款冬花多糖调控miR-889-3p/TLR4对IL-6诱导的乳腺癌细胞增殖、迁移及侵袭的影响

Effect of polysaccharides in Tussilago farfara on the proliferation，migration and invasion of breast cancer cells induced by IL-6 by regulating miR-889-3p/TLR4

关键词:  乳腺癌  款冬花多糖  miR-889-3p  TLR4  白细胞介素6  细胞增殖  迁移

Key words:  breast cancer  polysaccharides in Tussilago farfara  miR-889-3p  TLR4  interleukin 6  proliferation  migration

摘要:

[目的] 探讨款冬花多糖对白细胞介素-6（IL-6）诱导的乳腺癌细胞恶性行为作用机制。[方法] IL-6诱导人乳腺癌细胞MCF-7后，不同浓度的款冬花多糖处理。转染miR-NC、miR-889-3p mimics的MCF-7细胞经IL-6处理；转染NC-inhibitor、miR-889-3p-inhibitors的MCF-7细胞经款冬花多糖与IL-6处理；噻唑蓝（MTT）、平板克隆形成以及Transwell^TM实验分析细胞增殖、迁移及侵袭；实时荧光定量聚合酶链反应（qRT-PCR）检测miR-889-3p、Toll 样受体 4（TLR4）的表达量；蛋白免疫印迹（Western blot）法检测基质金属蛋白酶（MMP）-2、MMP-9、TLR4蛋白表达量；双荧光素酶报告实验验证miR-889-3p与TLR4的靶向关系。[结果] 款冬花多糖可降低IL-6诱导的MCF-7细胞增殖（P＜0.05）、迁移及侵袭能力（P＜0.05），而提高miR-889-3p的表达（P＜0.05），呈剂量依赖性；TLR4是miR-889-3p的靶基因，miR-889-3p可靶向调控TLR4的表达；转染miR-889-3p mimics可降低IL-6诱导的MCF-7细胞增殖、迁移及侵袭（P＜0.05）；转染miR-889-3p-inhibitors可恢复款冬花多糖对IL-6诱导的MCF-7细胞增殖、迁移及侵袭。[结论] 款冬花多糖可通过调节miR-889-3p/TLR4表达而抑制IL-6诱导的乳腺癌细胞恶性行为。

Abstract:

[Objective] To explore the effect and mechanism of polysaccharides in Tussilago farfara on breast cancer cell proliferation，migration and invasion induced by interleukin 6（IL-6）. [Methods] After IL-6 stimulation，MCF-7 cells were treated with different concentrations of polysaccharides in Tussilago farfara. The miR-NC and miR-889-3p mimics transfected MCF-7 cells were treated with IL-6；NC-inhibitor，miR-889-3p-inhibitors-transfected MCF-7 cells were treated with polysaccharides in Tussilago farfara and IL-6. Proliferation，migration，and invasion were assessed using MTT，plate clone formation，and TranswellTM experiments，miR-889-3p and TLR4 expression were detected using qRT-PCR. Western blot method was used to analysis of protein expression quantity of MMP-2，MMP-9 and TLR4. Dual luciferase reporter experiment verified the targeting between miR-889-3p and TLR4. [Results] Polysaccharides in Tussilago farfara could reduce IL-6-induced MCF-7 cell proliferation（P＜0.05），migration and invasion（P＜0.05），while miR-889-3p was increased（P＜0.05），in a dose-dependent manner. The miR-889-3p targeted TLR4 and regulated its expression. The miR-889-3p mimics reduced IL-6-induced MCF-7 cell proliferation，migration and invasion （P＜0.05）. The miR-889-3p-inhibitors restored the effects of polysaccharides in Tussilago farfara on MCF-7 cell proliferation，migration and invasion induced by IL-6. [Conclusion] Polysaccharides in Tussilago farfara inhibited breast cancer cell malignant behaviors induced by IL-6 through regulating miR-889-3p/TLR4 axis.