miR-182 通过上调非小细胞肺癌细胞中的缺氧诱导因子 1α 促进葡萄糖代谢。
MiR-182 promotes glucose metabolism by upregulating hypoxia-inducible factor 1α in NSCLC cells.
机构信息
Department of Thoracic Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan City, Hubei Province, 430000, PR China.
Department of Thoracic Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan City, Hubei Province, 430000, PR China.
出版信息
Biochem Biophys Res Commun. 2018 Oct 2;504(2):400-405. doi: 10.1016/j.bbrc.2018.06.035. Epub 2018 Jun 23.
OBJECTIVE
This study aims to demonstrate the role of miR-182 in the glucose metabolism of NSCLC cells and the potential mechanism.
METHODS
MTT Cytotoxicity Assay was used to measure the function of differentially expressed miR-182 on two NSCLC cell lines proliferation. Metabolite analysis was introduced to monitor the glucose consumption, lactate release and glycolytic intermediate metabolites. The mRNA level of critical genes involved in glycolysis was detected by qRT-PCR. The 3'UTRs of predicted gene with a miR-182 binding site and their seed-sequence-mutated version were cloned downstream to the ORF of a Renilla luciferase reporter gene and the ability of miR-182 to downregulate luciferase expression was assessed.
RESULTS
MiR-182 had significantly improved proliferation of NSCLC cell lines. Metabolite analysis of the cells with strengthened miR-182 revealed significantly increased glucose consumption and lactate release, as well as glycolytic intermediate metabolites, or conversely. Among a panel of genes controlling glucose metabolism, miR-182 exhibited significantly influence on ENO1, GLUT1, HIF-1α, HK1, HK2, LDHA and PDK1, especially HIF-1α. For the predicted target gene HIF1AN, the wild-type but not mutated 3'UTR, responded to miR-182 b y directing ∼45% reduction of reporter gene expression.
CONCLUSION
MiR-182 promotes glucose metabolism by upregulating HIF-1α in NSCLC cells.
目的
本研究旨在探讨 miR-182 在非小细胞肺癌（NSCLC）细胞葡萄糖代谢中的作用及其潜在机制。
方法
采用 MTT 细胞毒性检测法测量两种 NSCLC 细胞系中差异表达的 miR-182 对细胞增殖的功能。代谢物分析用于监测葡萄糖消耗、乳酸释放和糖酵解中间代谢物。通过 qRT-PCR 检测参与糖酵解的关键基因的 mRNA 水平。克隆预测基因的 3'UTR 及其与 miR-182 结合位点的种子序列突变版本，下游连接到 Renilla 荧光素酶报告基因的 ORF，评估 miR-182 下调荧光素酶表达的能力。
结果
miR-182 显著改善了 NSCLC 细胞系的增殖能力。对增强 miR-182 的细胞进行代谢物分析表明，葡萄糖消耗和乳酸释放以及糖酵解中间代谢物显著增加，或者相反。在控制葡萄糖代谢的一组基因中，miR-182 对 ENO1、GLUT1、HIF-1α、HK1、HK2、LDHA 和 PDK1 表现出显著影响，尤其是 HIF-1α。对于预测的靶基因 HIF1AN，野生型而非突变型 3'UTR 对 miR-182 作出反应，导致报告基因表达减少约 45%。
结论
miR-182 通过上调 NSCLC 细胞中的 HIF-1α 促进葡萄糖代谢。
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