ERb-dependent effects on uterine endothelial cells are cell specific and mediated via Sp1

General rights Copyright for the publications made accessible via the Edinburgh Research Explorer is retained by the author(s) and / or other copyright owners and it is a condition of accessing these publications that users recognise and abide by the legal requirements associated with these rights. Take down policy The University of Edinburgh has made every reasonable effort to ensure that Edinburgh Research Explorer content complies with UK legislation. If you believe that the public display of this file breaches copyright please contact [email protected] providing details, and we will remove access to the work immediately and investigate your claim. study question: What are the in vitro effects of estrogen receptor b (ERb) activation on the function of endothelial cells (ECs) from different vascular beds: human endometrial ECs (HEECs; endometrium), uterine myometrial microvascular ECs (UtMVECs; myometrium) and human umbilical vein ECs (HUVECs)? summary answer: Studies conducted in vitro demonstrate that the ERb agonist 2,3-bis(4-hydroxy-phenyl)-propionitrile (DPN) has EC type-specific effects on patterns of gene expression and network formation. Identification of a key role for the transcription factor Sp1 in ERb-dependent signaling in uterine ECs offers new insights into cell-specific molecular mechanisms of estrogen action in the human uterus. what is known already: Estrogens, acting via ERs (ERa and ERb), have important, body-wide impacts on the vasculature. The human uterus is an estrogen target organ, the endometrial lining of which exhibits physiological, cyclical angiogenesis. In fixed tissue sections, human endo-metrial ECs are immunopositive for ERb. study design, size, duration: Cells were treated with a vehicle control or the ERb agonist, DPN, for 2 h or 24 h (n ¼ 5) followed by gene expression analysis. Functional assays were analyzed after a 16 h incubation with ligand (n ¼ 5). participant/materials, setting, methods: Analysis of DPN-treated ECs using Taqman gene array cards focused on genes involved in angiogenesis and inflammation identified cell type-specific ERb-dependent changes in gene expression, with validation using qPCR and immunohistochemistry. Molecular mechanisms involved in ERb signaling were investigated using bioinformatics, reporter assays, immunopreci-pitation, siRNA and a specific inhibitor blocking Sp1-binding sites. The endometrium and myometrium from women with regular menses were used to validate the protein expression of candidate genes. main results and the role of chance: HEECs and UtMVECs were ERb+/ERa2. Treatment of ECs with DPN had opposite effects on network formation: a decrease in network formation in HEECs (P ≤ 0.001) but an increase in UtMVECs (P ≤ …