Lija Swain1, Andrea Kesemeyer1, Stefanie Meyer-Roxlau1, Christiane Vettel1, Anke Zieseniss1, Annemarie Güntsch1, Aline Jatho1, Andreas Becker1, Maithily S Nanadikar1, Bruce Morgan1, Sven Dennerlein1, Ajay M Shah1, Ali El-Armouche1, Viacheslav O Nikolaev1, Dörthe M Katschinski2. 1. From the Institute of Cardiovascular Physiology, Georg August University Göttingen, Germany (L.S., A.K., A.Z., A.G., A.J., A.B., M.S.N., D.M.K.); Institute of Pharmacology, Technical University Dresden, Germany (S.M.-R., A.E.-A.); Institute of Experimental and Clinical Pharmacology and Toxicology, Medical Faculty Mannheim, University of Heidelberg, Mannheim, Germany (C.V.); Cellular Biochemistry, Department of Biology, University of Kaiserslautern, Germany (B.M.); Department of Cellular Biochemistry, University Medical Center Göttingen, Germany (S.D.); Cardiovascular Division, King's College London, British Heart Foundation Centre, United Kingdom (A.M.S.); and German Center for Cardiovascular Research, University Medical Center Hamburg-Eppendorf, Hamburg, Germany (V.O.N.); and Institute of Experimental Cardiovascular Research, Hamburg, Germany (V.O.N.). 2. From the Institute of Cardiovascular Physiology, Georg August University Göttingen, Germany (L.S., A.K., A.Z., A.G., A.J., A.B., M.S.N., D.M.K.); Institute of Pharmacology, Technical University Dresden, Germany (S.M.-R., A.E.-A.); Institute of Experimental and Clinical Pharmacology and Toxicology, Medical Faculty Mannheim, University of Heidelberg, Mannheim, Germany (C.V.); Cellular Biochemistry, Department of Biology, University of Kaiserslautern, Germany (B.M.); Department of Cellular Biochemistry, University Medical Center Göttingen, Germany (S.D.); Cardiovascular Division, King's College London, British Heart Foundation Centre, United Kingdom (A.M.S.); and German Center for Cardiovascular Research, University Medical Center Hamburg-Eppendorf, Hamburg, Germany (V.O.N.); and Institute of Experimental Cardiovascular Research, Hamburg, Germany (V.O.N.). doerthe.katschinski@med.uni-goettingen.de.
Abstract
RATIONALE: Changes in redox potentials of cardiac myocytes are linked to several cardiovascular diseases. Redox alterations are currently mostly described qualitatively using chemical sensors, which however do not allow quantifying redox potentials, lack specificity, and the possibility to analyze subcellular domains. Recent advances to quantitatively describe defined redox changes include the application of genetically encoded redox biosensors. OBJECTIVE: Establishment of mouse models, which allow the quantification of the glutathione redox potential (EGSH) in the cytoplasm and the mitochondrial matrix of isolated cardiac myocytes and in Langendorff-perfused hearts based on the use of the redox-sensitive green fluorescent protein 2, coupled to the glutaredoxin 1 (Grx1-roGFP2). METHODS AND RESULTS: We generated transgenic mice with cardiac myocyte-restricted expression of Grx1-roGFP2 targeted either to the mitochondrial matrix or to the cytoplasm. The response of the roGFP2 toward H2O2, diamide, and dithiothreitol was titrated and used to determine the EGSH in isolated cardiac myocytes and in Langendorff-perfused hearts. Distinct EGSH were observed in the cytoplasm and the mitochondrial matrix. Stimulation of the cardiac myocytes with isoprenaline, angiotensin II, or exposure to hypoxia/reoxygenation additionally underscored that these compartments responded independently. A compartment-specific response was also observed 3 to 14 days after myocardial infarction. CONCLUSIONS: We introduce redox biosensor mice as a new tool, which allows quantification of defined alterations of EGSH in the cytoplasm and the mitochondrial matrix in cardiac myocytes and can be exploited to answer questions in basic and translational cardiovascular research.
RATIONALE: Changes in redox potentials of cardiac myocytes are linked to several cardiovascular diseases. Redox alterations are currently mostly described qualitatively using chemical sensors, which however do not allow quantifying redox potentials, lack specificity, and the possibility to analyze subcellular domains. Recent advances to quantitatively describe defined redox changes include the application of genetically encoded redox biosensors. OBJECTIVE: Establishment of mouse models, which allow the quantification of the glutathione redox potential (EGSH) in the cytoplasm and the mitochondrial matrix of isolated cardiac myocytes and in Langendorff-perfused hearts based on the use of the redox-sensitive green fluorescent protein 2, coupled to the glutaredoxin 1 (Grx1-roGFP2). METHODS AND RESULTS: We generated transgenic mice with cardiac myocyte-restricted expression of Grx1-roGFP2 targeted either to the mitochondrial matrix or to the cytoplasm. The response of the roGFP2 toward H2O2, diamide, and dithiothreitol was titrated and used to determine the EGSH in isolated cardiac myocytes and in Langendorff-perfused hearts. Distinct EGSH were observed in the cytoplasm and the mitochondrial matrix. Stimulation of the cardiac myocytes with isoprenaline, angiotensin II, or exposure to hypoxia/reoxygenation additionally underscored that these compartments responded independently. A compartment-specific response was also observed 3 to 14 days after myocardial infarction. CONCLUSIONS: We introduce redox biosensor mice as a new tool, which allows quantification of defined alterations of EGSH in the cytoplasm and the mitochondrial matrix in cardiac myocytes and can be exploited to answer questions in basic and translational cardiovascular research.
Authors: Shan Lu; Zhandi Liao; Xiyuan Lu; Dörthe M Katschinski; Mark Mercola; Ju Chen; Joan Heller Brown; Jeffery D Molkentin; Julie Bossuyt; Donald M Bers Journal: Circ Res Date: 2020-03-05 Impact factor: 17.367
Authors: David M Ryba; Chad M Warren; Chehade N Karam; Robert T Davis; Shamim A K Chowdhury; Manuel G Alvarez; Maximilian McCann; Chong Wee Liew; David F Wieczorek; Peter Varga; R John Solaro; Beata M Wolska Journal: Circ Heart Fail Date: 2019-11-05 Impact factor: 8.790
Authors: Maithily S Nanadikar; Ana M Vergel Leon; Sergej Borowik; Annette Hillemann; Anke Zieseniss; Vsevolod V Belousov; Ivan Bogeski; Peter Rehling; Jan Dudek; Dörthe M Katschinski Journal: Redox Biol Date: 2019-02-23 Impact factor: 11.799