P6275Role of autophagy in imatinib-induced cardiotoxicity

Abstract

Abstract Background Cardiotoxicity is one of the severe adverse effects of chemotherapeutic agents. Imatinib, a therapeutic agent for chronic myelogenous leukemia, has been reported to induce cardiotoxicity. Autophagy is an intracellular bulk protein and organelle degradation process, but it is unclear whether autophagy functions as pro-death or pro-survival program during disease conditions. We examined whether imatinib induces myocyte autophagy and the role of autophagy in imatinib-induced cardiotoxicity in in vitro and in vivo experiments. Methods In in vitro experiments, neonatal rat cardiac myocytes were treated with imatinib (1, 5, 10 μM; 1–6 hrs). Inhibition of autophagy was performed using 3-methyl-adenine (3MA), an autophagic inhibitor, and transfection with Atg5-targeted siRNA. Myocyte apoptosis was detected by morphological change in nuclei and caspase 3 activity. Mitochondria-derived reactive oxygen species production was detected using MitoSOX and mitochondrial membrane potential was assessed by TMRM staining. Expressions of cytochrome c in mitochondria and cytosole were examined by Werstern blotting. Myocyte autophagy was assessed by monodansylcadaverine staining and microtubule-associated protein light chain (LC) 3-II expression. In in vivo experiments, C57BL6 mice were treated with imatinib (50 and 200 mg/kg/day) for 5 weeks in the presence or absence of 3MA. Cardiac function was examined by echocardiography. In cardiac tissue, apoptotic myocytes were examined by TUNEL assay and autophagy was examined by LC3-II expression. Results In in vitro experiments, imatinib increased apoptotic nuclei and caspase 3 activity, in a dose-dependent manner. Consequently, imatinib augmented production of mitochondria-derived reactive oxygen species, loss of mitochondrial membrane potential, and the release of cytochrome c from mitochondria to cytosole, suggesting that imatinib induced mitochondrial-apoptotic pathway. On the other hand, imatinib significantly increased monodansylcadaverine stained dots and LC3-II expression, suggesting that imatinib increased autophagy. 3MA and Atg5 siRNA augmented imatinib-induced apoptosis by 60% and 30%, respectively. In in vivo experiments, imatinib (200 mg) exhibited the dilatation of left ventricle by 15% and the depression of left ventricular fractional shortening by 23%. Ratio of apoptotic myocytes was significantly increased and LC3-II expression in cardiac tissue was enhanced by imatinib in a dose-dependent fashion. Co-treatment with 3MA and imatinib further impaired imatinib-induced myocyte apoptosis by 3 fold and LV dysfunction by 20%. Conclusion These results indicate that imatinib induced myocyte apoptosis, leading to cardiac dysfunction. Imatinib enhanced myocyte autophagy as a consequence of apoptosis and autophagy was a beneficial phenomenon in this condition.