Assessment of H2Sin vivousing the newly developed mitochondria-targeted mass spectrometry probe MitoA.
Hydrogen sulfide (H2S) is produced endogenouslyin vivoand has multiple effects on signaling pathways and cell function. Mitochondria can be both an H2S source and sink, and many of the biological effects of H2S relate to its interactions with mitochondria. However, the significance of mitochondrial H2S is uncertain, in part due to the difficulty of assessing changes in its concentrationin vivoAlthough a number of fluorescent H2S probes have been developed these are best suited to cells in culture and cannot be usedin vivoTo address this unmet need we have developed a mitochondria-targeted H2S probe, MitoA, which can be used to assess relative changes in mitochondrial H2S levelsin vivoMitoA comprises a lipophilic triphenylphosphonium (TPP) cation coupled to an aryl azide. The TPP cation leads to the accumulation of MitoA inside mitochondria within tissuesin vivoThere, the aryl azido group reacts with H2S to form an aryl amine (MitoN). The extent of conversion of MitoA to MitoN thus gives an indication of the levels of mitochondrial H2Sin vivoBoth compounds can be detected sensitively by liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis of the tissues, and quantified relative to deuterated internal standards. Here we describe the synthesis and characterization of MitoA and show that it can be used to assess changes in mitochondrial H2S levelsin vivoAs a proof of principle we used MitoA to show that H2S levels increasein vivoduring myocardial ischemia.
From the MRC Mitochondrial Biology Unit, University of Cambridge, Hills Road, Cambridge CB2 0XY, United Kingdom.
Sabine
S
Arndt
the WestCHEM School of Chemistry, University of Glasgow, Glasgow G12 8QQ, United Kingdom.
Carlos D
CD
Baeza-Garza
From the MRC Mitochondrial Biology Unit, University of Cambridge, Hills Road, Cambridge CB2 0XY, United Kingdom.
Angela
A
Logan
the Department of Medicine, University of Cambridge, Biomedical Campus, Cambridge, CB2 2QQ, United Kingdom.
Tiziana
T
Rosa
the Department of Chemistry and Pharmacy, Friedrich-Alexander University of Erlangen-Nuremberg, Egerlandstrasse,1, 91058 Erlangen, Germany.
Rudolf
R
Wedmann
From the MRC Mitochondrial Biology Unit, University of Cambridge, Hills Road, Cambridge CB2 0XY, United Kingdom.
Tracy A
TA
Prime
the Department of Surgery and Cambridge NIHR Biomedical Research Centre, Biomedical Campus, University of Cambridge, Cambridge CB2 2QQ, United Kingdom.
Jack L
JL
Martin
the Department of Surgery and Cambridge NIHR Biomedical Research Centre, Biomedical Campus, University of Cambridge, Cambridge CB2 2QQ, United Kingdom.
Kourosh
K
Saeb-Parsy
the Department of Medicine, University of Cambridge, Biomedical Campus, Cambridge, CB2 2QQ, United Kingdom.
Thomas
T
Krieg
the Department of Chemistry and Pharmacy, Friedrich-Alexander University of Erlangen-Nuremberg, Egerlandstrasse,1, 91058 Erlangen, Germany.|the University of Bordeaux, IBGC, UMR 5095, F-33077 Bordeaux, France, and.
Milos R
MR
Filipovic
the WestCHEM School of Chemistry, University of Glasgow, Glasgow G12 8QQ, United Kingdom, Richard.Hartley@glasgow.ac.uk.
Richard C
RC
Hartley
From the MRC Mitochondrial Biology Unit, University of Cambridge, Hills Road, Cambridge CB2 0XY, United Kingdom, mpm@mrc-mbu.cam.ac.uk.
Michael P
MP
Murphy
D000818:Animals; D002412:Cations; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D002853:Chromatography, Liquid; D005260:Female; D045325:HCT116 Cells; D006571:Heterocyclic Compounds / Q000737:chemistry; D006801:Humans; D006862:Hydrogen Sulfide / Q000737:chemistry*; D000860:Hypoxia; D008099:Liver / Q000378:metabolism; D008297:Male; D013058:Mass Spectrometry / Q000379:methods*; D051379:Mice; D008810:Mice, Inbred C57BL; D008928:Mitochondria / Q000378:metabolism*; D017202:Myocardial Ischemia / Q000378:metabolism; D009943:Organophosphorus Compounds / Q000737:chemistry; D017208:Rats, Wistar; D053719:Tandem Mass Spectrometry; D013696:Temperature; D014466:Ultraviolet Rays
D002412:Cations; D006571:Heterocyclic Compounds; D009943:Organophosphorus Compounds; C528765:tris(o-phenylenedioxy)cyclotriphosphazene; D006862:Hydrogen Sulfide
analytical chemistry; chemical biology; energy metabolism; hydrogen sulfide; hypoxia; mass spectrometry (MS); mitochondria