Up-Regulation of miR-204 Enhances Anoikis Sensitivity in Epithelial Ovarian Cancer Cell Line Via Brain-Derived Neurotrophic Factor Pathway In Vitro
Up-Regulation of miR-204 Enhances Anoikis Sensitivity in Epithelial Ovarian Cancer Cell Line Via Brain-Derived Neurotrophic Factor Pathway In Vitro
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DOI：
10.1097/igc.0000000000000456
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发表时间：
2015-07-01
影响因子：
4.8
通讯作者：
Wang, Zheng
中科院分区：
文献类型：
--
作者：
Yan, Hongliang;Wu, Weiguang;Wang, Zheng
miR-204是一种预测靶向脑源性神经营养因子（BDNF）的基因位点，在包括上皮性卵巢癌（EOC）在内的多种癌症中经常丢失。在本研究中，我们旨在发现miR-204表达水平对EOC细胞anoikis敏感性的影响，并探讨这一过程的可能机制。方法首先，我们筛选了能维持anoikis抗性并形成anoikis模式的EOC细胞。通过定量逆转录酶聚合酶链反应和Annexin-V-R-PE/7-氨基放线菌素法分别检测miR-204表达水平和细胞凋亡。然后我们通过转染pre-miR-204恢复miR-204的表达水平。检测miR-204表达水平和细胞凋亡；采用transwell侵袭法和创面愈合法检测细胞的侵袭和迁移能力。定量逆转录聚合酶链反应检测BDNF信使RNA水平；Western blot检测pAKT的表达。结果miR-204的表达呈显著下调模式。miR-204表达水平的恢复使细胞对anoikis更加敏感，侵袭转移行为减少，并导致BDNF下调表达，通过PI3K/AKT信号通路抑制线粒体依赖通路的激活，导致EOC细胞anoikis。结论miR-204的上调可能通过促进BDNF的下调与EOC细胞损伤的敏感性直接相关。我们的研究结果提供了一种新的机制来治疗性地操纵miR-204水平以恢复anoikis的敏感性。
Objective Genomic loci encoding miR-204, which was predicted to target brain-derived neurotrophic factor (BDNF), were frequently lost in multiple cancer, including epithelial ovarian cancer (EOC). In this study, we aimed to find out the influence of miR-204 expression level on EOC cell anoikis sensitivity and to explore possible mechanisms of this process.Methods First, we screened EOC cells, which maintain anoikis resistance forming an anoikis pattern. miR-204 expression level and apoptosis were measured, respectively, by quantitative reverse transcriptase polymerase chain reaction and Annexin-V-R-PE/7-amino-actinomycin assay. Then we restored the expression level of miR-204 by transfection with pre-miR-204. miR-204 expression level and apoptosis were measured as before; cell invasion and migration ability were detected by transwell invasion assay and wound-healing assay. The messenger RNA level of BDNF was also detected by quantitative reverse transcriptase polymerase chain reaction; Western blot analysis was performed to assess pAKT expression.Results Expression of miR-204 is significantly down-regulated in an anoikis pattern. Restored expression level of miR-204 enables cells to acquire more sensitivity to anoikis and decrease invasive and metastatic behavior, and also results in BDNF down-expression and inhibits activation of mitochondria-dependent pathway through the PI3K/AKT signaling pathway leading to cancer cell anoikis in EOC cells.Conclusions miR-204 up-regulation may be linked directly to the sensitivity of EOC cell anoikis by contributing to BDNF down-regulation. Our findings provide a novel mechanism for manipulating miR-204 levels therapeutically to restore anoikis sensitivity.