ODP403 Diversity of Estrogen-Responsive Genes in Endometriotic Lesions

BACKGROUND: Endometriosis has been accepted as an estrogen-dependent disease for more than two decades. So far, estrogen modulators that targeting estrogen depletion in endometriotic lesions have been expected as the first option of treatments to improve the symptoms associated with endometriosis. However, the limits of treatment are becoming apparent. OBJECTIVE: As the background of limits, we hypothesized the heterogeneity of genetic and/or epigenetic environments in endometriotic lesions (1). To test our hypothesis, we examined the expression profile of estrogen-responsive genes using endometriotic stromal cells as a model. Patients: Institutional Review Boards approved this project. We obtained informed consent from all patients. The chocolate cyst lining in the ovaries of patients with endometriosis was the source of endometriotic tissue. As a control, the eutopic endometrial tissues were obtained from uteri of premenopausal women who had uterine leiomyoma. METHODS: Stromal cells were prepared from endometriotic and endometrial tissues. ESR1 and ESR2 expression were evaluated using RT-PCR (2). For RNA-seq analysis of estrogen-responsive genes, total cellular RNAs from endometriotic cells treated with or without 10nM estradiol for 8hr were prepared. Following the quality control (concentration>50ng/ul, RIN>9), RNA samples were used for polyA+ selected library preparation and the strand-specific RNA-seq (NovaSeq, DNBSEQ-G400, 20M paired- end (PE) reads and 2×150bp PE/sample). Estrogen-responsive genes were extracted using RNA-seq bioinformatics analysis. Genes with adjusted p-values < 0. 05 and absolute log2 fold changes > 1 were extracted as estrogen-responsive genes (DESeq2 BP). Principal component analysis (PCA) was used to visualize sample-to-sample distances. Differential gene GO enrichment analysis was performed (GOseq. v1.34.1). RESULTS: 1) wild type ERα mRNA expression in endometriotic cells was one-tenth of that in endometrial cells. wild type ERβ1 and the splice variant ERβ2 mRNAs were expressed in endometriotic cells at a comparable level of the ERα. 2) PCA suggested the presence of at least two cell types in endometriotic cells. 3) Cluster analysis of gene expression suggested the presence of 2 cell types: endometriotic cells with or without GATA6 expression (3). 4) Estrogen-responsive genes were significantly different among the cells examined. 4) Differential gene GO enrichment analysis suggested the diversity of gene expression profiles depending on the cells. CONCLUSION: In the hope of overcoming the limits of endocrine-targeted therapy in endometriosis, we examined the estrogen-responsive genes in endometriotic cells. The finding suggests the diversity of estrogen-responsive genes in endometriotic lesions. References(1) Fertil Steril (2019) 111: 327–39. (2) Reprod Sci (2016) 23: 871-6. (3) Am J Reprod Immunol (2019) 81: e13078. Presentation: No date and time listed