Isolation and Characterization of Testis Stem Cells

描述（由申请人提供）：我们的长期目标是研究精原干细胞（A/单个精原细胞）的生物学。有很大的兴趣在A/单精原细胞，因为它们的重要性，了解细胞自我更新与分化的基本机制，治疗不孕症，避孕药的发展和理解的病因睾丸癌，特别是睾丸癌。然而，对这些细胞的生物学研究受到严重阻碍，因为它们的数量很少，没有独特的标记物存在，并且很难将它们与其他更成熟的精原细胞分离。我们最近开发了一种方法，使我们能够分离纯种群的A/单精原细胞，我们已经开始研究他们的行为在体外。使用微阵列，我们还分析了A/单个精原细胞的胶质细胞源性神经营养因子（GDNF），支持细胞产生的生长因子的存在下的转录程序。第一个目的是分离单个精原细胞，通过生殖细胞移植评价其克隆形成能力。在第二个目标中，我们将确定哪些信号通路被GDNF诱导，导致单个精原细胞自我更新或分化。在第三个目标中，我们将研究GDNF如何与其他生长因子/受体相互作用以促进自我更新或分化。 总之，这条线的工作将扩大我们的知识的功能特性的A/单精原细胞，让我们更好地了解分子机制，驱动精子发生的第一步。特别地，将阐明生长因子GDNF在调节自我更新和/或分化中的作用。

DESCRIPTION (provided by applicant): Our long-term goal is to study the biology of spermatogonial stem cells, the A/single spermatogonia. There is great interest in the A/single spermatogonia because of their importance for understanding basic mechanisms of cell self-renewal versus differentiation, for the treatment of infertility, for the development of contraceptives and for the understanding of the etiology of testicular cancer, in particular seminoma. However, studies on the biology of these cells have been severely hampered because their number is low, no unique marker exists and it has been difficult to physically separate them from the other more mature spermatogonia. We recently developed a method that allows us to isolate pure populations of A/single spermatogonia, and we have started to study their behavior in vitro. Using microarrays, we also have analyzed the transcriptional program of the A/single spermatogonia in the presence of glial cell line-derived neurotrophic factor (GDNF), a growth factor produced by Sertoli cells. In the first aim, we will isolate Asingle spermatogonia and evaluate their clonogenic potential by germ cell transplantations. In the second aim, we will determine which signaling pathways are induced by GDNF, leading to Asingle spermatogonia self-renewal or differentiation. In the third aim, we will study how GDNF interacts with other growth factors/receptors to promote self-renewal or differentiation. In summary, this line of work will expand our knowledge of the functional characteristics of the A/single spermatogonia and give us better insight into the molecular mechanisms that drive the first steps of spermatogenesis. In particular, the role of the growth factor GDNF in regulating self-renewal and/or differentiation will be elucidated.