Cellular heterogeneity of the human endometrium : transcriptomic deconvolution and characterization of stem/progenitor cells

During a woman’s reproductive years the human endometrium becomes a highly dynamic tissue that sheds and regenerates on a cyclic basis. Apart from the main cell populations (i.e. stromal, endothelial, epithelial and immune cells), the endometrium also comprises a hierarchy of stem and more committed cells that are responsible for the remarkable regenerative ability of this tissue. This heterogeneity hinders the interpretation of bulk RNA-seq data and subsequently, the identification of specific stem cell transcriptomic signatures. Hence, the aim of this thesis was to assess and interpret endometrial heterogeneity with a focus on the characterization of stem/progenitor cells. Firstly, a reference matrix with the transcriptome profile of epithelial, uNK and stromal subpopulations was generated and used in the deconvolution of whole tissue transcriptome data across the menstrual cycle. This analysis showed that the transition from early- to mid-secretory is followed by an adjustment in the stromal:epithelial ratio. Additionally, it was demonstrated that endometrial mesenchymal stem cells (eMSC) encompass a heterogeneous population, and that their microenvironment may share common stemness properties with other adult stem cells. Secondly, single cell RNA-seq was employed to analyse endometrial cell heterogeneity. Apart from stromal, endothelial, epithelial and diverse immune cell populations, a discrete population of highly proliferative mesenchymal cells was identified and further characterized. Through several lines of evidence, I demonstrated that these cells correspond to a putative progenitor cell population and that anillin is a candidate marker for endometrial clonogenicity. Lastly, single-nucleus transcriptomic analysis was optimized for archived endometrial samples in order to bypass the disadvantages inherent to single cell approaches that are dependent on fresh tissues. In summary, this thesis contributed to the characterization of endometrial stem cell biology and identified a progenitor cell population that might be related to the decidua remodelling during pregnancy

Impact of sitagliptin on endometrial mesenchymal stem-like progenitor cells : a randomised, double-blind placebo-controlled feasibility trial

Background: Recurrent pregnancy loss (RPL) is associated with the loss of endometrial mesenchymal stem-like progenitor cells (eMSC). DPP4 inhibitors may increase homing and engraftment of bone marrow-derived cells to sites of tissue injury. Here, we evaluated the effect of the DPP4 inhibitor sitagliptin on eMSC in women with RPL, determined the impact on endometrial decidualization, and assessed the feasibility of a full-scale clinical trial. Methods: A double-blind, randomised, placebo-controlled feasibility trial on women aged 18 to 42 years with a history of 3 or more miscarriages, regular menstrual cycles, and no contraindications to sitagliptin. Thirty-eight subjects were randomised to either 100 mg sitagliptin daily for 3 consecutive cycles or identical placebo capsules. Computer generated, permuted block randomisation was used to allocate treatment packs. Colony forming unit (CFU) assays were used to quantify eMSC in midluteal endometrial biopsies. The primary outcome measure was CFU counts. Secondary outcome measures were endometrial thickness, study acceptability, and first pregnancy outcome within 12 months following the study. Tissue samples were subjected to explorative investigations. Findings: CFU counts following sitagliptin were higher compared to placebo only when adjusted for baseline CFU counts and age (RR: 1.52, 95% CI: 1.32–1.75, P<0.01). The change in CFU count was 1.68 in the sitagliptin group and 1.08 in the placebo group. Trial recruitment, acceptability, and drug compliance were high. There were no serious adverse events. Explorative investigations showed that sitagliptin inhibits the expression of DIO2, a marker gene of senescent decidual cells. Interpretation: Sitagliptin increases eMSCs and decreases decidual senescence. A large-scale clinical trial evaluating the impact of preconception sitagliptin treatment on pregnancy outcome in RPL is feasible and warranted. Funding: Tommy's Baby Charity. Clinical trial registration: EU Clinical Trials Register no. 2016-001120-54