Endometriosis is a chronic, painful condition with unknown etiology. A differential expression of microRNAs in the endometriotic tissues from women with endometriosis with pain compared to those without suggested a plausible role for miRNA or epigenetic mechanisms in the etiology of endometriotic pain. The peritoneal milieu is involved in maintenance of endometriotic lesion and nociception. We recently showed the mechanistic role for oxidized-lipoproteins (ox-LDLs) present in peritoneal fluid (PF) in endometriosis and pain. We explored the possibility of ox-LDLs modulating the expression of miRNAs in a manner similar to PF from women with endometriosis. Expression levels of miRNAs and their predicted nociceptive and inflammatory targets were determined in PF and ox-LDL treated human endometrial cell-lines. Samples from IRB-approved and consented patients with and without endometriosis or pain were used. These were compared to endometrial cell-lines treated with various forms of oxidized-lipoproteins. RNA (including miRNAs) were isolated from treated endometrial cells and expression levels were determined using commercial miRNome arrays. Cell lysates were used in immunoblotting for inflammatory proteins using a protein array. Twenty miRNAs including isoforms of miR-29, miR-181 and let-7 were mutually differentially expressed in cells treated with PF from endometriosis patients with pain and those treated with ox-LDL components. The ox-LDLs and endo-PF treatment also produced significant overexpression of microRNA predicted target genes nerve growth factor, interleukin-6 and prostaglandin E synthase and overexpression of their downstream protein targets Mip1α and MCP1. This study showed similarities between miRNA regulation in PF from endometriotic women and ox-LDLs present in abundance in the PF of these women. Key miRNAs responsible for targeting nociceptive and inflammatory molecules were downregulated in the presence of ox-LDLs and endo-PF, thus playing a role in the etiology of endometriotic pain. These redox-sensitive miRNAs can be of potential use as targets in the treatment of endometriosis-associated pain.

Activation of trimethylation of histone 3 lysine 27 (H3K27me3) by EZH2, a component of the Polycomb repressive complex 2 (PRC2), is suggested to play a role in endometriosis. However, the mechanism by which this complex is dysregulated in endometriosis is not completely understood. Here, using eutopic and ectopic tissues, as well as peritoneal fluid (PF) from IRB-approved and consented patients with and without endometriosis, the expression of PRC2 complex components, JARID2, miR-155 (known regulators of EZH2), and a key inflammatory modulator, FOXP3, was measured. A higher expression of EZH2, H3K27me3, JARID2, and FOXP3 as well as miR-155 was noted in both the patient tissues and in endometrial PF treated cells. Gain-or-loss of function of miR-155 showed an effect on the PRC2 complex but had little effect on JARID2 expression, suggesting alternate pathways. Chromatin immunoprecipitation followed by qPCR showed differential expression of PRC2 complex proteins and its associated binding partners in JARID2 vs. EZH2 pull down assays. In particular, endometriotic PF treatment increased the expression of PHF19 (p = 0.0474), a gene silencer and co-factor that promotes PRC2 interaction with its targets. Thus, these studies have identified the potential novel crosstalk between miR-155-PRC2 complex-JARID2 and PHF19 in endometriosis, providing an opportunity to test other epigenetic targets in endometriosis.

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