Estrogen- and Progesterone (P4)-Mediated Epigenetic Modifications of Endometrial Stromal Cells (EnSCs) and/or Mesenchymal Stem/Stromal Cells (MSCs) in the Etiopathogenesis of Endometriosis

AbstractEndometriosis is a common chronic inflammatory condition in which endometrial tissue appears outside the uterine cavity. Because ectopic endometriosis cells express both estrogen and progesterone (P4) receptors, they grow and undergo cyclic proliferation and breakdown similar to the endometrium. This debilitating gynecological disease affects up to 15% of reproductive aged women. Despite many years of research, the etiopathogenesis of endometrial lesions remains unclear. Retrograde transport of the viable menstrual endometrial cells with retained ability for attachment within the pelvic cavity, proliferation, differentiation and subsequent invasion into the surrounding tissue constitutes the rationale for widely accepted implantation theory. Accordingly, the most abundant cells in the endometrium are endometrial stromal cells (EnSCs). These cells constitute a particular population with clonogenic activity that resembles properties of mesenchymal stem/stromal cells (MSCs). Thus, a significant role of stem cell-based dysfunction in formation of the initial endometrial lesions is suspected. There is increasing evidence that the role of epigenetic mechanisms and processes in endometriosis have been underestimated. The importance of excess estrogen exposure and P4 resistance in epigenetic homeostasis failure in the endometrial/endometriotic tissue are crucial. Epigenetic alterations regarding transcription factors of estrogen and P4 signaling pathways in MSCs are robust in endometriotic tissue. Thus, perspectives for the future may include MSCs and EnSCs as the targets of epigenetic therapies in the prevention and treatment of endometriosis. Here, we reviewed the current known changes in the epigenetic background of EnSCs and MSCs due to estrogen/P4 imbalances in the context of etiopathogenesis of endometriosis.

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Raf-1, a Potential Therapeutic Target, Mediates Early Steps in Endometriosis Lesion Development by Endometrial Epithelial and Stromal Cells

Endocrinology ◽

10.1210/en.2011-1879 ◽

2012 ◽

Vol 153 (8) ◽

pp. 3911-3921 ◽

Cited By ~ 6

Author(s):

Elizabeth M. De La Garza ◽

Peter A. Binkley ◽

Manonmani Ganapathy ◽

Naveen K. Krishnegowda ◽

Rajeshwar R. Tekmal ◽

...

Keyword(s):

Growth Factor ◽

Epidermal Growth Factor ◽

Stromal Cells ◽

Pelvic Cavity ◽

Lesion Formation ◽

Endometrial Stromal Cells ◽

Peritoneal Mesothelial Cells ◽

Endometrial Cells ◽

Retrograde Menstruation ◽

Epidermal Growth

Endometriosis is a hormone-sensitive gynecological disorder characterized by the benign growth of endometrial-like tissue in the pelvic cavity. Endometriotic lesions composed of endometrial stromal cells (ESC) and glandular epithelial cells (EEC) are thought to arise from menstrual endometrial tissue reaching the pelvic cavity via retrograde menstruation. The cause of endometriotic lesion formation is still not clear. Recent evidence suggest that cytokines may play a role in the early development of endometriosis lesions. Because cytokines and growth factors signal via the v-raf-1 murine leukemia viral oncogene homolog 1 (Raf-1) kinase pathway, we have examined the role of Raf-1 in early steps of endometriosis lesion formation, specifically attachment of endometrial cells to peritoneal mesothelial cells (PMC) and invasion of endometrial cells through PMC (trans-mesothelial invasion). Raf-1 antagonist GW5074 decreased attachment to PMC and trans-mesothelial invasion by primary EEC and ESC. Raf-1 also mediated TGFβ-induced trans-mesothelial invasion by the established, low-invasive EEC line EM42. TGFβ treatment of EEC resulted in Raf-1 phosphorylation at S338 and phosphorylation of ERK, suggesting that TGFβ activates Raf-1 signaling in these cells. GW5074 had little effect on ESC proliferation but inhibited EEC growth significantly under reduced serum conditions. Antagonizing Raf-1 activity and expression via GW5074 and specific Raf-1 small interfering RNA, respectively, did not alter EEC resistance to growth inhibition by TGFβ. Raf-1 inhibition blocked induction of EEC growth by epidermal growth factor. Our data suggest that Raf-1 may mediate pathologic steps involved in early endometriosis lesion formation and may be a mediator of TGFβ and epidermal growth factor actions in endometriosis.

