Skin Stem Cells: Purification and Characterization

项目摘要 这项研究的总体目标是阐明组织稳态的机制， 研究哺乳动物皮肤的再生，并了解这一过程在人类疾病中是如何出错的， 癌的实现这一目标的核心是表征皮肤内的不同干细胞（SC）， 确定它们对组织稳态和伤口修复的相对贡献，并阐明它们如何改变 利基微环境影响这些事件。过去的AR 050452研究导致毛囊的纯化 (HF)隆突和基底滤泡间表皮（Epd）细胞，并建立他们作为长寿，自我更新 在组织再生和伤口修复中起作用的SC。然而，在它们的生物学和组织中， 在再生任务中，这些SC显示出不同的行为，这些行为似乎是基于它们独特的 微环境（niches）。该领域仍然缺乏对这些小生境的组成的全面了解， SC的性质：利基相互作用，以及它们如何帮助干细胞科普压力情况。过去的AR 050452 研究为解决以下关键问题奠定了基础：（1）。什么是利基组件 以及它们如何随着毛发周期而变化？(2)Epd-和HF-的相对贡献是什么？ 创伤修复的治疗方法？这是否与浅表和深部伤口不同？在年轻的老鼠和年老的老鼠之间？(3)有哪些 SC-生态位串扰的变化，发生在响应损伤和老化，什么是他们的生理 相关性？(4)供应链如何应对和协调修复其生态位屏障中的地方缺口？这 细胞将损伤信号传递给周围的免疫细胞，它们是如何被触发这样做的？怎么 SC存活？(5)在毛发周期结束时，当大部分HF经历了 凋亡和终末分化？为了回答这些问题，我们将使用FACS，单细胞RNA测序，ChIP- seq、条件性基因敲除和RNAi体内筛选，并利用这些方法探索皮肤干细胞 在它们的原生、突变和创伤诱导的环境中。

Project Summary The global objective of this research is to elucidate the mechanisms underlying tissue homeostasis and regeneration in mammalian skin and to understand how this process goes awry in human disorders, including cancers. Central to achieving this goal is the characterization of the different stem cells (SCs) within skin, determining their relative contributions to tissue homeostasis and wound-repair, and elucidating how changes in niche microenvironments impact these events. Past AR050452 research led to purification of hair follicle (HF) bulge and basal inter-follicular epidermal (Epd) cells, and established them as long-lived, self-renewing SCs that function in tissue regeneration and wound-repair. However, both in their biology and their tissue regenerative tasks, these SCs display distinct behaviors that appear to be predicated on their unique microenvironments (niches). The field still lacks a comprehensive knowledge of the constituents of these niches, the nature of SC:niche interactions, and how they help stem cells cope with stressful situations. Past AR050452 research has set the foundations to tackle the following key questions: (1). What are the niche components of HFSCs and how do they change with the hair cycle? (2) What are the relative contributions of Epd- and HF- SCs to wound repair? Does this differ in superficial vs deep wounds? In young vs aging mice? (3) What are the changes in SC-niche crosstalk that occur in response to injury and aging and what is their physiological relevance? (4) How do SCs respond to and orchestrate the repair of local breaches in their niche barrier? Which cells transmit damage signals to the surrounding immune cells and how are they triggered to do so? How do SCs survive? (5) How do SCs protect themselves at the end of the hair cycle, when the bulk of the HF undergoes apoptosis and terminal differentiation? To answer these questions, we’ll use FACS, single cell RNA-seq, ChIP- seq, conditional gene knockout and RNAi screens in vivo and employ these methods to explore skin stem cells in their native, mutant and wound-induced environments.