Rationale: Pulmonary arterial hypertension (PAH) is a cardiopulmonary disorder characterized by persistent elevation of pulmonary arterial (PA) pressure and premature death. PA smooth muscle cells (PASMCs) from PAH patients present a cancer-like hyperproliferative and apoptosis-resistant phenotype contributing to vasoconstriction and remodeling of distal PAs.
Although epigenetic alterations are increasingly appreciated as a contributing factor of PAH development, one important challenge in epigenetic therapy is defining which genes are the drivers.
G9a is an epigenetic factor, known to be overexpressed in many cancers, promoting cell proliferation and survival. Given that PAH and cancer cells share numerous similarities, it is crucial to determine if G9a is implicated in PAH. We thus hypothesized that the inhibition of G9a could reduce the pro-proliferative and apoptosis resistance phenotype of PAH-PASMCs.
Methods and Results: Using Western blot (WB) and immunofluorescence, we demonstrated that G9a is significantly overexpressed in distal PAs and isolated PASMCs from PAH patients (n= 6-14, p<0.01). Similarly, G9a was increased (WB and immunofluorescence, p<0.05) in two models mimicking the disease; namely the monocrotaline rat model and mice exposed to chronic hypoxia. In vitro, we found that pharmacological inhibition of G9a using BIX01294 and UNC0642 dose-dependently reduces PAH-PASMC proliferation (Ki67 labeling and EdU assay, p<0.001) and survival (Annexin V assay p<0.001). Inhibition of G9a was accompanied by the appearance of numerous cytoplasmic vacuoles. As detected by WB and immunofluorescence, inhibition of G9a in PAH-PASMCs resulted in a significant increase in LC3-II accumulation which was in parallel with an increase in p62; thus, suggesting that inhibition of G9a interferes with autophagic flux.
Conclusion: We showed for the first time that G9a is overexpressed in human and experimental PAH contributing to the pro-proliferative and anti-apoptotic phenotype of PAH-PASMCs. Further research is required to understand precisely the role of G9a in the abnormal phenotype of PAH-PASMCs.