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Endometrial Stromal Cells Circulate in the Bloodstream of Women with Endometriosis: A Pilot Study

International Journal of Molecular Sciences ◽

10.3390/ijms20153740 ◽

2019 ◽

Vol 20 (15) ◽

pp. 3740 ◽

Cited By ~ 3

Author(s):

Júlia Vallvé-Juanico ◽

Carlos López-Gil ◽

Agustín Ballesteros ◽

Xavier Santamaria

Keyword(s):

Pilot Study ◽

Epithelial Cells ◽

Stromal Cells ◽

Diagnostic Tools ◽

Pelvic Cavity ◽

Endometrial Stromal Cells ◽

Endometrial Cells ◽

Retrograde Menstruation ◽

The Brain ◽

Endometrial Epithelial Cells

Endometriosis is characterized by the presence of endometrial tissue outside the uterus. While endometriotic tissue is commonly localized in the pelvic cavity, it can also be found in distant sites, including the brain. The origin and pathophysiology of tissue migration is poorly understood; retrograde menstruation is thought to be the cause, although the presence of endometrium at distant sites is not explained by this hypothesis. To determine whether dissemination occurs via the bloodstream in women with endometriosis, we analyzed circulating blood for the presence of endometrial cells. Circulating endometrial stromal cells were identified only in women with endometriosis but not in controls, while endometrial epithelial cells were not identified in the circulation of either group. Our results support the hypothesis that endometrial stromal cells may migrate through circulation and promote the pathophysiology of endometriosis. The detection of these cells in circulation creates avenues for the development of less invasive diagnostic tools for the disease, and opens possibilities for further study of the origin of endometriosis.

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Activation of Blood Vessel Development in Endometrial Stromal Cells In Vitro Cocultured with Human Peri-Implantation Embryos Revealed by Single-Cell RNA-Seq

Life ◽

10.3390/life11050367 ◽

2021 ◽

Vol 11 (5) ◽

pp. 367

Author(s):

Bo Lv ◽

Xiaoyu Xu ◽

Xunyi Zhang ◽

Lingbin Qi ◽

Wen He ◽

...

Keyword(s):

Blood Vessel ◽

Single Cell ◽

Stromal Cells ◽

Differentially Expressed Gene ◽

Trophoblast Cells ◽

Endometrial Stromal Cells ◽

Endometrial Cells ◽

Blood Vessel Development ◽

Vessel Development

In humans, the maternal endometrium participates in the physical and physiological interaction with the blastocyst to begin implantation. A bidirectional crosstalk is critical for normal implantation and then a successful pregnancy. While several studies have used animal models or cell lines to study this step, little knowledge was acquired to address the role of endometrial cells in humans. Here, we analyzed single-cell sequencing data from a previous study including 24 non-coculture endometrial stromal cells (EmSCs) and 57 EmSCs after coculture with embryos. We further explored the transcriptomic changes in EmSCs and their interactions with trophoblast cells after coculture. Differentially expressed gene (DEG) analysis showed 1783 upregulated genes and 569 downregulated genes in the cocultured embryos. Weight gene coexpression network and gene ontology analysis of these DEGs showed a higher expression of RAMP1, LTBP1, and LRP1 in EmSCs after coculture, indicating the enrichment of biological processes in blood vessel development and female pregnancy. These data imply that EmSCs start blood vessel development at the implantation stage. Compared with endometrium data in vivo at the implantation window, key pathways including epithelial cell development and oxygen response were involved at this stage. Further analysis using CellphoneDB shed light on the interactions between EmSCs and embryonic trophoblasts, suggesting the important role of integrins and fibroblast growth factor pathways during implantation. Taken together, our work reveals the synchronization signaling and pathways happening at the implantation stage involving the acquisition of receptivity in EmSCs and the interaction between EmSCs and trophoblast cells.

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LIM Kinase 1 Mediates Estradiol Effects on the Phosphorylation of Cofilin1 in Eutopic Endometrial Stromal Cells During the Invasion and Proliferation of Endometriosis

Reproductive Sciences ◽

10.1177/1933719119828076 ◽

2019 ◽

Vol 26 (11) ◽

pp. 1499-1505 ◽

Cited By ~ 3

Author(s):

Jing Liu ◽

Zhifang Zhang ◽

Jiamei Liu ◽

Danbo Wang

Keyword(s):

Rna Interference ◽

Cell Invasion ◽

Stromal Cells ◽

Endometrial Stromal Cells ◽

Lim Kinase ◽

Endometrial Cells ◽

Lim Kinase 1 ◽

Gynecological Disease ◽

Cell Invasiveness

Endometriosis is an estrogen-dependent gynecological disease; however, the mechanism by which estradiol promotes the development of endometriosis, including invasion and proliferation, remains unclear. Estradiol is involved in cell invasion and proliferation by regulating the cytoskeleton. The abnormally high expression of cytoskeletal regulators (LIM kinase 1 [LIMK1] and cofilin1) is closely related to increased invasiveness and proliferation of eutopic endometrial stromal cells from endometriosis patients compared to normal eutopic endometrial cells. The aim of this study was to analyze the role of estradiol during invasion and proliferation through the LIMK1/cofilin1 pathway in the endometrium of women with endometriosis. To address this, primary eutopic endometrial stromal cells were isolated from the uteri of patients with endometriosis and cultured without estradiol. The phosphorylation of cofilin1 was analyzed by western blotting. Cell invasiveness and proliferation were evaluated following LIMK1 knockdown by RNA interference technology. We found that, before LIMK1silencing, the phosphorylation levels of cofilin1 and LIMK1 of eutopic endometrial stromal cells from endometriosis patients treated with estradiol were higher than cells not treated with estradiol ( P < .05 and P < .01, respectively). The total levels of cofilin1 and LIMK1 protein did not change ( P > .05 and P > .05, respectively). After LIMK1 silencing, the phosphorylation of cofilin1 by estradiol was significantly reduced, and invasiveness and proliferation were clearly and concurrently decreased ( P < .05 and P < .05, respectively). Thus, the phosphorylation of cofilin1 by estradiol is mediated by LIMK1, and estradiol is involved in regulating cell invasion and proliferation in endometriotic patients through the LIMK1/cofilin1 pathway.

