Pulmonary artery NADPH-oxidase is activated in hypoxic pulmonary vasoconstriction.

Abstract

An NADPH-oxidase complex containing at least two protein components (gp91-phox and p22-phox) and a unique low redox potential (-245 mV) cytochrome b-245 is the source of superoxide generated for bacterial killing in neutrophils and has been suggested as the oxygen sensor in the carotid body. In pure cultures of smooth muscle cells from calf small pulmonary arteries (300 microns diameter) the presence of the 91 kD protein specific to this cytochrome was demonstrated by Western blot analysis with monoclonal antibody 48. Low-temperature-difference spectrophotometry of homogenates of these cells demonstrated the characteristic cytochrome b-245 spectrum when titrated between redox potentials of -152 and -345 mV, consistent with the low redox potential form. When these same cells were exposed to hypoxia (approximately 40 mmHg), superoxide production increased significantly from 1.4 +/- 0.2 to 73 +/- 12 nmoles.min-1 mg-1 protein. Hypoxic generation of superoxide was inhibited by the NADPH-oxidase inhibitor diphenyleneiodonium (DPI: 10 microM) but not by the mitochondrial inhibitor myxathiazole (10 microM). The hypoxic superoxide increase was significantly greater than that observed from smooth muscle cells from large pulmonary arteries or from large or small systemic arteries. Fluorescence immunocytochemistry revealed the presence of the NADPH-oxidase protein in the walls of pulmonary arteries in rat lung slices, and confocal microscopy showed the complex to be widely distributed in the vicinity of the arterial smooth muscle walls. In hypoxia or norepinephrine (NP)-induced vasoconstriction of pulmonary artery rings from cats, the sensitivity to inhibition by DPI was observed to be significantly greater for hypoxia (ED50 = 0.8 microM) than for NP-induced (ED50 = 13.4 microM) constriction. Together these observations demonstrate that the unique cytochrome b-245 containing NADPH-oxidase is present in pulmonary artery smooth muscle and that an NADPH-oxidase or NADH-oxidoreductase complex is activated to release superoxide by hypoxic conditions. It is concluded that a trans-membrane NADPH-oxidase is the most likely and that activation of this system may be involved in the initiation of hypoxic pulmonary vasoconstriction.

Impairment by Hypoxia or Hypoxia/Reoxygenation of Nitric Oxide–Mediated Relaxation in Isolated Monkey Coronary Artery: the Role of Intracellular Superoxide

“…Moreover, hypoxia-induced increases in the tissue level of superoxide and NADPH oxidase activity are potently suppressed by apocynin. As the support of our results, Marshall et al have shown that hypoxia increases superoxide production caused by NADPH oxidase in bovine arterial smooth muscle (36). Rathore et al have also demonstrated that hypoxia significantly increases NADPH oxidase activity by translocation of the cytosolic subunit to the plasma membrane in smooth muscle cells (37).…”

Contribution of xanthine oxidase-derived superoxide to chronic hypoxic pulmonary hypertension in neonatal rats

“…This possibility warrants exploration in future studies. Recent studies have also identified a potentially major role for vascular cell-derived NAD(P)H oxidase in ROS formation (42,45,48). While not explored to any extent in the present study, our observations using in situ measurement of vascular ROS suggested that smooth muscle-derived NAD(P)H oxidase may be a major source of O 2 •Ϫ in the remodeled pulmonary vasculature.…”

Reactive oxygen species and cellular oxygen sensing

“…Early theories on the subject suggested that NADPH oxidase might be an important ROS-generating cellular O 2 sensor since this multi-subunit membrane-bound enzyme is expressed in tissues implicated in systemic hypoxic responses and affects cellular redox status depending on cellular O 2 concentrations [69,70]. However more in-depth studies did not support a requirement for NADPH in the hypoxic adaptive response [71].…”