Hypoxic neonatal pulmonary arterial myocytes are sensitized to ROS-generated 8-isoprostane.
8-Isoprostane, a ROS-derived prostanoid that acts via the thromboxane receptor (TP), is implicated in neonatal pulmonary hypertension. The purpose of this study was to examine the effect of hypoxia on vascular smooth muscle ROS generation, 8-isoprostane activity, and TP binding. First-passage neonatal porcine pulmonary artery myocytes were exposed to 10% O(2) (hypoxic myocytes; HM) or 21% O(2) (normoxic myocytes) for 72 h. Hypoxia increased in vitro generation of ROS, superoxide, and 8-isoprostane. ROS generation was ablated by inhibition of mitochondrial complex III. SOD1 and 3 activities were increased, but SOD2 activity decreased by 45% in HM. 8-Isoprostane generation was driven by the addition of peroxide and nitric oxide; incubation with permeative PEG-SOD, but not PEG-catalase or impermeative SOD, attenuated hypoxia-induced 8-isoprostane generation. 8-Isoprostane affinity for TP was markedly increased in HM. Myocyte 8-isoprostane challenge caused TP internalization and calcium release only in HM; this was sensitive to TP blockade and was normalized by activation of adenylyl cyclase. We propose that hypoxia induces superoxide accumulation in pulmonary artery myocytes through inhibition of mitochondrial SOD2 activity, promoting peroxynitrite-induced generation of 8-isoprostane. 8-Isoprostane binds to sensitized TP receptors, causing receptor internalization and signaling to calcium release in hypoxic myocytes. 8-Isoprostane may be an important pulmonary vasoconstrictor during neonatal hypoxia.
Gong
Y
Y
Manitoba Institute of Child Health, University of Manitoba, Winnipeg, MAN, Canada.
Yi
M
M
Fediuk
J
J
Lizotte
P P
PP
Dakshinamurti
S
S
D000262:Adenylyl Cyclases / Q000378:metabolism; D020013:Calcium Signaling; D015687:Cell Hypoxia; D002478:Cells, Cultured; D015237:Dinoprost / Q000031:analogs & derivatives* / Q000378:metabolism; D014450:Electron Transport Complex III / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D007231:Infant, Newborn; D008928:Mitochondria / Q000378:metabolism; D032389:Myocytes, Smooth Muscle / Q000378:metabolism* / Q000473:pathology; D010547:Persistent Fetal Circulation Syndrome / Q000378:metabolism / Q000473:pathology; D011485:Protein Binding; D011651:Pulmonary Artery / Q000378:metabolism / Q000473:pathology; D017382:Reactive Oxygen Species / Q000378:metabolism*; D017482:Receptors, Thromboxane / Q000378:metabolism; D013482:Superoxide Dismutase / Q000378:metabolism*; D014661:Vasoconstriction
D017382:Reactive Oxygen Species; D017482:Receptors, Thromboxane; C075750:8-epi-prostaglandin F2alpha; D015237:Dinoprost; D013482:Superoxide Dismutase; D000262:Adenylyl Cyclases; D014450:Electron Transport Complex III