RAB7 deficiency impairs pulmonary artery endothelial function and promotes pulmonary hypertension

Pulmonary arterial hypertension (PAH) is a devastating and progressive disease with limited treatment options. Endothelial dysfunction plays a central role in development and progression of PAH, yet the underlying mechanisms are incompletely understood. The endosome-lysosome system is important to maintain cellular health and the small GTPase RAB7 regulates many functions of this system. Here, we explored the role of RAB7 in endothelial cell (EC) function and lung vascular homeostasis. We found reduced expression of RAB7 in ECs from PAH patients. Endothelial haploinsufficiency of RAB7 caused spontaneous PH in mice. Silencing of RAB7 in ECs induced broad changes in gene expression revealed via RNA sequencing and RAB7 silenced ECs showed impaired angiogenesis, expansion of a senescent cell fraction, combined with impaired endolysosomal trafficking and degradation, which suggests inhibition of autophagy at the pre-degradation level. Overall design: Human primary PAECs were cultured in complete EGM-2MV media (Lonza CC-3162) in a cell culture incubator at 37°C with 5% CO2 and 100% humidity. For siRNA-mediated knockdown of RAB7, PAECs were transfected with 50 nM RAB7 or control siRNA using GenMute Reagent (SL100568, SignaGen). The following siRNAs were used: RAB7 siRNA (IDT, NM_004637 13.2), control siRNA (IDT, 51-01-14-04). After 24h, siRNA was removed, and RNA was isolated 48h after transfection.