title: AP39, a novel mitochondria-targeted hydrogen sulfide donor, stimulates cellular bioenergetics, exerts cytoprotective effects and protects against the loss of mitochondrial DNA integrity in oxidatively stressed endothelial cells in vitro.
abstract: The purpose of the current study was to investigate the effect of the recently synthesized mitochondrially-targeted H2S donor, AP39 [(10-oxo-10-(4-(3-thioxo-3H-1,2-dithiol-5yl)phenoxy)decyl) triphenylphosphonium bromide], on bioenergetics, viability, and mitochondrial DNA integrity in bEnd.3 murine microvascular endothelial cells in vitro, under normal conditions, and during oxidative stress. Intracellular H2S was assessed by the fluorescent dye 7-azido-4-methylcoumarin. For the measurement of bioenergetic function, the XF24 Extracellular Flux Analyzer was used. Cell viability was estimated by the combination of the MTT and LDH methods. Oxidative protein modifications were measured by the Oxyblot method. Reactive oxygen species production was monitored by the MitoSOX method. Mitochondrial and nuclear DNA integrity were assayed by the Long Amplicon PCR method. Oxidative stress was induced by addition of glucose oxidase. Addition of AP39 (30-300 nM) to bEnd.3 cells increased intracellular H2S levels, with a preferential response in the mitochondrial regions. AP39 exerted a concentration-dependent effect on mitochondrial activity, which consisted of a stimulation of mitochondrial electron transport and cellular bioenergetic function at lower concentrations (30-100 nM) and an inhibitory effect at the higher concentration of 300 nM. Under oxidative stress conditions induced by glucose oxidase, an increase in oxidative protein modification and an enhancement in MitoSOX oxidation was noted, coupled with an inhibition of cellular bioenergetic function and a reduction in cell viability. AP39 pretreatment attenuated these responses. Glucose oxidase induced a preferential damage to the mitochondrial DNA; AP39 (100 nM) pretreatment protected against it. In conclusion, the current paper documents antioxidant and cytoprotective effects of AP39 under oxidative stress conditions, including a protection against oxidative mitochondrial DNA damage.
authors.affiliation: Department of Anesthesiology, University of Texas Medical Branch, Galveston, TX, USA.
authors.forename: Bartosz
authors.initials: B
authors.lastname: Szczesny
authors.affiliation: Department of Anesthesiology, University of Texas Medical Branch, Galveston, TX, USA.
authors.forename: Katalin
authors.initials: K
authors.lastname: Módis
authors.affiliation: Department of Anesthesiology, University of Texas Medical Branch, Galveston, TX, USA.
authors.forename: Kazunori
authors.initials: K
authors.lastname: Yanagi
authors.affiliation: Department of Anesthesiology, University of Texas Medical Branch, Galveston, TX, USA.
authors.forename: Ciro
authors.initials: C
authors.lastname: Coletta
authors.affiliation: University of Exeter Medical School, St. Luke's Campus, Exeter, England, United Kingdom.
authors.forename: Sophie
authors.initials: S
authors.lastname: Le Trionnaire
authors.affiliation: Biosciences, College of Life and Environmental Science, University of Exeter, England, United Kingdom.
authors.forename: Alexis
authors.initials: A
authors.lastname: Perry
authors.affiliation: Biosciences, College of Life and Environmental Science, University of Exeter, England, United Kingdom.
authors.forename: Mark E
authors.initials: ME
authors.lastname: Wood
authors.affiliation: University of Exeter Medical School, St. Luke's Campus, Exeter, England, United Kingdom. Electronic address: m.whiteman@exeter.ac.uk.
authors.forename: Matthew
authors.initials: M
authors.lastname: Whiteman
authors.affiliation: Department of Anesthesiology, University of Texas Medical Branch, Galveston, TX, USA. Electronic address: szabocsaba@aol.com.
