Role of extracellular vesicles in trophoblast invasive migration

胎盘是胎儿适当发育的重要器官，其成功植入取决于滋养层细胞对子宫的入侵，直接影响动物和人类健康，导致显著的福祉和健康老龄化问题。流行病学研究表明，只有大约30%的怀孕得到临床认可，而大多数是自发或在妊娠并发症期间丢失的，植入过程是这一过程的关键调节器。 尽管胎盘对胎儿发育的重要性，我们对介导胎盘植入的机制知之甚少-这是发育健康，功能性胎盘的关键步骤，但滋养层侵入性迁移到接受性子宫内膜是至关重要的。这项拟议的工作计划将提供世界领先的，新的分子机制的见解，在植入过程中的滋养层侵入运动的过程，以及它是如何调节从周围子宫内膜释放的微泡。绒毛外滋养层细胞（EVT）从胎盘的锚定绒毛中的细胞滋养层增殖和迁移，并侵入母体蜕膜和子宫肌层。目前对细胞运动重要的分子途径知之甚少，但子宫内膜分泌的细胞外囊泡对这种调节很重要。本项目将利用我们实验室常规使用的一系列细胞生物学检测方法，分析子宫内膜通过释放细胞外囊泡来调节滋养细胞侵袭性迁移的分子机制，入侵，并确定这些过程中涉及的因素。细胞外囊泡的分离和定位于其中的因子也是常规进行的。常规使用的人细胞系最初将从子宫内膜谱系中分离EV及其对滋养层细胞运动性和侵袭性的影响，但也将检测离体样品。在该项目的第一部分，将建立最佳条件，以确定最敏感的滋养层细胞和最活跃的子宫内膜细胞外囊泡。该研究的第二部分旨在通过使用在我们目前BBSRC资助的工作中进行的协议和方法进行无偏倚的质谱分析来获得微泡的完整签名。该项目的最后一部分将旨在通过基因调控或通过使用可用的抑制剂（例如抗体和/或化学抑制剂），通过使用功能获得和功能丧失研究，确定一些新发现的标记物，并确定它们在细胞外囊泡中的存在是否对调节滋养层侵入性迁移很重要。

The placenta is an essential organ for appropriate foetal development and its successful implantation depends upon invasion of the uterus by trophoblast cells, impacting directly on both animal and human health leading to significant well-being and healthy ageing issues. Epidemiological studies have shown that only around 30% of all conceptions get clinically recognized whilst the majority are lost either spontaneously or during pregnancy complication and that the process of implantation is a key regulator in this process. Despite the importance of the placenta to foetal development, we know little of the mechanisms that mediate placental implantation - the key step in developing a healthy, functional placenta, but the invasive migration of the trophoblast into the receptive endometrium is paramount. This proposed programme of work will provide world-leading, novel molecular mechanistic insight to the process of trophoblast invasive motility during implantation and how it is regulated by microvesicles released from the surrounding endometrium. Extravillous trophoblast cells (EVTs) proliferate and migrate from the cytotrophoblast in the anchoring villi of the placenta and invade the maternal decidua and myometrium. Little is known about the molecular pathways that are important for cellular motility but extracellular vesicle secretion by the endometrium has been shown to be important for such regulation.This project will dissect the molecular mechanisms by which the endometrium regulates trophoblast invasive migration through the release of extracellular vesicles using an array of cellular biology assays that are routinely used in our laboratories to measure cell motility and invasion, and determine the factors involved in these process. The isolation of extracellular vesicles and the factors that are localised within is also routinely done. Human cell lines routinely used will initially be used from endometrial linage to isolate EV and their effects on trophoblast cells for motility and invasion but ex vivo samples will also be tested. In the first part of the project, the best conditions will be established to determine the most responsive trophoblast cells and the most active endometrial extracellular vesicles. The second part of the study will aim to get a full signature of the microvesicles through an unbiased mass spectrometry analysis using protocols and approaches that have been conducted within our current BBSRC-funded work. The last part of the project will aim to characterise some of the newly identified markers and determine whether their presence in extracellular vesicles are important to regulate the trophoblast invasive migration through the use of gain of function and loss of function studies either by gene regulation or through the use of available inhibitors (e.g. antibodies and or chemical inhibitors).