View Abstract

A6967 - Neonatal Hyperoxia Perturbs Gene Expression in Murine PDGFR-Alpha-Expressing Pulmonary Interstitial Fibroblasts
Author Block: S. Dautel1, J. Snowball2, J. A. Whitsett2, A. T. Perl2, S. K. Ahlfeld2; 1University of Washington, Seattle, WA, United States, 2Perinatal Institute, Division of Pulmonary Biology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH, United States.
Rationale: Bronchopulmonary dysplasia (BPD) is a common chronic lung disorder affecting infants born prematurely. Impaired lung function results from inhibited alveolar septation causing alveolar simplification. Although the etiology is multifactorial, BPD is strongly associated with exposure of the immature lung to inflammation and hyperoxia. While the mechanisms governing alveolarization are poorly understood, alveolar interstitial fibroblasts that express Platelet Derived Growth Factor Receptor Alpha (PDGFRα) play an important role in matrix production and tissue remodeling during septation. In the present study we used a transcriptomic approach to assess changes in lung gene expression using a murine model of neonatal hyperoxia to elucidate acute and persistent effects of hyperoxia on PDGFRα+ pulmonary cells.
Methods: Newly-delivered C57BL6 mouse pups were exposed continuously to either hyperoxia (>90% O2) or room air (RA) from birth through the first 7 postnatal days (P0-P7). On P7, hyperoxia-exposed mice were returned to RA and allowed to recover until P10. Using magnetic bead separation from single lung cell suspensions, cells expressing PDGFRα were obtained at P4 and P7 (acute hyperoxia exposure) and P10 (recovery from hyperoxia), processed for bulk RNA-Seq, and compared to RA controls. Transcriptomic data analysis identified differentially expressed genes between time-matched samples, during normal development (RA), as well as under hyperoxic conditions. Pathway analysis revealed enriched pathways relevant to septation. Changes in gene expression were used to predict upstream transcriptional regulators.
Results: Neonatal hyperoxia caused significant, global changes in PDGFRα+ fibroblast gene expression: RNAs associated with oxidative stress (super oxygen dismutase, glutathione metabolism) were significantly increased, RNAs encoding key lung developmental growth factors (TGF-β, PDGF, BMP, VEGF, FGF, IGF) were significantly decreased, and RNAs governing extracellular matrix production and remodeling (collagen, elastin, glycosaminoglycans, and proteinases) were significantly inhibited. Additionally, mis-expression of genes known to be important regulators of cell cycle (P21, CDKs, cyclins) indicated that hyperoxia inhibited cell cycle progression. Upstream analysis of perturbed genes predicted that Foxm1 serves as a master regulator of the cell cycle during alveolarization by inhibiting the G2/M checkpoint.
Conclusions: Taken together, this study demonstrates that neonatal hyperoxia causes global, persistent changes in the gene expression in murine PDGFRα+ interstitial fibroblasts. We have identified key regulators of alveolar septation as potential therapeutic targets to optimize lung development following preterm birth.