title: Intracellular oxidation of hydroethidine: compartmentalization and cytotoxicity of oxidation products.
abstract: Hydroethidine (HE) is a blue fluorescent dye that is intracellularly converted into red-emitting products on two-electron oxidation. One of these products, namely 2-hydroxyethidium, is formed as the result of HE superoxide anion-specific oxidation, and so HE is widely used for the detection of superoxide in cells and tissues. In our experiments we exploited three cell lines of different origin: K562 (human leukemia cells), A431 (human epidermoid carcinoma cells), and SCE2304 (human mesenchymal stem cells derived from endometrium). Using fluorescent microscopy and flow cytometry analysis, we showed that HE intracellular oxidation products accumulate mostly in the cell mitochondria. This accumulation provokes gradual depolarization of mitochondrial membrane, affects oxygen consumption rate in HE-treated cells, and causes cellular apoptosis in the case of high HE concentrations and/or long cell incubations with HE, as well as a high rate of HE oxidation in cells exposed to some stimuli.
authors.affiliation: Department of Intracellular Signaling and Transport, Institute of Cytology, Russian Academy of Sciences, Tikhoretsky pr. 4, St. Petersburg 94064, Russia. Electronic address: o.lyublinskaya@mail.ru.
authors.forename: O G
authors.initials: OG
authors.lastname: Lyublinskaya
authors.affiliation: Department of Intracellular Signaling and Transport, Institute of Cytology, Russian Academy of Sciences, Tikhoretsky pr. 4, St. Petersburg 94064, Russia. Electronic address: vvzenin@yandex.ru.
authors.forename: V V
authors.initials: VV
authors.lastname: Zenin
authors.affiliation: Department of Intracellular Signaling and Transport, Institute of Cytology, Russian Academy of Sciences, Tikhoretsky pr. 4, St. Petersburg 94064, Russia. Electronic address: shatrova@mail.ru.
authors.forename: A N
authors.initials: AN
authors.lastname: Shatrova
authors.affiliation: Department of Intracellular Signaling and Transport, Institute of Cytology, Russian Academy of Sciences, Tikhoretsky pr. 4, St. Petersburg 94064, Russia. Electronic address: aksenovn@gmail.com.
authors.forename: N D
authors.initials: ND
authors.lastname: Aksenov
authors.affiliation: Department of Intracellular Signaling and Transport, Institute of Cytology, Russian Academy of Sciences, Tikhoretsky pr. 4, St. Petersburg 94064, Russia. Electronic address: vzemelko@mail.ru.
authors.forename: V I
authors.initials: VI
authors.lastname: Zemelko
authors.affiliation: Department of Intracellular Signaling and Transport, Institute of Cytology, Russian Academy of Sciences, Tikhoretsky pr. 4, St. Petersburg 94064, Russia. Electronic address: aldomnina@mail.ru.
authors.forename: A P
authors.initials: AP
authors.lastname: Domnina
authors.affiliation: Department of Intracellular Signaling and Transport, Institute of Cytology, Russian Academy of Sciences, Tikhoretsky pr. 4, St. Petersburg 94064, Russia. Electronic address: anyasun@mail.ru.
authors.forename: A P
authors.initials: AP
authors.lastname: Litanyuk
authors.affiliation: Department of Intracellular Signaling and Transport, Institute of Cytology, Russian Academy of Sciences, Tikhoretsky pr. 4, St. Petersburg 94064, Russia. Electronic address: lenbur87@mail.ru.
authors.forename: E B
authors.initials: EB
authors.lastname: Burova
authors.affiliation: Department of Medical Physics and Bioengineering, Institute of Physics, Nanotechnology and Telecommunications, St. Petersburg State Polytechnical University, Polytechnicheskaya st. 29, St. Petersburg 195251, Russia. Electronic address: svyatoslav_gubarev@mail.ru.
authors.forename: S S
authors.initials: SS
authors.lastname: Gubarev
authors.affiliation: Department of Intracellular Signaling and Transport, Institute of Cytology, Russian Academy of Sciences, Tikhoretsky pr. 4, St. Petersburg 94064, Russia; Department of Medical Physics and Bioengineering, Institute of Physics, Nanotechnology and Telecommunications, St. Petersburg State Polytechnical University, Polytechnicheskaya st. 29, St. Petersburg 195251, Russia. Electronic address: yurineg@mail.cytspb.rssi.ru.
authors.forename: Y A
authors.initials: YA
authors.lastname: Negulyaev
authors.affiliation: Department of Intracellular Signaling and Transport, Institute of Cytology, Russian Academy of Sciences, Tikhoretsky pr. 4, St. Petersburg 94064, Russia; Department of Medical Physics and Bioengineering, Institute of Physics, Nanotechnology and Telecommunications, St. Petersburg State Polytechnical University, Polytechnicheskaya st. 29, St. Petersburg 195251, Russia. Electronic address: nikolsky@mail.cytspb.rssi.ru.
authors.forename: N N
authors.initials: NN
authors.lastname: Nikolsky
mesh_terms: D017209:Apoptosis / Q000502:physiology; D045744:Cell Line, Tumor; D004996:Ethidium / Q000031:analogs & derivatives / Q000737:chemistry; D005434:Flow Cytometry; D005456:Fluorescent Dyes / Q000737:chemistry / Q000494:pharmacology*; D006801:Humans; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000378:metabolism*; D051336:Mitochondrial Membranes / Q000502:physiology*; D010084:Oxidation-Reduction; D010101:Oxygen Consumption / Q000502:physiology; D010617:Phenanthridines / Q000737:chemistry / Q000494:pharmacology*; D013481:Superoxides / Q000737:chemistry / Q000378:metabolism*
chemical_list: C506392:2-hydroxyethidium; D005456:Fluorescent Dyes; D010617:Phenanthridines; D013481:Superoxides; C058428:hydroethidine; D004996:Ethidium
keywords: 2-Hydroxyethidium; Ethidium; Flow cytometry; Fluorescent probes; Hydroethidine; Superoxide detection