authors.forename: Csaba
authors.initials: C
authors.lastname: Szabo
mesh_terms: D000818:Animals; D002460:Cell Line; D004260:DNA Repair / Q000187:drug effects; D004272:DNA, Mitochondrial / Q000187:drug effects*; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism; D005949:Glucose Oxidase / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000032:analysis / Q000378:metabolism; D006862:Hydrogen Sulfide / Q000494:pharmacology*; D042541:Intracellular Space / Q000187:drug effects; D051379:Mice; D010755:Organophosphates / Q000494:pharmacology*; D009943:Organophosphorus Compounds / Q000494:pharmacology*; D018384:Oxidative Stress / Q000187:drug effects*; D020011:Protective Agents / Q000494:pharmacology*; D013871:Thiones / Q000494:pharmacology*
chemical_list: C000610725:AP39 compound; D004272:DNA, Mitochondrial; D010755:Organophosphates; D009943:Organophosphorus Compounds; D020011:Protective Agents; D013871:Thiones; D006861:Hydrogen Peroxide; D005949:Glucose Oxidase; D006862:Hydrogen Sulfide
keywords: Bioenergetics; Cytoprotection; DNA repair; Mitochondria; Oxidative stress
references.citation: Life Sci. 2009 Mar 13;84(11-12):328-36
references.pmid: 19159629
references.citation: J Biol Chem. 2012 Jan 2;287(1):81-89
references.pmid: 22086924
references.citation: Nitric Oxide. 2013 Nov 30;35:5-20
references.pmid: 23850632
references.citation: J Smooth Muscle Res. 2012;48(1):1-26
references.pmid: 22504486
references.citation: Nitric Oxide. 2014 Sep 15;41:146-56
references.pmid: 24667534
references.citation: Diabetologia. 2010 Aug;53(8):1722-6
references.pmid: 20414636
references.citation: Proc Natl Acad Sci U S A. 2013 Jul 30;110(31):12679-84
references.pmid: 23858469
references.citation: Free Radic Biol Med. 2012 Aug 1;53(3):595-609
references.pmid: 22595027
references.citation: Aging Male. 2012 Sep;15(3):153-8
references.pmid: 22455401
references.citation: Biochimie. 2013 Nov;95(11):2069-75
references.pmid: 23896375
references.citation: Antioxid Redox Signal. 2011 Jul 15;15(2):405-19
references.pmid: 20812864
references.citation: Antioxid Redox Signal. 2010 May 15;12(10):1203-17
references.pmid: 19769484
references.citation: PLoS One. 2013;8(2):e53147
references.pmid: 23393548
references.citation: Mol Cells. 2006 Aug 31;22(1):21-9
references.pmid: 16951546
references.citation: Free Radic Biol Med. 2011 Jan 1;50(1):196-205
references.pmid: 21034811
references.citation: Proc Natl Acad Sci U S A. 2007 Sep 25;104(39):15560-5
references.pmid: 17878306
references.citation: Biochem Biophys Res Commun. 2006 Apr 28;343(1):303-10
references.pmid: 16540095
references.citation: Nat Rev Drug Discov. 2007 Nov;6(11):917-35
references.pmid: 17948022
references.citation: Proc Natl Acad Sci U S A. 2013 Jul 23;110(30):12474-9
references.pmid: 23836652
references.citation: Diabetes. 2013 Mar;62(3):953-64
references.pmid: 23223176
references.citation: Anal Chem. 2009 Aug 15;81(16):6868-78
references.pmid: 19555051
references.citation: Bioorg Med Chem Lett. 2013 Nov 1;23(21):5878-81
references.pmid: 24055044
references.citation: Mech Ageing Dev. 2012 Apr;133(4):157-68
references.pmid: 22313689
references.citation: Shock. 2010 Sep;34 Suppl 1:4-14
references.pmid: 20523270
references.citation: Biochem Biophys Res Commun. 2012 May 4;421(2):164-9
references.pmid: 22503984
references.citation: J Cell Mol Med. 2009 Mar;13(3):488-507
references.pmid: 19374684
references.citation: FEBS Lett. 2009 Jun 18;583(12):1945-50
references.pmid: 19427865
references.citation: Circulation. 2013 Jun 25;127(25):2523-34
references.pmid: 23704252
references.citation: Methods Enzymol. 2013;526:159-73
references.pmid: 23791100
references.citation: Drug Discov Today. 2008 Mar;13(5-6):268-74
references.pmid: 18342804
references.citation: Curr Atheroscler Rep. 2012 Jun;14(3):264-76
references.pmid: 22350585
references.citation: Curr Pharm Des. 2011;17(22):2253-63
references.pmid: 21736540
references.citation: Curr Med Chem. 2012;19(20):3325-36
references.pmid: 22680638
references.citation: Br J Pharmacol. 2014 Apr;171(8):2099-122
references.pmid: 23991830
references.citation: Infect Disord Drug Targets. 2009 Aug;9(4):376-89
