), monocyte chemoattractant protein-1 (MCP-1)] in the lungs. Lung histology revealed inflammatory cell infiltration, alveolar simplification, and decreased microvascular density in LPS-exposed lungs. VEGF and VEGFR2 expression was decreased in the lungs of LPS-exposed neonatal mice. Furthermore, we detected elevated levels of the inflammatory mediators IL-1β, TNF-α, MIP-1α, and MCP-1 in the lungs, which are associated with the activation of NF-κB. Intranasal instillation of LPS inhibits lung development in newborn mice, and postnatal pulmonary inflammation may participate in the pathogenesis of BPD. The mechanism is related to the inhibition of VEGF and VEGFR2 and the upregulation of inflammatory mediators through activation of NF-κB.

Mesenchyme-specific deletion of Tgf-β1 in the embryonic lung disrupts branching morphogenesis and induces lung hypoplasia

Proper lung development depends on the precise temporal and spatial expression of several morphogenic factors, including Fgf10, Fgf9, Shh, Bmp4, and Tgf-β. Over- or under-expression of these molecules often leads to aberrant embryonic or postnatal lung development. Herein, we deleted the Tgf-β1 gene specifically within the lung embryonic mesenchymal compartment at specific gestational stages to determine the contribution of this cytokine to lung development. Mutant embryos developed severe lung hypoplasia and died at birth due to the inability to breathe. Despite the markedly reduced lung size, proliferation and differentiation of the lung epithelium was not affected by the lack of mesenchymal expression of the Tgf-β1 gene, while apoptosis was significantly increased in the mutant lung parenchyma. Lack of mesenchymal expression of the Tgf-β1 gene was also associated with reduced lung branching morphogenesis, with accompanying inhibition of the local FGF10 signaling pathway as well as abnormal development of the vascular system. To shed light on the mechanism of lung hypoplasia, we quantified the phosphorylation of 226 proteins in the mutant E12.5 lung compared with control. We identified five proteins, Hrs, Vav2, c-Kit, the regulatory subunit of Pi3k (P85), and Fgfr1, that were over- or under-phosphorylated in the mutant lung, suggesting that they could be indispensable effectors of the TGF-β signaling program during embryonic lung development. In conclusion, we have uncovered novel roles of the mesenchyme-specific Tgf-β1 ligand in embryonic mouse lung development and generated a mouse model that may prove helpful to identify some of the key pathogenic mechanisms underlying lung hypoplasia in humans.

Mitochondrial DNA variation modulates alveolar development in newborn mice exposed to hyperoxia.

Hyperoxia-induced oxidant stress contributes to the pathogenesis of bronchopulmonary dysplasia (BPD) in preterm infants. Mitochondrial functional differences due to mitochondrial DNA (mtDNA) variations are important modifiers of oxidant stress responses. The objective of this study was to determine whether mtDNA variation independently modifies lung development and mechanical dysfunction in newborn mice exposed to hyperoxia. Newborn C57BL6 wild type (C57n/C57mt, C57WT) and C3H/HeN wild type (C3Hn/C3Hmt, C3HWT) mice and novel Mitochondrial-nuclear eXchange (MNX) strains with nuclear DNA (nDNA) from their parent strain and mtDNA from the other-C57MNX (C57n/C3Hmt) and C3HMNX (C3Hn/C57mt)-were exposed to 21% or 85% O2 from birth to postnatal day 14 (P14). Lung mechanics and histopathology were examined on P15. Neonatal mouse lung fibroblast (NMLF) bioenergetics and mitochondrial superoxide (O2-) generation were measured. Pulmonary resistance and mitochondrial O2- generation were increased while alveolarization, compliance, and NMLF basal and maximal oxygen consumption rate were decreased in hyperoxia-exposed C57WT mice (C57n/C57mt) versus C57MNX mice (C57n/C3Hmt) and in hyperoxia-exposed C3HMNX mice (C3Hn/C57mt) versus C3HWT (C3Hn/C3Hmt) mice. Our study suggests that neonatal C57 mtDNA-carrying strains have increased hyperoxia-induced hypoalveolarization, pulmonary mechanical dysfunction, and mitochondrial bioenergetic and redox dysfunction versus C3H mtDNA strains. Therefore, mtDNA haplogroup variation-induced differences in mitochondrial function could modify neonatal alveolar development and BPD susceptibility.

Adverse early-life environment impairs postnatal lung development in mice.

