An insight into the mechanism of cytotoxicity of ricin to hepatoma cell:: Roles of Bcl-2 family proteins, caspases, Ca2+-dependent proteases and protein kinase C
An insight into the mechanism of cytotoxicity of ricin to hepatoma cell:: Roles of Bcl-2 family proteins, caspases, Ca2+-dependent proteases and protein kinase C
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DOI：
10.1002/jcb.1076
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发表时间：
2001-01-01
影响因子：
4
通讯作者：
Liu, XY
中科院分区：
文献类型：
--
作者：
Hu, RG;Zhai, QW;Liu, XY
采用荧光显微镜、流式细胞术和DNA片段化实验检测了II型核糖体失活蛋白蓖麻毒素体外诱导肝癌细胞（BEL7404）凋亡的能力。作为缺乏Bcl-2的模型，BEL7404具有独特的优势，可以研究过表达Bcl-2对蛋白合成抑制剂诱导的细胞凋亡的影响。通过建立Bcl-2过表达细胞株BEL7404/ Bcl-2，我们发现Bcl-2能促进肝癌细胞抗蓖麻毒素损伤的存活。蓖麻毒素诱导的BEL7404细胞凋亡伴随着Bak的表达增加，Bcl-xl和Bax的表达降低。发现半胱天冬酶和PARP切割活性与死亡过程有关。通过抑制剂测试，我们的结果排除了钙依赖性蛋白酶或蛋白激酶C参与蓖麻毒素诱导的凋亡过程，尽管细胞内钙水平升高确实是对蓖麻毒素治疗的直接反应。另一种蛋白质合成抑制剂环己亚胺对蓖麻毒素对肝癌细胞BEL7404的细胞毒性具有增效作用，而不是抑制作用。实际上，仅环己亚胺就能诱导肝癌细胞BEL7404死亡，而过表达Bcl-2也能抑制肝癌细胞的死亡。我们讨论了凋亡蛋白Bak的升高，以挑战蓖麻毒素通过非特异性抑制所有新生蛋白合成来发挥细胞毒性的观点。生物化学学报，31(2):583-593,2001。(C) 2001 Wiley-Liss, Inc。
The ability of ricin, a type II ribosome-inactivating protein, to induce hepatoma cell (BEL7404) to apoptosis in vitro was examined by fluorescence microscopy, flow cytometry, and DNA fragmentation assay. As a Bcl-2 lacking model, BEL7404 bore unique advantage to study the effect of over-expressing Bcl-2 on the apoptosis induced by the inhibitor of protein synthesis. By establishing a Bcl-2 over-expressing cell line (BEL7404/ Bcl-2), we found that Bcl-2 could promote the survival of the hepatoma cell against ricin insult. The ricin-induced apoptosis of BEL7404 was accompanied by increased expression of Bak and decreased levels of Bcl-xl and Bax. Caspases and PARP cleavage activity were found to be implicated in the death process. Through the inhibitor tests, our results excluded the participation of calcium-dependent proteases or protein kinase C in the apoptotic process induced by ricin, though an elevation of intracellular calcium did occur as an immediate response to ricin treatment. Cycloheximide, another protein synthesis inhibitor, did synergistically enhance rather than inhibit the cytotoxicity of ricin to hepatoma cell BEL7404. Actually, cycloheximide alone was able to induce hepatoma cell BEL7404 to death that could also be inhibited by over-expressing Bcl-2. The elevation of apoptotic protein Bak was discussed to challenge the notion that ricin exerted its cytotoxicity through nonspecific inhibition of all the de novo protein synthesis, J. Cell. Biochem. 81: 583-593, 2001. (C) 2001 Wiley-Liss, Inc.