Zhang Kejun, Chen Dong, Wang Xingang, Zhang Shaoyan, Wang Jigang, Gao Yuan, Yan Bomin
General Surgery of the Affiliated Hospital of Medical College, Qingdao University, Qingdao, Shan Dong Province 266003, China; E-Mails:
Int J Mol Sci. 2011 Jan 14;12(1):385-400. doi: 10.3390/ijms12010385.
Slug is an E-cadherin repressor and a suppressor of PUMA (p53 upregulated modulator of apoptosis) and it has recently been demonstrated that Slug plays an important role in controlling apoptosis. In this study, we examined whether Slug's ability to silence expression suppresses the growth of cholangiocarcinoma cells and/or sensitizes cholangiocarcinoma cells to chemotherapeutic agents through induction of apoptosis. We targeted the Slug gene using siRNA (Slug siRNA) via full Slug cDNA plasmid (Slug cDNA) transfection of cholangiocarcinoma cells. Slug siRNA, cisplatin, or Slug siRNA in combination with cisplatin, were used to treat cholangiocarcinoma cells in vitro. Western blot was used to detect the expression of Slug, PUMA, and E-cadherin protein. TUNEL, Annexin V Staining, and cell cycle analysis were used to detect apoptosis. A nude mice subcutaneous xenograft model of QBC939 cells was used to assess the effect of Slug silencing and/or cisplatin on tumor growth. Immunohistochemical staining was used to analyze the expression of Slug and PUMA. TUNEL was used to detect apoptosis in vivo. The results showed that PUMA and E-cadherin expression in cholangiocarcinoma cells is Slug dependent. We demonstrated that Slug silencing and cisplatin both promote apoptosis by upregulation of PUMA, not by upregulation of E-cadherin. Slug silencing significantly sensitized cholangiocarcinoma cells to cisplatin through upregulation of PUMA. Finally, we showed that Slug silencing suppressed the growth of QBC939 xenograft tumors and sensitized the tumor cells to cisplatin through PUMA upregulation and induction of apoptosis. Our findings indicate that Slug is an important modulator of the therapeutic response of cholangiocarcinoma cells and is potentially useful as a sensitizer in cholangiocarcinoma therapy. One of the mechanisms is the regulation of PUMA by Slug.
Slug是一种E-钙黏蛋白抑制因子，也是PUMA（p53上调凋亡调节因子）的抑制因子，最近有研究表明Slug在调控细胞凋亡中发挥重要作用。在本研究中，我们检测了Slug沉默表达的能力是否通过诱导细胞凋亡来抑制胆管癌细胞的生长和/或使胆管癌细胞对化疗药物敏感。我们通过胆管癌细胞的全长Slug cDNA质粒（Slug cDNA）转染，使用小干扰RNA（Slug siRNA）靶向Slug基因。Slug siRNA、顺铂或Slug siRNA与顺铂联合用于体外处理胆管癌细胞。蛋白质免疫印迹法用于检测Slug、PUMA和E-钙黏蛋白的表达。TUNEL法、膜联蛋白V染色和细胞周期分析用于检测细胞凋亡。使用QBC939细胞的裸鼠皮下异种移植模型评估Slug沉默和/或顺铂对肿瘤生长的影响。免疫组织化学染色用于分析Slug和PUMA的表达。TUNEL法用于检测体内细胞凋亡。结果表明，胆管癌细胞中PUMA和E-钙黏蛋白的表达依赖于Slug。我们证明，Slug沉默和顺铂均通过上调PUMA而非上调E-钙黏蛋白来促进细胞凋亡。Slug沉默通过上调PUMA使胆管癌细胞对顺铂显著敏感。最后，我们表明Slug沉默抑制了QBC939异种移植瘤的生长，并通过上调PUMA和诱导细胞凋亡使肿瘤细胞对顺铂敏感。我们的研究结果表明，Slug是胆管癌细胞治疗反应的重要调节因子，在胆管癌治疗中作为增敏剂具有潜在用途。其中一个机制是Slug对PUMA的调控。
