Natascha Sommer1, Maik Hüttemann1, Oleg Pak1, Susan Scheibe1, Fenja Knoepp1, Christopher Sinkler1, Monika Malczyk1, Mareike Gierhardt1, Azadeh Esfandiary1, Simone Kraut1, Felix Jonas1, Christine Veith1, Siddhesh Aras1, Akylbek Sydykov1, Nasim Alebrahimdehkordi1, Klaudia Giehl1, Matthias Hecker1, Ralf P Brandes1, Werner Seeger1, Friedrich Grimminger1, Hossein A Ghofrani1, Ralph T Schermuly1, Lawrence I Grossman2, Norbert Weissmann1. 1. From the Excellence Cluster Cardiopulmonary System, University of Giessen and Marburg Lung Center (UGMLC), Member of the German Center for Lung Research (DZL), Justus-Liebig-University, Giessen, Germany (N.S., O.P., S.S., F.K., M.M., M.G., A.E., S.K., F.J., C.V., A.S., N.A., K.G., M.H., W.S., F.G., H.A.G., R.T.S., N.W.); Center for Molecular Medicine and Genetics, Wayne State University School of Medicine, Detroit, MI (M.H., C.S., S.A., L.I.G.); Institut für Kardiovaskuläre Physiologie, Goethe-Universität, German Center for Cardiovascular Research (DZHK), Partner Site RheinMain, Frankfurt am Main, Germany (R.P.B.); and Max Planck Institute for Heart and Lung Research, Bad Nauheim, Germany (W.S.). 2. From the Excellence Cluster Cardiopulmonary System, University of Giessen and Marburg Lung Center (UGMLC), Member of the German Center for Lung Research (DZL), Justus-Liebig-University, Giessen, Germany (N.S., O.P., S.S., F.K., M.M., M.G., A.E., S.K., F.J., C.V., A.S., N.A., K.G., M.H., W.S., F.G., H.A.G., R.T.S., N.W.); Center for Molecular Medicine and Genetics, Wayne State University School of Medicine, Detroit, MI (M.H., C.S., S.A., L.I.G.); Institut für Kardiovaskuläre Physiologie, Goethe-Universität, German Center for Cardiovascular Research (DZHK), Partner Site RheinMain, Frankfurt am Main, Germany (R.P.B.); and Max Planck Institute for Heart and Lung Research, Bad Nauheim, Germany (W.S.). lgrossman@wayne.edu Norbert.Weissmann@innere.med.uni-giessen.de.
Abstract
RATIONALE: Acute pulmonary oxygen sensing is essential to avoid life-threatening hypoxemia via hypoxic pulmonary vasoconstriction (HPV) which matches perfusion to ventilation. Hypoxia-induced mitochondrial superoxide release has been suggested as a critical step in the signaling pathway underlying HPV. However, the identity of the primary oxygen sensor and the mechanism of superoxide release in acute hypoxia, as well as its relevance for chronic pulmonary oxygen sensing, remain unresolved. OBJECTIVES: To investigate the role of the pulmonary-specific isoform 2 of subunit 4 of the mitochondrial complex IV (Cox4i2) and the subsequent mediators superoxide and hydrogen peroxide for pulmonary oxygen sensing and signaling. METHODS AND RESULTS: Isolated ventilated and perfused lungs from Cox4i2-/- mice lacked acute HPV. In parallel, pulmonary arterial smooth muscle cells (PASMCs) from Cox4i2-/- mice showed no hypoxia-induced increase of intracellular calcium. Hypoxia-induced superoxide release which was detected by electron spin resonance spectroscopy in wild-type PASMCs was absent in Cox4i2-/- PASMCs and was dependent on cysteine residues of Cox4i2. HPV could be inhibited by mitochondrial superoxide inhibitors proving the functional relevance of superoxide release for HPV. Mitochondrial hyperpolarization, which can promote mitochondrial superoxide release, was detected during acute hypoxia in wild-type but not Cox4i2-/- PASMCs. Downstream signaling determined by patch-clamp measurements showed decreased hypoxia-induced cellular membrane depolarization in Cox4i2-/- PASMCs compared with wild-type PASMCs, which could be normalized by the application of hydrogen peroxide. In contrast, chronic hypoxia-induced pulmonary hypertension and pulmonary vascular remodeling were not or only slightly affected by Cox4i2 deficiency, respectively. CONCLUSIONS: Cox4i2 is essential for acute but not chronic pulmonary oxygen sensing by triggering mitochondrial hyperpolarization and release of mitochondrial superoxide which, after conversion to hydrogen peroxide, contributes to cellular membrane depolarization and HPV. These findings provide a new model for oxygen-sensing processes in the lung and possibly also in other organs.
RATIONALE: Acute pulmonary oxygen sensing is essential to avoid life-threatening hypoxemia via hypoxic pulmonary vasoconstriction (HPV) which matches perfusion to ventilation. Hypoxia-induced mitochondrial superoxide release has been suggested as a critical step in the signaling pathway underlying HPV. However, the identity of the primary oxygen sensor and the mechanism of superoxide release in acute hypoxia, as well as its relevance for chronic pulmonary oxygen sensing, remain unresolved. OBJECTIVES: To investigate the role of the pulmonary-specific isoform 2 of subunit 4 of the mitochondrial complex IV (Cox4i2) and the subsequent mediators superoxide and hydrogen peroxide for pulmonary oxygen sensing and signaling. METHODS AND RESULTS: Isolated ventilated and perfused lungs from Cox4i2-/- mice lacked acute HPV. In parallel, pulmonary arterial smooth muscle cells (PASMCs) from Cox4i2-/- mice showed no hypoxia-induced increase of intracellular calcium. Hypoxia-induced superoxide release which was detected by electron spin resonance spectroscopy in wild-type PASMCs was absent in Cox4i2-/- PASMCs and was dependent on cysteine residues of Cox4i2. HPV could be inhibited by mitochondrial superoxide inhibitors proving the functional relevance of superoxide release for HPV. Mitochondrial hyperpolarization, which can promote mitochondrial superoxide release, was detected during acute hypoxia in wild-type but not Cox4i2-/- PASMCs. Downstream signaling determined by patch-clamp measurements showed decreased hypoxia-induced cellular membrane depolarization in Cox4i2-/- PASMCs compared with wild-type PASMCs, which could be normalized by the application of hydrogen peroxide. In contrast, chronic hypoxia-induced pulmonary hypertension and pulmonary vascular remodeling were not or only slightly affected by Cox4i2 deficiency, respectively. CONCLUSIONS:Cox4i2 is essential for acute but not chronic pulmonary oxygen sensing by triggering mitochondrial hyperpolarization and release of mitochondrial superoxide which, after conversion to hydrogen peroxide, contributes to cellular membrane depolarization and HPV. These findings provide a new model for oxygen-sensing processes in the lung and possibly also in other organs.
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