Abstract 10068: Oxidative Modification of Endothelin-B Receptor Cysteinyl Thiol(s) by Aldosterone Decreases Pulmonary Endothelial Nitric Oxide Levels in Pulmonary Arterial Hypertension
Hyperaldosteronism (ALDO) induces a vasculopathy in systemic blood vessels by increasing reactive oxygen species (ROS) to decrease nitric oxide (NO) levels. In pulmonary arterial hypertension (PAH), the role of ALDO in pulmonary artery endothelial cell (PAEC) NO synthesis is unknown. We hypothesized that in PAH, ALDO increases ROS levels in PAECs to disrupt endothelin B receptor (ETB)-dependent NO synthesis through oxidative modification of ETB cysteinyl thiols important for NO generation. To test this hypothesis, we confirmed that ALDO levels were elevated in PAH. In monocrotaline (60 mg/ml)-treated rats, PAH was associated with decreased pulmonary artery acceleration time (34.5 ± 4.4 vs.13.4 ± 2.6 ms, p<0.02), increased right ventricular wall thickness (1.0 ± 0.1 vs. 0.6 ± 0.1 mm, p<0.01), and pulmonary vascular remodeling compared to vehicle (V)-treated rats at 4 weeks. These changes were associated with a 213% increase in plasma ALDO levels (p<0.04) in the absence of left ventricular systolic dysfunction. To determine the mechanism by which ALDO influenced pulmonary vascular function, we exposed human PAECs to ALDO (10-7 mol/L) for 24 hr. Compared to V, ALDO increased NADPH oxidase (NOX4) protein expression (109.0 ± 7.9 vs. 153.6 ± 12.6 arb. units, p<0.01) and hydrogen peroxide (H2O2) levels by 51% (p<0.05). ALDO-induced ROS stimulated the formation of two de novo ETB disulfide bonds demonstrated by immunoblotting for conjugated maleimide-labeled ETB cysteinyl thiols. In PAECs treated with V or endothelin-1 (10 nM) for 10 min to stimulate ETB signaling, ALDO significantly decreased ETB-dependent NOx (nitrite + nitrate) formation (100.2 ± 9.1 vs. 202.3 ± 20.8 vs.148.9 ± 12.6% control, respectively, p<0.05). These effects were inhibited by coincubation with the ALDO antagonist spironolactone (10 µM). Spironolactone decreased ALDO-induced H2O2 formation by 61% (p<0.05) and fully restored ETB-dependent NOx formation in ALDO-treated cells (p<0.05). These data suggest that in PAH, ALDO-induced ROS oxidatively modifies ETB cysteinyl thiols (disulfide formation) to decrease ETB-dependent NO synthesis. Identifying ALDO as a prospective target to improve pulmonary vascular tone has potentially useful therapeutic implications for patients with PAH.
- © 2011 by American Heart Association, Inc.