During the implantation window, the endometrium becomes poised to transition to a pregnant state, a process driven by differentiation of stromal cells into decidual cells (DC). Perturbations in this process, termed decidualization, leads to breakdown of the feto-maternal interface and miscarriage, but the underlying mechanisms are poorly understood. Here, we reconstructed the decidual pathway at single-cell level in vitro and demonstrate that stromal cells first mount an acute stress response before emerging as DC or senescent DC (snDC). In the absence of immune cell-mediated clearance of snDC, secondary senescence transforms DC into progesterone-resistant cells that abundantly express extracellular matrix remodelling factors. Additional single-cell analysis of midluteal endometrium identified DIO2 and SCARA5 as marker genes of a diverging decidual response in vivo. Finally, we report a conspicuous link between a pro-senescent decidual response in peri-implantation endometrium and recurrent pregnancy loss, suggesting that pre-pregnancy screening and intervention may reduce the burden of miscarriage.

SummaryThe fate of the human endometrium is determined during the mid-luteal window of implantation, coinciding with differentiation of endometrial stromal cells (EnSCs) into specialized decidual cells.In response to successful embryo implantation, differentiating EnSCs transform the endometrium into a decidua that maintains the placenta throughout gestation; whereas falling progesterone levels in the absence of pregnancy lead to tissue destruction and menstrual shedding. We used single-cell

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Pregnancy depends on the wholesale transformation of the endometrium, a process driven by differentiation of endometrial stromal cells (EnSC) into specialist decidual cells. Upon embryo implantation, decidual cells impart the tissue plasticity needed to accommodate a rapidly growing conceptus and invading placenta, although the underlying mechanisms are unclear. Here we characterize a discrete population of highly proliferative mesenchymal cells (hPMC) in midluteal human endometrium, coinciding with the window of embryo implantation. Single-cell transcriptomics demonstrated that hPMC express genes involved in chemotaxis and vascular transmigration. Although distinct from resident EnSC, hPMC also express genes encoding pivotal decidual transcription factors and markers, most prominently prolactin. We further show that hPMC are enriched around spiral arterioles, scattered throughout the stroma, and occasionally present in glandular and luminal epithelium. The abundance of hPMC correlated with the in vitro colony-forming unit activity of midluteal endometrium and, conversely, clonogenic cells in culture express a gene signature partially conserved in hPMC. Cross-referencing of single-cell RNA-sequencing data sets indicated that hPMC differentiate into a recently discovered decidual subpopulation in early pregnancy. Finally, we demonstrate that recurrent pregnancy loss is associated with hPMC depletion. Collectively, our findings characterize midluteal hPMC as novel decidual precursors that are likely derived from circulating bone marrow-derived mesenchymal stem/stromal cells and integral to decidual plasticity in pregnancy.

Decidualizing endometrial stromal cells (EnSC) critically determine the maternal response to an implanting conceptus, triggering either menstruation-like disposal of low-fitness embryos or creating an environment that promotes further development. However, the mechanism that couples maternal recognition of low-quality embryos to tissue breakdown remains poorly understood. Recently, we demonstrated that successful transition of the cycling endometrium to a pregnancy state requires selective elimination of pro-inflammatory senescent decidual cells by activated uterine natural killer (uNK) cells. Here we report that uNK cells express CD44, the canonical hyaluronan (HA) receptor, and demonstrate that high-molecular weight HA (HMWHA) inhibits uNK cell-mediated killing of senescent decidual cells. By contrast, low-molecular weight HA (LMWHA) did not attenuate uNK cell activity in co-culture experiments. Killing of senescent decidual cells by uNK cells was also inhibited upon exposure to medium conditioned by IVF embryos that failed to implant, but not successful embryos. Embryo-mediated inhibition of uNK cell activity was reversed by recombinant hyaluronidase 2 (HYAL2), which hydrolyses HMWHA. We further report a correlation between the levels of HYAL2 secretion by human blastocysts, morphological scores, and implantation potential. Taken together, the data suggest a pivotal role for uNK cells in embryo biosensing and endometrial fate decisions at implantation.