Embryonic Development of the Mammalian Epididymis

描述（由申请人提供）：附睾对精子成熟过程至关重要，因为它提供了一个独特的腔液微环境，允许精子成熟和存活，而这种功能的破坏会导致男性不育。然而，附睾功能的破坏也可能是胎儿发育异常的结果，尽管对附睾发育的过程或导致男性不育的先天性缺陷的性质和原因知之甚少。沃尔夫管/附睾管胚胎发育过程中的一个主要事件是伸长和卷曲，这可能是细胞增殖的结果。然而，在这个应用中，我们假设一个额外的机制，细胞嵌入有助于伸长和卷曲。伸长和盘绕不是一个微不足道的事件，但必须与其提供独特的腔液微环境的特殊功能高度协调，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盘绕已经开始，这是一个可能发生潜在缺陷的时期。研究细胞嵌入作为一种延长和缠绕沃尔夫氏管的手段的机制是新颖的，我们特别感兴趣的是研究平面细胞极性途径（PCP）成员Ptk7在沃尔夫氏管形态发生过程中作为这一重要事件的调节因子的贡献。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，将用于验证以下三个特定目的的假设：(1)验证Ptk7在细胞形状和细胞定位中起重要作用的假设，这进而使沃尔夫氏管经历伸长和卷曲的形态发生事件。(2)验证在Wolffian导管胚胎发育过程中，Ptk7通过调节基底外侧突起活动和细胞分裂方向调控特异性细胞重排，同时维持细胞增殖的假说。(3)验证Wolffian导管伸长和卷曲过程中的细胞嵌入是由细胞骨架组织的极化变化驱动的，并由Ptk7通过激活Wnt/PCP通路进行调控。总之，沃尔夫管可以通过细胞嵌入延长的假设是非常新颖和创新的，因为它一直被认为附睾管的延长只是通过细胞增殖。此外，我们还发现了一种不寻常的调节分子Ptk7，它被假设在沃尔夫氏管伸长和卷曲中发挥作用。本研究的预期结果不仅将对生殖生物学领域的研究产生重大影响，而且将有助于我们对试管形态发生的基本过程的理解。具体来说，他们将提供一个理解，如何调节生长的附睾在发育过程中是重要的临床。

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.