Shibayama-Imazu Toshiko, Masuda Yutaka, Shinki Toshimasa, Nakajo Shigeo, Nakaya Kazuyasu
Laboratory of Biological Chemistry, School of Pharmaceutical Sciences, Showa University, Tokyo 142-8555, Japan.
Anticancer Res. 2007 Jan-Feb;27(1A):245-9.
It has been previously demonstrated that bufalin, an active agent in the Chinese medicine chan'su, induces apoptosis in human leukemia cells by altering the expression of apoptosis-related genes, such as bcl-2 and c-myc. Tiam1 was also found to play a critical role in bufalin-induced apoptosis through the activation of the Rac1, PAK and JNK pathway in human leukemia cell lines. In the present study, the involvement of the Tiam1 gene products in bufalin-induced apoptosis in human solid tumor HeLa cells was examined.
HeLa cells were treated with 10(-8) M bufalin and apoptosis was measured by ELISA quantification of nucleosomes. Tiam1 mRNA levels were quantified by real-time PCR analysis and inhibited by transfected siRNA specific for Tiam1.
Apoptosis was induced in HeLa cells by treatment with 10(-8) M bufalin. Expression of both Tiam1 mRNA and its protein was induced 0.5 h after the start of the bufalin treatment. Transfection of Tiam1-specific siRNA into HeLa cells markedly inhibited bufalin-induced apoptosis.
Our results suggest that Tiam1 is a downstream mediator of bufalin-induced apoptosis in the human solid tumor HeLa cell line, as well as in leukemia cell lines.
先前已证明，中药蟾酥中的活性成分蟾毒灵通过改变凋亡相关基因（如bcl-2和c-myc）的表达来诱导人白血病细胞凋亡。还发现Tiam1通过激活人白血病细胞系中的Rac1、PAK和JNK途径在蟾毒灵诱导的凋亡中起关键作用。在本研究中，检测了Tiam1基因产物在蟾毒灵诱导人实体瘤HeLa细胞凋亡中的作用。
用10^(-8) M蟾毒灵处理HeLa细胞，通过ELISA定量核小体来检测凋亡。通过实时PCR分析定量Tiam1 mRNA水平，并通过转染针对Tiam1的特异性siRNA来抑制其表达。
用10^(-8) M蟾毒灵处理HeLa细胞可诱导凋亡。蟾毒灵处理开始后0.5小时，Tiam1 mRNA及其蛋白的表达均被诱导。将Tiam1特异性siRNA转染到HeLa细胞中可显著抑制蟾毒灵诱导的凋亡。
我们的结果表明，Tiam1是蟾毒灵诱导人实体瘤HeLa细胞系以及白血病细胞系凋亡的下游介质。
