Salinomycin inhibits proliferation and induces apoptosis of human hepatocellular carcinoma cells in vitro and in vivo.
Salinomycin inhibits proliferation and induces apoptosis of human hepatocellular carcinoma cells in vitro and in vivo.
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Salinomycin 在体外和体内抑制人肝细胞癌细胞的增殖并诱导其凋亡。
DOI：
10.1371/journal.pone.0050638
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
Guo CY
中科院分区：
文献类型：
--
作者：
Wang F;He L;Dai WQ;Xu YP;Wu D;Lin CL;Wu SM;Cheng P;Zhang Y;Shen M;Wang CF;Lu J;Zhou YQ;Xu XF;Xu L;Guo CY
抗肿瘤抗生素盐霉素（Salinomycin，Sal）是近年来发现的一种选择性乳腺癌干细胞抑制剂，但其对肝细胞癌（Hepatocellular carcinoma，HCC）的作用尚不清楚。本研究旨在探讨丹参对肝癌的抗肿瘤作用及其机制。用Sal处理HCC细胞系（HepG 2、SMMC-7721和BEL-7402）。绘制细胞生长曲线测定细胞倍增时间，采用Cell Counting Kit 8评价细胞活力。通过流式细胞术评估CD 133+细胞亚群的分数。我们发现，Sal抑制增殖，降低PCNA水平以及HCC细胞中HCC CD 133+细胞亚群的比例。流式细胞仪分析细胞周期，结果表明，Sal引起不同肝癌细胞系的细胞周期停滞在不同时期。流式细胞术和Hoechst 33342染色检测细胞凋亡。Sal诱导细胞凋亡，其特征在于Bax/Bcl-2比率的增加。选择几种信号通路，使用真实的时间PCR和Western印迹分析进行进一步的机制分析。与对照相比，β-连环蛋白表达在添加Sal后显著下调。流式细胞仪检测肝癌细胞内Ca 2+浓度，Sal处理组细胞内Ca 2+浓度较高。使用肝癌原位肿瘤模型在体内进一步验证了Sal的抗肿瘤作用，并且所获得的数据显示，与对照相比，Sal处理组中的肝肿瘤的大小减小。免疫组化和TUNEL染色也表明，Sal抑制增殖和诱导凋亡在体内。最后，通过Western blot和免疫组织化学评估Sal对体内Wnt/β-catenin信号传导的作用。本研究表明，Sal在体外和体内抑制HCC细胞增殖并诱导细胞凋亡，一个潜在的机制是通过增加细胞内Ca 2+水平抑制Wnt/β-catenin信号传导。
The anti-tumor antibiotic salinomycin (Sal) was recently identified as a selective inhibitor of breast cancer stem cells; however, the effect of Sal on hepatocellular carcinoma (HCC) is not clear. This study aimed to determine the anti-tumor efficacy and mechanism of Sal on HCC. HCC cell lines (HepG2, SMMC-7721, and BEL-7402) were treated with Sal. Cell doubling time was determinated by drawing growth curve, cell viability was evaluated using the Cell Counting Kit 8. The fraction of CD133+ cell subpopulations was assessed by flow cytometry. We found that Sal inhibits proliferation and decreases PCNA levels as well as the proportion of HCC CD133+cell subpopulations in HCC cells. Cell cycle was analyzed using flow cytometry and showed that Sal caused cell cycle arrest of the various HCC cell lines in different phases. Cell apoptosis was evaluated using flow cytometry and Hoechst 33342 staining. Sal induced apoptosis as characterized by an increase in the Bax/Bcl-2 ratio. Several signaling pathways were selected for further mechanistic analyses using real time-PCR and Western blot assays. Compared to control, β-catenin expression is significantly down-regulated upon Sal addition. The Ca2+ concentration in HCC cells was examined by flow cytometry and higher Ca2+ concentrations were observed in Sal treatment groups. The anti-tumor effect of Sal was further verified in vivo using the hepatoma orthotopic tumor model and the data obtained showed that the size of liver tumors in Sal-treated groups decreased compared to controls. Immunohistochemistry and TUNEL staining also demonstrated that Sal inhibits proliferation and induces apoptosis in vivo. Finally, the role of Sal on in vivo Wnt/β-catenin signaling was evaluated by Western blot and immunohistochemistry. This study demonstrates Sal inhibits proliferation and induces apoptosis of HCC cells in vitro and in vivo and one potential mechanism is inhibition of Wnt/β-catenin signaling via increased intracellular Ca2+ levels.
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