Fetal growth restriction (FGR) is a major risk factor for bronchopulmonary dysplasia (BPD). Maternal stress and poor diet are linked to FGR. Effect of perinatal stress on lung development remains unknown. Using a murine model of adverse early life environment (AELE), we hypothesized that maternal exposure to perinatal environmental stress and high-fat diet (Western diet) lead to impaired lung development in the offspring. Female mice were placed on either control diet or Western diet before conception. Those exposed to Western diet were also exposed to perinatal environmental stress, the combination referred to as AELE. Pups were either euthanized at postnatal day 21 (P21) or weaned to control diet and environment until adulthood (8-14 wk old). Lungs were harvested for histology, gene expression by quantitative RT-PCR, microRNA profiling, and immunoblotting. AELE increased the mean linear intercept and decreased the radial alveolar count and secondary septation in P21 and adult mice. Capillary count was also decreased in P21 and adult mice. AELE lungs had decreased vascular endothelial growth factor A (VEGFA), VEGF receptor 2, endothelial nitric oxide synthase, and hypoxia inducible factor-1α protein levels and increased expression of genes that regulate DNA methylation and upregulation of microRNAs that target genes involved in lung development at P21. AELE leads to impaired lung alveolar and vascular growth, which persists into adult age despite normalizing the diet and environment at P21. AELE also alters the expression of genes involved in lung remodeling.

Autophagy is required for lung development and morphogenesis.

Bronchopulmonary dysplasia (BPD) remains a major respiratory illness in extremely premature infants. The biological mechanisms leading to BPD are not fully understood, although an arrest in lung development has been implicated. The current study aimed to investigate the occurrence of autophagy in the developing mouse lung and its regulatory role in airway branching and terminal sacculi formation. We found 2 windows of epithelial autophagy activation in the developing mouse lung, both resulting from AMPK activation. Inhibition of AMPK-mediated autophagy led to reduced lung branching in vitro. Conditional deletion of beclin 1 (Becn1) in mouse lung epithelial cells (Becn1Epi-KO), either at early (E10.5) or late (E16.5) gestation, resulted in lethal respiratory distress at birth or shortly after. E10.5 Becn1Epi-KO lungs displayed reduced airway branching and sacculi formation accompanied by impaired vascularization, excessive epithelial cell death, reduced mesenchymal thinning of the interstitial walls, and delayed epithelial maturation. E16.5 Becn1Epi-KO lungs had reduced terminal air sac formation and vascularization and delayed distal epithelial differentiation, a pathology similar to that seen in infants with BPD. Taken together, our findings demonstrate that intrinsic autophagy is an important regulator of lung development and morphogenesis and may contribute to the BPD phenotype when impaired.

Insights into the Signal Transduction Pathways of Mouse Lung Type II Cells Revealed by Transcription Factor Profiling in the Transcriptome.

Alveolar type II cells constitute a small fraction of the total lung cell mass. However, they play an important role in many cellular processes including trans-differentiation into type I cells as well as repair of lung injury in response to toxic chemicals and respiratory pathogens. Transcription factors are the regulatory proteins dynamically modulating DNA structure and gene expression. Transcription factor profiling in microarray datasets revealed that several members of AP1, ATF, NF-kB, and C/EBP families involved in diverse responses were expressed in mouse lung type II cells. A transcriptional factor signature consisting of Cebpa, Srebf1, Stat3, Klf5, and Elf3 was identified in lung type II cells, Sox9+ pluripotent lung stem cells as well as in mouse lung development. Identification of the transcription factor profile in mouse lung type II cells will serve as a useful resource and facilitate the integrated analysis of signal transduction pathways and specific gene targets in a variety of physiological conditions.

A critical role for miR-142 in alveolar epithelial lineage formation in mouse lung development.

The respiratory epithelium arises from alveolar epithelial progenitors which differentiate into alveolar epithelial type 1 (AT1) and type 2 (AT2) cells. AT2 cells are stem cells in the lung critical for the repair process after injury. Mechanisms regulating AT1 and AT2 cell maturation are poorly defined. We report that the activation of the glucocorticoid pathway in an in vitro alveolar epithelial lineage differentiation assay led to increased AT2 marker Sftpc and decreased miR-142 expression. Using miR-142 KO mice, we demonstrate an increase in the AT2/AT1 cell number ratio. Overexpression of miR-142 in alveolar progenitor cells in vivo led to the opposite effect. Examination of the KO lungs at E18.5 revealed enhanced expression of miR-142 targets Apc, Ep300 and Kras associated with increased β-catenin and p-Erk signaling. Silencing of miR-142 expression in lung explants grown in vitro triggers enhanced Sftpc expression as well as increased AT2/AT1 cell number ratio. Pharmacological inhibition of Ep300-β-catenin but not Erk in vitro prevented the increase in Sftpc expression triggered by loss of miR-142. These results suggest that the glucocorticoid-miR-142-Ep300-β-catenin signaling axis controls pneumocyte maturation.

