Regenerative Airway Epithelium by Embryonic Stem Cells

胚胎干细胞再生气道上皮

• 批准号: 7342093
• 负责人: Reen Wu
• 金额: $ 19万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-02-01 至 2010-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7342093
• 关键词: Adult; Animals; Area; Asthma; C57BL/6 Mouse; Cell Lineage; Cell Therapy; Cell Transplants; Cells; Cellular Morphology; Chronic Obstructive Airway Disease; Condition; Development; Developmental Biology; Differentiation Antigens; Disease; ES Cell Line; Embryo; Epithelial Cells; Fibrinogen; Goals; Growth Factor; Hormonal; Hormones; Human; Implant; In Vitro; Inner Cell Mass; Mus; Naphthalene; Naphthalenes; Natural regeneration; Nude Mice; Pattern; Process; Protocols documentation; Rattus; Regenerative Medicine; Regulation; Research; Respiratory System; Risk; Serum; Staging; Stem cells; System; Teratoma; Testing; Therapeutic; Time; Tissues; Trachea; Training; Transplantation; United States National Institutes of Health; airway epithelium; base; blastocyst; caN protocol; cell growth; cell type; embryonic stem cell; genetic manipulation; growth hormone regulating factor; human embryonic stem cell line; in vitro Model; in vivo; innovation; mouse model; repaired; stem; toxicant

DESCRIPTION (provided by applicant): A major area of research in regenerative medicine is the potential application of cell therapy for various disorders. Embryonic stem (ES) cells are self-renewable and pluripotent cells derived from the inner cell mass of a blastocyst-stage embryo. ES cells have the potential to provide innovative therapeutic options for a wide variety of degenerative and non-degenerative human disorders, including the replacement of dysfunctional airway epithelia. To achieve this goal, it would require the development of well defined and efficient protocols for directing the commitment and differentiation of stem cells into the airway epithelial cell lineage, together with their selective purification and proliferation in vitro. The development of such protocols would reduce the likelihood of spontaneous differentiation of ES cells into divergent lineages upon transplantation, as well as reduce the risk of teratoma formation. Additional, such protocols could provide useful in vitro models for studying the developmental biology, of respiratory tract epithelial cells, as well as facilitate the genetic manipulation of stem cells for therapeutic application. The hypothesis of this application is that ES cells can be in vitro trained to differentiate to the airway epithelial cell lineages that have the full potential to regenerate airway epithelium in vivo. To test this hypothesis, three approaches are undertaken. 1) Define "minimally optimized serum-free culture conditions" that will direct both mouse (D3 and Bruce4) and human (H9) ES cells to differentiate to the airway epithelial cell lineages based on differentiation marker expression that are specific for airway epithelial cells, cell growth, and cell morphology using minus/plus one factor approach at various culture time points. 2) Assess morphologically and immunohistochemically the potential of the airway epithelial cell lineages derived from mouse (D3 and Bruce4) and human (H9) ES cells under the "minimally optimized serum-free culture conditions" to regenerate airway epithelium through the repopulation of denuded rat tracheal graft implanted in SCID/nude mice. 3) Characterize the patterns of localization/differentiation of in vivo transplanted cell lineages derived from GFP-tagged Bruce4 ES cells under the "minimally optimized serum-free culture conditions" during the repair process of the airway epithelium in C57BL/6 mice that have been previously treated with an airway epithelium-specific toxicant, naphthalene.

描述（由申请人提供）：再生医学研究的一个主要领域是细胞疗法在各种疾病中的潜在应用。胚胎干细胞（Embryonic stem cell，ES细胞）是从囊胚期胚胎的内细胞团衍生的可自我更新的多能细胞。ES细胞有潜力为多种退行性和非退行性人类疾病提供创新的治疗选择，包括功能障碍的气道上皮细胞的替代。为了实现这一目标，需要开发明确定义的和有效的方案，用于指导干细胞向气道上皮细胞谱系的定型和分化，以及它们的体外选择性纯化和增殖。这种方案的发展将降低ES细胞在移植时自发分化成不同谱系的可能性，以及降低畸胎瘤形成的风险。另外，这样的方案可以提供用于研究呼吸道上皮细胞的发育生物学的有用的体外模型，以及促进用于治疗应用的干细胞的遗传操作。本申请的假设是ES细胞可以在体外训练以分化成气道上皮细胞谱系，其具有在体内再生气道上皮的全部潜力。为了检验这一假设，采取了三种方法。1)定义“最低限度优化的无血清培养条件”，该条件将指导小鼠（D3和Bruce 4）和人（H9）ES细胞分化为气道上皮细胞谱系，其基于分化标志物表达，该分化标志物表达对气道上皮细胞、细胞生长和细胞形态具有特异性，在不同培养时间点使用负/加一因子方法。2)从形态学和生物化学上评估来源于小鼠（D3和Bruce 4）和人（H9）ES细胞的气道上皮细胞谱系在“最低限度优化的无血清培养条件”下通过植入SCID/裸鼠的裸露大鼠气管移植物的再增殖再生气道上皮的潜力。3)在C57 BL/6小鼠气道上皮修复过程中，在“最低限度优化的无血清培养条件”下，表征来自GFP标记的Bruce 4 ES细胞的体内移植细胞系的定位/分化模式，这些小鼠先前已接受气道上皮特异性毒物萘处理。