references.pmid: 19689380
references.citation: Chin Med J (Engl). 2011 Nov;124(21):3445-9
references.pmid: 22340156
references.citation: Acta Diabetol. 2014 Feb;51(1):155-7
references.pmid: 23852531
references.citation: Oxid Med Cell Longev. 2012;2012:878052
references.pmid: 23304257
references.citation: Biochim Biophys Acta. 2014 Apr;1840(4):1404-12
references.pmid: 24246956
references.citation: J Biol Chem. 2011 Sep 16;286(37):31975-83
references.pmid: 21768646
references.citation: J Intern Med. 2013 Jun;273(6):529-43
references.pmid: 23432181
references.citation: Bioorg Med Chem Lett. 2011 Apr 1;21(7):2000-2
references.pmid: 21376590
references.citation: Proc Natl Acad Sci U S A. 2003 Apr 29;100(9):5407-12
references.pmid: 12697897
references.citation: Clin Chem Lab Med. 2012 Feb 01;50(8):1287-95
references.pmid: 22868792
references.citation: Front Biosci (Landmark Ed). 2011 Jan 01;16:1986-95
references.pmid: 21196278
references.citation: Physiol Rev. 2012 Apr;92(2):791-896
references.pmid: 22535897
references.citation: Antioxid Redox Signal. 2010 May 15;12(10):1147-54
references.pmid: 19769459
references.citation: Angew Chem Int Ed Engl. 2013 Apr 22;52(17):4641-4
references.pmid: 23512751
references.citation: Biol Chem. 2012 Dec;393(12):1485-1512
references.pmid: 23092819
references.citation: Biochim Biophys Acta. 2013 Jan;1832(1):174-82
references.pmid: 22846607
references.citation: Antioxid Redox Signal. 2011 Dec 15;15(12):3021-38
references.pmid: 21395490
references.citation: Ann N Y Acad Sci. 2010 Jul;1201:96-103
references.pmid: 20649545
references.citation: FASEB J. 2013 Feb;27(2):601-11
references.pmid: 23104984
references.citation: Neurochem Int. 2013 Dec;63(8):826-31
references.pmid: 23770272
references.citation: J Med Chem. 2013 Nov 27;56(22):9170-9
references.pmid: 24147900
references.citation: Oxid Med Cell Longev. 2012;2012:324256
references.pmid: 22811764
references.citation: Free Radic Biol Med. 2013 Jul;60:325-35
references.pmid: 23485584
references.citation: Mutat Res. 1997 Nov;385(2):139-49
references.pmid: 9447235
references.citation: Antioxid Redox Signal. 2011 Jul 15;15(2):379-91
references.pmid: 21028947
references.citation: Am J Physiol Heart Circ Physiol. 2010 May;298(5):H1310-9
references.pmid: 20154265
references.citation: Mol Pharmacol. 2009 Jan;75(1):27-34
references.pmid: 18832435
references.citation: Biochem J. 2012 Jan 15;441(2):609-21
references.pmid: 21950347
references.citation: Br J Pharmacol. 2011 Oct;164(3):853-65
references.pmid: 21198548
references.citation: Biochem Biophys Res Commun. 2013 Apr 19;433(4):401-7
references.pmid: 23537657
references.citation: Chem Commun (Camb). 2013 Nov 14;49(88):10361-3
references.pmid: 24072060
references.citation: FASEB J. 2007 Jun;21(8):1699-706
references.pmid: 17314140
references.citation: Toxicology. 2004 Oct 15;203(1-3):69-76
references.pmid: 15363583
references.citation: PLoS One. 2013 Sep 16;8(9):e75201
references.pmid: 24066171
references.citation: Clin Sci (Lond). 2011 Dec;121(11):459-88
references.pmid: 21843150
references.citation: Free Radic Biol Med. 2010 Apr 1;48(7):905-14
references.pmid: 20093177
references.citation: Biochem Pharmacol. 2013 Nov 1;86(9):1311-9
references.pmid: 24012591
references.citation: Antioxid Redox Signal. 2012 Jul 1;17(1):68-80
references.pmid: 22149162
references.citation: Free Radic Biol Med. 2010 Apr 15;48(8):1064-70
references.pmid: 20100568
references.citation: Antioxid Redox Signal. 2010 Apr;12(4):537-77
references.pmid: 19650713
references.citation: Proc Natl Acad Sci U S A. 2011 Aug 16;108(33):13829-34
references.pmid: 21808008
references.citation: Exp Physiol. 2011 Sep;96(9):847-52
references.pmid: 21666034
references.citation: Chem Biol. 2012 Oct 26;19(10):1237-46
references.pmid: 23102218
references.citation: Antioxid Redox Signal. 2012 Jul 1;17(1):119-40
references.pmid: 22432697
references.citation: Exp Biol Med (Maywood). 2013 May;238(5):450-60
references.pmid: 23856898
references.citation: Circulation. 2013 Jun 25;127(25):2514-22
references.pmid: 23704251
references.citation: Nephrol Dial Transplant. 2009 Dec;24(12):3756-63
references.pmid: 19654230