Transcriptional profiling of lung cell populations in idiopathic pulmonary arterial hypertension

Abstract

Despite recent improvements in management of idiopathic pulmonary arterial hypertension, mortality remains high. Understanding the alterations in the transcriptome–phenotype of the key lung cells involved could provide insight into the drivers of pathogenesis. In this study, we examined differential gene expression of cell types implicated in idiopathic pulmonary arterial hypertension from lung explants of patients with idiopathic pulmonary arterial hypertension compared to control lungs. After tissue digestion, we analyzed all cells from three idiopathic pulmonary arterial hypertension and six control lungs using droplet-based single cell RNA-sequencing. After dimensional reduction by t-stochastic neighbor embedding, we compared the transcriptomes of endothelial cells, pericyte/smooth muscle cells, fibroblasts, and macrophage clusters, examining differential gene expression and pathways implicated by analysis of Gene Ontology Enrichment. We found that endothelial cells and pericyte/smooth muscle cells had the most differentially expressed gene profile compared to other cell types. Top differentially upregulated genes in endothelial cells included novel genes: ROBO4, APCDD1, NDST1, MMRN2, NOTCH4, and DOCK6, as well as previously reported genes: ENG, ORAI2, TFDP1, KDR, AMOTL2, PDGFB, FGFR1, EDN1, and NOTCH1. Several transcription factors were also found to be upregulated in idiopathic pulmonary arterial hypertension endothelial cells including SOX18, STRA13, LYL1, and ELK, which have known roles in regulating endothelial cell phenotype. In particular, SOX18 was implicated through bioinformatics analyses in regulating the idiopathic pulmonary arterial hypertension endothelial cell transcriptome. Furthermore, idiopathic pulmonary arterial hypertension endothelial cells upregulated expression of FAM60A and HDAC7, potentially affecting epigenetic changes in idiopathic pulmonary arterial hypertension endothelial cells. Pericyte/smooth muscle cells expressed genes implicated in regulation of cellular apoptosis and extracellular matrix organization, and several ligands for genes showing increased expression in endothelial cells. In conclusion, our study represents the first detailed look at the transcriptomic landscape across idiopathic pulmonary arterial hypertension lung cells and provides robust insight into alterations that occur in vivo in idiopathic pulmonary arterial hypertension lungs.

Vascular Homeostasis and Inflammation in Health and Disease—Lessons from Single Cell Technologies

“…ScRNA-Seq analyses in human PAH patients have uncovered the upregulation of angiogenesis-associated genes in ECs such as endoglin (ENG), NOTCH1 and KDR, together with genes involved in SMC migration and proliferation such as PDGFB and FGFR1, as well as vasoconstrictor endothelin 1 (EDN1) [74]. Interestingly, apelin (APLN) and apelin cleaving enzyme lysosomal pro-X carboxypeptidase (PRCP) were elevated in PAH samples [74], contradicting the previously reported reduction in APLN and PRCP expression in pulmonary artery ECs isolated from PAH patients in vitro and the regression of PAH in mice after APLN administration [75,76]. Genes upregulated in SMCs and pericytes were related to ECM components that support vessel wall remodeling and increased ECM stiffness [74].…”

“…Interestingly, apelin (APLN) and apelin cleaving enzyme lysosomal pro-X carboxypeptidase (PRCP) were elevated in PAH samples [74], contradicting the previously reported reduction in APLN and PRCP expression in pulmonary artery ECs isolated from PAH patients in vitro and the regression of PAH in mice after APLN administration [75,76]. Genes upregulated in SMCs and pericytes were related to ECM components that support vessel wall remodeling and increased ECM stiffness [74]. Furthermore, bioinformatic analyses of regulatory networks derived a number of transcription factors such as SOX18, STRA13, LYL1 and TFDP1 [74], which are likely to play an important role in ECs in PAH.…”

“…Genes upregulated in SMCs and pericytes were related to ECM components that support vessel wall remodeling and increased ECM stiffness [74]. Furthermore, bioinformatic analyses of regulatory networks derived a number of transcription factors such as SOX18, STRA13, LYL1 and TFDP1 [74], which are likely to play an important role in ECs in PAH.…”

Molecular Basis of Cardiac and Vascular Injuries Associated With COVID-19

“…The cellular composition of the lung is 40-50% endothelial cells, which differentiate in parallel with epithelial cells to form gas exchange units which are in contact with the external environment and thus need to ensure a rapid immune response (30). In lung diseases, including infections, the transcriptomes of endothelial cells, pericyte/smooth muscle cells, fibroblasts, and macrophage clusters showed that endothelial cells had the most differentially expressed gene profile compared to other cell types (31). We speculated that if we found common differentially expressed genes shared between the two cell types and which could be affected by SARS-CoV-2 infection, then we may be able further to understand the link between COVID-19 and associated endothelium injuries.…”

Essential genes and miRNA-mRNA network contributing to the pathogenesis of idiopathic pulmonary arterial hypertension

“…In the past few decades, microarray analysis has been used for gene-wide expression pro ling in lung tissues or peripheral blood from PAH patients or experimental animals to de ne PAH's pathobiology [6][7][8][9][10]. However, the ndings are inconsistent between different researches, which may be caused by various analysis platforms, limited sample sizes, different methods, no classi cation of PAH, and not for IPAH.…”