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On the use of fluorescence lifetime imaging and dihydroethidium to detect superoxide in intact animals and ex vivo tissues: A reassessment |
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| Institution: | 1. Department of Biophysics and Free Radical Research Center, Medical College of Wisconsin, Milwaukee, WI 53226, USA;2. Institute of Applied Radiation Chemistry, Lodz University of Technology, 90-924 Lodz, Poland;1. Department of Chemistry, College of Science, University of Tikrit, Tikrit, Iraq;2. Department of Chemistry, College of Education for Pure Science, University of Tikrit, Tikrit, Iraq;3. Department of Nursing, Al-Hadi University College, Baghdad, Iraq;4. Center of Excellence on Solar Cells & Renewable Energy, Department of Physics, School of Basic Sciences and Research, Sharda University, Great er Noida, Uttar Pradesh 201308, India;1. Department of Pathology, Columbia University, NY, NY 10032, USA;2. Dipartimento di Medicina Sperimentale e Clinica, Università degli Studi di Firenze, Firenze 50134, Italy;1. Department of Endocrinology, Institute of Zoology, Jagiellonian University, Krakow, Poland;2. Department of Cell Biology, Poznan University of Medical Sciences, Poznan, Poland;1. Key Laboratory for Advanced Materials and Institute of Fine Chemicals, East China University of Science & Technology, Shanghai 200237, PR China;2. School of Chemical Biology and Pharmaceutical Science, Capital Medical University, Beijing 100069, PR China;1. Whitehead Institute for Biomedical Research, 9 Cambridge Center, Cambridge, MA 02142, USA;2. Department of Biology, Massachusetts Institute of Technology (MIT), Cambridge, MA 02139, USA;3. Broad Institute, Seven Cambridge Center, Cambridge, MA 02142, USA;4. David H. Koch Institute for Integrative Cancer Research at MIT, 77 Massachusetts Avenue, Cambridge, MA 02139, USA;5. Howard Hughes Medical Institute, MIT, Cambridge, MA 02139, USA;6. Department of Cell Biology, New York University School of Medicine, New York, NY 10016, USA;7. Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305, USA;8. Department of Biochemistry, Netherlands Cancer Institute, Plesmanlaan 121 1066 CX, Amsterdam, the Netherlands |
| Abstract: | Recently, D.J. Hall et al. reported that ethidium (E+) is formed as a major product of hydroethidine (HE) or dihydroethidium reaction with superoxide (O2 ?) in intact animals with low tissue oxygen levels (J. Cereb. Blood Flow Metab. 32:23–32, 2012). The authors concluded that measurement of E+ is an indicator of O2? formation in intact brains of animals. This finding is in stark contrast to previous reports using in vitro systems showing that 2-hydroxyethidium, not ethidium, is formed from the reaction between O2? and HE. Published in vivo results support the in vitro findings. In this study, we performed additional experiments in which HE oxidation products were monitored under different fluxes of O2?. Results from these experiments further reaffirm our earlier findings (H. Zhao et al., Free Radic. Biol. Med. 34:1359, 2003). We conclude that whether in vitro or in vivo, E+ measured by HPLC or by fluorescence lifetime imaging is not a diagnostic marker product for O2? reaction with HE. |
| Keywords: | 2-hydroxyethidium DE"} {"#name":"keyword" "$":{"id":"key0020"} "$$":[{"#name":"text" "_":"deuteroethidine DHE"} {"#name":"keyword" "$":{"id":"key0030"} "$$":[{"#name":"text" "_":"dihydroethidium dtpa"} {"#name":"keyword" "$":{"id":"key0040"} "$$":[{"#name":"text" "_":"diethylenetriaminepentaacetate ethidium HE"} {"#name":"keyword" "$":{"id":"key0060"} "$$":[{"#name":"text" "_":"hydroethidine HX"} {"#name":"keyword" "$":{"id":"key0070"} "$$":[{"#name":"text" "_":"hypoxanthine X"} {"#name":"keyword" "$":{"id":"key0080"} "$$":[{"#name":"text" "_":"xanthine XO"} {"#name":"keyword" "$":{"id":"key0090"} "$$":[{"#name":"text" "_":"xanthine oxidase HRP"} {"#name":"keyword" "$":{"id":"key0100"} "$$":[{"#name":"text" "_":"horseradish peroxidase TFA"} {"#name":"keyword" "$":{"id":"key0110"} "$$":[{"#name":"text" "_":"trifluoroacetic acid |
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