Characterization of Epidermal Stem Cells

表皮干细胞的表征

• 批准号: 6973188
• 负责人: RUBY GHADIALLY
• 金额: $ 36.3万
• 依托单位: NORTHERN CALIFORNIA INSTITUTE/RES/EDU
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-09-01 至 2010-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6973188
• 关键词: biotechnology; cell differentiation; cell population study; cell proliferation; cell type; embryonic stem cell; epithelium; flow cytometry; gene therapy; genetic transcription; green fluorescent proteins; hair follicle; keratinocyte; laboratory mouse; microarray technology; polymerase chain reaction; sebaceous glands; skin; skin neoplasms; stem cell transplantation; therapy design /development; tissue /cell culture; wound healing

DESCRIPTION (provided by applicant): The long-term objective of this project is to obtain epidermal stem cells in a relatively pure population of known stem cell content in order to provide improved therapy for healing of wounds, for stem cell-targeted gene therapy, and to better understand epidermal differentiation and epidermal carcinogenesis. In Aim 1, we will first characterize the number, multipotency, and relationship of follicular vs. interfollicular epidermal stem cells. Specifically, we will determine the number of follicular vs. interfollicular stem cells, using limiting dilution analysis in an in vivo competitive repopulation assay. The multipotency of follicular vs. interfollicular stem cells will then be determined by using isolated cell populations (follicular vs. interfollicular) and determining the ability of a GFP marked stem cell and its progeny to produce interfollicular epidermis in a primary transplantation chamber, followed by a follicle or part thereof in a subsequent secondary transplantation chamber. In Aim 2 further studies using both established enrichment methods, as well as new combinations of selection strategies, will be used to determine the most effective epidermal stem cell selection strategy for long-term repopulating cells (stem cells) of the epidermis. Each potential individual isolation technique will be tested by performing a limiting dilution study of the enriched population of kertinocytes. We will then determine the best combination of positive and negative markers to effectively isolate epidermal stem cells. This population of highly enriched stem cells will be used to determine epidermal stem cell-specific gene transcription using microarray analysis. Finally, in Aim 3 we will expand an epidermal stem cell population using over-expression of HOX B4 or sonic hedghog, two proteins associated with increased stem cell renewal in other organs.These studies will help clarify the relationship between follicular and interfollicular stem cells and their degree of multipotency, as well as move closer to isolating true long-term repopulating cells of the epidermis. Enriched populations of epidermal stem cells will aid stem cell-directed gene therapy, and when expanded will serve as a potential wound/burn healing therapy.

描述（由申请人提供）：该项目的长期目标是在已知干细胞含量的相对纯净的群体中获得表皮干细胞，以便为伤口愈合、干细胞靶向基因治疗提供改进的治疗方法，并更好地了解表皮分化和表皮癌变。在目标 1 中，我们将首先描述毛囊与毛囊间表皮干细胞的数量、多能性和关系。具体来说，我们将在体内竞争性再增殖测定中使用有限稀释分析来确定滤泡与滤泡间干细胞的数量。然后，通过使用分离的细胞群（滤泡与滤泡间）并确定 GFP 标记的干细胞及其后代在初级移植室中产生滤泡间表皮，然后在随后的次级移植室中产生滤泡或其部分的能力，来确定滤泡与滤泡间干细胞的多能性。在目标 2 中，使用已建立的富集方法以及新的选择策略组合的进一步研究将用于确定表皮干细胞长期再生的最有效的表皮干细胞选择策略。将通过对角质细胞富集群进行有限稀释研究来测试每种潜在的单独分离技术。然后我们将确定阳性和阴性标记的最佳组合，以有效分离表皮干细胞。该高度富集的干细胞群将用于通过微阵列分析确定表皮干细胞特异性基因转录。最后，在目标 3 中，我们将利用 HOX B4 或 sonic hedghog 的过度表达来扩大表皮干细胞群，这两种蛋白质与其他器官中干细胞更新的增加有关。这些研究将有助于阐明滤泡和滤泡间干细胞之间的关系及其多能性程度，并更接近于分离真正的表皮长期再生细胞。丰富的表皮干细胞群将有助于干细胞导向的基因治疗，并且当扩大时将作为潜在的伤口/烧伤愈合疗法。