Bronchopulmonary Dysplasia with Pulmonary Hypertension Associates with Loss of Semaphorin Signaling and Functional Decrease in FOXF1 Expression

Lung injury in preterm infants leads to lifelong structural and functional respiratory deficits, with a risk for bronchopulmonary dysplasia (BPD). In its most severe form BPD is accompanied by pulmonary hypertension (PH). While impaired alveologenesis and vasculogenesis in BPD are well-described, the molecular mechanisms driving these phenotypes and the cellular dynamics associated with evolving BPD and BPD+PH in humans are not well-described. We performed single-cell RNA sequencing on preterm infant lungs in early stages of BPD, BPD+PH and term infants. Analysis of the endothelium revealed an aberrant capillary cell-state primarily in BPD+PH marked by ANKRD1 expression and a transcriptomic signature not previously described in previously analyzed human lung disease, a finding validated in an expanded repository of human infant lung tissue. Predictive signaling analysis identified deficits in the semaphorin guidance-cue signaling pathway and decreased expression of pro-angiogenic transcription factor FOXF1 within the alveolar parenchyma of the human neonatal lung samples with BPD and BPD+PH. We observed similar trends in the in decreased semaphoring signaling in a murine BPD mouse model and in analysis of ACDMPV single-nuclear sequencing data. These transcriptional deficits in semaphorin signaling common to BPD+PH and ACDMPV suggest a mechanistic link between the developmental phenotype in BPD and ACDMPV, providing a foundation for further exploration of the role of semaphorins in normal alveolar development. Further, these data fill a critical gap in currently available transcriptomic analysis of the normal and diseased lung across the lifespan as searchable atlas of early human lung development and injury. To examine the cellular dynamics and mediators of neonatal lung injury, single-cell RNA sequencing was performed on human neonatal lungs of varying stages of lung injury and chronic lung disease: 1 acute preterm infant with lung injury, 2 BPD samples at term corrected gestational age (CGA), 2 BPD+PH at term CGA, and 2 term infants in the first 3 weeks of life.