敲低 lncRNA XIST 通过调控 miR-381-3p/NEK5 轴抑制鼻咽癌细胞缺氧诱导的糖酵解、迁移和侵袭。
Knockdown of lncRNA XIST inhibits hypoxia-induced glycolysis, migration and invasion through regulating miR-381-3p/NEK5 axis in nasopharyngeal carcinoma.
机构信息
Department of Otorhinolaryngology, the First People's Hospital of Tongxiang City, Tongxiang, P.R. China.
出版信息
Eur Rev Med Pharmacol Sci. 2020 Mar;24(5):2505-2517. doi: 10.26355/eurrev_202003_20518.
OBJECTIVE
Hypoxia is an important feature of nasopharyngeal carcinoma (NPC). Growing evidence demonstrated that long non-coding RNAs (lncRNAs) could participate in cancer progression and hypoxia regulation. However, the exact roles and underlying mechanism of lncRNA X-inactive specific transcript (XIST) in NPC under hypoxia are still unclear.
MATERIALS AND METHODS
The expressions of XIST, microRNA-381-3p (miR-381-3p) and NIMA related kinase 5 (NEK5) were detected by quantitative Real-time polymerase chain reaction (qRT-PCR). The glucose consumption and lactate production were measured using the glucose assay kit and lactate assay kit, respectively. Western blot assay was used to determine the protein levels of hexokinase II (HK2) and NEK5. Transwell assay was employed to evaluate cell migration and invasion. The interaction between miR-381-3p and XIST or NEK5 was predicted by bioinformatics analysis and verified by dual-luciferase reporter assay. The mice xenograft model was established to investigate the roles of XIST in NPC progression in vivo.
RESULTS
XIST and NEK5 were highly expressed while miR-381-3p was lowly expressed in NPC (tissues and cells) and hypoxia-induced NPC cells. Deficiency of XIST or NEK5 suppressed hypoxia-induced glycolysis and metastasis in NPC cells. Moreover, miR-381-3p could directly bind to XIST and its inhibition reversed the inhibitory effects of XIST knockdown on glycolysis and metastasis under hypoxia. NEK5 was a direct target of miR-381-3p and its interference attenuated the promotive effects of miR-381-3p downregulation on glycolysis and metastasis under hypoxic conditions. Besides, interference of XIST decreased tumor growth by upregulating miR-381-3p and downregulating NEK5.
CONCLUSIONS
XIST knockdown inhibited glycolysis and metastasis in hypoxia-induced NPC cells through regulating miR-381-3p/NEK5 axis, providing new insights into the pathogenesis of NPC.
目的
缺氧是鼻咽癌（NPC）的一个重要特征。越来越多的证据表明，长非编码 RNA（lncRNA）可以参与癌症的进展和缺氧的调节。然而，lncRNA X 失活特异性转录物（XIST）在 NPC 中的具体作用及其在缺氧下的潜在机制仍不清楚。
材料与方法
采用实时定量聚合酶链反应（qRT-PCR）检测 XIST、microRNA-381-3p（miR-381-3p）和 NIMA 相关激酶 5（NEK5）的表达。葡萄糖检测试剂盒和乳酸检测试剂盒分别用于检测葡萄糖消耗和乳酸生成。Western blot 检测用于测定己糖激酶 II（HK2）和 NEK5 的蛋白水平。Transwell 实验用于评估细胞迁移和侵袭。通过生物信息学分析预测 miR-381-3p 与 XIST 或 NEK5 的相互作用，并通过双荧光素酶报告基因实验进行验证。建立小鼠异种移植模型，研究 XIST 在体内 NPC 进展中的作用。
结果
在 NPC（组织和细胞）和缺氧诱导的 NPC 细胞中，XIST 和 NEK5 高表达，而 miR-381-3p 低表达。XIST 或 NEK5 的缺失抑制了 NPC 细胞在缺氧下的糖酵解和转移。此外，miR-381-3p 可直接与 XIST 结合，其抑制作用逆转了 XIST 敲低对缺氧下糖酵解和转移的抑制作用。NEK5 是 miR-381-3p 的直接靶标，其干扰减弱了 miR-381-3p 下调对缺氧下糖酵解和转移的促进作用。此外，XIST 的干扰通过上调 miR-381-3p 和下调 NEK5 来抑制肿瘤生长。
结论
XIST 敲低通过调节 miR-381-3p/NEK5 轴抑制缺氧诱导的 NPC 细胞中的糖酵解和转移，为 NPC 的发病机制提供了新的见解。
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