234 - Endothelial nitric oxide synthase is uncoupled and reactive oxygen species generation is increased in the right ventricle of newborn piglets with chronic hypoxia-induced pulmonary hypertension

Poster Number: 234
Publication Number: 234.222

Matthew S. Douglass, University of Utah School of Medicine, Salt Lake City, UT, United States; Yongmei Zhang, University of Utah, Salt lake city, UT, United States; Mark R. Kaplowitz, University of Utah School of Medicine, Salt Lake City, UT, United States; Candice D. Fike, University of Utah School of Medicine, Salt Lake City, UT, United States

Background: Adult models of pulmonary hypertension (PH) have demonstrated that excessive generation of reactive oxygen species (ROS) contribute to right ventricular hypertrophy (RVH) and that augmenting cyclic guanosine 3′,5′-monophosphate (cGMP) signaling contributes to an anti-hypertrophic response. Uncoupled endothelial nitric oxide synthase (eNOS) was shown to be a source of ROS underlying cardiac hypertrophy in adult mice after chronic transverse aortic constriction. Whether these mechanisms are involved in the development of RVH in newborns with PH is not yet understood.

Objective: We wanted to determine if ROS generation, eNOS coupling, and cGMP levels are altered in the right ventricle of newborn pigs with chronic hypoxia-induced pulmonary hypertension (CHPH).

Design/Methods: Newborn pigs were obtained on DOL 2, placed in a normobaric normoxic or hypoxic chamber (10-11% oxygen) for 10 days. Hemodynamic measurements of pulmonary artery pressure, left ventricle end diastolic pressure, and cardiac output were obtained to calculate pulmonary vascular resistance (PVR). Fulton's index, the ratio of the weight of the right ventricular (RV) free wall to the weight of the left ventricular wall and septum, was calculated to assess RV mass. RV tissue was obtained and frozen. eNOS coupling was assessed in RV tissue by measuring eNOS dimers and monomers by western blot where uncoupled eNOS is demonstrated by a loss of dimer formation and an increase in monomer formation. ROS were measured using lucigenin-enhanced chemiluminescence assay. cGMP levels were measured by immunoassay.

Results: Both PVR and Fulton’s index were increased in piglets with CHPH (n=9) compared to normoxic controls (n=6) (p < 0.05). ROS generation was greater in the RVs of CHPH piglets (p < 0.05) compared to normoxic piglets. Consistent with eNOS uncoupling, the RVs of CHPH piglets had decreased eNOS dimer-to-monomer ratios compared to normoxic piglets (p < 0.05). cGMP levels did not differ between CHPH and normoxic piglet RVs.Conclusion(s): Increases in ROS but not alterations in cGMP levels are likely to be involved with the RVH that develops in piglets with CHPH after 10 days of hypoxia. Moreover, uncoupled eNOS is a source of ROS that could contribute to RVH in piglets with CHPH. These findings suggest that therapies that recouple eNOS and reduce ROS generation in the RV merit evaluation as strategies to protect the myocardium from pathologic RV remodeling and perhaps prevent progression to heart failure in newborns who develop PH in chronic cardiopulmonary conditions associated with hypoxia.