Effects of estradiol, progesterone or cAMP on expression of PGRMC1 and progesterone receptor in a xenograft model of human endometrium and in endometrial cell culture.

Estradiol and progesterone are key regulators of the menstrual cycle. In the human endometrium, progesterone induces morphological changes required for blastocyst implantation. Dysregulated response to progesterone can lead to endometrial pathologies including uterine bleeding and endometriosis. Besides the canonical nuclear progesterone receptor (encoded by the PGR gene), alternative response pathways include Progesterone Receptor Membrane Component 1 (PGRMC1), suspected to be involved in pathogenesis of endometrial diseases. We previously reported the spatiotemporal profile of PGRMC1 expression in the human endometrium along the menstrual cycle, highlighting progressive increase and decrease during the proliferative and secretory phases, respectively. Here we directly addressed its regulation by estradiol and progesterone, with systematic comparison with regulation of PGR expression. We found a direct correlation between expression of both genes during the proliferative and secretory phases in the cycling endometrium, but not during the menstrual phase. In a xenograft model mimicking the cycle phases, estradiol significantly increased and progesterone significantly decreased PGR expression but changes were not significant for PGRMC1. Finally, we did not find any significant effect of the ovarian steroids on expression of PGR or PGRMC1 in primary culture of endometrial stromal cells, except for a small increase in PGR expression by estradiol. Altogether, our experiments do not allow a major advance in our understanding of the mechanisms of cyclic variation of PGRMC1 expression, in particular regarding potential regulation by the ovarian steroids.

Spatiotemporal expression pattern of Progesterone Receptor Component (PGRMC) 1 in endometrium from patients with or without endometriosis or adenomyosis.

The endometrium plays a crucial role in reproduction and, in humans, is cyclically remodeled under hormonal control. Estradiol favors tissue proliferation whereas progesterone inhibits tissue growth and induces morphological changes. Endometriosis is often associated with fertility issues and with exacerbated estrogen and reduced progesterone concentration or response in the eutopic endometrium. However, underlying mechanisms remain unclear. Progesterone Receptor Membrane Component (PGRMC) 1 is a protein able to modulate progesterone response and its murine knockout reduced fertility. However, the precise spatiotemporal pattern of PGRMC1 expression in the human endometrium is still poorly characterized. We investigated variations of eutopic endometrial PGRMC1 expression by combining RT-qPCR, immunofluorescence and in situ hybridization. We found that PGRMC1 expression progressively increases during the proliferative phase and decreases during the secretory phase. However, immunolabeling and identification of mRNA-containing cells were regularly heterogeneous in samples, according to tissue depth, with a gradient extending from the surface epithelium towards the basalis. There was no significant difference in PGRMC1 mRNA amounts between patients with or without endometriosis or adenomyosis, for any phase of the menstrual cycle, but cells with strong or moderate PGRMC1 immunolabeling were reduced during the proliferative phase in endometriotic patients. In conclusion, although the cyclical variation of PGRMC1 expression globally follows fluctuation of ovarian steroids, further work is required to precisely characterize hormonal control and identify the additional levels of regulation responsible for local adjustment of PGRMC1 concentration. This is particularly important in the light of recent studies emphasizing the correlation between adequate PGRMC1 amounts and fertility.

AG-205 Upregulates Enzymes Involved in Cholesterol Biosynthesis and Steroidogenesis in Human Endometrial Cells Independently of PGRMC1 and Related MAPR Proteins.

An inappropriate response to progestogens in the human endometrium can result in fertility issues and jeopardize progestin-based treatments against pathologies such as endometriosis. PGRMC1 can mediate progesterone response in the breast and ovaries but its endometrial functions remain unknown. AG-205 is an alleged PGRMC1 inhibitor but its specificity was recently questioned. We added AG-205 in the cultures of two endometrial cell lines and performed a transcriptomic comparison. AG-205 significantly increased expression of genes coding enzymes of the cholesterol biosynthetic pathway or of steroidogenesis. However, these observations were not reproduced with cells transfected with siRNA against PGRMC1 or its related proteins (MAPRs). Furthermore, AG-205 retained its ability to increase expression of selected target genes even when expression of PGRMC1 or all MAPRs was concomitantly downregulated, indicating that neither PGRMC1 nor any MAPR is required to mediate AG-205 effect. In conclusion, although AG-205 has attractive effects encouraging its use to develop therapeutic strategies, for instance against breast cancer, our study delivers two important warning messages. First, AG-205 is not specific for PGRMC1 or other MAPRs and its mechanisms of action remain unclear. Second, due to its effects on genes involved in steroidogenesis, its use may increase the risk for endometrial pathologies resulting from imbalanced hormones concentrations.