Embryonic Development of the Mammalian Epididymis

描述(申请人提供)：附睾对精子成熟过程至关重要，因为它提供了一个独特的管腔液体微环境，允许精子成熟和存活，而这一功能的中断会导致男性不育。然而，尽管人们对导致男性不育的先天性缺陷的性质和原因知之甚少，但胎儿发育异常也可能导致附睾功能中断。在Wolffian/附睾管胚胎发育过程中的一个主要事件是伸长和卷曲，可能是细胞增殖的结果。然而，在这一应用中，我们假设另一种机制，细胞插层有助于伸长和卷曲。伸长和卷曲不是一件微不足道的事情，但必须与其提供独特的管腔液体微环境的专门功能高度协调，这对精子成熟非常重要。我们之所以选择胚胎时期，是因为细胞增殖和导管卷曲开始启动，而这是一个可能发生潜在缺陷的时期。研究细胞插入作为延长和卷曲Wolffian管的一种手段的机制(S)是新颖的，我们特别感兴趣的是考察平面细胞极性途径(PCP)的一个成员Ptk7作为这一重要事件在Wolffian管形态发生过程中的调节作用。结合转基因小鼠、现代成像技术、体外器官培养技术和基因修饰方法，将检验以下三个具体目标中概述的假设：(1)检验Ptk7在细胞形状和细胞定位中扮演重要角色的假设，这反过来又允许Wolffian管经历延长和卷曲的形态发生事件。(2)验证Ptk7在Wolffian管胚胎发育过程中通过调节细胞基侧突起活动和细胞分裂方向，维持细胞增殖而调节特定细胞重排的假说。(3)验证假设，在Wolffian管延长和卷曲过程中，细胞插入是由细胞骨架组织的极化变化驱动的，并受Ptk7通过激活Wnt/PCP途径调节。综上所述，Wolffian管可以通过细胞插入延长的假说是非常新颖和创新的，因为一直认为附睾管的延长只通过细胞增殖。此外，我们还发现了一种不寻常的调控分子，Ptk7，它被认为在沃尔夫管的延长和卷曲中发挥作用。这项研究的预期结果不仅将对生殖生物学这个一直知之甚少的领域产生重大影响，而且还将有助于我们理解管子形态发生的基本过程。具体地说，他们将提供关于附睾体发育过程中的生长调节在临床上是如何重要的理解。

DESCRIPTION (provided by applicant): The epididymis is critical to the process of sperm maturation because it provides a unique luminal fluid microenvironment that allows for sperm maturation and survival, and disruptions to this function lead to male infertility. However, disruptions to epididymal function may also arise as a consequence of abnormal fetal development, although very little is known either of the process of epididymal development or of the nature and causes of congenital defects that lead to male infertility. A major event during Wolffian/ epididymal duct embryonic development is elongation and coiling, presumably as a result of cell proliferation. However, in this application we hypothesize that an additional mechanism, cell intercalation contributes to elongation and coiling. Elongation and coiling is not a trivial event but must be highly coordinated with its specialized function of providing a unique luminal fluid microenvironment that is so important for sperm maturation. We have chosen the embryonic time period because cell proliferation and ductal coiling are initiated and it is a time when potential defects can occur. Examining the mechanism(s) of cell intercalation as a means to elongate and coil the Wolffian duct is novel, and we are especially interested in examining the contribution of a member of the planar cell polarity pathway (PCP), Ptk7, as a regulator of this important event during Wolffian duct morphogenesis. A combination of genetically modified mice, modern imaging techniques, in vitro organ culture techniques, and gene modifying approaches will be used to test the hypotheses outlined in the following three specific aims: (1) To test the hypothesis that Ptk7 plays an important role in cell shape and cell positioning, which in turn allow the Wolffian duct to undergo the morphogenic events of elongation and coiling. (2) To test the hypothesis that Ptk7 regulates specific cell rearrangements by modulating the basolateral protrusive activity and the orientation of cell division yet maintaining cell proliferaion during Wolffian duct embryonic development. (3) To test the hypothesis that cell intercalation during Wolffian duct elongation and coiling is driven by polarized changes in cytoskeleton organization, and is regulated by Ptk7 through activation of the Wnt/PCP pathway. In summary, the hypothesis that the Wolffian duct can elongate via cell intercalation is highly novel and innovative because it has always been assumed that elongation of the epididymal duct is via cell proliferation only. In addition, we have uncovered an unusual regulatory molecule, Ptk7, which is hypothesized to play a role in Wolffian duct elongation and coiling. The anticipated outcomes of this study will not only have a major impact on an area of reproductive biology that has been poorly understood, but will also contribute to our understanding of the fundamental process of tube morphogenesis. Specifically they will provide an understanding as to how the regulation of growth of the epididymis during development is important clinically.