Career Enchancement Award for Stem Cell Research in Urology

描述（由申请人提供）： NIDDK的使命包括泌尿和消化系统疾病的研究，以改善美国人的健康。最近的研究表明，膀胱和胃等器官可能含有能够再生受损组织的干细胞。该奖项将培训博士莫妮卡利伯特，在尿路上皮（膀胱）生物学，在干细胞技术的高级研究员。Liebert将由乳腺癌干细胞生物学研究领导者Max Wicha和胃肠器官发生专家Deborah Gumucio共同指导。Wicha将在基本的干细胞分离、表征和培养技术方面对Liebert进行培训。然后，Liebert将应用这种训练来研究胃干细胞。Gumucio先前发现，由绒毛蛋白启动子驱动的转基因在位于胃干细胞所在组织中的胃细胞亚群中高水平表达。我们的假设是，这些细胞是一个更大的胃干细胞群的一个子集。目的1：分离和鉴定胃干细胞。将使用磁性或流式细胞术分离从组织中富集具有干细胞特征的细胞。这些细胞将被培养以形成不依赖于锚定的"胃球"，并评估干细胞特性和基因表达。采用抗体染色、基因芯片和实时荧光定量PCR技术对这些细胞进行检测，并与villin-promoter驱动的EGFP表达鉴定的细胞进行比较。目标2将测试这些干细胞样细胞是否可以在体外或体内产生肠或胃谱系。细胞将在分化条件下培养。将来自Rosa-26小鼠和Villin-EGFP x Rosa 26杂交的胃干细胞的富集群体以及未分选细胞的对照群体分离并移植到裸鼠上。将评价异种移植物的EGFP、β-gal表达和肠或胃组织的形态发育。这些研究将证实不同的潜在干细胞群的繁殖和分化能力。这项研究将帮助我们了解胃如何对溃疡等疾病作出反应，以及胃癌如何发展。在Liebert的培训结束时，她将把这些技术应用于膀胱发育，组织工程和膀胱癌的研究。她未来在泌尿科的工作将开发新的治疗尿路疾病和膀胱癌。

DESCRIPTION (provided by applicant): NIDDK's mission includes research in urologic and digestive diseases to improve the health of Americans. Recent studies indicate that organs, such as the urinary bladder and stomach, may contain stem cells capable of regenerating damaged tissue. This award will train Dr. Monica Liebert, a senior investigator in urothelial (urinary bladder) biology, in stem cell techniques. Liebert will be co-mentored by Max Wicha, leader in breast cancer stem cell biology research, and Deborah Gumucio, a expert in gastrointestinal organogenesis. Wicha will train Liebert in basic stem cell isolation, characterization and culture techniques. Liebert will then apply that training to study gastric stem cells. Gumucio previously found that transgenes driven by the villin promoter are expressed at high levels in a subset of gastric cells located in tissue where gastric stem cells are thought to reside. Our hypothesis is that these cells are a subset of a larger gastric stem cell population. Aim 1 is to isolate and characterize stem-like gastric stem cells. Cells with stem cell characteristics will be enriched from tissues using magnetic or flow cytometric separation. These cells will be cultured to form anchorage-independent "gastrospheres," and evaluated for stem cell properties and gene expression. Antibody staining, microarray methods and real-time quantitative PCR will be used to examine the these cells and compare them to the cells identified by villin-promoter driven EGFP expression. Aim 2 will test whether these stem-like cells can give rise to intestinal or gastric lineages in vitro or in vivo. Cells will be cultured under differentiating conditions. Enriched populations of gastric stem cells from Rosa-26 mice and the Villin-EGFP x Rosa 26 cross, as well as a control population of unsorted cells, will be isolated and grafted on nude mice. Xenografts will be evaluated for EGFP, beta-gal expression and for morphologic development into intestinal or gastric tissues. These studies will confirm the ability of the different potential stem cell populations to reproduce and to differentiate. This research will help us understand how the stomach responds to diseases, such as ulcers, and how stomach cancer may develop. At the end of Liebert's training, she will apply these techniques to the study of urinary bladder development, tissue engineering, and bladder cancer. Her future work in urology will develop new treatments for urinary tract disorders and bladder cancer.