Towards an in vitro model of human hypoblast

众所周知，绝大多数在辅助受孕计划后开始的妊娠在植入后很早就失败了。虽然这些失败中的许多可以归因于与母亲的子宫不相容，但大约三分之一是由发育中的胚胎缺陷引起的。在植入时，胚胎必须由三个组织组成：滋养外胚层，它将与子宫进行第一次连接并形成胎盘；亚原始细胞，它是确定胎儿的前后和形成卵黄囊所必需的；以及外胚层，它产生胎儿的所有组织。为了随后的正常发育，每个早期胚胎谱系必须被适当和成比例地表现出来。根据我们的观察，我们假设，未能指定足够的外胚层细胞或低胚层细胞，或两者兼而有之，可能是产生可存活妊娠的主要问题。然而，对这些血统的适当分配的调控过程却知之甚少。很难获得统计能力来回答有关胚胎这一过程的问题。同样，不可能将血统的分配与最终成功的子宫植入联系起来。因此，为了了解早期胚胎谱系是如何分配的，有必要用代表早期胚胎谱系的细胞系构建人工胚胎模型。已验证的细胞系代表上胚层和滋养层细胞，但不代表低胚层细胞。我们使用了特殊配方的支持性凝胶和一种培养制度，可以从小鼠胚胎中捕获低母细胞，并在它们扩展到细胞系的过程中将它们保持在早期胚胎状态。在这项研究中，我们将优化机械和化学条件，特别是从人类胚胎中产生自我更新的低分化细胞系。用代表所有三种胚胎谱系的细胞系武装起来，我们将使用专门建造的3D水凝胶来构建人造胚胎。通过我们的模型人工胚胎，结合我们的合作者Margherita Turco开发的新的子宫内膜器官，我们可以定量地测试关于每个谱系启动正常发育(包括植入)所需的细胞数量的预测。我们的研究将提供有价值的信息，为扩大人类低母细胞群体的特定因素提供有价值的信息，这些因素可能有助于改善辅助受孕计划的培养制度。

It is widely known that the vast majority of pregnancies initiated following assisted conception programmes fail very early, at around the time of implantation. Although many of these failures can be attributed to incompatibility with the mother's uterus, around one third are caused by defects in the developing embryo. At the time of implantation, the embryo must consist of three tissues: the trophectoderm that will make the first connection with the uterus and give rise to the placenta; the hypoblast that is essential for specifying the anterior and posterior of the foetus and forming the yolk sac; and the epiblast that produces all the tissues of the foetus. In order for normal development to ensue, each early embryonic lineage must be appropriately and proportionately represented. Based on our observations, we hypothesise that failure to specify enough cells of either epiblast or hypoblast, or both, is likely to be a major problem for generating a viable pregnancy. However, the process by which appropriate allocation of these lineages is regulated is poorly understood. It is difficult to attain statistical power to answer questions about this process from embryos. Similarly, it is not possible to correlate apportionment of the lineages with eventual successful uterine implantation. Therefore, in order to understand how early embryonic lineages are allocated, it is essential to have model artificial embryos constructed from cell lines representing the early embryonic lineages. There are validated cell lines representing the epiblast and trophoblast, but not the hypoblast. We have used specially formulated supportive gels and a culture regime that can capture hypoblast cells from mouse embryos and keep them in an early embryonic state as they expand into cell lines. In this study, we will optimise the mechanical and chemical conditions specifically to generate self-renewing hypoblast cell lines from human embryos. Armed with cell lines representing all three embryonic lineages, we will use purpose-built 3D hydrogels to construct artificial embryos. With our model artificial embryos, in combination with the new endometrial organoids developed by our collaborator Margherita Turco, we can quantifiably test predictions concerning, for example, the number of cells of each lineage needed to initiate normal development, including implantation. Our study will provide valuable information on the requirements for specific factors for expanding the human hypoblast population that may enable improvement of culture regimes for assisted conception programmes.