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Research on regulation mechanisms of development and differentiation of mouse primordial germ cells

小鼠原始生殖细胞发育分化调控机制研究

基本信息

批准号：
08458241
负责人：
NAKATSUJI Norio
金额：
$ 4.03万
依托单位：
National Institute of Genetics
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1996
资助国家：
日本
起止时间：
1996 至 1997
项目状态：
已结题

项目摘要

Mouse primordial germ cells appear at the base of allantois in the posterior extraembryonic region, migrate to the genital ridges, and differentiate into oocytes or prospermatogonia in the fetal ovary or testis. We carried out the following experiments to study mechanisms in the development and differentiation of fetal germ cells.There have been no culture system of fetal germ cells after arrival at gonads, because they go into apoptosis and disappear in usual culture conditions. We developed novel methods of sexing embryos, isolation of germ cells with magnetic beads, and detection of meiotic cells with specific antibodies. As a result, we detected many germ cells entering into meiotic prophase in our culture system.Mammalian sex-determination and differentiation is controlled by several genes, such as Sry, Sox-9, Dax-1 and Ad4BP/SF-1, but their upstream and downstream genes are largely unknown. In order to identify these genes involved in the sex-differentiation, we carried out the differential hybridization of the mouse embryonic gonad cDNA library using presumably male-specific probes. We identified nephgonadin, which encodes a basic helix-loop-helix motif. The earliest expression of nephgonadin was observed at 8.5 days post coitum (dpc) in the intermediate mesodermal tissues of anterior and posterior parts of the mouse embryo. The posterior expression continued until this region forms the urogenital ridge. From 13.5 dpc to 2 weeks postnatal, nephgonadin was expressed at higher levels in the male than female gonad. In adults, however, expression of nephgonadin was drastically decreased in the testis, while it was increased in the ovary. These sex- and stage-dependent expression patterns are similar to that of Ad4BP/SF-1, which is important for the development and sex-differentiation of the gonad.

小鼠原始生殖细胞出现在胚外区后部的尿囊基部，迁移到生殖嵴，并在胎儿卵巢或睾丸中分化为卵母细胞或受精卵。为了研究胎儿生殖细胞发育和分化的机制，我们进行了以下实验：胎儿生殖细胞到达性腺后没有培养系统，因为它们在通常的培养条件下进入凋亡并消失。我们开发了新的方法性别鉴定胚胎，分离生殖细胞的磁珠，并检测减数分裂细胞的特异性抗体。哺乳动物的性别决定和分化受多种基因控制，如Sry、Sox-9、Dax-1和Ad 4 BP/SF-1，但其上下游基因尚不清楚。为了鉴定这些参与性分化的基因，我们用雄性特异性探针对小鼠胚胎性腺cDNA文库进行了差异杂交。我们确定了nephgonadin，它编码一个基本的螺旋-环-螺旋基序。肾促性腺素在小鼠胚胎发育后8.5天（dpc），在胚胎前、后部的中间中胚层组织中最早表达。后部的表达一直持续到该区域形成泌尿生殖嵴。从出生后13.5天到2周，nephgonadin在雄性性腺中的表达水平高于雌性性腺。然而，在成年人中，nephgonadin的表达在睾丸中急剧下降，而在卵巢中则增加。这些性别和阶段依赖的表达模式与Ad 4 BP/SF-1相似，这对性腺的发育和性别分化很重要。