The derivation of pituitary cells from human ES cells

 描述(申请人提供)：本提案旨在设计一种以细胞疗法为基础的治疗垂体功能减退症的方法，特别关注辐射损伤所致的垂体功能障碍。脑下垂体是一个主要的内分泌器官，在下丘脑的直接控制下，协调有机体中多个腺体的功能。脑垂体功能低下是脑部放射治疗后的一个非常普遍的问题。它是与剂量相关的、渐进性的和不可逆转的；它还导致癌症幸存者的生活质量下降和其他实质性问题。甲状旁腺功能减退也是感染、创伤性脑损伤或先天性疾病的一个重要的永久性后果。我们已经开发了一种方案，用于从人类ES细胞中分离出垂体细胞，并进一步分化为产生激素的细胞。我们之前也优化了一个大鼠的脑辐射模型，使用了临床上相关的分次放射疗程。这些动物的生长、体重和行为参数都得到了很好的研究，我们有一些初步证据表明它们存在垂体功能障碍。在这里，我们建议优化人类ES和iPS分化方案，以允许浓缩到特定的垂体细胞亚型。通过以下途径获得细胞表型的验证 基因表达和标记图谱，激素分泌的测量，以及体内功能。将人ES来源的垂体细胞移植到去垂体的裸鼠体内，在不同时间点检测血清中的基础激素水平。移植物将被放置在下丘脑/垂体界面的皮下或原位。我们还将把这些细胞移植到有垂体损伤证据的受照大鼠体内。分析将集中在血浆中激素水平的恢复，以及通过刺激测试移植物整合到下丘脑-脑垂体轴的证据。此外，功能分析将尝试询问接受细胞移植的HJR雌性大鼠是否可以恢复其生殖和乳酸盐的能力。该提案解决了一个迄今为止鲜有人关注的新问题。随着我们成功地在癌症患者中实现长期缓解，幸存者中生活质量较差的问题变得更加紧迫。该项目将为基于细胞的治疗的可行性提供亟需的见解，以改善脑辐射的这些后果。它还将承担从人类细胞或iPS细胞中衍生出特定的垂体细胞亚型的工作，这是迄今为止在人类细胞中尚未报道的一项成就。

 DESCRIPTION (provided by applicant): This proposal aims at designing a cell-therapy based approach to hypopituitarism , with special focus on pituitary dysfunction due to radiation injury. The pituitary gland is a master endocrine organ that orchestrates the function of multiple glands in the organism, under direct control by the hypothalamus. Hypopituitarism is a very prevalent problem following radiation treatment to the brain. It is dose-related, progressive and irreversible; it also contributes to poor quality of life and other substantial problems among cancer survivors. Hypopituitarism is also an important permanent consequence of infection, traumatic brain injury or congenital conditions. We have developed a protocol for the specification of pituitary cells from human ES cells and further differentiation into hormone producing cells. We have also previously optimized a brain radiation model in the rat, using a clinically relevant fractionated radiation course. The animals are well studied for growth, weight and behavioral parameters and we have some preliminary evidence of pituitary dysfunction. Here we propose to optimize the human ES and iPS differentiation protocol to allow the enrichment into specific pituitary cell subtypes. Validation of cell phenotypes is obtained through gene expression and marker profiles, measurements of hormone secretions, as well as in vivo function. Human ES derived pituitary cells are grafted into hypophysectomized nude rats and basal hormone levels are measured in the serum at different time points. Grafts will be placed subcutaneously or orthotopically into the hypothalamus / pituitary interface. We will also graft the cells into the irradiated rats with evidence of pituitary damage. Analysis will focus on revovery of hormone levels in the plasma, as well as evidence of integration of the graft into the hypothalamic - pituitary axis, using stimulation tests. Furthermore, functional analysis will be attempted by asking whetehjr female rats with cell grafts can recover their ability to reproduce and lactate.The proposal addresses a novel problem that has received little attention to date. As we succeed in achieving long term remission among cancer patients, questions of poor quality of life among survivors have acquired a more pressing nature. This project will provide much needed insight into the feasibility of cell based therapy to ameliorate these consequences of brain radiation. It will also undertake the derivation of specific pituitary cell subtypes from humn ES or iPS cells, an unreported achievement in human cells to date.