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EPR detection of cellular and mitochondrial superoxide using cyclic hydroxylamines Full article

Journal Free Radical Research
ISSN: 1071-5762
Output data Year: 2011, Volume: 45, Number: 4, Pages: 417-430 Pages count : DOI: 10.3109/10715762.2010.540242
Tags Superoxide; spin trap; cyclic hydroxylamine; intracellular; extracellular; EPR
Authors Dikalov Sergey I. 1 , Kirilyuk Igor A. 2 , Voinov Maxim 3 , Grigor'ev Igor A. 2
Affiliations
1 (Данные Web of science) Emory Univ, Sch Med, Div Cardiol, FRIMCORE, Atlanta, GA 30322 USA
2 (Данные Web of science) Novosibirsk Organ Chem Inst, Novosibirsk, Russia
3 (Данные Web of science) N Carolina State Univ, Raleigh, NC 27695 USA

Abstract: Superoxide (O(2)(center dot-)) has been implicated in the pathogenesis of many human diseases, but detection of the O(2)(center dot-) radicals in biological systems is limited due to inefficiency of O(2)(center dot-) spin trapping and lack of site-specific information. This work studied production of extracellular, intracellular and mitochondrial O(2)(center dot-) in neutrophils, cultured endothelial cells and isolated mitochondria using a new set of cationic, anionic and neutral hydroxylamine spin probes with various lipophilicity and cell permeability. Cyclic hydroxylamines rapidly react with O(2)(center dot-), producing stable nitroxides and allowing site-specific O(2)(center dot-) detection in intracellular, extracellular and mitochondrial compartments. Negatively charged 1-hydroxy-4-phosphono-oxy-2,2,6, 6-tetramethylpiperidine (PP-H) and positively charged 1-hydroxy-2,2,6,6-tetramethylpiperidin-4-yl-trimethylammonium (CAT1-H) detected only extramitochondrial O(2)(center dot-). Inhibition of EPR signal by SOD2 over-expression showed that mitochondria targeted mitoTEMPO-H detected intramitochondrial O(2)(center dot-) both in isolated mitochondria and intact cells. Both 1-hydroxy-3-carboxy-2,2,5,5-tetramethylpyrrolidine (CP-H) and 1-hydroxy-3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine (CM-H) detected an increase in cytoplasm O(2)(center dot-) stimulated by PMA, but only CM-H and mitoTEMPO-H showed an increase in rotenone-induced mitochondrial O(2)(center dot-). These data show that a new set of hydroxylamine spin probes provide unique information about site-specific production of the O(2)(center dot-) radical in extracellular or intracellular compartments, cytoplasm or mitochondria.
Cite: Dikalov S.I. , Kirilyuk I.A. , Voinov M. , Grigor'ev I.A.
EPR detection of cellular and mitochondrial superoxide using cyclic hydroxylamines
Free Radical Research. 2011. V.45. N4. P.417-430. DOI: 10.3109/10715762.2010.540242 WOS Scopus РИНЦ OpenAlex
Files: Full text from publisher
Dates:
Published online: Dec 3, 2010
Published print: Apr 1, 2011
Identifiers:
Web of science: WOS:000288217400005
Scopus: 2-s2.0-79952406501
Elibrary: 16644061
OpenAlex: W2170048907
Citing:
DB Citing
Web of science 134
Scopus 143
Elibrary 135
OpenAlex 162
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