MicroRNA-99a induces G1-phase cell cycle arrest and suppresses tumorigenicity in renal cell carcinoma.
MicroRNA-99a induces G1-phase cell cycle arrest and suppresses tumorigenicity in renal cell carcinoma.
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DOI：
10.1186/1471-2407-12-546
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发表时间：
2012-11-23
期刊：
影响因子：
3.8
通讯作者：
He X
中科院分区：
文献类型：
--
作者：
Cui L;Zhou H;Zhao H;Zhou Y;Xu R;Xu X;Zheng L;Xue Z;Xia W;Zhang B;Ding T;Cao Y;Tian Z;Shi Q;He X
越来越多的证据表明，microRNAs （miRNAs）在癌症诊断和治疗中发挥着重要作用。MicroRNA-99a （miR-99a）是一种潜在的肿瘤抑制因子，在几种人类恶性肿瘤中下调。然而，miR-99a在人肾细胞癌（RCC）中的表达和功能迄今尚未被研究。因此，我们检测了miR-99a在RCC细胞系和组织中的表达，并评估了miR-99a对RCC肿瘤发生的影响。通过实时定量反转录PCR （qRT-PCR）评估40对RCC和匹配的邻近非肿瘤组织中的MiR-99a水平。用miR-99a模拟物转染RCC细胞系786-O和OS-RC-2以恢复miR-99a的表达。然后通过细胞增殖、细胞周期、transwell和集落形成试验评估miR-99a的作用。通过小鼠RCC异种移植模型证实miR-99a对体内致瘤性的影响。利用western blotting和荧光素酶报告基因法鉴定潜在靶基因。我们发现miR-99a在RCC中显著下调，miR-99a的低表达水平与RCC患者的低生存率相关。在体外实验中，miR-99a的修复显著抑制了RCC细胞的生长、克隆、迁移和侵袭，并诱导g1期细胞周期阻滞。此外，在人RCC小鼠异种移植模型中，瘤内递送miR-99a可抑制肿瘤生长。此外，我们还发现哺乳动物雷帕霉素靶点（mTOR）是RCC细胞中miR-99a的直接靶点。此外，sirna介导的mTOR敲低部分地表型化了miR-99a过表达的作用，这表明miR-99a的肿瘤抑制作用可能主要通过mTOR调控介导。总之，据我们所知，这些结果首次表明miR-99a的失调部分通过直接靶向mTOR途径参与了RCC的病因学，这表明miR-99a可能为RCC的诊断和治疗干预提供了一个有吸引力的新靶点。
A growing body of evidence suggests that microRNAs (miRNAs) play an important role in cancer diagnosis and therapy. MicroRNA-99a (miR-99a), a potential tumor suppressor, is downregulated in several human malignancies. The expression and function of miR-99a, however, have not been investigated in human renal cell carcinoma (RCC) so far. We therefore examined the expression of miR-99a in RCC cell lines and tissues, and assessed the impact of miR-99a on the tumorigenesis of RCC. MiR-99a levels in 40 pairs of RCC and matched adjacent non-tumor tissues were assessed by real-time quantitative Reverse Transcription PCR (qRT-PCR). The RCC cell lines 786-O and OS-RC-2 were transfected with miR-99a mimics to restore the expression of miR-99a. The effects of miR-99a were then assessed by cell proliferation, cell cycle, transwell, and colony formation assay. A murine xenograft model of RCC was used to confirm the effect of miR-99a on tumorigenicity in vivo. Potential target genes were identified by western blotting and luciferase reporter assay. We found that miR-99a was remarkably downregulated in RCC and low expression level of miR-99a was correlated with poor survival of RCC patients. Restoration of miR-99a dramatically suppressed RCC cells growth, clonability, migration and invasion as well as induced G1-phase cell cycle arrest in vitro. Moreover, intratumoral delivery of miR-99a could inhibit tumor growth in murine xenograft models of human RCC. In addition, we also fond that mammalian target of rapamycin (mTOR) was a direct target of miR-99a in RCC cells. Furthermore, siRNA-mediated knockdown of mTOR partially phenocopied the effect of miR-99a overexpression, suggesting that the tumor suppressive role of miR-99a may be mediated primarily through mTOR regulation. Collectively, these results demonstrate for the first time, to our knowledge, that deregulation of miR-99a is involved in the etiology of RCC partially via direct targeting mTOR pathway, which suggests that miR-99a may offer an attractive new target for diagnostic and therapeutic intervention in RCC.
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