FGF Signaling in Lung Development and Disease: Human Versus Mouse.

Fibroblast growth factor 10 (FGF10) plays an important role in mouse lung development, injury, and repair. It is considered the main morphogen driving lung branching morphogenesis in rodents. While many studies have found FGF10 SNPs associated with COPD and branch variants in COPD smokers, there is no evidence of a causative role for FGF10 or these SNPs in human lung development and pediatric lung diseases. We and others have shown divergent roles for FGF10 in mouse lung development and early human lung development. Herein, we only review the existing literature on FGF signaling in human lung development and pediatric human lung diseases, comparing what is known in mouse lung to that in human lung.

Smooth muscle differentiation shapes domain branches during mouse lung development.

During branching morphogenesis, a simple cluster of cells proliferates and branches to generate an arborized network that facilitates fluid flow. The overall architecture of the mouse lung is established by domain branching, wherein new branches form laterally off the side of an existing branch. The airway epithelium develops concomitantly with a layer of smooth muscle that is derived from the embryonic mesenchyme. Here, we examined the role of smooth muscle differentiation in shaping emerging domain branches. We found that the position and morphology of domain branches are highly stereotyped, as is the pattern of smooth muscle that differentiates around the base of each branch. Perturbing the pattern of smooth muscle differentiation genetically or pharmacologically causes abnormal domain branching. Loss of smooth muscle results in ectopic branching and decreases branch stereotypy. Increased smooth muscle suppresses branch initiation and extension. Computational modeling revealed that epithelial proliferation is insufficient to generate domain branches and that smooth muscle wrapping is required to shape the epithelium into a branch. Our work sheds light on the physical mechanisms of branching morphogenesis in the mouse lung.

Control Interventions Can Impact Alveolarization and the Transcriptome in Developing Mouse Lungs.

There is currently much interest in understanding the mechanisms of normal and aberrant lung alveolarization, particularly in the context of bronchopulmonary dysplasia, a common complication of preterm birth where alveolarization is impeded. To this end, the parenteral administration of pharmacological agents that modulate biochemical pathways, or facilitate modulation of gene expression in transgenic animals, has facilitated the discovery and validation of mechanisms that direct lung development. Such studies include control interventions, where the solvent vehicle, perhaps containing an inactive form of the agent applied, is administered; thereby providing a well-controlled point of reference for the analysis of the partner experiment. In the present study, the impact of several widely used control interventions in developing C57Bl/6J mouse pups was examined for effects on lung structure and the lung transcriptome. Parenteral administration of scrambled microRNA inhibitors (called antagomiRs) that are used to control in vivo microRNA neutralization studies, impacted lung volume, septal thickness, and the transcriptome of developing mouse lungs; with some effects dependent upon nucleotide sequence. Repeated intraperitoneal isotonic saline injections altered lung volume, with limited impact on the transcriptome. Parenteral administration of the tamoxifen solvent Miglyol accelerated mouse pup growth, and changed the abundance of 73 mRNA transcripts in the lung. Tamoxifen applied in Miglyol-in the absence of Cre recombinase-decreased pup growth, lung volume, and lung alveolarization and changed the abundance of 298 mRNA transcripts in the lung. These data demonstrate that widely used control interventions can directly impact lung alveolarization and the lung transcriptome in studies on lung development. Anat Rec, 302:346-363, 2019. © 2018 Wiley Periodicals, Inc.

Glucocorticoid receptor-mediated signalling inhibits mesenchymal cell proliferation via repression of the V1 isoform of versican during mouse lung development

Glucocorticoid receptor-mediated signalling inhibits mesenchymal cell proliferation via repression of the V1 isoform of versican during mouse lung development

Myh10 deficiency leads to defective extracellular matrix remodeling and pulmonary disease

Impaired alveolar formation and maintenance are features of many pulmonary diseases that are associated with significant morbidity and mortality. In a forward genetic screen for modulators of mouse lung development, we identified the non-muscle myosin II heavy chain gene, Myh10. Myh10 mutant pups exhibit cyanosis and respiratory distress, and die shortly after birth from differentiation defects in alveolar epithelium and mesenchyme. From omics analyses and follow up studies, we find decreased Thrombospondin expression accompanied with increased matrix metalloproteinase activity in both mutant lungs and cultured mutant fibroblasts, as well as disrupted extracellular matrix (ECM) remodeling. Loss of Myh10 specifically in mesenchymal cells results in ECM deposition defects and alveolar simplification. Notably, MYH10 expression is downregulated in the lung of emphysema patients. Altogether, our findings reveal critical roles for Myh10 in alveologenesis at least in part via the regulation of ECM remodeling, which may contribute to the pathogenesis of emphysema.

