During early human pregnancy, the fetal placenta implants into the uterine mucosa (decidua)where placental trophoblast cells intermingle and communicate with maternal cells. Trophoblastdecidual interactions underlie common diseases of pregnancy including pre-eclampsia and stillbirth. Here, we profile transcriptomes of ~70,000 single cells from first trimester placentas with matched maternal blood and decidual cells. The cellular composition of human decidua reveals new subsets of perivascular and stromal cells, which are located in distinct decidual layers.There are three major subsets of decidual NK cells, with distinctive immunomodulatory and chemokine profiles. We develop a repository of ligand-receptor complexes (https://cellphonedb.org/) and a statistical tool to predict the cell-type specificity of cell-cell communication via these molecular interactions. This identifies many regulatory interactions that prevent any damaging innate or adaptive immune responses in this environment. Our single cell atlas of the maternal-fetal interface reveals the cellular organization and interactions critical for placentation and reproductive success.During early pregnancy, the uterine mucosal lining, the endometrium, is transformed into decidua under the influence of progesterone. Decidualisation results from a complex and well-orchestrated differentiation program that involves all cellular elements of the mucosa: stromal, glandular, and immune cells, including the distinctive decidual Natural Killer cells (dNK) 1,2 . The blastocyst implants into the decidua and initially, before arterial connections are established, uterine glands are the source of histotrophic nutrition in the placenta 3,4 . Following implantation, placental extravillous trophoblast cells (EVT) invade through the decidua and move towards the spiral arteries, where they destroy the smooth muscle media and transform the arteries into high conductance vessels 5 . Balanced regulation of EVT invasion is critical to pregnancy success: arteries must be sufficiently transformed, but excessive invasion prevented, to ensure correct allocation of resources to both mother and baby 6 . The pivotal regulatory role of the decidua is obvious from the life-threatening, uncontrolled, trophoblast invasion that occurs when the decidua is absent as when the placenta implants on a previous cesarean section scar 7 .EVT have a unique HLA profile: they do not express the dominant T cell ligands, class I HLA-A and HLA-B or class II molecules 8,9 , but do express HLA-G and HLA-E and polymorphic HLA-C class I molecules. These trophoblast HLA ligands have receptors expressed by the dominant decidual immune cells, dNK, including maternal killer immunoglobulin-like receptors (KIR), that bind HLA-C molecules 10,11 . Certain combinations of maternal KIR and fetal HLA-C genetic variants are associated with pregnancy disorders such as pre-eclampsia, where trophoblast invasion is deficient 12 . However, detailed understanding of the cellular interactions in the decidua supporting early...

In humans, the endometrium, the uterine mucosal lining, undergoes dynamic changes throughout the menstrual cycle and pregnancy. Despite the importance of the endometrium as the site of implantation and nutritional support for the conceptus, there are no long-term culture systems that recapitulate endometrial function in vitro. We adapted conditions used to establish human adult stem cell-derived organoid cultures to generate 3D cultures of normal and decidualised human endometrium. These organoids expand long-term, are genetically stable and differentiate following treatment with reproductive hormones. Single cells from both endometrium and decidua can generate a fully functional organoid. Transcript analysis confirmed great similarity between organoids and the primary tissue of origin. On exposure to pregnancy signals, endometrial organoids develop characteristics of early pregnancy. We also derived organoids from malignant endometrium, and so provide a foundation to study common diseases, such as endometriosis and endometrial cancer, as well as the physiology of early gestation.

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SummaryBMP is thought to induce hESC differentiation toward multiple lineages including mesoderm and trophoblast. The BMP-induced trophoblast phenotype is a long-standing paradox in stem cell biology. Here we readdressed BMP function in hESCs and mouse epiblast-derived cells. We found that BMP4 cooperates with FGF2 (via ERK) to induce mesoderm and to inhibit endoderm differentiation. These conditions induced cells with high levels of BRACHYURY (BRA) that coexpressed CDX2. BRA was necessary for and preceded CDX2 expression; both genes were essential for expression not only of mesodermal genes but also of trophoblast-associated genes. Maximal expression of the latter was seen in the absence of FGF but these cells coexpressed mesodermal genes and moreover they differed in cell surface and epigenetic properties from placental trophoblast. We conclude that BMP induces human and mouse pluripotent stem cells primarily to form mesoderm, rather than trophoblast, acting through BRA and CDX2.

The endometrium, the mucosal lining of the uterus, undergoes dynamic changes throughout the menstrual cycle in response to ovarian hormones. We have generated dense single-cell and spatial reference maps of the human uterus and three-dimensional endometrial organoid cultures. We dissect the signaling pathways that determine cell fate of the epithelial lineages in the lumenal and glandular microenvironments. Our benchmark of the endometrial organoids reveals the pathways and cell states regulating differentiation of the secretory and ciliated lineages both in vivo and in vitro. In vitro downregulation of WNT or NOTCH pathways increases the differentiation efficiency along the secretory and ciliated lineages, respectively. We utilize our cellular maps to deconvolute bulk data from endometrial cancers and endometriotic lesions, illuminating the cell types dominating in each of these disorders. These mechanistic insights provide a platform for future development of treatments for common conditions including endometriosis and endometrial carcinoma.