Endocrine Abstracts

Somatic cells of the seminiferous epithelium, called Sertoli cells, play a key role in germ cell development and maintenance of proper course of spermatogenesis. Although androgens are considered the main regulators of Sertoli cell activity, recent studies indicate that their metabolites, estrogens, also influence Sertoli cell function. Estrogens, act mainly through nuclear estrogen receptors α and β (ERα, ERβ), however non-classical signaling via membrane G protein-coupled estrogen receptor 1 (GPER) was also confirmed. It is well established that direct interactions between the cells in the seminiferous epithelium, including the Notch pathway, are essential for undisrupted spermatogenesis. Notch pathway is activated by binding membranous ligands Delta-like (DLL) or Jagged (JAG) of one cell to surface Notch receptors of the neighboring cell. The aim of this study was to explore the role of estrogens and their receptors in the control of the expression of Notch pathway ligands in Sertoli cells. Experiments were performed on primary Sertoli cells cultures isolated from rat testis (PSC) and mouse Sertoli cell line (TM4). First, the effect of estrogenic stimulation or estrogen action inhibition were examined using 17β-estradiol or estrogen receptor antagonists (ICI 182.780; G-15), respectively. Next, to determine the precise role of each receptor, siRNA silencing was conducted to knockdown the expression of ERα, ERβ, or GPER. qRT-PCR, western blot and immunofluorescence were employed to analyze the expression of DLL1, DLL4 and JAG1. The expression of all studied ligands was increased after stimulation by 17β-estradiol. The increase of either JAG1 or DLL1 expression in estrogen-stimulated cells was inhibited only by ICI 182.780. Knockdown experiments revealed that ERα silencing entirely abolished the effect of 17β-estradiol on both JAG1 and DLL1 expression. Exposure to ICI 182.780 and G-15 decreased DLL4 protein expression in 17β-estradiol-stimulated Sertoli cells. Silencing experiments demonstrated that ERβ and GPER knockdown effectively blocked estrogen influence on DLL4 protein expression. In summary, our results indicate that the expression of Notch pathway ligands in Sertoli cells is regulated by estrogens. JAG1 and DLL1 expression is controlled mainly via ERα, while DLL4 expression is dependent on ERβ and GPER signaling. Thus, the cooperation between classical and non-classical estrogen signaling pathways may be important for the communication within the seminiferous epithelium via Notch signaling, and thereby for proper spermatogenesis. This study was supported by a grant N18/MNW/000022 (Jagiellonian University, Faculty of Biology). The conference attendance supported by “Jagiellonian Interdisciplinary PhD Programme” POWR.03.05.00-00-Z309/17-00 (The European Social Fund).