INTEGRATIVE PHYSIOLOGY Glutathione peroxidase 1 protects mitochondria against hypoxia/reoxygenation damage in mouse hearts Vu Thi Thu & Hyoung Kyu Kim & Seung Hee Ha & Ji-Young Yoo & Won Sun Park & Nari Kim & Goo Taeg Oh & Jin Han Received: 22 January 2010 / Revised: 16 February 2010 / Accepted: 18 February 2010 / Published online: 20 March 2010 # Springer-Verlag 2010 Abstract Glutathione peroxidase 1 (GPx1) plays an im- portant role in preventing cardiac dysfunction following ischemia-reperfusion injury. However, its role in protecting cardiac mitochondria against reoxygenation-induced reac- tive oxygen species (ROS) generation in vivo is unclear. We examined the role of GPx1 in protecting cardiac mitochondria against hypoxia–reoxygenation (HR) damage by testing for alterations in cardiac mitochondrial function. We used a two-dimensional gel electrophoresis proteomics analysis to examine the effects of reoxygenation on cardiac protein in wild-type (GPx1 +/+ ) and GPx1 knockout (GPx1 -/- ) mouse hearts. We identified 42 protein spots showing differential expression in the two groups. Sixteen of the proteins identified were located in mitochondria and were involved in a number of key metabolic pathways. To verify our proteomics findings functionally, we performed NADH autofluorescence measurements and ATP production assays. The reduced expression of oxidative phosphorylation pro- teins in GPx1 -/- mice following HR treatment resulted in loss of the mitochondrial membrane potential and decreased mitochondrial respiration. Mitochondrial ROS production and oxidative mtDNA damage were increased markedly during reoxygenation in GPx1 -/- hearts. We also found morphological abnormalities in cardiac mitochondria and myocytes in HR-treated GPx1 -/- . This is the first report of the role of GPx1 in protecting cardiac mitochondria against reoxygenation damage in vivo. These findings will help clarify the mechanisms of HR injury and will aid in the development of antioxidant therapies to prevent cardiac mitochondrial dysfunction associated with reoxygenation. Keywords Mitochondria . Glutathione peroxidase 1 (GPx1) . Hypoxia–reoxygenation . Reactive oxygen species Abbreviations 2-DE Two-dimensional gel electrophoresis ATP Adenosine triphosphate BSA Bovine serum albumin PBS Phosphate-buffered saline COX Complex EM Electron microscopy GPx1 Glutathione peroxidase 1 H 2 DCFDA Dichlorodihydrofluorescein diacetate H 2 O 2 Hydrogen peroxidase HR Hypoxia/reoxygenation IR Ischemia-reperfusion MALDI- TOF Matrix-assisted laser desorption/ionization– time of flight mass spectrometry mtDNA Mitochondrial DNA MS Mass spectrometry NADH Nicotinamide adenine dinucleotide ROS Reactive oxygen species Goo Taeg Oh and Jin Han contributed equally to this study V. T. Thu : H. K. Kim : S. H. Ha : N. Kim : J. Han (*) National Research Laboratory for Mitochondrial Signaling, Department of Physiology, College of Medicine, Cardiovascular and Metabolic Disease Center, FIRST Mitochondrial Research Group, Inje University, 633-165 Gaegeum-Dong, Busanjin-Gu, Busan 613-735, Korea e-mail: phyhanj@inje.ac.kr W. S. Park Department of Physiology, Kangwon National University School of Medicine, Chuncheon 200-701, Korea J.-Y. Yoo : G. T. Oh Department of Molecular Life Science, Ewha Women’ s University, Seoul, Korea Pflugers Arch - Eur J Physiol (2010) 460:55–68 DOI 10.1007/s00424-010-0811-7