Endocrine Abstracts

White adipose tissue (WAT) is a dynamic and heterogeneous organ composed of different cell types involved in a wide array of biological processes. We know that women with polycystic ovary syndrome (PCOS) suffer from insulin resistance and type 2 diabetes which is associated with pathological white adipose tissue (WAT) function and expansion characterized by hypertrophic adipocytes, altered production and release of lipids and adipokines, and chronic low-grade tissue inflammation. We hypothesize that cell-type-specific WAT dysfunction contributes to insulin resistance and subsequent development of type 2 diabetes in women with PCOS and that treatment aimed at improving insulin sensitivity and reducing androgen excess has the potential to reverse these changes. Because our subcutaneous WAT are frozen and adipocytes are too large and fragile for traditional single-cell sequencing, in order to unravel the cellular complexity, we extracted single-nuclei (sn) from snap frozen WAT from controls (n=4) and from hyperandrogenic and insulin-resistant women with PCOS (n=10) at baseline and in PCOS after 16 weeks of metformin (n=6) or lifestyle management (n=3) for snRNA-sequencing using the 10x Genomics protocol. 200 million reads were sequenced from ~5,000 nuclei/sample. Pre-processing and quality control (QC) of the data was performed using CellRanger and Seurat 4.0 was used for downstream QC, filtering, integration, clustering and differential gene expression analyses.

Preliminary results: First we mapped all sequenced cells (n=109,739) from all subjects and identified four main canonical cell types based on the expression of known markers: 1) adipocytes (n=17,896); 2) adipose stem and progenitor cells (ASPCs) (n=12,438); 3) immune cells (n=15,925); and 4) vascular cells (n=20,486). Subclustering and differential gene expression within each subcluster, pathway- and functional analyses as well as defining the effect of metformin and lifestyle management are ongoing and will be presented.

Summary: These comprehensive analyses will significantly increase our understanding of the cellular complexity and heterogeneity in specific cell types underlying WAT dysfunction in PCOS, and define whether cell-type-specific molecular dysfunctions can be reversed by current first-line treatment.