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Abstract
The current studies were designed to explore the protective effects of pifithrin-α and melatonin against doxorubicin-induced cardiotoxicity. Doxorubicin was injected at a dose of 22.5 mg/kg (i.p.) in mice to induce cardiotoxic effects. Meanwhile, doxorubicin caused a significant increase of cardiac cell apoptosis following injection (14.2 ± 1.1% for doxorubicin-5 d vs. 1.8 ± 0.12% for control, P < 0.01). Ribonuclease protection assays and Western blot analyses revealed that doxorubicin upregulated the p53-dependent genes Bax, BclxL, and MDM2 at least 2-fold. p53 was phosphorylated at Ser 15 in mouse hearts 1 h following doxorubicin injection, and p38 and ERK1/2 MAPKs mediated the phosphorylation of p53. In addition, caspases-3 and -9 were activated 24 h after doxorubicin injection. A p53 inhibitor, pifithrin-α inhibited doxorubicin-induced apoptosis when administered at a dose of 2.2 mg/kg. Pifithrin-a abolished p53 transactivation activity but did not influence doxorubicin-induced phosphorylation at Ser 15. By effectively inhibiting the expression of p53-dependent genes, pifithrin-α blocked doxorubicin-induced activation of caspases-3 and -9, thereby preventing cardiac apoptosis. In addition, pifithrin-α attenuated doxorubicin-induced structural and functional damages, without diminishing its anti-tumor efficacy on p53-null PC-3 cancer cells. The protective effects of melatonin and its metabolite 6-hydroxymelatonin on doxorubicin-induced cardiac dysfunction were evaluated in an isolated perfused mouse hearts and in vivo doxorubicin-treated mice. While perfusion of mouse hearts with 5 μM doxorubicin for 60 min resulted in a 50% suppression of HR×LVDP and a 50% reduction of coronary flow, pre-exposure of hearts to 1 μM melatonin or 6-hydroxymelatonin eased the cardiac dysfunction. In addition, administration of melatonin or 6-hydroxymelatonin (2 mg/kg/d) significantly attenuated doxorubicin-induced cardiac dysfunction, myocardial lesions, and cardiac cell apoptosis. Melatonin and 6-hydroxymelatonin significantly improved the survival rate of doxorubicin-treated mice. Another melatonin analog, 8-methoxy-2-propionamidotetralin, did not show any convincing protection on either animal survival or on in vitro cardiac function, presumably due to its lack of free radical-scavenging activity. Finally, neither melatonin nor 6-hydroxymelatonin compromised the anti-tumor activity of doxorubicin in cultured PC-3 cells. These studies suggest that pifithrin-α and melatonin have significant therapeutic potential for patients suffering doxorubicin-induced cardiotoxicity.
