原发性肝癌肝肾阴虚证患者外周血单个核细胞的特征性差异基因  

Characteristic genomics of peripheral blood mononuclear cells of hepatocellular carcinoma patients with liver-kidney yin deficiency syndrome

作　　者：翁莉[1] 杜鹃[1] 何文婷[1] 凌昌全[1] 

机构地区：[1]第二军医大学长海医院中医科,上海200433

出　　处：《中西医结合学报》2012年第4期406-415,共10页Journal of Chinese Integrative Medicine

基　　金：国家自然科学基金重点项目(No.30730114)

摘　　要：目的:探讨原发性肝癌肝肾阴虚证患者外周血单个核细胞的特征性差异基因。方法:选取肝肾阴虚证与非肝肾阴虚证肝癌患者作为研究对象,以肝肾阴虚证肝癌患者为试验组,以非肝肾阴虚证肝癌患者作为对照组;选用全基因组表达谱芯片,研究肝肾阴虚证的基因表达谱,并进行差异表达基因分析,基于Gene Ontology(http://www.geneontology.org/)和KEGG(http://www.genome.jp/kegg/)数据库对差异基因进行功能(gene ontology,GO)和通路(pathway)注释,并选取显著性差异基因构建共表达网络图。扩大样本量,运用实时荧光定量聚合酶链反应与蛋白质印迹法分别从mRNA及蛋白层面对显著性差异基因进行验证。结果:肝肾阴虚证组和非肝肾阴虚证组间存在差异基因表达特征图谱,获得差异表达的基因共615个。GO注释后,差异基因表达具有278个显著性功能差异,主要涉及跨膜转运、细胞周期停滞、细胞转录、诱导凋亡等。Pathway标记后,上调差异基因参与显著性信号转导通路16项,下调差异基因参与显著性信号转导通路10项,包括抗原加工与递呈转导通路、代谢途径转导通路、细胞周期转导通路、蛋白质输出转导通路等。用差异基因构建共表达网络图,筛选出3个显著性差异基因(SEC62[SEC62 homolog(S.cerevisiae)]、CCNBl[cyclin B1]、BIRC3[baculoviral IAPrepeat-containing 3])。扩大样本量进行验证,肝肾阴虚证肝癌患者SEC62、CCNB1、BIRC3基因的mRNA(P<0.01)和蛋白(P<0.01)表达水平均显著低于非肝肾阴虚证肝癌患者。结论:本研究从基因组学的角度反映出肝肾阴虚证肝癌患者与非肝肾阴虚证肝癌患者之间存在着差异表达基因,进一步证实了"证"是多基因在mRNA和(或)蛋白质水平发生改变并导致人体偏离正常状态的假说。OBJECTIVE： To explore the characteristic genomics of syndrome of liver-kidney yin deficiency in peripheral blood mononuclear cells of hepatocellular carcinoma （HCC） patients. METHODS： HCC patients with or without syndrome of liver-kidney yin deficiency were enrolled into the experimental group and the control group, respectively; their gene expression profiles were evaluated by a whole-genome Affymetrix GeneChip Human Genome U133 Plus 2.0 Array. The differentially expressed mRNAs were then selected by Gene ~ntology （GO） and pathway analyses, respectively. Based on the results of GO and pathway analyses, gene coexpression networks were built according to the normalized signal intensity of specifically expressed genes. Finally, the results from microarray were confirmed by real-time fluorescence quantitative polymerase chain reaction and Western blot methods. RESULTS： The results showed that a set of 615 mRNAs were differentially expressed in the HCC patients with syndrome of liver-kidney yin deficiency. By GO enrichment analysis, the genes for anti-apoptosis, regulation of cell cycle, transmembrane transport, etc. were up-regulated or down-regulated in the experimental group. Another functional analysis of mRNAs by KEGG revealed that 10 signal transduction pathways were up-regulated and ]_6 were down-regulated, such as antigen processing and presentation, cell cycle, and protein export. Based on the above results, we constructed coexpression networks to determine which genes may play pivotal role in HCC patients with syndrome of liver-kidney yin deficiency. Some critical genes, including SEC62 （SEC62 homolog （S. cerevisiae））, CCNB1 （cyclin B1） and BIRC3 （baculoviral lAP repeat- containing 3）, which rank the top 3 in ｜△ normalized degree｜ were chosen. Of another 60 samples, we found that the mRNA expressions of SEC62, CCNB1 and BIRC3 were significantly lower in HCC patients with syndrome of liver-kidney yin deficiency than those without syndrome of liver-kidney yin deficiency （

关 键 词：肝肿瘤 肝肾阴虚 基因表达 基因组学 

分 类 号：R735.7[医药卫生—肿瘤] R273[医药卫生—临床医学]