浸润性膀胱癌差异基因的互作网络分析及验证研究

Analysis and verification of the interaction network of differentially expressed genes in invasive bladder cancer

目的采用蛋白互作网络分析研究表达谱基因芯片数据,构建浸润性膀胱癌基因互作网络图并对筛选出的网络中心节点进行实验验证。方法将表达谱芯片筛选到的浸润性膀胱癌152个共同差异表达基因,导入STRING在线数据库进行分析,绘制差异基因互作网络图,并将互作网络数据导出到Cytoscape2.6.2软件中,筛选出网络中心节点。采用KEGG数据库等进行信号通路分类及基因功能研究,进而采用实时定量RT-PCR对其基因表达进行验证,筛选出基因表达在膀胱癌组与正常膀胱粘膜组差异最大的基因,并对浸润性膀胱癌发生的分子机制进行初步探讨。结果共有103个膀胱癌共同差异表达基因编码的蛋白经STRING筛选存在相互作用,并构成一个复杂的互作网络图;Cytoscape共筛选出26个网络中心节点,广泛参与肿瘤发生的多条信号通路;实时定量RT-PCR结果表明UBE2C、VEGF、TGFBR2、CAV1四个基因表达量在膀胱癌组与正常膀胱粘膜组的差异最大,分别为9.45、4.17、0.13、0.18倍(以GAPHD作为内参,计算2-△△Ct),与芯片检测结果的趋势一致。结论本研究构建出的浸润性膀胱癌差异基因互作网络图,尤其是其中的网络中心节点基因,对于浸润性膀胱癌发生分子机制的研究、早期诊断及分子靶向治疗研究具有较好的提示作用。

Objective To study the data of gene expression mieroarray by protein interaction network analysis, establish an interaction network of differentially expressed genes in invasive bladder cancer and verify the central nodes of the network. Methods A total of 152 differentially expressed genes in invasive bladder cancer detected by gene expression microarray were inputted into STRING database online for analysis and establishment of the interaction network. The interaction data were imported into Cytoscape 2.6.2 software for screening the central nodes of the network. KEGG database was exploited for pathway analysis and functional study of the central node genes. Real-time RT-PCR was used for verification, and the genes with maximal differential expressions were screened for exploring the molecular mechanism of carcinogenesis of invasive bladder cancer. Results The protein products of 103 differentially expressed genes in bladder cancer had interactions, forming a complicated interaction network. Twenty-six nodes involved in several signal pathways were confirmed by Cytoscape as the central nodes of the network, among which UBE2C, VEGF, TGFBR2, and CAV 1 nodes were verified by real-time RT-PCR as the genes with maximal differential expressions between the bladder cancer and normal tissues, and the 2aAQ of these genes were 9.45, 4.17, 0.13 and 0.18 （GAPHD as the internal control）, respectively. Conclusion The interaction network of the differentially expressed genes, especially the central nodes of this network, can provide clues to the carcinogenesis, early diagnosis and molecular targeted therapy of invasive bladder cancer