Abstract
The cardiotoxicity of adriamycin limits its clinical use as a powerful drug for solid tumors and malignant hematological disease. Although the precise mechanism by which it causes cardiac damage is not yet known, it has been suggested that apoptosis is the principal process in adriamycin-induced cardiomyopathy, which involves DNA fragmentation, cytochrome C release, and caspase activation. However, there has been no direct evidence for the critical involvement of caspase-3 in adriamycin-induced apoptosis. To determine the requirements for the activation of caspase-3 in adriamycin-treated cardiac cells, the effect of a caspase inhibitor on the survival of and apoptotic changes in H9c2 cells was examined. Exposure of H9c2 cells to adriamycin resulted in a time- and dose-dependent cell death, and the cleavage of pro-caspase-3 and of the nuclear protein poly (ADP-ribose) polymerase (PARP). However, neither the reduction of cell viability nor the characteristic morphological changes induced by adriamycin were prevented by pretreatment with the general caspase inhibitor z-VAD.FMK. In contrast, caspase inhibition effectively blocked the apoptosis induced by H2O2 in H9c2 cells, as determined by an MTT assay or microscopy. We also observed that p53 expression was increased by adriamycin, and this increase was not affected by the inhibition of caspase activity, suggesting a role for p53 in adriamycin-induced caspase-independent apoptosis in cardiac toxicity. (Mol Cell Biochem 270: 13–19, 2005)
Similar content being viewed by others
References
Mott MG: Anthracycline cardiotoxicity and its prevention. Ann NY Acad Sci 824: 221–228, 1997
Doroshow JH: Effect of anthracycline antibiotics on oxygen radical formation in rat heart. Cancer Res 43: 460–472, 1983
Olson RD, Mushlin PS: Doxorubicin cardiotoxicity: Analysis of prevailing hypotheses. FASEB J 4: 3076–3086, 1990
Tong J, Ganguly PK, Singal PK: Myocardial adrenergic changes at two stages of heart failure due to adriamycin treatment in rats. Am J Physiol 260: 909–916, 1991
Bristow MR, Sageman WS, Scott RH: Acute and chronic cardiovascular effects of doxorubicin in the dog: The cardiovascular pharmacology of drug-induced histamine release. J Cardiovasc Pharmacol 2: 487–515, 1980
Singal PK, Panagia V: Direct effects of adriamycin on the rat heart sarcolemma. Res Commun Chem Pathol Pharmacol 43: 67–77,1984
Doroshow JH, Davies KJ: Redox cycling of anthracyclines by cardiac mitochondria. II. Formation of superoxide anion, hydrogen peroxide, and hydroxyl radical. J Biol Chem 261: 3068–3074, 1986
Kurabayashi M, Jeyaseelan R, Kedes L: Doxorubicin represses the function of the myogenic helix-loop-helix transcription factor MyoD: Involvement of Id gene induction. J Biol Chem 269: 6031–6039, 1994
Andrieu-Abadie N, Jaffrezou JP, Hatem S, Laurent G, Levade T, Mercaider JJ: L-carnitine prevents doxorubicin-induced apoptosis of cardiac myocytes: Role of inhibition of ceramide generation. FASEB J 13: 1501–1510, 1999
Wang L, Ma W, Markovich R, Lee WL, Wang PH: Insulin-like growth factor I modulates induction of apoptotic signaling in H9c2 cardiac muscle cells. Endocrinology 139: 1354–1360, 1998
Martin SJ, Green, DR: Protease activation during apoptosis: Death by a thousand cuts? Cell 82: 349–352, 1995
Nicholson DW, Thornberry NA: Caspases: Killer proteases. Trends Biochem Sci 22: 299–306, 1997
Cohen GM: Caspases: The executioners of apoptosis. Biochem J 326: 1–16, 1997
Lampidis TJ, Moreno G, Salet C, Vinzens F: Nuclear and mitochondrial effects of adriamycin in singly isolated pulsating myocardial cells. J Mol Cell Cardiol 11: 415–422, 1979
Kaufmann SH, Desnoyers S, Ottaviano Y, Davidson NE, Poirier GG: Specific proteolytic cleavage of poly(ADP-ribose) polymerase: An early marker of chemotherapy-induced apoptosis. Cancer Res 53: 3976–3985, 1993
Soldani C and Scovasse AI: Poly(ADP-ribose) polymerase-1 cleavage during apoptosis: An update. Apoptosis 7: 321–328, 2002
Turner NA, Xia F, Azhar G, Zhang X, Liu L, Wei JY: Oxidative stress induces DNA fragmentation and caspase activation via the c-Jun NH2-terminal kinase pathway in H9c2 cardiac muscle cells. J Mol Cell Cardiol 30: 1789–1801, 1998
Dipietrantonio AM, Hsieh TC, Wu-JM: Activation of caspase-3 in HL-60 cells exposed to hydrogen peroxide. Biochem Biophys Res Commun 255: 477–482, 1999
Zaidi AU, McDonough JS, Klocke BJ, Latham CB, Korsmeyer SJ, Flavell RA, Schmidt RE, Roth KA: Chloroquine-induced neuronal cell death is p53 and Bcl-2 family-dependent but caspase-independent. J Neuropathol Exp Neurol 60: 937–945, 2001
