Clinical Laboratory, The First Affiliated Hospital, Wenzhou Medical College, Wenzhou, China.
Urol Oncol. 2013 Feb;31(2):219-27. doi: 10.1016/j.urolonc.2010.11.007. Epub 2011 Jul 27.
The goal of this study was to extend the known repertoire of microRNAs (miRNAs) expressed in urothelial bladder cancer (BCa) of the Chinese population and further understand the molecular events of miRNAs underlying urothelial bladder tumorigenesis at the global genome level.
We separated well-characterized epithelial tumor cells from 20 moderately differentiated or poorly differentiated BCa specimens by laser capture microdissection (LCM) and pooled these cells of interest prior to RNA analysis. Ten normal bladder epithelia (NBE) samples were pooled as the control. After preparation of small RNAs library, the 2 samples were sequenced simultaneously by the next generation high through-put Solexa sequencing technology.
We employed the next generation high through-put Solexa sequencing technology to clone and identify miRNAs in BCa and NBE, and generated 11,146,610, and 10,263,845 high quality sequence reads, respectively. According to the analysis of size distribution, 22 nt class was the most abundant group of small RNAs in the BCa. Likewise, the 20 and 22 nt sequences were significantly greater than shorter or longer sequences, and accounted for 59.55% of the total sequence number of NBE library. The whole-genome-scale data mining suggested that BCa and NBE libraries both contained multiple and heterogeneous small RNA species. On further analysis, the sequencing data revealed that different miRNAs showed clearly in-house differential expression levels in BCa and NBE and 74 miRNAs aberrantly expressed between BCa and NBE at the global genome level. We also predicted 13 novel miRNAs in both BCa and NBE libraries.
Our results suggest that BCa miRNAs include a large proportion of conserved miRNAs and a set of non-conserved miRNAs with low expression levels. These known and newly identified miRNAs at the population level significantly enhance our knowledge of BCa miRNAs expression profiling and provide insights into miRNAs oncogenesis and oncotherapy in BCa. Further studies are necessary to elucidate the roles of miRNAs in urothelial bladder tumorigenesis and determine the potential of miRNAs as diagnostic and prognostic tools or therapeutic targets for BCa in the future.
本研究旨在扩展中国人群尿路上皮膀胱癌（BCa）中表达的 microRNAs（miRNAs）的已知谱，并进一步从全基因组水平了解 miRNA 在尿路上皮膀胱癌发生中的分子事件。
我们通过激光捕获显微切割（LCM）从 20 例中-低分化 BCa 标本中分离出特征明确的上皮肿瘤细胞，并在进行 RNA 分析之前将这些感兴趣的细胞进行汇集。10 例正常膀胱上皮（NBE）样本被汇集作为对照。在制备小 RNA 文库后，我们采用新一代高通量 Solexa 测序技术对这 2 个样本进行了同时测序。
我们采用新一代高通量 Solexa 测序技术克隆和鉴定了 BCa 和 NBE 中的 miRNAs，并分别生成了 11,146,610 和 10,263,845 条高质量序列读段。根据大小分布分析，22nt 类是 BCa 中小 RNA 最丰富的一组。同样，20nt 和 22nt 序列明显大于较短或较长的序列，占 NBE 文库总序列数的 59.55%。全基因组规模的数据挖掘表明，BCa 和 NBE 文库均包含多种异质性小 RNA 种类。进一步分析显示，不同的 miRNAs 在 BCa 和 NBE 中表现出明显的差异表达水平，并且在全基因组水平上有 74 个 miRNAs 在 BCa 和 NBE 之间存在异常表达。我们还预测了这两个文库中的 13 个新的 miRNAs。
我们的结果表明，BCa miRNAs 包括大量保守的 miRNAs 和一组低表达水平的非保守 miRNAs。这些在人群水平上已知和新鉴定的 miRNAs 显著增强了我们对 BCa miRNAs 表达谱的认识，并为 miRNA 在 BCa 中的致癌作用和肿瘤治疗提供了新的见解。进一步的研究有必要阐明 miRNAs 在尿路上皮膀胱癌发生中的作用，并确定 miRNAs 在未来作为 BCa 的诊断和预后工具或治疗靶点的潜力。
