Orientin-Induced Cardioprotection against Reperfusion Is Associated with Attenuation of Mitochondrial Permeability Transition

Abstract

In this study, we provide new evidence that orientin from bamboo leaves (Phyllostachys nigra) protect H9c2 cardiomyocytes against ischemia/reperfusion (I/R) injury through the mitochondrial apoptotic pathway. A previous work has identified that orientin could protect myocardium against ischemia/reperfusion injury. Mitochondria are both critical determinants of cardioprotection and crucial targets of cardioprotective signaling. Their role during reperfusion is conspicuously critical because the conditions promote apoptosis through the mitochondrial pathway and necrosis though irreversible damage to mitochondria, which is in association with mitochondrial permeability transition (MPT). After myocardial ischemia, opening of the mPTP is a critical determinant of cell death. The relationship of orientin and mPTP in mediating reperfusion-induced cardiomyocytes injury is still elusive. Here, our results indicate that the protective effect of orientin in H9c2 cells subjected to I/R injury is associated with depression of the mPTP opening, resultant mitochondrial dysfunction, and apoptosis. Further investigation of cellular mechanisms revealed that these effects were associated with inhibition of reactive oxygen species (ROS) generation, repolarization of mitochondrial membrane potential (Δψ _m), suppression of mitochondrial cytochrome c release, enhancement of the Bcl-2 level, and inhibition of Bax and Smac/DIABLO levels. Furthermore, these beneficial effects of orientin were blocked by the phosphatidylinositol 3-kinase (PI3K) inhibitor wortmannin, and orientin could enhance Akt phosphorylation. In summary, we demonstrate that orientin protects H9c2 cardiomytocytes against I/R-induced apoptosis by modulating the mPTP opening, and this role of orientin may involve the PI3K/Akt signaling pathway.

Key words

orientin - mitochondrial permeability transition - cardiomyocytes - ischemia/reperfusion (I/R) - apoptosis