Sola S, Ma X, Castro RE, Kren BT, Steer CJ, Rodrigues CM: Ursodeoxycholic acid modulates E2F-1 and p53 expression through caspase-independent mechanism in transforming growth factor beta1-induced apoptosis of rat hepatocytes. J Biol Chem 278: 48831–48838, 2003
Godefroy N, Lemaire C, Renaud F, Rincheval V, Perez S, Parvu-Ferecatu I, Mignotte B, Vayssiere J-L: p53 can promote mitochondria-and caspase-independent apoptosis. Cell death and Differentiation 11: 785–787, 2004
Lorenzo E, Ruiz-Ruiz C, Quesada AJ, Hernandez G, Rodriguez A, Lopez-Rivas A, Redondo JM: Doxorubicin induces apoptosis and CD95 gene expression in human primary endothelial cells through a p53-dependent mechanism. J Biol Chem 277: 10883–10892, 2002
Liem AA, Appleyard MV, O’Neill MA, Hupp TR, Chamberlain MP, Thompson AM: Doxorubicin and vinorelbine act independently via p53 expression and p38 activation respectively in breast cancer cell lines. Br J Cancer 88: 1281–1284, 2003
Yeh PY, Chuang SE, Yeh KH, Song YC, Chang LL, Cheng AL: Phosphorylation of p53 on Thr55 by ERK2 is necessary for doxorubicin-induced p53 activation and cell death. Oncogene 23: 3589–3588, 2004
L’Ecuyer T, Horenstein MS, Thomas R, Vander Heide R: Anthracycline-induced cardiac injury using a cardiac cell line: Potential for gene therapy studies. Mol Genet Metab 74: 370–379, 2001
Green PS, Leeuwenburgh C: Mitochondrial dysfunction is an early indicator of doxorubicin-induced apoptosis. Biochim Biophys Acta 1588: 94–101, 2002
Okuno S, Shimizu S, Ito T, Nomura M, Hamada E, Tsujimoto Y, Matsuda H: Bcl-2 prevents caspase-independent cell death. J Biol Chem 273: 34272–34277, 1998
D’Mello SR, Kuan CY, Flavell RA, Rakic P: Caspase-3 is required for apoptosis-associated DNA fragmentation but not for cell death in neurons deprived of potassium. J Neurosci Res 59: 24–31, 2000
Stefanis L, Park DS, Friedman WJ, Greene LA: Caspase-dependent and -independent death of camptothecin-treated embryonic cortical neurons. J Neurosci 19: 6235–6247, 2002
Kobayashi T, Sawa H, Morikawa J, Zhang W, Shiku H: Bax induction activates apoptotic cascade via mitochondrial cytochrome-c release and Bax overexpression enhances apoptosis induced by chemotherapeutic agents in DLD-1 colon cancer cells. Jpn J Cancer Res 91: 1264–1268, 2000
Hong F, Kwon SJ, Jhun BS, Kim SS, Ha J, Kim SJ, Sohn NW, Kang C, Kang I: Insulin-like growth factor-1 protects H9c2 cardiac myoblasts from oxidative stress-induced apoptosis via phosphatidylinositol 3-kinase and extracellular signal-regulated kinase pathways. Life Sci 68: 1095–1105, 2001
Panaretakis T, Pokrovskaja K, Shoshan MC, Grander D: Activation of Bak, Bax, and BH3-only proteins in the apoptotic response to doxorubicin. J Biol Chem 277: 44317–44326, 2002
Johnson MD, Xiang H, London S, Kinoshita Y, Knudson M, Mayberg M, Korsmeyer SJ, Morrison RS: Evidence for involvement of Bax and p53, but not caspases, in radiation-induced cell death of cultured postnatal hippocampal neurons. J Neurosci Res 54: 721–733, 1998
Selznick LA, Zheng TS, Flavell RA, Rakic P, Roth KA: Amyloid beta-induced neuronal death is bax-dependent but caspase-independent. J Neuropathol Exp Neurol 59: 271–279, 2000
Schuler M, Green DR: Mechanisms of p53-dependent apoptosis. Biochem Soc Trans 29: 684–688, 2001
Wu X, Deng Y: Bax and BH3-domain-only proteins in p53-mediated apoptosis. Front Biosci 7: d151–d156, 2002
Cande C, Cecconi F, Dessen P and Kroemer G: Apoptosis-inducing factor (AIF): Key to the conserved caspase-independent pathways of cell death? J Cell Sci 115: 4727–4734, 2002
Liu X, Chua CC, Gao J, Chen Z, Landy CL, Hamdy R, Chua BH: Pifithrin-alpha protects against doxoruicin-induced apoptosis and acute cardiotoxicity in mice. Am J Physiol Heart Circ Physiol 286: H933–H936, 2004
DeAtley SM, Aksenov MY, Aksenova MV, Harris B, Hadley R, Cole Harper P, Carney JM, Butterfield DA: Antioxidants protect against reactive oxygen species associated with adriamycin-treated cardiomyocytes. Cancer Lett 136: 41–46, 1999
Tsang WP, Chau SP, Kong SK, Fung KP, Kwok TT: Reactive oxygen species mediate doxorubicin induced p53-independent apoptosis. Life Sci 73: 2047–2058, 2003
Author information
Authors and Affiliations
Corresponding author
Additional information
These authors contributed equally to this work
Rights and permissions
About this article
Cite this article
Youn, HJ., Kim, HS., Jeon, MH. et al. Induction of caspase-independent apoptosis in H9c2 cardiomyocytes by adriamycin treatment. Mol Cell Biochem 270, 13–19 (2005). https://doi.org/10.1007/s11010-005-2541-2
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/s11010-005-2541-2