The effect of oxygen in Sirt3-mediated myocardial protection: a proof-of-concept study in cultured cardiomyoblasts
- 21 Downloads
Sirtuin 3 is a nicotinamide adenine dinucleotide dependent mitochondrial deacetylase that governs mitochondrial metabolism and oxidative defense. The demise in myocardial function following myocardial ischemia has been associated with mitochondrial dysfunction. Sirt3 maintains myocardial contractile function and protects from cardiac hypertrophy. The role of Sirt3 in ischemia is controversial. Our objective was to understand, under what circumstances Sirt3 is protective in different facets of ischemia, using an in vitro proof-of-concept approach based on simulated ischemia in cultured cardiomyoblasts. Cultured H9c2 cardiomyoblasts were subjected to hypoxia and/or serum deprivation, the combination of which we refer to as simulated ischemia. Apoptosis, as assessed by Annexin V staining in life-cell imaging and propidium-iodide inclusion in flow cytometry, was enhanced following simulated ischemia. Interestingly, serum deprivation was a stronger trigger of apoptosis than hypoxia. Knockdown of Sirt3 further increased apoptosis upon serum deprivation, whereas no such effect occurred upon additional hypoxia. Similarly, only upon serum deprivation but not upon simulated ischemia, silencing of Sirt3 led to a deterioration of mitochondrial function in extracellular flux analysis. In the absence of oxygen these Sirt3-dependent effects were abolished. These data indicate, that Sirt3-mediated myocardial protection is oxygen-dependent. Thus, mitochondrial respiration takes center-stage in Sirt3-dependent prevention of stress-induced myocardial damage.
KeywordsSirt3 Cardiomyoblasts Ischemia Oxygen Mitochondrial function
This work was funded by the National Health and Medical Research Council (NHMRC), the Victorian Government’s Operational Infrastructure Support Program, the Swiss National Science Foundation (310030, 146923), Matching Funds at the University of Zurich and the Zurich Heart House, Zurich, Switzerland. PD was supported by the German Research Foundation. JS was supported by the German Society of Cardiology. KP is a Principal Research Fellow of the NHMRC.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
- 1.Moran AE, Oliver JT, Mirzaie M, Forouzanfar MH, Chilov M, Anderson L, Morrison JL, Khan A, Zhang N, Haynes N, Tran J, Murphy A, Degennaro V, Roth G, Zhao D, Peer N, Pichon-Riviere A, Rubinstein A, Pogosova N, Prabhakaran D, Naghavi M, Ezzati M, Mensah GA (2012) Assessing the global burden of ischemic heart disease: part 1: methods for a systematic review of the global epidemiology of ischemic heart disease in 1990 and 2010. Glob Heart 7(4):315–329. https://doi.org/10.1016/j.gheart.2012.10.004 CrossRefPubMedPubMedCentralGoogle Scholar
- 3.Roes SD, Kelle S, Kaandorp TA, Kokocinski T, Poldermans D, Lamb HJ, Boersma E, van der Wall EE, Fleck E, de Roos A, Nagel E, Bax JJ (2007) Comparison of myocardial infarct size assessed with contrast-enhanced magnetic resonance imaging and left ventricular function and volumes to predict mortality in patients with healed myocardial infarction. Am J Cardiol 100(6):930–936. https://doi.org/10.1016/j.amjcard.2007.04.029 CrossRefPubMedGoogle Scholar
- 4.Wallace DC (2000) Mitochondrial defects in cardiomyopathy and neuromuscular disease. Am Heart J 139(2 Pt 3):S70–S85Google Scholar
