Modelling the impact of decidual senescence on embryo implantation in human endometrial assembloids.

Decidual remodelling of midluteal endometrium leads to a short implantation window after which the uterine mucosa either breaks down or is transformed into a robust matrix that accommodates the placenta throughout pregnancy. To gain insights into the underlying mechanisms, we established and characterized endometrial assembloids, consisting of gland-like organoids and primary stromal cells. Single-cell transcriptomics revealed that decidualized assembloids closely resemble midluteal endometrium, harbouring differentiated and senescent subpopulations in both glands and stroma. We show that acute senescence in glandular epithelium drives secretion of multiple canonical implantation factors, whereas in the stroma it calibrates the emergence of anti-inflammatory decidual cells and pro-inflammatory senescent decidual cells. Pharmacological inhibition of stress responses in pre-decidual cells accelerated decidualization by eliminating the emergence of senescent decidual cells. In co-culture experiments, accelerated decidualization resulted in entrapment of collapsed human blastocysts in a robust, static decidual matrix. By contrast, the presence of senescent decidual cells created a dynamic implantation environment, enabling embryo expansion and attachment, although their persistence led to gradual disintegration of assembloids. Our findings suggest that decidual senescence controls endometrial fate decisions at implantation and highlight how endometrial assembloids may accelerate the discovery of new treatments to prevent reproductive failure

Characterisation of endometrial gland defects associated with recurrent missed miscarriage

Recurrent pregnancy loss, defined as multiple miscarriages, is a distressing condition associated with significant physical trauma and psychological morbidity. Miscarriage often causes bleeding in early gestation, reflecting the breakdown of the emerging maternal-fetal interface. However, bleeding is not a prominent feature in missed miscarriage, which typically involves very early-onset fetal growth restriction, fetal bradycardia and ultimately fetal demise. I hypothesised that recurrent missed miscarriage (RMM), defined here as 3 or more ‘silent’ miscarriages, reflects an endometrial glandular defect that compromises histotrophic nutrition before the onset of placental perfusion around 12 weeks of pregnancy. To test this hypothesis, RNA-sequencing data of laser-captured mid-luteal endometrial glands from RMM patients and control subjects were subjected to metabolic modelling using flux variability and balance analysis. Computational modelling revealed a notable metabolic signature in RMM, characterised by heightened oxidative phosphorylation and fatty acid oxidation and decreased glycolysis, which was not attributable to glandular asynchrony. To explore the drivers of this endometrial defect in RMM, I optimised a robust organoid protocol to assess glandular differentiation in vitro. RMM organoids not only recapitulated the in vivo metabolic signature but displayed a multitude of defects caused by more naïve progenitor cells, including attenuated Wnt and Notch signalling, and impaired specification of epithelial cells into differentiated and senescent subpopulations upon progesterone-induced cell cycle arrest and differentiation. Importantly, senescent epithelial cells, which are a major source of growth factors and extracellular matrix proteinases, may critically regulate endoglandular trophoblast invasion and access to glandular secretions in early gestation. Quantification of p16INK4 immuno reactivity in 48 biopsies confirmed that RMM is associated with lack of glandular senescence during the peri-implantation window. Collectively, the data indicate that aberrant metabolic programming of endometrial epithelial progenitor cells compromises glandular function, thus revealing a novel therapeutic target for RMM prevention

Endometrial assembloids to model human embryo implantation in vitro

Understanding the process of human embryo implantation is impeded by the inability to study this phenomenon in vivo, thus limiting opportunities to gain knowledge to in vitro modeling. Previous models have relied on monolayer co-cultures, which do not replicate the complexity of endometrial tissue. Here, we detail the establishment of three-dimensional endometrial assembloids, comprising gland-like epithelial organoids in a stromal matrix. Endometrial assembloids mimic endometrial tissue structure more faithfully and can be used to study human embryo-endometrial interactions. Co-cultures of human embryos and endometrial assembloids will enhance our fundamental understanding of these processes as well as allowing us to study the mechanisms of persistent reproductive failure

EndoTime : non-categorical timing estimates for luteal endometrium

STUDY QUESTION Can the accuracy of timing of luteal phase endometrial biopsies based on urinary ovulation testing be improved by measuring the expression of a small number of genes and a continuous, non-categorical modelling approach