References

• 1 Zhang Y, Wu X Q, Yu Z Y. Comparison study on total flavonoid content and anti-free radical activity of the leaves of bamboo, Phyllostachys nigra, and Ginkgo biloba.  Zhongguo Zhong Yao Za Zhi. 2002;  27 254-257
• 2 Dietrych-Szostak D, Oleszek W. Effect of processing on the flavonoid content in buckwheat (Fagopyrum esculentum Möench) grain.  J Agric Food Chem. 1999;  47 4384-4387
• 3 Suzuki K, Sawa Y, Kaneda Y, Ichikawa H, Shirakura R, Matsuda H. In vivo gene transfection with heat shock protein 70 enhances myocardial tolerance to ischemia-reperfusion injury in rat.  J Clin Invest. 1997;  99 1645-1650
• 4 Sun Y, Deng T, Lu N, Yan M, Zheng X. B-type natriuretic peptide protect cardiomyocytes at reperfusion via mitochondrial calcium uniporter.  Biomed Pharmacother. 2010;  64 170-176
• 5 Halestrap A P, Clarke S J, Javadov S A. Mitochondrial permeability transition pore opening during myocardial reperfusion–a target for cardioprotection.  Cardiovasc Res. 2004;  61 372-385
• 6 Lemasters J J, Nieminen A L, Qian T, Trost L C, Elmore S P, Nishimura Y, Crowe R A, Cascio W E, Bradham C A, Brenner D A, Herman B. The mitochondrial permeability transition in cell death: a common mechanism in necrosis, apoptosis and autophagy.  Biochim Biophys Acta. 1998;  1366 177-196
• 7 Sun Y, Zhang Y, Yan M, Wu Y, Zheng X. B-type natriuretic peptide-induced cardioprotection against reperfusion is associated with attenuation of mitochondrial permeability transition.  Biol Pharm Bull. 2009;  32 1545-1551
• 8 Redegeld F A, Moison R M, Koster A S, Noordhoek J. Depletion of ATP but not of GSH affects viability of rat hepatocytes.  Eur J Pharmacol. 1992;  228 229-236
• 9 Green D R, Reed J C. Mitochondria and apoptosis.  Science. 1998;  281 1309-1312
• 10 Kang P M, Haunstetter A, Aoki H, Usheva A, Izumo S. Morphological and molecular characterization of adult cardiomyocyte apoptosis during hypoxia and reoxygenation.  Circ Res. 2000;  87 118-125
• 11 Baines C P. The mitochondrial permeability transition pore and ischemia-reperfusion injury.  Basic Res Cardiol. 2009;  104 181-188
• 12 Gustafsson A B, Gottlieb R A. Heart mitochondria: gates of life and death.  Cardiovasc Res. 2008;  77 334-343
• 13 Kominsky D J, Bickel R J, Tyler K L. Reovirus-induced apoptosis requires mitochondrial release of Smac/DIABLO and involves reduction of cellular inhibitor of apoptosis protein levels.  J Virol. 2002;  76 11414-11424
• 14 Abas L, Bogoyevitch M A, Guppy M. Mitochondrial ATP production is necessary for activation of the extracellular-signal-regulated kinases during ischaemia/reperfusion in rat myocyte-derived H9c2 cells.  Biochem J. 2000;  349 119-126
• 15 Akao M, Teshima Y, Marban E. Antiapoptotic effect of nicorandil mediated by mitochondrial ATP-sensitive potassium channels in cultured cardiac myocytes.  J Am Coll Cardiol. 2002;  40 803-810
• 16 Deng T, Zhang L, Ge Y, Lu M, Zheng X. Redistribution of intracellular calcium and its effect on apoptosis in macrophages: Induction by oxidized LDL.  Biomed Pharmacother. 2008;  63 267-274
• 17 Braunwald E, Kloner R A. Myocardial reperfusion: a double-edged sword?.  J Clin Invest. 1985;  76 1713-1719
• 18 Ambrosio G, Zweier J L, Duilio C, Kuppusamy P, Santoro G, Elia P P, Tritto I, Cirillo P, Condorelli M, Chiariello M, Flaherty J T. Evidence that mitochondrial respiration is a source of potentially toxic oxygen free radicals in intact rabbit hearts subjected to ischemia and reflow.  J Biol Chem. 1993;  268 18532-18541
• 19 Kim J S, Ohshima S, Pediaditakis P, Lemasters J J. Nitric oxide protects rat hepatocytes against reperfusion injury mediated by the mitochondrial permeability transition.  Hepatology. 2004;  39 1533-1543
• 20 Robin E, Guzy R D, Loor G, Iwase H, Waypa G B, Marks J D, Hoek T L, Schumacker P T. Oxidant stress during simulated ischemia primes cardiomyocytes for cell death during reperfusion.  J Biol Chem. 2007;  282 19133-19143
• 21 Borutaite V, Brown G C. Mitochondria in apoptosis of ischemic heart.  FEBS Lett. 2003;  541 1-5
• 22 Suleiman M S, Halestrap A P, Griffiths E J. Mitochondria: a target for myocardial protection.  Pharmacol Ther. 2001;  89 29-46
• 23 Crompton M. The mitochondrial permeability transition pore and its role in cell death.  Biochem J. 1999;  341 233-249
• 24 Fu X C, Wang M W, Li S P, Wang H L. Anti-apoptotic effect and mechanism of orientin on ischaemic/reperfused myocardium.  J Asian Nat Prod Res. 2006;  8 265-272
• 25 Kroemer G, Dallaporta B, Resche-Rigon M. The mitochondrial death/life regulator in apoptosis and necrosis.  Annu Rev Physiol. 1998;  60 619-642
• 26 Hausenloy D J, Yellon D M. New directions for protecting the heart against ischaemia-reperfusion injury: targeting the Reperfusion Injury Salvage Kinase (RISK)-pathway.  Cardiovasc Res. 2004;  61 448-460
• 27 Zhu M, Feng J, Lucchinetti E, Fischer G, Xu L, Pedrazzini T, Schaub M C, Zaugg M. Ischemic postconditioning protects remodeled myocardium via the PI3K-PKB/Akt reperfusion injury salvage kinase pathway.  Cardiovasc Res. 2006;  72 152-162
• 28 Armstrong S C. Protein kinase activation and myocardial ischemia/reperfusion injury.  Cardiovasc Res. 2004;  61 427-436
• 29 Miyamoto S, Rubio M, Sussman M A. Nuclear and mitochondrial signalling Akts in cardiomyocytes.  Cardiovasc Res. 2009;  82 272-285
• 30 Venkatapuram S, Wang C, Krolikowski J G, Weihrauch D, Kersten J R, Warltier D C, Pratt Jr P F, Pagel P S. Inhibition of apoptotic protein p 53 lowers the threshold of isoflurane-induced cardioprotection during early reperfusion in rabbits.  Anesth Analg. 2006;  103 1400-1405