Ndufs2, A Core Subunit of Mitochondrial Complex I, Is Essential for Acute Oxygen-Sensing and Hypoxic Pulmonary Vasoconstriction.

RATIONALE: Hypoxic pulmonary vasoconstriction (HPV) optimizes systemic oxygen delivery by matching ventilation to perfusion. HPV is intrinsic to pulmonary artery smooth muscle cells (PASMC). Hypoxia dilates systemic arteries, including renal arteries. Hypoxia is sensed by changes in mitochondrial-derived reactive oxygen species, notably H2 O2 ([H2 O2 ]mito ). Decreases in [H2 O2 ]mito elevate pulmonary vascular tone by increasing intracellular calcium ([Ca2+ ]i ) through redox regulation of ion channels. Although HPV is mimicked by the Complex I inhibitor, rotenone, the molecular identity of the O2-sensor is unknown.

OBJECTIVE: To determine the role of NADH dehydrogenase [ubiquinone] iron-sulfur protein 2 (Ndufs2), Complex I's rotenone binding site, in pulmonary vascular oxygen-sensing.

METHODS AND RESULTS: Mitochondria-conditioned media (MCM) from pulmonary and renal mitochondria isolated from normoxic and chronically hypoxic rats, were infused into an isolated lung bioassay. MCM from normoxic lungs contained more H2O2 than MCM from chronic hypoxic lungs or kidneys and uniquely attenuated HPV via a catalase-dependent mechanism. In PASMC acute hypoxia decreased H2 O2 within 112{plus minus}7s, followed, within 205{plus minus}34s, by increased intracellular calcium concentration, [Ca2+ ]i . Hypoxia had no effects on [Ca2+ ]i in renal artery SMC (RASMC). Hypoxia decreases both cytosolic and mitochondrial H2 O2 in PASMC while increasing cytosolic H2 O2 in RASMC. Ndufs2 expression was greater in PASMC versus RASMC. Lung Ndufs2 cysteine residues became reduced during acute hypoxia and both hypoxia and reducing agents caused functional inhibition of Complex I. In PASMC, siNdufs2 decreased normoxic H2 O2 , prevented hypoxic increases in [Ca2+ ]i , and mimicked aspects of chronic hypoxia, including decreasing Complex I activity, elevating the NADH/NAD+ ratio and decreasing expression of the O2 -sensitive ion channel, Kv1.5. Knocking down another Fe-S center within Complex I (Ndufs1) or other mitochondrial subunits proposed as putative oxygen sensors (Complex III's Rieske Fe-S center and COX4i2 in Complex IV) had no effect on hypoxic increases in [Ca2+ ]i . In vivo, siNdufs2 significantly decreased hypoxia- and rotenone-induced constriction while enhancing phenylephrine-induced constriction.

CONCLUSIONS: Ndufs2 is essential for oxygen-sensing and HPV.