OR22-3 Uterine TGFβ Signaling Controls Endometrial Cell Homeostasis And Regeneration

The transforming growth factor beta (TGFβ) signaling pathway has critical roles in the transduction of paracrine and autocrine signals that control development, reproduction, and cancer. To study the tissue-specific contribution of TGFβ signaling in the endometrium, we developed mice with conditional deletion SMAD2 and SMAD3 in the endometrial epithelium (using Lactoferrin-cre, "Ltf-cre"). We identified that double conditional deletion of SMAD2 and SMAD3 in the endometrial epithelium resulted in endometrial hyperplasia by 12-weeks of age, with myometrial invasion, metastasis, and death by 6-9-months of age. Smad2/3-Ltf-cre mice also lost epithelial progesterone receptor expression in the endometrium by 12 weeks of age. To study the signaling mechanism that controlled the malignant transformation of the glandular epithelium in Smad2/3-Ltf-cre mice, we developed 3-dimensional (3D) endometrial organoid cultures from the uterine epithelial tissues of control and mutant mice. To test how suppression of TGFβ signaling affected endometrial regeneration, 3D endometrial organoids from WT or Smad2/3-Ltf-cre mice were grown in the presence or absence of the TGFβ receptor inhibitor, A83-01. Using these conditions, we identified that WT endometrial organoids cultured with the SB505124 inhibitor or from Smad2/3-Ltf-cre mice, developed abnormal morphology. Histological analysis of the endometrial organoids showed that organoids cultured with A83-01 or from Smad2/3-Ltf-cre mice, developed increased secretory-like cells with increased mucin-1 expression. RNA-sequencing of the endometrial organoids revealed that a large number of differentially expressed genes were conserved between the WT organoids cultured with A83-01 and those from Smad2/3-Ltf-cre mice. Gene ontology analysis of the differentially expressed genes indicated that pharmacological or genetic inhibition of TGFβ signaling, resulted in over-representation of gene families related to BMP/SMAD1/5 signaling and retinoic acid signaling, with decreased enrichment of WNT/β-catenin signaling. These studies indicate that TGFβ-mediated signals are critical to endometrial function and that perturbation of this balance results in endometrial cancer. Funding support NICHD grants R00-HD096057 (to DM) and R01-HD032067 (to MMM), and grants NGP10125 and 1016187 from the Burroughs Wellcome Fund (to DM). Presentation: Monday, June 13, 2022 11:30 a.m. - 11:45 a.m.