胶质细胞源性神经营养因子对胶质瘤促增殖作用的基因表达谱研究  

The Study of Gene Expression Profile of Glial Cell Line-derived Neurotrophic Factor for the Proliferation of Glioma

作　　者：王莉 肖成华 高殿帅 

机构地区：[1]徐州医科大学附属医院神经内科,221002

出　　处：《中国临床神经科学》2017年第6期629-640,共12页Chinese Journal of Clinical Neurosciences

基　　金：国家自然科学基金课题(编号:31271358)

摘　　要：目的研究胶质细胞源性神经营养因子(GDNF)促进胶质瘤增殖的差异表达基因。方法体外培养的大鼠C6胶质瘤细胞分为3组,正常对照组(对照组)和实验组(GDNF 70μg·L-1干预6 h组和GDNF 70μg·L-1干预12 h组)。利用全基因组表达谱芯片技术进行分析,实时定量PCR验证芯片结果,并对差异基因进行聚类分析和信号转导通路分析等。结果与对照组比较,GDNF干预6 h组和GDNF干预12 h组分别上调772和664个基因,下调282和259个基因。其中,最显著的上调基因为NTN1、SLC7A5、NDST1、CD24、NFIC、KDM4A、CEACAM1、SOCS2。下调基因主要有Sctr、C4-2、GNAL、HSD3B1、Stfa2l1等。基于KEGG数据库进行生物学通路分析显示上调基因主要参与葡糖胺聚糖半乳糖基转移酶-硫酸盐肝素的生物合成、细胞的黏附连接、癌症的转录失活、JAK-STAT信号通路等。选取8个差异基因(CD24,CEACAM1、Synpo、Ndst1、Nfic、Ntn1、Socs2、Ccdc6)进行RT-PCR验证,GDNF干预12 h组与对照组比较,CEACAM1、Nfic、Socs2、Ccdc6基因表达倍数显著增加(P<0.05)。提示差异基因的mRNA变化趋势与表达谱结果一致。结论应用全基因组表达谱芯片并结合生物信息学软件分析差异表达基因,为GDNF促进胶质瘤细胞增殖的分子生物学机制的研究提供了依据。Aim To study the gene expression profile of glial cell line-derived neurotrophic factor（GDNF） in promoting the proliferating of gliomas. Methods The C6 glioma cells cultured in vitro were divided into 3 groups： a normal control group and two treatment groups（with the concentration of GDNF was 70 μg·L^（-1） and the time lasted 6 h, 12 h respectively）. Then the technique of Nimble Gen was used to analyze the data. The differentially expressed genes identified from the microarray results were verified by real-time PCR（RT-PCR）. These differentially expressed genes were analyzed using cluster analysis and Kyoto Encyclopedia of Genes and Genomes（KEGG）pathway analysis.Results Compared with the control group,772 genes at 6 h and 664 genes at 12 h after GDNF were up-regulated and 282 genes at 6 h and 259 genes at 12 h were down-regulated.Among them,the most significant up-regulated genes were NTN1（Netrin-1）,SLC7A5,NDST1,CD24,NFIC,KDM4A,CEACAM1,SOCS2.The expression of Sctr（secretin receptor）,C4-2,GNAL,HSD3B1,Stfa2l1 was down-regulated.The latest KEGG database for biological pathway analysis showed that the up-regulated genes were mainly involved in the biosynthesis of glycosaminoglycan-heparan sulfate,cell adhesion junction,transcriptional misregulation in cancer,and JAK-STAT signaling pathway.Eight differential genes（CD24,CEACAM1,Synpo,Ndst1,Nfic,Ntn1,Socs2,Ccdc6）were selected for RT-PCR verification.Compared with the control group,the expression of CEACAM1,Nfic,Socs2,Ccdc6 increased significantly in GDNF at 12 h.These results suggested that the trend of differential genes mRNA should be in line with the microarray data.Conclusion Differentially expressed genes were found by analyzing the whole genome oligonucleotide microarray combined with the bioinformatics approaches,which may serve as a basis for studying the mechanism of GDNF in promoting the proliferation of gliomas.

关 键 词：胶质细胞源性神经营养因子 大鼠C6胶质瘤细胞 全基因表达谱 细胞增殖 差异基因 

分 类 号：Q189[生物学—神经生物学] R730.264[生物学—普通生物学]