Cell shape cell adhesion and regulation of ovarian folliculogenesis

雌性哺乳动物出生时就拥有它们所拥有的所有卵子，这一种群是不可替代的。卵巢中的这些卵子大多保存在小的未成熟卵子的休息库中。每个卵子被一层扁平的细胞包围，称为颗粒细胞，并形成一种称为毛囊的结构。在生殖过程中，稳定的细流卵泡离开休眠阶段并开始生长。这种情况一直持续到卵子储备耗尽，对于人类女性来说，这会导致更年期，通常是在50岁左右。必须严格控制卵泡从静止期到生长期的进程，以确保正常的生殖寿命。卵母细胞过早耗尽，导致更年期提前，是女性不孕不育的常见原因。人们对控制卵泡开始生长的因素知之甚少。卵巢局部产生的生长因子似乎起着重要作用，但有几个可能的候选因素，目前尚不清楚哪些是关键因素，以及为什么一些卵泡开始生长，而邻近的其他卵泡则不生长。在本项目中，我们以小鼠卵巢为模型，重点研究颗粒细胞形状发生的变化，这是卵泡生长的第一个迹象。这将使我们对颗粒细胞之间以及与卵母细胞之间的交流方式有一个新的了解。在显微镜下，当卵泡开始生长时，我们看到的最戏剧性的变化之一是扁平的颗粒细胞变得更胖，形状更立方体。在颗粒细胞改变形状后，它们开始分裂，导致数量增加，此时卵母细胞开始生长。对皮肤细胞的研究表明，细胞形状的这种关键变化必须包括将细胞粘合在一起的蛋白质(所谓的黏附分子)的增加，以及细胞内部蛋白质“支架”或细胞骨架的变化。黏附分子的显著之处在于，除了将细胞粘在一起外，它们还与细胞骨架紧密相连，此外，它们还向细胞核发送分子信号。如果细胞改变形状，黏附分子可以发出信号，产生细胞分裂所需的新蛋白质和其他分子。令人惊讶的是，人们对黏附分子和卵泡中的细胞骨架知之甚少。因此，我们想要绘制出静止的卵泡中存在哪些黏附分子和细胞骨架蛋白，并查看是否可以在细胞核附近看到一些信号分子。我们将研究当卵泡开始生长时这些分子是如何变化的。如果我们在培养皿中培养卵巢，同时阻止黏附，我们可以看到黏附分子通过阻断它们将细胞粘连在一起的能力，在颗粒细胞形状、细胞分裂和卵母细胞生长的改变中是多么重要。总体而言，通过定义参与细胞形状变化的蛋白质并显示它们在卵泡生长的不同阶段的定位，我们将能够更好地识别导致颗粒细胞形状变化的环境中最重要的输入信号。然后，我们还可以检查从变化的颗粒细胞到卵子的结果输出信号，这些信号刺激了卵子的生长。换句话说，我们的目标是确定决定哺乳动物生殖寿命的关键因素。

Female mammals are born with all the eggs they will ever have, and this stock is irreplaceable. The majority of these eggs in the ovary are held in a resting stock of small immature eggs. Each egg is surrounded by a layer of a few flattened cells, called granulosa cells, and forms a structure known as a follicle. During reproductive life a steady trickle of follicles leave the resting stage and start to grow. This continues until the stock of eggs is exhausted and, in the human female, this results in the menopause, normally at the age of about 50 years. The progression of follicles from the resting to the growing phase has to be tightly regulated to ensure a normal reproductive lifespan. Premature depletion of oocytes, leading to an early menopause, is a common cause of infertility in women. Little is known about the factors that control the start of follicle growth. Growth factors produced locally in the ovary seem to have an important role but there are several possible candidates and it is not clear which ones are the key factors, and why some follicles start growing while others which are close by don't. In this project, using the mouse ovary as a model, we focus on the change that occurs in the shape of the granulosa cells, which is the first indication of follicle growth. This will give us a new insight into the way in which granulosa cells communicate with each other and with the oocyte. Using microscopy, one of the most dramatic changes that we see as the follicles start to grow is that the flattened granulosa cells become fatter, and more cuboidal in shape. After the granulosa cells have changed shape, they begin to divide, leading to an increase in number and at this point the oocyte begins to grow. Studies in skin cells have shown that this critical change in cell shape must involve an increase in the proteins that stick the cells together (so called adhesion molecules) along with changes in the internal protein 'scaffold', or cytoskeleton, of the cell. Adhesion molecules are remarkable in that, as well as sticking cells together, they also link tightly to the cytoskeleton and, in addition, send molecular signals to the nucleus. If the cell changes shape, the adhesion molecules can send signals to produce new proteins and other molecules needed for cell division. Surprisingly little is known about adhesion molecules and the cytoskeleton in follicles. We therefore want to map which adhesion molecules and cytoskeleton proteins are present in resting follicles and look to see if some of the signalling molecules can be seen near the nucleus. We will investigate how these molecules change as the follicle starts to grow. If we culture ovaries in dishes whilst blocking adhesion, we can see how important the adhesion molecules are in the change in granulosa cell shape, cell division and oocyte growth by blocking their ability to stick the cells together. Overall, by defining the proteins involved in the change in cell shape and showing where they are localized during the various stages of follicle growth, we will be better able to identify the most important input signals from the environment that lead to the change in shape of granulosa cells. We can also then examine the resultant output signals from the changing granulosa cells to the egg which stimulate its growth. In other words, we aim to identify the key factors that determine the reproductive lifespan of the mammal.