Introduction: Pulmonary Arterial Hypertension (PAH) is characterized by progressive pulmonary arteries (PAs) obstruction leading to increased blood flow resistance and heart failure. PASMCs of PAH patients display a “cancer-like” phenotype that contribute to PA remodeling. However, PAH-PASMCs very stressfull environment should normally trigger excessive DNA damage accumulation and cell death. This suggests that PAH-PASMCs have developed adaptive mechanisms to face stress which might be interesting targets in PAH. NUDT1 is a DNA protective enzyme that hydrolyzes oxidized guanines (8OHdG), preventing DNA damage overload and cell death. Given the antitumoral effects of NUDT1 inhibition, we hypothesized that NUDT1 overexpression is essential for the cancer-like phenotype of PAH-PASMCs and vascular remodeling.
Methods: We used primary cell lines and tissues from control and PAH patients as well as animals’ models of PAH and assess the effect of NUDT1 inhibition by siRNA and (S)-Crizotinib both in vitro and in vivo.
Results: We found by a proteomic study that NUDT1 is one of the most overexpressed proteins in PAH-PASMCs. Accordingly, NUDT1 expression measured by immunoblot was increased in PAH-tissues and PASMCs (p<0.05) (n=9), compared to control donors (n=9), and also in animal models (MCT: monocrotaline, Sugen/Hypoxia, and Fawn Hooded rats, n>5) (p<0.05). NUDT1 inhibition in PAH-PASMCs resulted in (i) marked accumulation of 8OHdG in both nuclear and mitochondrial DNA (Immunofluorescence, p<0,01); (ii) excessive DNA damage (Comet Assays, p<0,05); (iii) compromised bioenergetic functions (Seahorse, p<0,05); (iv) impaired autophagic flux (Immunofluorescence/blot, p<0,001), and finally (v) decreased proliferation/apoptosis resistance (Ki67/TUNEL, p<0,01). In vivo, nebulization of (S)-Crizotinib to MCT rats with established PAH improved hemodynamics (right heart catheterization: RVSP, mPAP, CO, p<0,05) and vascular remodeling (Elastica van Giesen staining, p<0,05). Conclusions: NUDT1 overexpression in PAH-PASMCs represents a stress-buffering mechanism supporting cell survival and proliferation. Inhibition of NUDT1 may represents a novel therapeutic avenue for PAH.