沉默STAT3增强SNX-2112诱导食管癌干细胞凋亡的研究

徐单单; 陈素红; 王颖

doi:10.3971/j.issn.1000-8578.2019.18.1004

沉默STAT3增强SNX-2112诱导食管癌干细胞凋亡的研究

Silence of STAT3 Enhances SNX-2112-induced Apoptosis of Esophageal Cancer Stem Cells

摘要

摘要:
目的通过沉默STAT3探讨Hsp90抑制剂SNX-2112对食管癌干细胞的凋亡诱导效果。
方法构建干扰STAT3的慢病毒载体shSTAT3并转染食管癌干细胞，同时设计阴性对照组，SNX-2112作用于细胞，CCK8法分析shSTAT3与SNX-2112联用对食管癌干细胞的活性抑制效果；软琼脂平板克隆实验研究二者联用对细胞的增殖抑制效果；流式细胞仪检测并分析二者联用对食管癌干细胞的凋亡诱导效果；Western blot分析凋亡蛋白变化。
结果设计并构建的针对STAT3的慢病毒载体shSTAT3能在蛋白（P < 0.01）及mRNA水平（P < 0.05）有效干扰其表达；经过shSTAT3及0.015625、0.03125、0.0625、0.125、0.25、0.5和1.0 μmol/L的SNX-2112作用24 h后，细胞活性较单用组明显下降（P < 0.05），同时SNX-2112的IC₅₀为（0.28±0.01）μmol/L；克隆形成实验显示，二者联用导致细胞克隆形成效率明显下降（P < 0.01）；流式细胞仪结果显示，二者联用后细胞凋亡比例以及subG₁比例细胞数量显著提高（P < 0.01, P < 0.01）；Western blot结果显示，shSTAT3及SNX-2112联用后抗凋亡蛋白Bcl2表达水平下调（P < 0.05），促凋亡蛋白Bax水平上调（P < 0.05）。
结论沉默STAT3增强了Hsp90抑制剂SNX-2112诱导食管癌干细胞凋亡的效果。

Abstract:
Objective To investigate the effect of STAT3 silence on SNX-2112-induced apoptosis of esophageal cancer stem cells.
Methods shSTAT3 lentiviral vector was designed and constructed. Esophageal cancer stem cells were transfected with shSTAT3 vector and then these cells were treated by Hsp90 inhibitor SNX-2112. CCK8 assay and soft agar plate assay were employed to evaluate the inhibitory effect of shSTAT3 and SNX-2112 on the viability and proliferation of esophageal cancer stem cells. Flow cytometry assay was used to detect the induction effect of shSTAT3 and SNX-2112 on the apoptosis of esophageal cancer stem cells. The expression levels of Bcl2 and Bax in esophageal cancer stem cells were analyzed by Western blot.
Results shSTAT3 significantly silenced the expression of STAT3 at mRNA(P < 0.05) and protein(P < 0.01) levels. shSTAT3 and different concentrations (0.015625, 0.03125, 0.0625, 0.125, 0.25, 0.5, 1.0 μmol/L) of SNX-2112 inhibited notably the viability of cancer stem cells after 24h. And the IC₅₀ value was (0.28±0.01) μmol/L. The colony formation efficiency was decreased after cancer stem cells were treated by shSTAT3 and SNX-2112(P < 0.01). The apoptotic cells and the percentage of subG₁ were increased significantly by shSTAT3 and SNX-2112(both P < 0.01). Bcl2 expression was decreased while Bax expression was increased by shSTAT3 and SNX-2112(both P < 0.05).
Conclusion Silence of STAT3 enhances SNX-2112-induced apoptosis of esophageal cancer stem cells.

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