Abstract 316: Angiotensin 1-7 Regulation Of Endothelin-1 System In Pulmonary Arterial Hypertension

Abstract

Studies suggest that activation of ACE2-Ang-(1-7)-Mas ameliorates vascular, cardiac and lung damage observed in pulmonary hypertension (PAH); a condition where endothelin-1 (ET-1) is important. Here we assessed whether Ang 1-7 regulates ET-1 system in pulmonary hypertension and putative mechanisms. Hypobaric hypoxia was used to induce hypoxic PAH in mice, which were divided in 4 groups: normoxic controls (NC), hypoxic PAH (HP), normoxic (NA) and hypoxic PAH (HA) treated with Ang 1-7 (hydroxypropyl-βcyclodextrin-Ang(1-7) 30μg/kg/day given by oral gavage for 14 days after established hypoxia-induced PAH. In HP mice, RVSP (18.7 NC vs. 47.6mmHg HP, p<0.001), RVH by Fulton Index (0.198 NC vs. 0.279 HP, p<0.01) and urinary ET-1 levels (0.88 NC vs 2.48pg/ml HP, p<0.05 vs NC) were increased, an effect blocked by Ang1-7 treatment. In NA, Ang 1-7 increased urinary levels of ET-1 (2.24pg/ml, p<0.05 vs NC). No changes were observed on ETBR protein expression between all groups. Human microvascular endothelial cells (HMEC) were also used and stimulated with ET-1 in the absence/presence of Ang 1-7. Ang 1-7 stimulation increased preproET-1 mRNA levels (250%), ET-1 release (125%), and ETBR protein levels (50%), p<0.05 vs vehicle. To examine whether the regulation of ET-1 system by Ang 1-7 is beneficial, HMEC were stimulated with ET-1 in the absence/presence of Ang 1-7. ET-1 increased e-selectin mRNA (400%) and VCAM-1 protein (38%) levels, as well as TNFα production (30%); all effects blocked by Ang 1-7 (p<0.05). Ang 1-7 inhibition of ET-1 pro-inflammatory effects was dependent on NO production, since it was blocked by L-NAME and LY294002 (inhibitors of NO pathway). Interestingly, Ang 1-7 increased NO production (257%) through Mas and ETBR-dependent manner. An interaction between Mas and ETbR was observed in HMEC by immunoprecipitation and peptide array protocols. In conclusion, Ang 1-7 modulates ET-1 system in pulmonary hypertension and HMEC. Our findings indicate that Ang 1-7 negatively modulates proinflammatory signalling in human endothelial cells. The protective effect may involve upregulation/crosstalk of/with ETBR. These data identify the ET-1 system as a novel molecular mechanism involved in the putative protective action of Ang 1-7 in pulmonary hypertension.