Abstract 11961: Ductus Arteriosus Oxygen Response is Mediated by Mitochondrial Subunit NDUFS2

• Published in: Circulation (New York, N.Y.) N.Y.), 2022-11, Vol.146 (Suppl_1), p.A11961-A11961
• Main Authors: Bentley, Rachel E, Hindmarch, Charles C, Lima, Patricia, Mewburn, Jeffrey, Martin, Ashley, Dunham-Snary, Kimberly J, Archer, Stephen L
• Format: Article
• Language: English
• ISSN: 0009-7322; 1524-4539
• DOI: 10.1161/circ.146.suppl_1.11961

IntroductionThe ductus arteriosus (DA) is a fetal vessel connecting the pulmonary artery (PA) and aorta, diverting oxygenated blood away from the developing lungs to the systemic circulation. The DA rapidly constricts in response to the increase in arterial oxygen (O2) following the first breath. Failure of the DA to close in response to O2 results in persistent ductus arteriosus, one of the most common complications of preterm birth. Mitochondria are known to play an important role in the O2 response of DA smooth muscle cells (DASMC), however the mechanism is not fully understood. A subunit of mitochondrial Complex I, NDUFS2 (NADH:ubiquinone oxidoreductase core subunit 2) mediates the O2 response of other specialized O2 sensing tissues (PA and carotid body); we assess whether NDUFS2 also plays a role in DA O2 sensing. MethodsDASMC were isolated from male term (29 days gestation) fetal New Zealand white rabbits. Cell lines were purified via fluorescence activated cell sorting, excluding CD90+ and CD31+ cells (fibroblasts and endothelial cells, respectively). Smooth muscle cell purity was confirmed using flow cytometry for α-smooth muscle actin (α-SMA). Primary cultures were 97.65% ± 0.65% CD90- CD31-, and were 100% α-SMA+ post-sorting. We measured the O2 responsiveness of DASMC using confocal microscopy, measuring changes in intracellular calcium with Cal-520AM. Cells were cultured in hypoxia (2.5% O2, pO2 = 41 mmHg), with O2-induced increase in calcium relative to hypoxic baseline serving as a surrogate for constriction. DASMC were treated for 48 hours with 30 pmol siRNA and 90 pmol Lipofectamine® RNAiMAX to knockdown gene expression of NDUFS2 or a negative control. Knockdown was confirmed using qPCR. Results & ConclusionsThere was a 14.7% (±2.6%, n=33, p