The lncRNA DSCR9 is modulated in pulmonary arterial hypertension endothelial cell models and is associated with alterations in the nitric oxide pathway

Abstract

Long non-coding RNA (lncRNA) may be involved in dysfunction of pulmonary artery endothelial cells (PAEC) and, thus, in pulmonary arterial hypertension (PAH) pathobiology.

We screened the RNA expression profile of commercial human PAEC (hPAEC) exposed to increased hydrostatic pressure, and found that the lncRNA Down syndrome critical region 9 (DSCR9) was the most regulated transcript (log2FC 1.89 vs control). We confirmed by RT-qPCR that DSCR9 levels were higher in PAEC isolated from patients with idiopathic PAH (iPAH-PAEC), as well as in induced pluripotent stem cell-derived endothelial cells (iPSC-EC) from a patient with BMPR2-mutated PAH, than in relevant controls. Moreover, a re-analysis of the publicly available GSE117261 microarray dataset revealed that DSCR9 was upregulated in the lung tissue of PAH patients. In silico simulation indicated that DSCR9 would be mainly located in the nucleus and could interact with calcium/calmodulin-dependent protein kinase II beta (CAMK2B) and nitric oxide synthase 3 (NOS3, encoding eNOS). CAMK2B levels resulted 3.4-fold higher (p < 0.05) in iPAH-PAEC transfected with a DSCR9-GFP carrying plasmid than with a GFP-only-carrying one. A trend for higher NOS3 expression was also noted. GFP immunostaining was predominantly nuclear and cytoplasmic upon DSCR9-GFP or GFP-only transfection, respectively. CAMK2B and NOS3 mRNA were also higher in iPAH-PAEC than control-PAEC in basal conditions. Instead, variations in total and phosphorylated CAMK2B, eNOS, and NO synthesis were inconsistent. We conclude that DSCR9 is upregulated in PAH-related endothelial cell models and influences CAMK2B and NOS3 expression. Future studies are necessary to determine whether DSCR9 affects NO availability, including in PAH.