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Background: Melatonin is a potential therapeutic agent for endometriosis, but its molecular mechanism is unclear. Here, we investigated the effect of melatonin on the epithelial-mesenchymal transition (EMT) in endometriotic endometrial epithelial cells and explored the pathway that might be involved., Methods: This hospital-based study included 60 women of reproductive age using the endometrium for immunohistochemistry, 6 women of reproductive age undergoing bilateral tubal ligation and 6 patients with endometriosis for isolation of endometrial epithelial cells or subsequent analysis, respectively. We examined the expression of Notch1/Numb signaling and EMT markers by immunohistochemistry analysis and western blot analysis, the invasion and migration of endometrial epithelial cells by transwell assays, and the cell proliferation by CCK8 assays., Results: Compared with normal endometrium, the endometriotic eutopic endometrium showed increased expression of Notch1, Slug, Snail, and N-cadherin, and decreased expression of E-cadherin and Numb. Melatonin or Notch inhibition by specific inhibitor blocked 17β-estradiol-induced cell proliferation, invasion, migration and EMT-related markers in both normal and endometriotic epithelial cells., Conclusions: Our data suggest that aberrant expression of Notch1/Numb signaling and the EMT is present in endometriotic endometrium. Melatonin may block 17β-estradiol-induced migration, invasion and EMT in normal and endometriotic epithelial cells by upregulating Numb expression and decreasing the activity of the Notch signaling pathway.

In previous studies, we found that endometriotic stromal cells lose the ability to regulate cell survival signaling in endometriotic epithelial cells. Here, we invested the effect of Metformin on the stromal-epithelial cells crosstalk in endometriosis and explored the pathway that might be involved. We found that ectopic endometriotic stromal cells (ESC) expressed and secreted higher Wnt2 protein compared with normal endometrial stromal cells (NSC). Conditioned medium (CM) from ESC supplemented with Wnt2 antibody significantly inhibited the growth of normal endometrial epithelial cells (NEC), while CM from ESC per se showed no significant effect on the growth of NEC. Metformin decreased the expression and secretion of Wnt2 in ESC. CM from Metformin-pretreated ESC significantly inhibited the growth of NEC. In conclusion, Wnt2/β-catenin signaling was involved in stromal-epithelial cells interaction in endometriosis. Metformin might regulate the stroma-epithelium communication via Wnt2-mediated signaling in endometriosis., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Objective: To explore the stroma-epithelium interactions in endometriosis and to identify the possible signalling pathways involved in this cross-talk., Design: Laboratory study via primary cultured endometrial stromal and epithelial cells., Setting: University Hospital., Patients: Fifteen patients with endometriosis confirmed by histopathology were recruited in the study, and 12 women free of endometriosis were used as control group., Intervention(s): Specific NFkappaB inhibitor 1-Pyrrolidinecarbodithioic acid ammonium salt (PDTC) was used in cell cultures., Main Outcome Measure(s): The expression and secretion of MMP-2, MMP-9, TIMP-1, TIMP-2 and the DNA-binding activity of NFkappaB in normal endometrial stromal cells or in co-cultures with normal or endometriotic epithelial cells from patients with endometriosis., Result(s): Endometrial epithelial cells induced MMP-9 and MMP-2 expression in normal stromal cells in vitro. In co-cultures with endometriotic epithelial cells, normal endometrial stromal cells expressed and secreted higher MMP-2 (p < 0.05) and MMP-9 (p < 0.05). Specific inhibition of NFkappaB pathway in stromal cells abolished this induction effect by epithelial cells., Conclusion(s): Endometriotic epithelial cells induce MMPs expression and secretion in normal endometrial stromal cells via an NFkappaB-dependent pathway in vitro. This cross-talk between epithelial cells and stromal cells may facilitate the implantation and extension of the ectopic foci and favour the development of the disease.

In normal endometrium, stromal factors regulate the growth of epithelial cells. However, epithelial cells in endometriotic lesions display increased proliferation and decreased apoptosis. This work tested the hypothesis that in endometriosis stromal cells lose the ability to regulate survival signaling and cell growth in epithelial cells. Primary normal, endometriotic eutopic and ectopic epithelial cells were cultured in the presence of medium conditioned by normal, eutopic and ectopic endometriotic endometrial stromal cells. Endometriotic epithelial cells showed higher Survivin expression than normal epithelial cells. Conditioned medium (CM) from normal or eutopic endometriotic stromal cells significantly inhibited the Survivin expression and AKt phosphorylation in normal or eutopic endometriotic epithelial cells. However, CM from ectopic endometriotic stromal cells did not have an inhibitory effect on normal or ectopic endometriotic epithelial cells. Inhibition of AKt phosphorylation and Survivin expression in normal or eutopic endometriotic epithelial cells in the presence of stromal factors from normal or eutopic endometriotic stromal cells was enhanced by progesterone, whereas progesterone had little effect in the presence of stromal factors from ectopic endometriotic stromal cells. The inability of ectopic endometriotic stromal cells to regulated PI3K/AKt/Survivin signaling and mediate the progesterone response in endometriotic epithelial cells may facilitate epithelial cell proliferation in endometriosis and promote the survival of endometriotic lesions.

The programmed death-1 (PD-1)/PD-L1 pathway plays important roles in immune responses and peripheral tolerance. Under pathological conditions, this pathway can suppress protective T cell responses and induce immune system disorders. Endometriosis (EM) is an estrogen-dependent, immune-associated disease. The expression of PD-1 and PD-L1 has been studied in a variety of cancers and autoimmune diseases. Few studies, however, have attempted to explore their expression and the possibility that they are regulated by estrogen in EM. Eutopic and ectopic endometria and blood samples were collected from 15 women with EM and 15 women without EM. PD-1/PD-L1 expression in endometrial tissues was detected using immunohistochemistry and western blot analysis. Their expression in blood was detected using flow cytometry. In addition, their expression was evaluated in endometrial cells after estrogen and cytokine treatment. We observed PD-1/PD-L1 expression in both eutopic and ectopic endometria, with elevated expression in EM. Their expression was also higher in CD4+/CD8+ T cells in blood from EM patients. In addition, treatment with 17β-estradiol upregulated PD-L1 expression in eutopic epithelial cells in EM. These data suggest that the PD-1/PD-L1 pathway may be involved in EM immune dysfunction and be regulated by estrogen. [ABSTRACT FROM AUTHOR]

Previous studies report that fibroblast growth factor 2 (FGF2) modulates Sproutys (SPRYs)/dual specificity phosphatase 6 (DUSP6)/extracellular signal-regulated kinase (ERK) signaling pathway in endometrial glandular epithelial cells. However, its role in endometriosis remains unclear. The expression patterns and localization of related proteins in endometrium patients' samples were determined using quantitative reverse transcription PCR, Western blotting, and immunohistochemistry, respectively. Human endometrial stromal cells (HESCs) were isolated and transfected with small interfering RNA (siRNA) targeting FGF2 (FGF2-siRNA). Cell viability was determined using 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. It was found that FGF2 mRNA and protein levels were increased in the ectopic endometrium, whilst the mRNA and protein levels of SPRYs/DUSP6/ERK signaling pathway related-genes were dysregulated. Spearman's rank correlation analysis revealed a negative correlation between FGF2 and SPRYs/DUSP6 signaling pathway-related proteins. In vitro study demonstrated that FGF2 silencing suppressed cell proliferation. Our results suggest that FGF2 upregulation might contribute to endometriosis via the regulation of the SPRYs/DUSP6/ERK signaling pathway. [ABSTRACT FROM AUTHOR]