Molecular & cellular biology of hair follicle stem cells

分子

• 批准号: 6473343
• 负责人: STEPHEN R LYLE
• 金额: $ 11.95万
• 依托单位: BETH ISRAEL DEACONESS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-06-01 至 2007-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6473343
• 关键词: athymic mouse; cell differentiation; cell migration; clinical research; confocal scanning microscopy; gene expression; guanine nucleotide binding protein; hair follicle; human genetic material tag; human tissue; immunocytochemistry; immunoprecipitation; in situ hybridization; keratin; microarray technology; molecular biology; nuclear receptors; phenotype; polymerase chain reaction; protein kinase; protein reconstitution; receptor expression; stem cells; tissue /cell culture; video microscopy; western blottings

DESCRIPTION (provided by applicant): The overall goal of this project is to characterize the expression of genes within hair follicle stem cells and to study the role of specific genes in maintaining the stem cell phenotype. The hair follicle is a model for studying epithelial stem cells and understanding the mechanisms which maintain the homeostasis of hair follicle stem cells and may lead to advances in the areas of tumorigenesis, wound healing and hair disorders. Similar to other self renewing tissues, such as bone marrow, gastrointestinal tract and epidermis, the hair follicle contains slowly cycling stem cells with a high proliferative capacity and the ability to produce differentiated cells to regenerate the tissue. Unlike other tissues, however, hair follicle growth (anagen) is interrupted by periods of regression (catagen) and rest (telogen), with subsequent regeneration. Since the lower follicle degenerates in the catagen phase, leaving only the telogen club structure, this residual epithelial cap surrounding the club hair represents an enriched source of stem cells which are responsible for regenerating a new lower follicle in anagen. They have extracted RNA from the stem cell rich area of telogen follicles as well as from the bulb area of anagen follicles dissected from human scalp. Through cDNA expression arrays, a number of genes appear to be expressed in the stem cell area that are not expressed in matrix keratinocytes of the anagen bulb. Several of the genes have been shown to mediate proliferative behavior and differentiation pathways in other experimental systems. The hypothesis that will be tested in this proposal is that specific genes expressed in the stem cells of the hair follicle are important in maintaining stem cell properties. The specific aims are: 1. To characterize the expression of specific genes identified by the expression array through in situ hybridization, immunohistochemistry and immunofluorescence with confocal microscopy. Three genes which likely have roles in maintaining stem cell properties will be examined. 2. To test the significance of these three genes in maintaining in vitro properties of hair follicle stem cells through retroviral transfection of human hair follicle stem cells with wild type and mutant gene constructs. Clonogenicity assays and other in vitro experiments of native and transfected stem cells will be used to study the role of these genes in clonality and other stem cell attributes. 3. To test the function of the specific genes during hair follicle development and cycling through in vitro and in vivo methods. Hair reconstitution assays performed with native and genetically altered stem cells will be used to analyze potential functions in folliculogenesis and cycling. In addition to the established mouse hair reconstitution system, they will attempt to develop a human hair follicle reconstitution system from hair follicle stem cells and microdissected dermal papillae using acellular dermal analogues with transplantation onto athymic mice.

描述（由申请人提供）：本项目的总体目标是 表征毛囊干细胞内基因的表达， 研究特定基因在维持干细胞表型中的作用。的 毛囊是研究上皮干细胞和理解 维持毛囊干细胞稳态的机制， 可能导致肿瘤发生、伤口愈合和毛发等领域的进展 紊乱与其他自我更新组织类似，如骨髓， 胃肠道和表皮，毛囊含有缓慢循环 干细胞具有高增殖能力， 分化的细胞来再生组织。然而，与其他组织不同， 毛囊生长（生长期）被退化期（退化期）打断 和休息（休止期），随后再生。因为下卵泡 在退行期阶段退化，只留下休止期俱乐部结构， 残留的上皮帽围绕着梅花毛， 负责再生新的下部卵泡的干细胞 生长期。他们从静止期富含干细胞的区域提取了RNA 毛囊以及从解剖的生长期毛囊的球部区域 人类头皮通过cDNA表达阵列，一些基因似乎是 在基质角质形成细胞中不表达的干细胞区域中表达 生长期灯泡。有几个基因已经被证明可以介导 增殖行为和分化途径在其他实验 系统. 在这项提议中将要检验的假设是，特定的基因 在毛囊干细胞中表达的蛋白在维持 干细胞特性具体目标是：1.描述表达式的特征 表达阵列通过原位杂交鉴定特定基因 用共聚焦显微镜进行杂交、免疫组织化学和免疫荧光 显微镜三个基因可能在维持干细胞中起作用 财产将被审查。2.为了检验这三个基因的重要性 在维持毛囊干细胞的体外特性方面， 逆转录病毒转染人毛囊干细胞的野生型和 突变基因构建体。克隆形成测定和其他体外实验 天然和转染的干细胞将被用来研究这些基因的作用 克隆性和其他干细胞属性。3.要测试 毛囊发育和体外循环过程中的特定基因 和体内方法。用天然的和天然的毛发进行毛发重建测定， 基因改变的干细胞将用于分析潜在的功能， 卵泡发生和周期。除了已经确定的老鼠毛发 重建系统，他们将尝试开发人类毛囊 毛囊干细胞和显微真皮的重建系统 使用脱细胞真皮类似物移植到无胸腺的乳头 小鼠