雄激素对大鼠睾丸间质祖细胞增殖分化的调控机制研究

Leydig cells are present in the interstitial compartment of the testes and their main function is to produce testosterone, which is essential for spermatogenesis and development of secondary sexual characteristics. The progenitor Leydig cell (PLC) is a founder cell in the Leydig cell lineage and is originated from undifferentiated stem Leydig cell. PLC is a very important developmental stage in the Leydig cell lineage, in which its proliferation and formation of steroidogenic capacity have a critical role in Leydig cell number and testosterone biosynthesis. A classic theory is that the proliferation, maturation and steroidogenesis of Leydig cells are controlled by hypothalamus-pituitary-testis axis. Pituitary-secreted luteinizing hormone (LH) is the key regulator of Leydig cell function and it is indispensable for functional differentiation and proliferation of Leydig cells, it acts on Leydig cells to stimulate testosterone biosynthesis. However, it has been found that PLC has very few LH receptors and is insensitive to LH stimulation while it has abundant androgen receptors (AR), suggesting that androgen can regulate PLC development. The objective of this application is to study the regulation of 7-methyl-19-nortestosterone (MENT), an potent androgen that is resistant to 5-reductase, on the earlier stage of development of PLC, including the analysis of the cell proliferation and the expression of testosterone biosynthetic and metabolizing enzymes.

睾丸间质祖细胞（PLC）是Leydig细胞的一种起源细胞，这种细胞从未分化的间质干细胞分化而来。其在Leydig细胞系发育过程中是非常重要的一个阶段，增殖能力以及类固醇酶系形成情况对后期的成年Leydig细胞的数量和睾酮合成能力起着决定性的作用。经典的观点认为垂体分泌的促黄体生成激素（LH）作用于Leydig细胞刺激睾酮的合成。但PLC中LH受体（LHR）表达很低，LH对其刺激不敏感，表明LH并不是leydig细胞发育中唯一起作用的激素，PLC中含有的大量的雄激素受体（AR），数量上远远超过LHR数量。我们推测除了LH，雄激素也可能参与了分化过程。因此在本项目中我们提出假说，雄激素可能刺激截断型LHR成为全长有功能的LHR，从而调节Leydig细胞早期发育。本课题主要运用现代生物学技术从体内和体外对这一假说进行验证，探索Leydig细胞早期发育中的作用机制，为治疗男性生殖障碍提供实验依据。

睾丸间质细胞是哺乳动物睾丸间质中的一种内分泌细胞，具有合成和分泌睾酮的功能。对男性而言，睾酮具有非常重要的生理功能。因此研究Leydig细胞的发育对于治疗男性生殖疾病具有非常重要的临床意义。在本项目中，我们利用特异性抑制剂乙烷二甲烷硫砜（Ethane Dimefhane sulfonaft, EDS）选择性诱导Leydig细胞凋亡，构建成年大鼠Leydig细胞“敲除”动物模型，以下丘脑-垂体-性腺轴为中心，以睾丸间质细胞中睾酮合成的关键酶及调节分子为靶标，应用分子生物学、细胞生物学和内分泌学等技术研究了Leydig再生细胞的发育特点以及外源性激素对成年Leydig细胞的诱导分化的作用及其机制，为Leydig细胞的发育和再生治疗男性生殖障碍提供重要的理论依据和实验基础。主要完成了如下研究工作：(1)成功的构建成年大鼠Leydig细胞“敲除”动物模型，并对Leydig再生细胞的发育特点做了深入的研究。(2) 用LH或MENT对分离纯化后的PLC进行培养，检测了雄激素的水平，并观察了PLC的分化情况。(3) 研究了蛋白Slc15a1在成年大鼠睾丸中发育中的作用。.通过本课题的实施，正式发表研究论文5篇，其中SCI论文4篇，另外有2篇中文核心期刊还在审稿中，以本课题为载体，培养青年教师1名，博士研究生1名，硕士研究生1名。