目的：探讨脑胶质瘤细胞中miR－21对FasL表达的调控作用以及对细胞生长和凋亡的影响，并研究其分子作用机制。方法：将miR－21模拟物（ miR－21 mimics ）、miR－21抑制物（ miR－21 inhibitor ）以及阴性对照（scramble）瞬时转染到U251细胞中，CCK－8法和流式细胞术检测细胞活力和凋亡情况。构建FasL 3’ UTR双萤光素酶报告载体，通过采用双萤光素酶报告实验验证miR－21的靶基因。构建表达载体pcDNA3．1－FasL，回复实验分析miR－21对细胞凋亡的影响。结果：miR－21过表达可促进U251细胞的活力，抑制细胞凋亡；miR－21表达下调则抑制细胞活力，促进细胞凋亡，和对照组比较差异有统计学意义（P＜0．05）。双萤光素酶报告实验和回复实验结果提示miR－21可以通过作用于FasL的3’ UTR区，负向调控其表达，从而抑制细胞的凋亡。结论： miR－21可以通过靶向调控FasL的表达进而促进U251细胞的生长。%AIM:ToinvestigatetheregulationofmiR-21onFasLexpressionanditseffectonthegrowthand apoptosis in glioma cells , and to evaluate the molecular mechanism .METHODS:Differential expression levels of miR-21 in human glioma U251 cells were achieved by transfecting with miR-21 mimics, miR-21 inhibitor or scramble .The viability and apoptosis of U251 cells were detected by CCK-8 assay and flow cytometry with Annexin V-FITC/PI double staining. The recombination vector pmirGLO-FasL was constructed .Dual-luciferase reporter experiment was performed to validate the target genes of miR-21.The expression vector pcDNA3.1-FasL was also constructed , and the biological activity and regula-tory role of miR-21 in U251 cell apoptosis were analyzed by a restore experiment .RESULTS:Exogenous overexpression of miR-21 increased the viability and decreased the apoptosis of U 251 cells ( P<0.05 ) , while miR-21 inhibitors generated the opposite results (P<0.05).Dual-luciferase reporter assay and restore experiment revealed that miR-21 negatively reg-ulated the expression of FasL gene which was regarded as the target gene , thus decreasing the apoptosis of U 251 cells. CONCLUSION:miR-21 increases the viability of glioma U251 cells, in which FasL may be one of the target genes .
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