目的 筛选出在膀胱癌组织中特异性表达的长链非编码RNA(lncRNA)及其共表达mRNA,并进行初步验证及功能预测.方法 采用lncRNA v4.0芯片筛选4对膀胱癌和癌旁组织中lncRNA及其共表达mRNA的差异表达谱,通过聚类分析比较二者表达差异;采用实时荧光定量PCR(qRT-PCR)法对5个异常调节的lncRNA(RP11-359E19.2、AL928768.3、AC002519.6、RP11-79H23.3、AK021804)和4个共表达的mRNA(HRAS、VEGFA、ITGB1、DNMT3B)进行验证.同时,对lncRNA共表达的mRNA进行GO及KEGG pathway富集分析.结果 差异表达的lncRNA共4 155条,其中2 045条高表达、2 110条低表达;与lncRNA共表达的mRNA共4 416条,其中2 472条高表达、1 944条低表达.|FC|≥10的lncRNA 345条,包括127条高表达和218条低表达.RP11-436F21.1和H19是上调最明显的lncRNA.|FC|≥10的共表达mRNA有75条,其中57条上调、18条下调.与癌旁组织相比,膀胱癌组织5种lncRNA中RP11-359E19.2表达上调,AL928768.3、AC002519.6、RP11-79H23.3及AK021804表达下调(P均<0.05);4种共表达的mRNA中HRAS、VEGFA、ITGB1和DNMT3B表达均上调(P均<0.05);qRT-PCR结果与芯片结果一致.GO分析显示,上调的共表达mRNA主要参与细胞代谢和有丝分裂的生物学过程,下调的共表达mRNAs主要参与免疫系统的刺激反应和免疫应答;KEGG pathway富集分析显示,10个通路与上调或下调的共表达mRNA有关,其中在上调的共表达mRNA中,p53信号通路和膀胱癌的富集度最高,在下调的共表达mRNA中细胞因子-因子受体相互作用富集度最高.结论 成功筛选出膀胱癌特异表达的lncRNA及其共表达mRNA,这些特异表达的lncRNA及共表达mRNA在膀胱癌的发生、发展和转移中发挥重要的生物学作用,有望成为膀胱癌新的标志物.%Objective To screen out differentially expressed long non-coding RNAs (lncRNAs) and co-expression mRNAs and to perform the preliminary validation and functional prediction in bladder carcinoma.Methods Four pairs of carcinoma and para-carcinoma tissues were used for microarray assay to screen out the differentially expressed lncRNAs and co-expression mRNAs.Hierarchical clustering was performed to show lncRNA and mRNA expression patterns among samples.Five lncRNAs (RP11-359E19.2, AL928768.3, AC002519.6, RP11-79H23.3, AK021804) and four co-expression mRNAs (HRAS, VEGFA, ITGB1, DNMT3B) were chosen for verification of the microarray results by quantitative real-time PCR.Gene Ontology (GO) and KEGG pathway were used to analyze the biological function of co-expression mRNA.Results A total of 4 155 lncRNAs and 4 416 co-expression mRNAs were detected to be differentially expressed, among which, 2 045 lncRNAs were up-regulated and and 2 110 were down-regulated, meanwhile, 2 472 co-expression mRNAs were up-regulated and and 1944 were down-regulated, respectively.Totally 345 lncRNAs displayed fold change ≥10, including 127 up-regulated lncRNAs and 218 down-regulated lncRNAs.RP11-436F21.1 and H19 were the most up-regulated lncRNAs.Overview of coding gene profile showed that 75 mRNAs had fold change ≥10 (up: 57;down: 18).Comapred with the para-carcinoma tissues, the expression of RP11-359E19.2 in 5 lncRNAs was up-regulated, whereas AL928768.3, AC002519.6, RP11-79H23.3, and AK021804 expression was down-regulated in the bladder carcinoma tissues (all P<0.05).Meanwhile, the expression of HRAS, VEGFA, ITGB1, and DNMT3B in four co-expression mRNAs was all up-regulated (all P<0.05).The result was consistent with the microarray assay.GO assay showed that the up-regulated co-expression mRNAs mainly participate in the biological processes, such as cellular metabolic and mitotic cell cycle.Meanwhile, the down-regulated co-expression mRNAs mainly participate in the immune system process of stimulus response and immune response.KEGG pathway assay showed that 10 pathways were associated with the up-regulated or down-regulated mRNAs.Among the co-expression mRNAs, the enrichment of P53 signal pathway and bladder cancer was the highest, whereas in the down-regulated co-expression mRNAs, the cytokine-factor receptor interaction was the highest.Conclusions The differentially expressed LncRNAs and co-expression mRNAs are found and screened out in the bladder cancer.The specially expressed lncRNAs and and mRNAs could play important roles in the pathogenesis and development of bladder carcinoma, which might be new markers of bladder cancer.
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