Bufalin induces G0/G1 phase arrest through inhibiting the levels of cyclin D, cyclin E, CDK2 and CDK4, and triggers apoptosis via mitochondrial signaling pathway in T24 human bladder cancer cells
Bufalin induces G0/G1 phase arrest through inhibiting the levels of cyclin D, cyclin E, CDK2 and CDK4, and triggers apoptosis via mitochondrial signaling pathway in T24 human bladder cancer cells
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DOI：
10.1016/j.mrfmmm.2011.09.010
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发表时间：
2012-04-01
影响因子：
2.3
通讯作者：
Chung, Jing-Gung
中科院分区：
文献类型：
--
作者：
Huang, Wen-Wen;Yang, Jai-Sing;Chung, Jing-Gung
大多数膀胱癌的化疗目的是杀死癌细胞，但药物治疗后的复发率仍然很高。从蟾蜍皮肤和腮腺毒腺中提取的蟾毒灵已被证明可诱导多种类型的癌细胞系的细胞凋亡。然而，目前还没有关于蟾酥能诱导人膀胱癌细胞死亡的报道。本研究旨在探讨蟾酥诱导人膀胱癌细胞株(T24)凋亡的机制。采用DAPI/TUNEL双染法、PI排除法和流式细胞仪分析方法，研究了蟾酥对T24细胞生长和凋亡的影响。用流式细胞仪检测蟾酥对T24细胞产生的活性氧(ROS)、线粒体膜电位(Delta Psi(M))和DNA含量(包括亚G1期细胞凋亡)的影响。用免疫印迹法检测蟾酥灵对T24细胞G(0)/G(1)期调控蛋白和凋亡相关蛋白表达的影响。结果表明，蟾酥能显著降低T24细胞的存活率，使其发生G(0)/G(1)期阻滞，并引发细胞凋亡。细胞周期蛋白D、CDK4、细胞周期蛋白E、CDK2、磷酸化Rb、磷酸化AKT和Bcl2蛋白表达下调，细胞色素c、APAF-1、AIF、caspase-3、-7、-9和Sax蛋白表达及caspase活性上调。根据我们的结果，蟾酥通过抑制AKT活性和抗凋亡的Bcl2蛋白以及诱导促凋亡的Bax蛋白来诱导T24细胞的凋亡。Caspase-3、-7和-9的表达也参与了蟾酥灵诱导的T24细胞的凋亡。因此，蟾酥有可能成为未来治疗人膀胱癌的药物。(C)2012爱思唯尔B.V.保留所有权利。
Most of the chemotherapy treatments for bladder cancer aim to kill the cancer cells, but a high recurrence rate after medical treatments is still occurred. Bufalin from the skin and parotid venom glands of toad has been shown to induce apoptotic cell death in many types of cancer cell lines. However, there is no report addressing that bufalin induced cell death in human bladder cancer cells. The purpose of this study was investigated the mechanisms of bufalin-induced apoptosis in a human bladder cancer cell line (T24). We demonstrated the effects of bufalin on the cell growth and apoptosis in T24 cells by using DAPI/TUNEL double staining, a PI exclusion and flow cytometric analysis. The effects of bufalin on the production of reactive oxygen species (ROS), the level of mitochondrial membrane potential (Delta Psi(m)), and DNA content including sub-G1 (apoptosis) in T24 cells were also determined by flow cytometry. Western blot analysis was used to examine the expression of G(0)/G(1) phase-regulated and apoptosis-associated protein levels in bufalin-treated T24 cells. The results indicated that bufalin significantly decreased the percentage of viability, induced the G(0)/G(1) phase arrest and triggered apoptosis in T24 cells. The downregulation of the protein levels for cyclin D, CDK4, cyclin E, CDK2, phospho-Rb, phospho-AKT and Bcl-2 with the simultaneous up-regulation of the cytochrome c, Apaf-1, AIF, caspase-3, -7 and -9 and Sax protein expressions and caspase activities were observed in T24 cells after bufalin treatment. Based on our results, bufalin induces apoptotic cell death in T24 cells through suppressing AKT activity and antiapoptotic Bcl-2 protein as well as inducing pro-apoptotic Bax protein. The levels of caspase-3, -7 and -9 are also mediated apoptosis in bufalin-treated T24 cells. Therefore, bufalin might be used as a therapeutic agent for the treatment of human bladder cancer in the future. (C) 2012 Elsevier B.V. All rights reserved.