Cellular Phenotyping of Endometriosis at the Single Cell Level Highlights Complex Heterogeneity

Abstract

Study Objective To understand how endometriosis cells work and interact with the surrounding cells by looking at cellular components and biomarkers. Design In this DoD funded study, both eutopic and peritoneal ectopic tissue were collected from patients with stage III-IV endometriosis. An unbiased single cell transcriptomic (scRNAseq) approach was used to comprehensively identify all the cellular components of endometriosis lesions and their microenvironment. Setting Tissue samples were obtained in the operating room and brought to the Laboratory for its fresh processing RNA sequencing Patients or Participants Women between 18-50 who were preoperatively diagnosed with stage III or IV endometriosis were consented prior to their scheduled laparoscopic surgery. Tissue banked samples of eutopic endometrium from “non-endometriosis patients” were used for comparison. Interventions Peritoneal endometriosis lesions and eutopic endometrium were collected during surgery from patients with stage III and IV endometriosis. These tissue samples were then used to generate scRNAseq data using a droplet based RNAseq platform (10x Genomics). Matched eutopic endometrium was also analyzed for comparison. Measurements and Main Results Our unbiased single cell approach allowed us to capture multiple cell types from ectopic, adjacent ectopic, and eutopic endometrium. We identified the major cell types composing the ectopic endometrium and its microenvironment, as well as in the adjacent peritoneum. Cell types identified within the microenvironment included T cells, B cells, macrophages, endothelial, fibroblast, muscle cells and pericytes. We identified endometrial epithelial cells, in both ectopic and matched eutopic tissues, and their single-cell differential expression analysis revealed substantial differences in their gene expression and main differences emerged from the macrophage populations. Conclusion Precise determination of the heterogeneous cellular composition of endometriosis lesion allows for characterization of important key players, such as epithelial cells and macrophages among others, for lesion formation and evolution. With this detailed approach, we hope to establish a list of potential biomarkers, a much needed effort for endometriosis diagnosis and treatment.