- 9.Sundaresan NR, Samant SA, Pillai VB, Rajamohan SB, Gupta MP (2008) SIRT3 is a stress-responsive deacetylase in cardiomyocytes that protects cells from stress-mediated cell death by deacetylation of Ku70. Mol Cell Biol 28(20):6384–6401. https://doi.org/10.1128/MCB.00426-08 CrossRefPubMedPubMedCentralGoogle Scholar
- 11.Cheung KG, Cole LK, Xiang B, Chen K, Ma X, Myal Y, Hatch GM, Tong Q, Dolinsky VW (2015) Sirtuin-3 (SIRT3) protein attenuates doxorubicin-induced oxidative stress and improves mitochondrial respiration in H9c2 cardiomyocytes. J Biol Chem 290(17):10981–10993. https://doi.org/10.1074/jbc.M114.607960 CrossRefPubMedPubMedCentralGoogle Scholar
- 15.Koentges C, Pfeil K, Schnick T, Wiese S, Dahlbock R, Cimolai MC, Meyer-Steenbuck M, Cenkerova K, Hoffmann MM, Jaeger C, Odening KE, Kammerer B, Hein L, Bode C, Bugger H (2015) SIRT3 deficiency impairs mitochondrial and contractile function in the heart. Basic Res Cardiol 110(4):36. https://doi.org/10.1007/s00395-015-0493-6 CrossRefPubMedGoogle Scholar
- 16.Koentges C, Pfeil K, Meyer-Steenbuck M, Lother A, Hoffmann MM, Odening KE, Hein L, Bode C, Bugger H (2016) Preserved recovery of cardiac function following ischemia-reperfusion in mice lacking SIRT3. Can J Physiol Pharmacol 94(1):72–80. https://doi.org/10.1139/cjpp-2015-0152 CrossRefPubMedGoogle Scholar
- 17.Bochaton T, Crola-Da-Silva C, Pillot B, Villedieu C, Ferreras L, Alam MR, Thibault H, Strina M, Gharib A, Ovize M, Baetz D (2015) Inhibition of myocardial reperfusion injury by ischemic postconditioning requires sirtuin 3-mediated deacetylation of cyclophilin D. J Mol Cell Cardiol 84:61–69. https://doi.org/10.1016/j.yjmcc.2015.03.017 CrossRefPubMedGoogle Scholar
- 18.Kuznetsov AV, Javadov S, Sickinger S, Frotschnig S, Grimm M (2015) H9c2 and HL-1 cells demonstrate distinct features of energy metabolism, mitochondrial function and sensitivity to hypoxia-reoxygenation. Biochim Biophys Acta 1853(2):276–284. https://doi.org/10.1016/j.bbamcr.2014.11.015 CrossRefPubMedGoogle Scholar
- 23.Charles I, Khalyfa A, Kumar DM, Krishnamoorthy RR, Roque RS, Cooper N, Agarwal N (2005) Serum deprivation induces apoptotic cell death of transformed rat retinal ganglion cells via mitochondrial signaling pathways. Invest Ophthalmol Vis Sci 46(4):1330–1338. https://doi.org/10.1167/iovs.04-0363 CrossRefPubMedGoogle Scholar
- 25.Hirschey MD, Shimazu T, Goetzman E, Jing E, Schwer B, Lombard DB, Grueter CA, Harris C, Biddinger S, Ilkayeva OR, Stevens RD, Li Y, Saha AK, Ruderman NB, Bain JR, Newgard CB, Farese RV Jr, Alt FW, Kahn CR, Verdin E (2010) SIRT3 regulates mitochondrial fatty-acid oxidation by reversible enzyme deacetylation. Nature 464(7285):121–125. https://doi.org/10.1038/nature08778 CrossRefPubMedPubMedCentralGoogle Scholar
- 26.Stride N, Larsen S, Hey-Mogensen M, Sander K, Lund JT, Gustafsson F, Kober L, Dela F (2013) Decreased mitochondrial oxidative phosphorylation capacity in the human heart with left ventricular systolic dysfunction. Eur J Heart Fail 15(2):150–157. https://doi.org/10.1093/eurjhf/hfs172 CrossRefPubMedGoogle Scholar
- 29.Argaud L, Gateau-Roesch O, Raisky O, Loufouat J, Robert D, Ovize M (2005) Postconditioning inhibits mitochondrial permeability transition. Circulation 111(2):194–197. https://doi.org/10.1161/01.CIR.0000151290.04952.3B CrossRefPubMedGoogle Scholar
- 30.Robich MP, Chu LM, Burgess TA, Feng J, Han Y, Nezafat R, Leber MP, Laham RJ, Manning WJ, Sellke FW (2012) Resveratrol preserves myocardial function and perfusion in remote nonischemic myocardium in a swine model of metabolic syndrome. J Am Coll Surg 215(5):681–689. https://doi.org/10.1016/j.jamcollsurg.2012.06.417 CrossRefPubMedPubMedCentralGoogle Scholar