MicroRNA．218通过靶向多梳家族Bmil基因抑制胶质瘤的侵袭、迁移、增殖和肿瘤干细胞样细胞的自我更新

MicroRNA-218 inhibits glioma invasion, mi gration, proliferation, and cancer stem-like cell self-renewal by targeting the polycomb group gene Bmil

目的：探讨microRNA-218抑制胶质瘤侵袭、迁移、增殖和抑制肿瘤干细胞样细胞自我更新的分子机制．方法：在胶质瘤细胞中过表达miR-218，利用定量逆转录PCR（qRT-PCR）和免疫印迹法（Westernblotting）检测Bmil的表达水平，并利用创伤修复和体外迁移实验检测胶质瘤细胞的迁移和增殖能力；通过裸鼠移植瘤实验，检测体内过表达miR-218对胶质瘤生长的影响；利用生物信息学手段预测miR-218的直接靶标，并通过qRT—PCR、蛋白质印记技术、荧光素酶报告系统和免疫荧光实验对预测的靶标基因进行验证；以胶质瘤细胞球形成实验和极限稀释法验证miR-218对胶质瘤干细胞样细胞自我更新能力的影响；构建Broil干扰载体下调胶质瘤细胞中Bmil的表达，检测对胶质瘤迁移和增殖的影响；利用基因表达芯片分析胶质瘤细胞中miR-218对基因表达的影响．结果：Bmil是miR-218的一个直接靶标，miR-218通过下调Broil的表达来削弱胶质瘤细胞的迁移和增殖能力，并抑制胶质瘤干细胞样细胞的自我更新．另外，miR-218还参与调控一系列与胶质瘤细胞发育相关的基因．结论：作为一种肿瘤抑制因子，miR-218能阻止胶质瘤细胞的迁移、侵袭、增殖，并削弱其干细胞样特征．

AIM： To study the molecular mechanism of microR- NA-218 inhibiting glioma invasion, migration, proliferation, and cancer stem-like cell serf-renewal. METHODS ： miR-218 was overexpressed in glioma cells. Then quantitative reverse transcrip tion PCR （qRT-PCR） and western blotting analysis were used to detect the expression level of Broil. Wound healing and in vitro migration assays were employed to study the glioma migration and proliferation. The effect of miR-218 on cancer growth was deter mined via animal studies. Bioinformatics anlysis was performed to predict the direct targets of miR-218. Predicted target genes were verified using qRT-PCR, western bloting, luciferase reporter and The effect of miR-218 on self-renewal ca- pacity of glioma stem-like cells was examed using tumor sphere formation and limiting dilution assay. To determin the function of Bmil in glioma migration and proliferation, a RNAi vector of Broil was constructed and introduced into glioma cells. Further more, effect of miR-218 on genes involved in glioma development were determined using microarray-based gene expression analysis. RESULTS： Broil is a direct target of miR-218, miR-218 inhibitglioma migration, proliferation, and cancer stem-like cell self-re newal by targeting Bmil. Moreover, miR-218 also regulates a se ries of genes involved in glioma development. CONCLUSION： As a tumor suppressor, miR-218 prevents migration, invasion, proliferation, and stemlike qualities in glioma cells.