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miR-212-3p靶向调控NAP1L1抑制胶质瘤细胞增殖、迁移和上皮-间充质转化

郭小兵 李晓文 李恒希 曹艳 李坪

郭小兵, 李晓文, 李恒希, 曹艳, 李坪. miR-212-3p靶向调控NAP1L1抑制胶质瘤细胞增殖、迁移和上皮-间充质转化[J]. 昆明医科大学学报. doi: 10.12259/j.issn.2095-610X.S20241104
引用本文: 郭小兵, 李晓文, 李恒希, 曹艳, 李坪. miR-212-3p靶向调控NAP1L1抑制胶质瘤细胞增殖、迁移和上皮-间充质转化[J]. 昆明医科大学学报. doi: 10.12259/j.issn.2095-610X.S20241104
Xiaobing GUO, Xiaowen LI, Hengxi LI, Yan CAO, Ping LI. miR-212-3p Targeted Regulation of NAP1L1 Inhibits Glioma Cell Proliferation,Migration and EMT[J]. Journal of Kunming Medical University. doi: 10.12259/j.issn.2095-610X.S20241104
Citation: Xiaobing GUO, Xiaowen LI, Hengxi LI, Yan CAO, Ping LI. miR-212-3p Targeted Regulation of NAP1L1 Inhibits Glioma Cell Proliferation,Migration and EMT[J]. Journal of Kunming Medical University. doi: 10.12259/j.issn.2095-610X.S20241104

miR-212-3p靶向调控NAP1L1抑制胶质瘤细胞增殖、迁移和上皮-间充质转化

doi: 10.12259/j.issn.2095-610X.S20241104
基金项目: 云南省科技厅-昆明医科大学应用基础研究联合专项基金资助项目(202101AY070001-037)
详细信息
    作者简介:

    郭小兵(1982~),男,湖南邵东人,医学硕士,讲师,主要从事神经系统疾病与损伤研究工作

    通讯作者:

    李坪,E-mail:liping@kmmu.edu.cn

  • 中图分类号: R735.7

miR-212-3p Targeted Regulation of NAP1L1 Inhibits Glioma Cell Proliferation,Migration and EMT

  • 摘要:   目的  探究miR-212-3p在胶质瘤细胞增殖、侵袭和上皮-间充质转化(EMT)中的分子机制。  方法  RT-qPCR用于衡量胶质瘤细胞中miR-212-3p和NAP1L1表达,构建NC mimic、miR-212-3p mimic、oe-NC和oe-NAP1L1转染至细胞中,利用CCK-8 、Transwell和伤口实验评估细胞生物学行为。通过Western blot分析EMT相关标志物的蛋白表达。miR-212-3p与NAP1L1的关系通过双荧光素酶报告基因和AgO2-RIP实验进行证实。  结果  miR-212-3p的表达水平在胶质瘤细胞中显著下调(P < 0.0001),过表达miR-212-3p 可以显著降低胶质瘤细胞增殖(P < 0.0001)、侵袭(P = 0.0011)和迁移能力(P < 0.0001),并且抑制了EMT标志物N-钙黏蛋白(N-cadherin)(P = 0.000861)和波形蛋白(Vimentin)(P = 0.007430)的表达,而上调了E-钙黏蛋白(E-cadherin)(P < 0.0001)的表达。miR-212-3p靶向负调控NAP1L1的表达。过表达NAP1L1逆转了miR-212-3p对胶质瘤细胞增殖(P < 0.0001)、迁移(P < 0.0001)和EMT(P < 0.0001)的抑制作用。  结论  miR-212-3p通过靶向负调节NAP1L1的表达抑制胶质瘤细胞增殖、迁移和EMT。
  • 图  1  miR-212-3p在胶质瘤细胞中低表达

    与NHA组比较,***P < 0.001。

    Figure  1.  Low expression of miR-212-3p in glioma cells

    图  2  miR-212-3p抑制胶质瘤细胞增殖、迁移和EMT

    A:RT- qPCR检测miR-212-3p转染效率;B:CCK-8分析;C:Transwell评估细胞侵袭;D:伤口愈合实验评估细胞迁移;E:Western blot分析EMT相关标志物的表达(N-cadherin、E-cadherin和Vimentin)的蛋白水平。与NC组比较,**P < 0.01,***P < 0.001。

    Figure  2.  miR-212-3p inhibits glioma cell proliferation,migration and EMT

    图  3  miR-212-3p靶向调控NAP1L1的表达

    A:miR-212-3p与NAP1L1的结合序列;B:双荧光素酶基因报告验证miR-212-3p与NAP1L1的靶向关系;C:AgO2-RIP检测miR-212-3p与NAP1L1的关系;D:生存率预测,high-risk样本33例,low-risk样本41例,P-value=0.023500;E:RT-qPCR检测细胞中NAP1L1的表达;与NC-mimic组比较,***P < 0.001。

    Figure  3.  miR-212-3p targets and regulates NAP1L1 expression

    图  4  miR-212-3p通过下调NAP1L1抑制胶质瘤细胞增殖、迁移和EMT

    A:RT- qPCR检测NAP1L1转染效率;B:RT- qPCR检测NAP1L1的表达;C:CCK-8分析;D:Transwell评估细胞侵袭;E:伤口愈合实验评估细胞迁移;F:Western blot检测N-cadherin、E-cadherin和Vimentin的蛋白水平。与NC组比较,**P < 0.01,***P < 0.001;与miR-212-3p mimic组比较,#P < 0.05,##P < 0.01,###P < 0.001。

    Figure  4.  miR-212-3p inhibits glioma cell proliferation,migration and EMT through down-regulation of NAP1L1

    表  1  PCR引物序列

    Table  1.   PCR primer sequences

    基因 引物 序列(5'-3'
    miR-212-3p forward 5'-GCGCGTAACAGTCTCCAGTC -3'
    reverse 5'-AGTGCAGGGTCCGAGGTATT -3'
    U6 forward 5'-CAAATTCGTGAAGCGTTCCA-3'
    reverse 5'-AGTGCAGGGTCCGAGGTATT-3'
    NAP1L1 forward 5'-GGCAGACATTGACAACAA-3'
    reverse 5'-TTCCTCCGAAATCTCATC-3'
    GAPDH forward 5'-TGACCACAGTCCATGCCATCAC-3'
    reverse 5'-CGCCTGCTTCACCACCTTCTT-3'
    下载: 导出CSV
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