Obesity alters the endometrial transcriptome in a cell context-dependent manner

Obesity significantly impacts fertility, is positively correlated with endometrial cancer occurrence, and negatively correlated with endometriosis incidence. The endometrial epithelia often harbor disease driver-mutations, while the endometrial stroma are highly regulative of the neighboring endometrial epithelia. Two cohorts of CD-1 mice were fed high-fat diet or control diet ad libitum for 18 weeks, beginning at 8 weeks of age. Weight measurements were taken weekly and serum was collected bi-weekly. After 17 weeks, a glucose tolerance test was performed on all mice. After 18 weeks, uteri were collected. We profiled the transcriptomes of positively-enriched endometrial epithelia (cohorts 1 and 2) and macrophages (cohort 2), and epithelia- and macrophage-depleted stroma populations (cohort 2). Pieces of uterine tissue were saved for histological analysis. Purity of cell populations were confirmed by flow cytometry. RNA was isolated from each cell type and libraries were synthesized for RNA-seq. Mice fed a high-fat diet displayed significantly increased body mass compared to control diet mice after three weeks. High-fat diet mice developed glucose intolerance, hyperinsulinemia, and fatty liver. Obese mice displayed differential gene expression in the endometrial epithelia for 105 genes (FDR < 0.05), including factors related to innate immunity and leukocyte chemotaxis, such as calprotectin. The endometrial stroma of obese mice differentially expressed 141 genes (FDR < 0.05) including several factors related to circadian rhythm. Expression of many of these genes was significantly correlated with glucose tolerance or body mass (p < 0.05). We observed correlations between F4/80 macrophage marker, Cleaved Caspase 3 (CC3) apoptosis marker in the luminal epithelia and glucose intolerance. Among obese mice, we observed a subgroup with high CC3 staining in the luminal epithelium by immunohistochemistry. This subgroup displayed differential gene expression among all cell types, with pathways related to immune escape observed in epithelial and macrophage compartments, and pathways related to epithelial regulation observed in the stromal compartments. The endometrial macrophages of obese mice compared to control mice displayed differential gene expression of only one gene, although the abundance of F4/80+ macrophages was shown to be increased in the obese mouse uterus. There may be species-specific differences between humans and mice which limit interpretations. The results suggest an important role for differential response of both the epithelia and stroma in their response to obese phenotypes, while macrophages are dysregulated in the context of apoptotic epithelium. The obesity-related gene expression programs in cells found within the uterine microenvironment may influence the ability of the endometrium to function during pregnancy and contribute to endometrial cancer pathogenesis. Overall design: EPCAM+ epithelia, F4/80+ macrophages and VIM+ stroma were isolated from the uteri of mice following 18 weeks on high-fat diet or control diet. RNA was collected from cels and used for RNA-seq analysis.