Digital gene expression (DGE) sequencing of 10 pairs samples between kidney normal tissue and cancer tissue. Homo sapiens

we combined their genome-wide profiles of tumors and normal adjacent tissues in 10 ccRCC patients. The results showed that 283 miRNAs were down-regulated and 187 up-regulated, meanwhile 7473 mRNAs were up-regulated and 3439 down-regulated in ccRCC. Expressions of 12 miRNAs and genes were validated in 10 patients we studied by RT-qPCR (validation rate from 60% to 100%). Differentially expressed gene analysis showed the collective change of miR-200 and SLC22A gene family, and pathway analysis revealed down-regulation of multiple metabolic pathways and up-regulation of focal adhesion, ECM-receptor interaction, etc. Moreover, A miRNA cluster located in chromosome Xq27.3 were significantly down-regulated in ccRCC, which were verified in 53 ccRCC patients by RT-qPCR (validation rate from 63.8% to 88.6%).56 and 586 potentially novel miRNAs and mRNA we detected according to statistical analyses. In addition, 14 miRNA candidates were randomly selected to validate using qRT-PCR and Sanger sequcencing technology, 10 of out 14 could be successfully amplified,.Plus, the sequences of all 5 randomly selected amplified products were confirmed by cloned Sanger sequencing.Our data demonstrated a combined profiling of miRNAs and mRNAs in ccRCC, which showed the decreased metabolism and the perturbation of renal cell function. Moreover, this study identified the expression alteration of a miRNA cluster on Xq27.3, which suggested its potential function in carcinogenesis of ccRCC Overall design: Examination of 10 pairs of kidney normal cells and cancer cells .

本研究整合了10例肾透明细胞癌（clear cell renal cell carcinoma, ccRCC）患者的肿瘤组织与癌旁正常组织的全基因组表达谱。结果显示，在ccRCC中，共有283个微小RNA（microRNA, miRNA）表达下调、187个miRNA表达上调；同时有7473个信使RNA（messenger RNA, mRNA）表达上调、3439个mRNA表达下调。本研究通过实时定量聚合酶链反应（real-time quantitative polymerase chain reaction, RT-qPCR）对纳入的10例患者的12个miRNA及基因的表达水平进行了验证，验证率介于60%至100%之间。差异表达基因（differentially expressed gene, DEG）分析显示，miR-200家族与SLC22A基因家族存在集体表达变化；通路（pathway）分析则揭示，多条代谢通路呈现下调趋势，而黏着斑（focal adhesion）、细胞外基质-受体相互作用（ECM-receptor interaction）等通路则呈现上调趋势。此外，位于X染色体q27.3区域的一个微小RNA簇在ccRCC中显著下调，该结果在53例ccRCC患者中通过RT-qPCR得到验证，验证率介于63.8%至88.6%之间。本研究通过统计学分析共检测到56个潜在新型miRNA与586个潜在新型mRNA。此外，本研究随机选取14个候选miRNA，采用qRT-PCR与桑格测序（Sanger sequencing）技术进行验证，其中10个可成功扩增；进一步对随机选取的5个扩增产物的序列进行克隆化桑格测序，均得到确认。本研究的数据证实了ccRCC中miRNA与mRNA的联合表达谱特征，揭示了代谢水平降低与肾细胞功能紊乱的现象。此外，本研究明确了Xq27.3区域miRNA簇的表达异常，提示其在ccRCC癌变过程中可能发挥潜在功能。整体实验设计：对10对肾正常细胞与癌细胞进行检测。