Regulation of mouse spermatogonial stem cell self-renewal

描述（由申请人提供）：精原干细胞（ssc），男性生殖系干细胞，通过其自我更新和产生子细胞的能力为精子发生提供基础，子细胞分化为精子。这些特性使ssc成为维持雄性生殖能力和保存遗传系的关键成分。小鼠SSC自我更新的信号是由生长因子胶质细胞系来源的神经营养因子（GDNF）介导的，可能通过c-Ret原癌基因受体。使用该生长因子和确定的中、长期（超过6个月）小鼠SSCs在体外发生增殖，这已被细胞表面抗原标记的维持和培养的SSCs移植到受体时产生供体细胞来源的精子发生和后代的能力所证实。因此，为研究小鼠SSC自我更新的分子机制提供了一个独特而有力的系统。我们最近使用微阵列分析在高度富集的SSC群体中鉴定gdnf调节基因，提供了可能构成调节SSC自我更新和生存的核心分子机制的基因数据库。同样的生长因子和类似的分子机制可能被其他哺乳动物ssc使用，因为许多物种（如大鼠、猪、狒狒和人）的干细胞已被证明在小鼠睾丸中增殖。此外，发现小鼠SSC自我更新的重要分子可能有助于理解其他成体组织干细胞使用的机制。利用小鼠SSC培养系统和干细胞移植技术分析小鼠SSC自我更新的分子调控。提出的两个具体目标是：1)研究参与SSC自我更新的GDNF信号通路；2)确定GDNF调控基因在SSC自我更新中的生物学意义。识别男性生殖系干细胞生长和分裂所需的信号将有助于理解人类男性不育，并建立体外培养或生长人类ssc的系统。这种培养系统可以用于在化疗期间维持男性的干细胞，然后干细胞在治疗后返回男性的睾丸以重建生育能力。

DESCRIPTION (provided by applicant): Spermatogonial stem cells (SSCs), male germ line stem cells, provide the foundation for spermatogenesis by their ability to both self-renew and generate daughter cells, which differentiate into spermatozoa. These attributes make SSCs the key component for sustained fertility in males and the preservation of genetic lines. Signaling for mouse SSC self-renewal is mediated by the growth factor glial cell line-derived neurotrophic factor (GDNF) likely through the c-Ret proto-oncogene receptor. Using this growth factor and a defined medium, long-term (more than 6-months) in vitro proliferation of mouse SSCs occurs, which has been confirmed by maintenance of cell surface antigenic markers and ability of cultured SSCs, when transplanted to recipients, to produce donor cell-derived spermatogenesis and progeny. Thus, a unique and powerful system is available to study the molecular mechanisms of mouse SSC self-renewal. We have recently used microarray profiling to identify GDNF-regulated genes in highly enriched populations of SSCs, providing a database of genes that may constitute the core molecular machinery regulating SSC self-renewal and survival. The same growth factor and similar molecular mechanisms are likely used by other mammalian SSCs, because stem cells from many species (e.g. rat, pig, baboon and human) have been shown to proliferate in mouse testes. In addition, important molecules identified for mouse SSC self-renewal may be helpful in understanding mechanisms used by other adult tissue stem cells. The mouse SSC culture system and stem cell transplantation technique will be used to analyze the molecular regulation of mouse SSC self- renewal. The two proposed specific aims are: 1) Investigate GDNF signaling pathways involved in SSC self- renewal, and 2) Determine the biological significance of GDNF-regulated genes in SSC self-renewal. The identification of the signals required for male germ line stem cells to grow and divide will help in understanding male infertility in humans and in establishing a system to culture or grow human SSCs outside the body. Such a culture system could be used to maintain stem cells of a male during chemotherapy, and the stem cells then returned to the male's testes to reestablish fertility after therapy.