miR-216b-5p靶向调控BTN3A2促进胶质瘤细胞LN-229的迁移以及侵袭能力

大量证据表明microRNA（miRNA）通过靶向调控靶基因的表达从而在肿瘤侵袭与转移中发挥重要作用。然而关于microRNA-216b-5p (miR-216b-5p )通过靶向嗜乳脂蛋白第3亚家族膜蛋白A2（butyrophilin subfamily 3 member A2，BTN3A2）促进胶质瘤侵袭与转移的机制尚不明确。本研究通过GSE15824与GSE4290差异表达分析筛选出同时在2个芯片中表达上调的BTN3A2（P＜0.05）。生存曲线结果显示，高表达BTN3A2病人总生存期明显下降（P＜0.001）。表达量分析结果显示，BTN3A2表达随WHO分级升高而升高（P＜0.05），同时1p/19q未联合缺失与IDH突变型病人BTN3A2表达升高（P＜0.001）。基因集富集分析（gene set enrichment analysis，GSEA）结果显示，BTN3A2与众多癌症相关通路有关（P＜0.05）；Western印迹结果显示，BTN3A2在7例胶质瘤组织和胶质瘤细胞系U87、U251和LN-229中表达上调，过表达miR-216b-5p （miR-216b-5p mimics）后BTN3A2蛋白表达水平降低；Transwell结果显示，转染BTN3A2干扰质粒（si-BTN3A2）和miR-216b-5p mimics后可以抑制LN 229细胞体外迁移与侵袭能力（P＜0.05）；在线预测网站证实，miR-216b-5p 为BTN3A2潜在靶基因；生存曲线结果显示，与低表达miR-216b-5p 病人相比，高表达病人生存率明显上调（P=0.025）；荧光定量RT PCR结果显示，miR-216b-5p 在胶质瘤U87、U251和LN-229细胞中表达下降（P＜0.05）；双荧光素酶结果显示，BTN3A2存在与miR-216b-5p 的结合靶点(P<005)；综上所述，BTN3A2可能通过结合miR-216b-5p 促进胶质瘤细胞LN 229的迁移以及侵袭。

miR-216b-5p Promotes the Migration and Invasion of LN-229 Glioma Cells by Targeting BTN3A2

Accumulating evidence indicated that microRNAs (miRNA) play an important role in tumor invasion and metastasis by regulating their target genes. However, whether the miRNA-216b-5p (miR-216b-5p ) and their target genes butyrophilin subfamily 3 member A2(BTN3A2) promote glioma invasion and metastasis is unclear. This study aims to study whether miR-216b-5p promoted migration and invasion in glioma cells by negatively regulating BTN3A2. The differential expression analysis of GSE15824 and GSE4290 was analyzed by GEO2R. We found that only BTN3A2 is up-regulated in both GSE15824 and GSE4290 (P<0.05). The gene set enrichment analysis (GSEA) analysis indicated BTN3A2 was related to many cancer-related pathways (P<0.05). The results of survival curves showed that the overall survival of patients with high expression of BTN3A2 decreased significantly (P <0.001). The expression of BTN3A2 was increased with the increase of WHO grade (P<0.05), while the expression of BTN3A2 was increased in 1p/19q uncombined deletion and IDH mutant patients (P<0.001). Western blotting results showed that BTN3A2 was up-regulated in seven glioma tissues and glioma cell lines U87, U251 and LN-229 and down regulated in the miR-216b-5p mimics group; Transwell results showed that transfection with BTN3A2 silencing plasmids(si-BTN3A2) or miR-216b-5p mimics plasmids could inhibit the ability of migration and invasion in LN-229 cells in vitro (P<0.05). The online websites predicted miR-216b-5p as a potential target gene of BTN3A2. The survival curve results show that compared with patients with low expression of miR-216b-5p , the survival rate of patients with high expression was significantly increased (P=0.025). The relative expression of miR-216b-5p was decreased in U87, U251 and LN 229 cells was detected by real time quantitative PCR (P<0.05). The results of dual luciferase assay showed that BTN3A2 could bind to miR-216b-5p (P<0.05). Transwell experiment results showed that overexpression of miR-216b-5p can inhibit the migration and invasion ability of LN 229 cells (P<0.05). In summary, miR-216b-5p promotes the migration and invasion by targeting BTN3A2 of LN-229 glioma cells.