Characterization of neural crest stem cells in human hair follicles.

描述（由申请人提供）：我们的长期目标是研究控制人类神经嵴干细胞（hNCSCs）增殖和分化的分子机制，并将这些细胞用于再生医学。胚胎NCSCs是多能性的，并且在成年后至少在两个分化组织中持续存在，即肠道的肠神经系统和皮肤和毛囊。毛囊是皮肤中NCSCs的生长环境。骨形态发生蛋白（BMPs）在胚胎NCSC的自我更新和分化中起着至关重要的作用。NCSCs在鸡和小鼠系统中已被很好地表征。尽管神经嵴细胞生物学在发育和疾病中的重要性，但人类来源的NCSCs的分离一直难以捉摸。我们假设人类毛囊NCSCs保留了多能性，并具有与胚胎NCSCs相似的功能。我们最近在人类毛囊中发现了一个干细胞群。这些细胞明显不同于上皮细胞或黑素细胞干细胞。它们具有克隆性、自我更新能力和多能性。它们表达NCSC标记物和BMP-4以及BMP受体，但不表达谱系特异性标记物。这些细胞位于人类毛囊的凸起区域。我们建议在体外和体内对膨大的hNCSCs进行表征。目的1：表征人类毛囊中的hNCSCs。我们将进一步表征这些膨出的hNCSCs的多能性，定义它们的遗传特征，并鉴定可用于直接从新鲜组织中分离这些细胞的表面标记物。我们将研究BMP信号通路在鼓包型hNCSCs增殖分化中的作用。目的2：研究hNCSCs在体内的生理功能。我们将验证凸起hNCSCs与胚胎NCSCs具有相似生物学功能的假设。我们将研究凸起型hNCSCs是否遵循微环境神经嵴线索发育成合适的细胞类型及其在体内皮肤和毛囊中的生理功能。目的3：确定hNCSCs向黑素细胞分化的信号。我们将研究hNCSCs谱系确定的机制，特别是hNCSCs向黑素母细胞到成熟黑素细胞的分期分化。在此过程中，我们将研究Wnt3a、干细胞因子和内皮素-3与Mitf的相互作用。我们期望膨出的hNCSCs保留胚胎NCSCs的多能性和功能。它们有助于正常的人体皮肤和毛囊的稳态。本研究将为研究人类体细胞组织中的NCSCs建立一个新的模型。该模型可用于研究hNCSC自我更新和谱系确定所必需的各种信号通路，hNCSC在成人组织中的生理功能，以及这些细胞用于再生治疗的潜在用途。此外，由于许多疾病，包括一些癌症，已知与神经嵴细胞有关，这些研究中开发的模型可能有助于测试这些疾病的病因。

DESCRIPTION (provided by applicant): Our long term objective is to investigate the molecular mechanisms that control the proliferation and differentiation of human neural crest stem cells (hNCSCs) and use these cells for regenerative medicine. Embryonic NCSCs are pluripotent and NCSCs have been shown to persist in the adult in at least two differentiated tissues, the enteric nervous system of the gut and the skin and hair follicles. Hair follicles are a niche for NCSCs in the skin. Bone morphogenic proteins (BMPs) play a critical role in embryonic NCSC self-renewal and differentiation. NCSCs have been well characterized in chick and murine systems. Despite the importance of neural crest cell biology in development and disease, the isolation of NCSCs of human origin has been elusive. We hypothesize that human hair follicle NCSCs retain pluripotency and have similar functions as embryonic NCSCs. We have recently identified a stem cell population in human hair follicles. These cells are distinctively different from epithelial or melanocyte stem cells. They are clonogenic, capable of self-renewal and pluripotent. They express NCSC markers and BMP-4 as well as BMP receptors but not lineage specific markers. These cells are located in the bulge region of human hair follicles. We propose to characterize the bulge hNCSCs in vitro and in vivo. Aim 1: Characterize hNCSCs in human hair follicles. We will further characterize the pluripotent of these bulge hNCSCs, define their genetic signature and identify surface markers that can be used to directly isolate these cells from fresh tissues. We will study the role of BMP signal pathway in the proliferation and differentiation of bulge hNCSCs. Aim 2: Study the physiological function of hNCSCs in vivo. We will test the hypothesis that bulge hNCSCs have a similar biological function as embryonic NCSCs. We will study whether bulge hNCSCs follow microenvironmental neural crest cues to develop into appropriate cell types and their physiological functions in the skin and hair follicles in vivo. Aim 3: Determine the signals for hNCSCs to differentiate to melanocytes. We will study the mechanism underlying lineage determination for hNCSCs, particularly staged differentiation of hNCSCs to melanoblasts to mature melanocytes. We will study the interaction of Wnt3a, stem cell factor and endothelin-3 with Mitf during this process. We expect that bulge hNCSCs retain the pluripotency and function of embryonic NCSCs. They contribute to normal human skin and hair follicle homeostasis. The proposed studies will establish a novel model to study human NCSCs in the somatic tissues. This model can be used to study various signalling pathways that are essential for hNCSC self-renewal and lineage determination, the physiological functions of hNCSCs in adult tissue, and the potential use of these cells for regenerative therapy. Also, because many diseases, including some cancers, are known to involve neural crest cells, the models developed in these studies may be useful in testing the etiologies of these diseases. PUBLIC HEALTH RELEVANCE. Stem cell research holds tremendous promise for regenerative medicine. The goal of this application is to reveal the molecular mechanisms underlying the proliferation and differentiation of human neural crest stem cells in the hair follicles. The proposed studies may lead to potential use of these cells in regenerative therapy and establish a novel model to study the etiology of neural crest cells related human diseases.