Role of Hypoxia in Regulating Stromal-Epithelial Communication during Pregnancy

当胚泡附着到子宫腔上皮并随后 渗透到基质中，牢固地嵌入子宫内膜。虽然许多方面都是 植入尚不清楚，新出现的证据表明，细胞间的通讯 子宫内膜上皮和间质细胞对妊娠的成功建立至关重要。旁长椅 调节这种串扰的信号仍然知之甚少。我们最近的研究表明， 转录因子缺氧诱导因子2α(HIF2α)在子宫内膜间质中的选择性诱导 着床时腔上皮下的细胞。这一发现意义重大，因为它是 很早就知道，植入期间的母体环境是低氧的。要解决这个问题 在植入期间母体对低氧的适应机制，我们产生了HIF2αd/d小鼠， 子宫内膜有条件地缺乏HIF2α。在这些小鼠中，胚泡与子宫紧密相对。 但不能牢固地附着在上皮细胞上，导致植入失败。大鼠子宫间质细胞 HIF2αd/d小鼠表现出代谢因素的下调，如乳酸和一组氨基酸，以及 一组独特的Rab GTP酶，调节细胞外小泡(EV)的分泌，包括微泡 和外显体。电动汽车被认为是一种新的细胞间通讯机制。低氧 促进某些细胞的EV分泌支持HIF2α驱动子宫内膜EV分泌的概念 基质细胞是一种关键的适应性反应，促进细胞间的通讯。 植入。在拟议的研究中，我们将使用hif2αd/d小鼠模型来检验假设 HIF2α在早孕低氧条件下调节一种新的旁分泌信号机制 控制EV在母体组织中的传播，并指导关键信号分子的转移和 在植入过程中，从基质到腔上皮细胞的代谢物影响上皮功能。 我们提出了三个具体目标来检验这一假设。在目标1中，我们将研究hif2α介导的 EV在子宫内膜间质细胞中的运输，并确定EVS中的驻留分子货物。在目标2中，我们将 探讨子宫内膜间质细胞分泌EVS和代谢因子对子宫内膜异位基因表达的影响 上皮细胞的功能。在目标3中，我们将研究hif2α在调节细胞分泌中的作用。 人子宫内膜间质细胞的功能。多管齐下，利用(I)独特的 适应性反应中存在特定缺陷的动物模型(II)用小鼠和 人类原代子宫内膜细胞，以及(Iii)了解细胞间的基因组和蛋白质组分析 贩运机制，将使我们能够为有关 低氧对早期控制子宫内膜功能的间质-上皮沟通的影响 怀孕了。

Implantation is initiated when the blastocyst attaches to the uterine luminal epithelium and subsequently penetrates into the underlying stroma to firmly embed into the endometrium. While many aspects of implantation remain unclear, emerging evidence indicates that intercellular communication between endometrial epithelial and stromal cells is vital for successful establishment of pregnancy. The paracrine signals that mediate this crosstalk remain poorly understood. Our recent studies revealed that the transcription factor, hypoxia-­‐inducible factor 2 alpha (HIF2α), is induced selectively in endometrial stromal cells subjacent to the luminal epithelium at the time of implantation. This finding is significant because it is known for a long time that the maternal environment during implantation is hypoxic. To address the mechanisms of maternal adaptation to hypoxia during implantation, we generated Hif2αd/d mice that conditionally lack HIF2α in the endometrium. In these mice, the blastocysts are closely apposed to the uterine epithelium but fail to sustain firm attachment to it, resulting in implantation failure. The uterine stromal cells of Hif2αd/d mice exhibited downregulation of metabolic factors, such as lactate and a subset of amino acids, and a distinct set of Rab GTPases that regulate secretion of extracellular vesicles (EVs), including microvesicles and exosomes. The EVs are known to mediate a novel mechanism of cell-cell communication. That hypoxia promotes EV secretion by certain cells support the concept that HIF2α-driven EV secretion by the endometrial stromal cells represents a critical adaptive response that promotes intercellular communication during implantation. In the proposed study, we will employ the Hif2αd/d mouse model to test the hypothesis that HIF2α regulates a novel paracrine signaling mechanism under the hypoxic conditions of early pregnancy by controlling EV trafficking in the maternal tissue and that it directs transfer of key signaling molecules and metabolites from the stromal to luminal epithelial cells to influence epithelial functionality during implantation. We have proposed three specific aims to test this hypothesis. In Aim 1, we will investigate HIF2α-mediated EV trafficking in endometrial stromal cells and identify the resident molecular cargo in EVs. In Aim 2, we will investigate the effects of EVs and metabolic factors secreted from endometrial stromal cells on the functionality of epithelial cells. In Aim 3, we will investigate the role of HIF2α in regulating the secretory function of human endometrial stromal cells. A multi-pronged approach, utilizing a combination of (i) a unique animal model harboring a specific defect in adaptive response (ii) functional analyses using mouse and human primary endometrial cells, and (iii) genomic and proteomic analyses to understand cell-to-cell trafficking mechanisms, will enable us to provide answers to important unresolved questions regarding the impact of hypoxia on stromal-epithelial communication controlling endometrial function during early pregnancy.