Role of the extracellular matrix during Wolffian/epididymal duct morphogenesis

细胞外基质在沃尔夫/附睾管形态发生过程中的作用

• 批准号: 9751347
• 负责人: Barry T. Hinton
• 金额: $ 33.51万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-01 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9751347
• 关键词: Area; Atomic Force Microscopy; Attention; Behavior; Biomechanics; Cardiomyopathies; Cells; Clinical; Computer software; Congenital Abnormality; Deposition; Development; Duct (organ) structure; Ductal; Elasticity; Embryo; Embryonic Development; Environment; Epididymis; Epithelial; Epithelial Cells; Event; Extracellular Matrix; Failure; Fetal Development; Genes; Goals; Growth; In Vitro; Lead; Liquid substance; Liver Fibrosis; Male Infertility; Mesenchymal; Mesenchyme; Microscopy; Modulus; Molecular; Morphogenesis; Movement; Mus; Nature; Neural Tube Development; Organ Culture Techniques; Outcome; Outcome Study; Pathologic; Play; Process; Property; Protein Tyrosine Kinase; Radial; Regulation; Reproductive Biology; Role; Sperm Maturation; Structure of mesonephric duct; Testing; Tissues; Tubular formation; cardiogenesis; cell motility; fetal; gastrulation; hearing impairment; human disease; intercalation; interest; male; male fertility; organ growth; real-time images; reproductive tract; second harmonic; tumor progression; two-photon

PROJECT SUMMARY The epididymis provides a unique luminal fluid microenvironment that allows for sperm maturation and survival, and disruptions to this function lead to male infertility. Further, disruptions to epididymal function may also arise as a consequence of abnormal fetal development, although very little is known either of the process of Wolffian duct/epididymal development or of the nature and causes of congenital defects that lead to male infertility. The central hypothesis of this application is that elongation and coiling of the Wolffian duct is crucial for the function of the resulting epididymis, and that failure to elongate and coil leads to male infertility. The overall goal is two-fold, to examine: (a) the mechanisms by which cells move to elongate the duct and (b) to dissect the underlying cellular and molecular mechanisms that regulate those cell movements. Although we have evidence that epithelial cells move by intercalating, we will test the hypothesis that intercalation-type movements of the mesenchyme cells that surround the duct and the en masse movement of mesenchymal cells in the interstitium contribute to ductal elongation and coiling. A combination of genetically modified mice, advanced microscopy including confocal, second harmonic two-photon, and atomic force microscopy, in vitro organ culture and contemporary software analyses will be used to test the hypotheses outlined in the following three specific aims: (1) To test the hypothesis that the stiffness (modulus) of the ECM undergoes dynamic changes during Wolffian duct morphogenesis thereby providing a biomechanical environment that promotes epithelial and mesenchymal cell intercalation and en masse movement of mesenchymal cells within the interstitium. (2) To test the hypothesis that radial intercalation of mesenchymal cells and the en mass movements of mesenchymal cells within the interstitium are major drivers of Wolffian duct elongation and coiling. (3) To test the hypothesis that Ptk7 and Rac1 regulate radial intercalation and en masse movement of mesenchymal cells via regulating ECM biomechanical properties through changes in its deposition and assembly during Wolffian duct development, which in turn are important for male fertility. Therefore, this application focuses on the role of the ECM during Wolffian duct morphogenesis paying special attention to the importance of its assembly, distribution and stiffness, and how this is coordinated to regulate mediolateral and radial intercalation of epithelial and mesenchymal cells respectively, and the en masse movement of mesenchymal cells within the interstitium. Coordination of these events is critical for Wolffian/epididymal duct development, and therefore, male fertility. 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 tubular morphogenesis. Specifically they will provide an understanding as to how the regulation of growth of the epididymis during development is important clinically.

项目概要 附睾提供了独特的管腔液体微环境，允许精子成熟和存活， 这一功能的破坏会导致男性不育。此外，附睾功能也可能受到干扰 作为胎儿发育异常的结果，尽管对沃尔夫过程的任何一个知之甚少 导管/附睾发育或导致男性不育的先天缺陷的性质和原因。这 该应用的中心假设是沃尔夫管的伸长和卷曲对于 由此产生的附睾的功能，以及未能伸长和卷曲会导致男性不育。 总体目标有两个，即检查：(a) 细胞移动以延长导管的机制；(b) 剖析调节这些细胞运动的潜在细胞和分子机制。虽然我们 有证据表明上皮细胞通过嵌入移动，我们将检验嵌入型的假设 导管周围的间充质细胞的运动以及间充质细胞的集体运动 间质细胞有助于导管伸长和卷曲。基因改造小鼠的组合， 先进的体外显微镜技术，包括共焦、二次谐波双光子和原子力显微镜 器官培养和当代软件分析将用于检验以下概述的假设 三个具体目标：(1) 检验 ECM 的刚度（模量）经历动态变化的假设 沃尔夫管形态发生过程中的变化，从而提供了一个生物力学环境，促进 上皮细胞和间充质细胞的嵌入以及间充质细胞在细胞内的集体运动 间质。 (2) 检验间充质细胞径向嵌入和团块的假设 间质内间质细胞的运动是沃尔夫管伸长的主要驱动力 卷绕。 (3)检验Ptk7和Rac1调节径向嵌入和集体运动的假设 间充质细胞通过改变其沉积和调节 ECM 生物力学特性 沃尔夫管发育过程中的组装，这反过来对男性生育能力很重要。因此，这 应用重点关注 ECM 在沃尔夫管形态发生过程中的作用，特别关注 它的组装、分布和刚度的重要性，以及如何协调它们来调节内侧和外侧 上皮细胞和间充质细胞分别呈放射状插入，以及细胞的集体运动 间质内有间质细胞。这些事件的协调对于沃尔夫/附睾管至关重要 发育，从而影响男性生育能力。这项研究的预期结果不仅具有重大意义 对生殖生物学领域的影响目前还知之甚少，但也将有助于我们 了解肾小管形态发生的基本过程。具体来说，他们将提供一个 了解发育过程中附睾生长的调节在临床上很重要。