Peritoneal Fluid Obtained from Women with Endometriosis Induces CYP19A1 Expression in Isolated Control Endometrial Cells.

Endometriosis patients commonly present altered volume and composition of peritoneal fluid (PF) and also, an endometrial estrogenic micro-environment as consequence of anomalous CYP19A1 expression. This gene is positively regulated by NR5A1 and negatively by NR2F. We studied the effect of PF obtained from women with endometriosis on CYP19A1 expression in endometrial epithelial cells in vitro (approved by Institutional Review Board). Eutopic endometrium was obtained during surgery and each patient signed a written informed consent. Culture of epithelial cells from eutopic endometrium of women with (n=12) and without (control; n=12) endometriosis were treated for 24 hours with: PF (0-1-10%, v/v, obtained from 10 women with endometriosis at proliferative phase of the menstrual cycle), charcoal adsorbed PF (10% v/v), cAMP (0.1 mmol/L) and/or indomethacin (0.1-10 μmol/L, prostaglandin synthesis inhibitor). CYP19A1, NR5A1, NR2F1 and NR2F2 mRNA and/or protein were studied by RT-PCR and immunoblot, respectively. Endometrial cells were transfected (GeneJammer) with pcDNA2NR5A1(kindly donated by Dr. S. Bulun) and incubated with and without 10%PF. In the absence of treatment, the mRNA and/or protein of NR5A1 and CYP19A1 were almost undetectable in control cells but they were 90 and 310% (mRNA) and 760 and 1400% (protein) higher in endometriosis cells (p<<0.05); NR2F1 and NR2F2 were slightly lower in endometriosis than in control cells. In control, PF increased mRNA and protein of CYP19A1 and NR5A1 in a dose-dependent manner (p<<0.01), cAMP increased both gene mRNAs (40 and 250%, respectively), effect enhanced by 10%PF (150 and 580%); this behaviour was not observed with 10%PF plus 10 mmol/L indomethacin or with charcoal adsorbed PF; PF did not modify NR2F1 and NR2F2 mRNAs. In endometriosis, the addition of PF did not affect the mRNA level of any molecule studied. CYP19A1 mRNA was increased 290% in control cells transfected with pcDNA2NR5A1 and the addition of 10%PF or cAMP enhanced it 836 and 440%, respectively (p<<0.01); indomethacin blocked almost completely these responses. The induction only in controls, and not in endometriosis cells, of NR5A1 and CYP19A1 (molecules strongly expressed in endometriosis), suggests a role of PF on implant maintenance and growth, involving probably prostaglandins, allowing the particular estrogenic microenvironment. (poster)