Multiple actions of pifithrin-α on doxorubicin- induced apoptosis in rat myoblastic H9c2 cells
Multiple actions of pifithrin-α on doxorubicin- induced apoptosis in rat myoblastic H9c2 cells
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DOI：
10.1152/ajpheart.01138.2005
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发表时间：
2006-06-01
影响因子：
4.8
通讯作者：
Chua, Balvin H. L.
中科院分区：
文献类型：
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作者：
Chua, Chu Chang;Liu, Xuwan;Chua, Balvin H. L.
多柔比星（Dox）是引起显著心脏毒性的化学治疗剂。我们先前表明，Dox激活p53并诱导小鼠心脏细胞凋亡。本研究旨在阐明导致p53稳定的分子事件，以检查参与Dox诱导的细胞凋亡的途径，并评估p53抑制剂pifithrin-alpha（PFT-alpha）在阻断大鼠H9 c2成肌细胞凋亡中的有效性。暴露于5 μ M Dox的H9 c2细胞具有升高水平的p53和丝氨酸15处磷酸化的p53。Dox还触发p38、p42/p44 ERK和p46/p54 JNK MAP激酶的瞬时激活。Caspase活性测定和Western blot分析显示，Dox处理H9 c2细胞16 h后，Caspase-2、-3、-8、-9、-12、Fas和裂解的聚ADP核糖聚合酶（PARP）水平明显升高。有p53结合活性，细胞色素c的释放，和细胞凋亡的伴随增加。这些结果表明，Dox可以触发内源性，外源性和内质网相关的凋亡途径。用PFT-α预处理细胞，然后给予Dox，减弱了Dox诱导的p53水平和p53结合活性的增加，并部分阻断了p46/p54 JNK和p42/p44 ERK的激活。PFT-α还导致半胱天冬酶-2、-3、-8、-9、-12、Fas、PARP、细胞色素c释放和细胞凋亡水平降低。我们的研究结果表明，p53的稳定是一个焦点的Dox诱导的细胞凋亡和PFT-α干扰多个步骤的Dox诱导的细胞凋亡。
Doxorubicin (Dox) is a chemotherapeutic agent that causes significant cardiotoxicity. We showed previously that Dox activates p53 and induces apoptosis in mouse hearts. This study was designed to elucidate the molecular events that lead to p53 stabilization, to examine the pathways involved in Dox-induced apoptosis, and to evaluate the effectiveness of pifithrin-alpha (PFT-alpha), a p53 inhibitor, in blocking apoptosis of rat H9c2 myoblasts. H9c2 cells that were exposed to 5 mu M Dox had elevated levels of p53 and phosphorylated p53 at Ser15. Dox also triggered a transient activation of p38, p42/p44ERK, and p46/p54JNK MAP kinases. Caspase activity assays and Western blot analysis showed that H9c2 cells treated with Dox for 16 h had marked increase in the levels of caspases-2, -3, -8, -9, -12, Fas, and cleaved poly(ADP ribose) polymerase (PARP). There was a concomitant increase in p53 binding activity, cytochrome c release, and apoptosis. These results suggest that Dox can trigger intrinsic, extrinsic, and endoplasmic reticulum-associated apoptotic pathways. Pretreatment of cells with PFT-alpha followed by Dox administration attenuated Dox-induced increases in p53 levels and p53 binding activity and partially blocked the activation of p46/p54JNK and p42/p44ERK. PFT-alpha also led to decreased levels of caspases-2, -3, -8, -9, -12, Fas, PARP, cytochrome c release, and apoptosis. Our results suggest that p53 stabilization is a focal point of Dox-induced apoptosis and that PFT-alpha interferes with multiple steps of Dox-induced apoptosis.