人脑胶质瘤细胞系miRNA表达谱初步研究

Preliminary study on the miRNA expression protfiles in several glioma cell lines

目的确定部分人脑胶质瘤细胞系miRNA表达谱的初步特征。方法提取人脑胶质母细胞瘤细胞系U251、TJ861、TJ905、TJ899和A172以及星形细胞瘤细胞系H4细胞的miRNA，miRNA微阵列芯片杂交，扫描后分析差异表达的miRNA。选择差异表达的miR-21设计反义寡核苷酸处理U251细胞后原位杂交和Western blot检查SEPT7的表达。结果在胶质瘤细胞系中hsa—miR-21等8种miRNA一致表达上调；hsa—miR-1等18种miRNA一致表达下调。miR-21锁核酸修饰反义寡核苷酸处理U251细胞72h后，原位杂交发现阳性信号集中在细胞核的miR-21表达显著下降，Western blot发现U251细胞中SEPT7的表达明显上调。结论miRNA的差异表达可能是胶质瘤的重要分子生物学标签，并在基因表达调控中具有潜在的研究价值。

Objective To study the fundamental characters of miRNA expression profiles in several human glioma cell lines. Methods Total miRNA was isolated from human glioblastoma cell line U251, TJ861, TJ905, TJ899 and A172 as well as human astrocytoma cell line H4. Hybridization was carried out on CapitalBio＇s expression profiling microarray chip , hybridization signals were detected using LuxScan 10K/A, scanner images were quantified and analyzed by SAM. Locked nucleic acid modified antisense oligonucleotide corresponding to miR-21 and scrambled oligonucleotide were transfected into U251 cells, in situ hydridization was used to detected the expression of miR-21, Western blotting was used to investigate the expression of SEPT 7. Result Compared with normal brain tissue, eight miRNAs were over-expressed in human glioma cell lines. However, eighteen miRNAs were down-expressed. MiR-21 located in U251 cell nuclei was detected by in situ hybridization after transfect by locked nucleic acid modified antisense oligonucleotide 72 hours, consequently, SEPT7 was up-regulated in the treated U251 group. Conclusion The aberrant expression of miRNA genes may be the molecular signature of human gliomas, thereby possibly to be the potential therapeutic targets of human gliomas.