Quantitative lipidomic analysis of mouse lung during postnatal development by electrospray ionization tandem mass spectrometry.

Lipids play very important roles in lung biology, mainly reducing the alveolar surface tension at the air-liquid interface thereby preventing end-expiratory collapse of the alveoli. In the present study we performed an extensive quantitative lipidomic analysis of mouse lung to provide the i) total lipid quantity, ii) distribution pattern of the major lipid classes, iii) composition of individual lipid species and iv) glycerophospholipid distribution pattern according to carbon chain length (total number of carbon atoms) and degree of unsaturation (total number of double bonds). We analysed and quantified 160 glycerophospholipid species, 24 sphingolipid species, 18 cholesteryl esters and cholesterol from lungs of a) newborn (P1), b) 15-day-old (P15) and c) 12-week-old adult mice (P84) to understand the changes occurring during postnatal pulmonary development. Our results revealed an increase in total lipid quantity, correlation of lipid class distribution in lung tissue and significant changes in the individual lipid species composition during postnatal lung development. Interestingly, we observed significant stage-specific alterations during this process. Especially, P1 lungs showed high content of monounsaturated lipid species; P15 lungs exhibited myristic and palmitic acid containing lipid species, whereas adult lungs were enriched with polyunsaturated lipid species. Taken together, our study provides an extensive quantitative lipidome of the postnatal mouse lung development, which may serve as a reference for a better understanding of lipid alterations and their functions in lung development and respiratory diseases associated with lipids.

Human lung development: recent progress and new challenges.

ABSTRACTRecent studies have revealed biologically significant differences between human and mouse lung development, and have reported new in vitro systems that allow experimental manipulation of human lung models. At the same time, emerging clinical data suggest that the origins of some adult lung diseases are found in embryonic development and childhood. The convergence of these research themes has fuelled a resurgence of interest in human lung developmental biology. In this Review, we discuss our current understanding of human lung development, which has been profoundly influenced by studies in mice and, more recently, by experiments using in vitro human lung developmental models and RNA sequencing of human foetal lung tissue. Together, these approaches are helping to shed light on the mechanisms underlying human lung development and disease, and may help pave the way for new therapies.

Iloprost attenuates hyperoxia-mediated impairment of lung development in newborn mice.

Cyclooxygenase-2 (COX-2/PTGS2) mediates hyperoxia-induced impairment of lung development in newborn animals and is increased in the lungs of human infants with bronchopulmonary dysplasia (BPD). COX-2 catalyzes the production of cytoprotective prostaglandins, such as prostacyclin (PGI2), as well as proinflammatory mediators, such as thromboxane A2. Our objective was to determine whether iloprost, a synthetic analog of PGI2, would attenuate hyperoxia effects in the newborn mouse lung. To test this hypothesis, newborn C57BL/6 mice along with their dams were exposed to normoxia (21% O2) or hyperoxia (85% O2) from 4 to 14 days of age in combination with daily intraperitoneal injections of either iloprost 200 µg·kg-1·day-1, nimesulide (selective COX-2 antagonist) 100 mg·kg-1·day-1, or vehicle. Alveolar development was estimated by radial alveolar counts and mean linear intercepts. Lung function was determined on a flexiVent, and multiple cytokines and myeloperoxidase (MPO) were quantitated in lung homogenates. Lung vascular and microvascular morphometry was performed, and right ventricle/left ventricle ratios were determined. We determined that iloprost (but not nimesulide) administration attenuated hyperoxia-induced inhibition of alveolar development and microvascular density in newborn mice. Iloprost and nimesulide both attenuated hyperoxia-induced, increased lung resistance but did not improve lung compliance that was reduced by hyperoxia. Iloprost and nimesulide reduced hyperoxia-induced increases in MPO and some cytokines (IL-1β and TNF-α) but not others (IL-6 and KC/Gro). There were no changes in pulmonary arterial wall thickness or right ventricle/left ventricle ratios. We conclude that iloprost improves lung development and reduces lung inflammation in a newborn mouse model of BPD.