骨肉瘤长链非编码RNA基因芯片检测及差异分析

Detection and differential gene analysis of long-chain non-coding RNA microarray in osteosarcoma tissues

目的采用长链非编码RNA(lncRNA)芯片检测在骨肉瘤与瘤旁组织中lncRNA表达谱,分析差异表达的lncRNA,探讨其功能。方法利用lncRNA芯片筛查2010年至2012年期间广州市第一人民医院和南部战区总医院7对骨肉瘤组织与瘤旁组织中lncRNA表达谱,实时荧光定量聚合酶链式反应(RT-qPCR)验证芯片结果;分层聚类和火山图分析特征性表达lncRNA,结合基因本体论数据库(GO)分析和通路分析探讨lncRNA参与肿瘤细胞的生物过程。使用Agilent Array平台对样品进行微阵列分析。Agilent Feature Extraction软件分析采集阵列图像。GeneSpring GX v11.5.1软件包进行分子标准化和随后数据处理。采用重复测量设计的方差分析评价各组之间的变化,多组间比较采用单因素方差分析。结果筛选得到7组瘤组织中差异lncRNA总数为4221个,其中上调的1995个,下调的2226个;RT-qPCR验证结果与lncRNA结果基本一致;分层聚类分析显示骨肉瘤组织(T)和瘤旁组织(C)中的lncRNA表达差异有统计学意义;GO分析显示lncRNA广泛参与细胞的生物过程,还参与细胞的细胞器组成。结论lncRNA芯片能有效检测骨肉瘤组织差异性表达的lncRNA,结合分子生物学分析技术发现lncRNA参与骨肉瘤细胞多种生物活性过程。

Objective To detect and analyze the expression profile of long noncoding RNA(lncRNA)that is differentially expressed in osteosarcoma and paratumoral tissues by lncRNA microarray.Simultaneously,functional research was carried out in order to provide basis and ideas for gene therapy of osteosarcoma.Methods The lncRNA expression profiles of 7 pairs of osteosarcoma and paratumoral tissues which were gotten inGuangzhou First People’s Hospital and General Hospital of Southern Theatre Command from 2010 to 2012,were screened by lncRNA microarray,and the results were verified by real-time quantitative reverse transcriptase-polymerase chain reaction(RT-qPCR).Hierarchical clustering and scatter plot analysis,combined with Gene Ontology(GO)and pathway analysis,were used to study the characteristic expression of lncRNA and the biological process of tumor cells.The samples were analyzed by microarray using Agilent Array platform.Agilent Feature Extraction software was used to analyze and collect array images.GeneSpring GX v11.5.1 software was used for molecular standardization and subsequent data processing.The statistical software Graphpam,and variance analysis were used to analyze the data.P<0.05 was considered statistically significant.Results The results showed that there were 4221 lncRNA differentially expressed in cancer tissues,of which 1995 were up-regulated and 2226 were down-regulated.RT-qPCR verification results were basically consistent with lncRNA results.Hierarchical cluster analysis indicated that there was significant difference in the expression of lncRNA between(T)and(C)in osteosarcoma.GO analysis showed that lncRNA was not only widely involved in the biological process of cells,but also in the organelle composition of cells.Conclusion lncRNA chip can effectively and reliably detect the differential expression of lncRNA in osteosarcoma.It also can be found that lncRNA is involved in a variety of biological activities of osteosarcoma cells by lncRNA combined with molecular biological techniques.This study may provide basis and ideas for further study on the roles of lncRNA in osteosarcoma.