Generating molecular markers that selectively label urothelial sub-populations

描述（由申请人提供）：泌尿流出道，包括肾盂、输尿管、膀胱和上尿道，由称为尿道上皮的特化复层上皮内衬，由基底层、中间层和由伞细胞组成的浅层组成。伞细胞是专门用于合成的大型多核细胞 以及尿斑蛋白的运输，尿斑蛋白是一个分泌蛋白家族，它组装成排列在尿道顶面的晶体蛋白斑，在尿液和血液之间形成防漏屏障。泌尿系统可能会因损伤或疾病（包括间质性膀胱炎、炎症和UTI）而功能失调，但目前还没有办法修复/再生该组织。此外，膀胱癌被认为是引起不同形式膀胱癌的干细胞/祖细胞的来源。进展的一个障碍是缺乏区分包括祖细胞在内的尿路上皮亚型的标志物。我们提出从我们已经使用表达RFP或GFP的转基因和谱系标记的小鼠模型定义的那些开始，产生对尿路上皮亚群特异的遗传标志物的集合。除了伞细胞和基底细胞，我们已经能够定义一种新的细胞类型，它位于中间层，可能是伞细胞祖细胞。Potter/Aronow实验室将使用RNA-Seq对流式分选的细胞群进行遗传分析，该技术已成功用于表达肾脏细胞类型。一旦确认候选人在手，我们将绘制他们的分布图， 用原位杂交和免疫组织化学检测LUT。我们还将绘制在GUDMAP 1中鉴定为潜在尿路上皮细胞特异性的现有候选物的分布。 在过去的3年里，我们一直是GUDMAP 1联盟的一员，在此期间，我们成功地生成了以LUT为重点的表达数据，这些数据已被纳入现有的Alas中。我们与GUDMAP合作者建立的关系和LUT中表达分析的经验将成为我们进行拟议研究的优势。

DESCRIPTION (provided by applicant): The urinary outflow tract, including the renal pelvis, ureters, bladder and upper urethra, is lined by a specialized stratified epithelium called the urothelium, composed of a basal layer, an intermediate layer and a superficial layer consisting of umbrella cells. Umbrella cells are large multinucleated cells that are specialized for synthesis and trafficking of Uroplakins, a family of secreted proteins that assemble into the crystallin plaque lining the apical surface ofthe urothelium that generates a leak-proof barrier between the urine and the blood. The urothelium can be rendered dysfunctional by damage, or disease, including interstitial cystitis, inflammation and UTI, but at present there is no way to repair/regenerate this tissue. In addition the urothelium is thought to be a source of stem/progenitor cells that give rise to different forms of bladder cancer. One impediment to progress has been the paucity of markers distinguishing urothelial sub-types including progenitors We propose to generate a collection of genetic markers specific for urothelial sub-populations beginning with those that we have defined using transgenic and lineage-tagged mouse models expressing RFP or GFP. In addition to umbrella cells and basal cells, we have been able to define a novel cell type that resides in the intermediate layer that is a likely umbrella cell progenitor. Flow sorted cell populations will be genetically profiled by the Potter/Aronow labs using RNA-Seq, a technique that they have successfully used to expression profile kidney cell types. Once validated candidates are in hand, we will map their distribution in the LUT using in situ hybridization and immunohistochemistry. We will also map the distribution of existing candidates identified in GUDMAP1 as potentially urothelial-cell specific. We have been part of the GUDMAP1 consortium for the past 3 years, and during this time we have successfully generated LUT-focused expression data which has been incorporated into the existing alas. The relationships that we have developed with GUDMAP collaborators and experience with expression analysis in the LUT will be an advantage as we undertake the proposed studies.