Poster Presentation 46th Lorne Genome Conference 2025

Long-read single-cell RNA sequencing of endometrial organoids derived from fertile and infertile women uncovers dysregulation of endometrial receptivity (#235)

Michaela Sacco 1 2 , Sefi Prawer 3 , Wei Zhou 1 2 , Changqing Wang 4 5 , Wan Tinn Teh 2 6 7 , Kate Stern 2 6 7 , Tarana Lucky 6 8 , Catarina Ang 2 6 , Matthew E Ritchie 4 5 , Michael Clark 3 , Evdokia Dimitriadis 1 2
  1. Gynaecology Research Centre, Royal Women's Hospital, Melbourne, Victoria, Australia
  2. Department of Obstetrics, Gynaecology and Newborn Health, The University of Melbourne, Melbourne, Victoria, Australia
  3. Department of Anatomy and Physiology, The University of Melbourne, Parkville, Victoria, Australia
  4. Department of Medical Biology, The University of Melbourne, Parkville, Victoria, Australia
  5. Epigenetics and Development Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, Australia
  6. Reproductive Services, Royal Women's Hospital, Melbourne, Victoria, Australia
  7. Melbourne IVF, Melbourne, Victoria, Australia
  8. Faculty of Medicine, Dentistry and Health Sciences, The University of Melbourne, Parkville, Victoria, Australia

The human endometrium undergoes remodelling throughout the menstrual cycle to become receptive to embryo implantation within a small time period called the window of implantation (WOI). The epithelial cells lining the endometrium are the point of contact during embryo implantation. Dysfunction in these cells produces poor endometrial receptivity leading to implantation failure and infertility; however, the causal factors are still poorly understood. Therefore, we aimed to determine gene expression in fertile and infertile endometrial epithelial organoids (EEOs) using long-read scRNA-seq to identify cell-type specific markers and changes associated with infertility.

We performed the first long-read scRNA-seq of EEOs from fertile (n=3) and primary unexplained infertile (n=3) endometrium modelling the WOI. Organoids were dissociated into single cells, processed using 10x Genomics and Nanopore protocols, and analyzed via FLAMES, a long-read single cell analysis pipeline. Further analyses were conducted in Seurat on a total of 7,923 cells.

Long-read scRNA-seq results identified five epithelial cell subtypes: unciliated, secretory, ciliated, pre-ciliated, and proliferative, classified according to endometrial marker genes. Comparison of pathway enrichment between fertile and infertile cells of each cluster identified dysregulated functions unique to cell subtypes that each impact implantation uniquely. Further gene analysis revealed 48 differentially expressed genes, each dysregulated in specific cell subtypes. This facilitated association of the dysregulated gene’s role within its respective cell and subsequent impacts on endometrial receptivity. More focused analysis of transcript data revealed isoform expression of fertility associated genes, including WDR61, and key intergenic non-coding regions regulating expression.

This study characterised endometrial epithelial cell subtype changes, exemplifying how dysregulated gene expression in different cell subtypes impact receptivity uniquely and highlighting the importance of non-coding regions in regulating said expression. Our study identified key pathways, genes, and transcripts dysregulated in infertile epithelial cell subtypes that present potential use as personalised biomarkers and treatment targets of infertility.