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6-(7-nitro-2,1,3-benzoxadiazol-4-ylthio) hexanol ameliorates the progression of endometriosis by regulating GSTM4 expression to inhibit proliferation and induce apoptosis

10.21203/rs.3.rs-861230/v1 ◽

2021 ◽

Author(s):

Wei Liu ◽

Na Zhang ◽

Yanbo Du ◽

Xiaoqiang Liu ◽

Jinlong Ma ◽

...

Keyword(s):

Cell Proliferation ◽

Stromal Cells ◽

Therapeutic Potential ◽

Benign Disease ◽

Glutathione S Transferase ◽

Endometrial Stromal Cells ◽

Endometrial Cells ◽

Oxidative Balance ◽

Induced Apoptosis

Abstract Background: Endometriosis is a chronic disease associated with disorder of the oxidative balance and chronic inflammation. Although endometriosis is a benign disease, it has the characteristics properties similar to malignant cancer.Methods: The present study aim to investigate the role of glutathione S-transferase Mu class 4 (GSTM4), and tested if 6-(7-nitro-2,1,3-benzoxadiazol-4-ylthio) hexanol (NBDHEX) could regulate GSTM4 expression to affect cell proliferation, migration, invasion and apoptosis in endometriosis. Expression of GSTM4 was detected by immunohistochemistry in 15 cases of endometriosis patients and compared with 15 healthy controls. Primary endometrial cells were analyzed by western blotting (WB) to determine expression of GSTM4, PCNA, MMP-9, Survivin, Bcl-xl, Bax, Keap1 and Nrf2. CCK8 and transwell assays were used to study the effects of GSTM4 and NBEHEX on endometrial cells. The effect on apoptosis was analysised by flow cytometry. Results: The expression of GSTM4 was significantly increased in endometriosis than those from controls (p<0.01). The results suggested that NBDHEX negatively regulates GSTM4 expression, induces cell proliferation, migration, invasion, and promotes cell apoptosis. NBDHEX decreased the expression of GSTM4 (p<0.05), PCNA (p<0.05), MMP-9 (p<0.01), Survivin (p<0.05) and Bcl-xl (p<0.05) , along with increased expression of Bax (p<0.05). The results also showed that NBDHEX decreased the expression of Nrf2 (p<0.05), but had no effect on the expression of Keap1(p>0.05). After transfection with si-GSTM4, the protein level was down-regulated by nearly 70% (p<0.05). Silencing of GSTM4 depressed the proliferation, migration, invasion and gene expression of endometrial stromal cells in patients with endometriosis and controls. Knockdown of GSTM4 interacting with Nrf2 induced apoptosis by decreasing the expression of Survivin (p<0.05), Bcl-xl (p<0.05) and increasing the expression of Bax (p<0.05) , but it did not affect the expression of Keap1(p>0.05) in endometriosis and controls. Conclusions: Inhibition of GSTM4 by NBDHEX suppresses the cell viability growth, migration, invasion and interact with Nrf2 to induce apoptosis, but has no effect on the expression of Keap1 in endometriosis. The use of siRNA to knockdown GSTM4 more accurately confirmed its ability to ameliorate the progression of endometriosis. NBDHEX may have therapeutic potential in the treatment of endometriosis.

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Increased Expression of Retinol-Binding Protein 4 in Ovarian Endometrioma and Its Possible Role in the Pathogenesis of Endometriosis

International Journal of Molecular Sciences ◽

10.3390/ijms22115827 ◽

2021 ◽

Vol 22 (11) ◽

pp. 5827

Author(s):

Jae Chul Lee ◽

Sung Hoon Kim ◽

Young Sang Oh ◽

Ju Hee Kim ◽

Sa Ra Lee ◽

...

Keyword(s):

Retinol Binding Protein ◽

Endometrial Stromal Cells ◽

Endometrial Cells ◽

Retinol Binding Protein 4 ◽

Ovarian Endometrioma ◽

Proliferation And Differentiation ◽

In Vitro Effects ◽

Ovarian Endometriomas

Although endometriosis is a benign disease characterized by the presence of endometrial tissues outside the uterus, ectopic endometrial cells can exhibit malignant biological behaviors. Retinol-binding protein4 (RBP4) is a novel adipocyte-derived cytokine, which has important roles in regulating insulin sensitivity and energy metabolism. RBP4 is a potent modulator of gene transcription, and acts by directly controlling cell growth, invasiveness, proliferation and differentiation. Here, we evaluated the possible role of RBP4 in the pathogenesis of endometriosis. We compared the levels of RBP4 in the tissues and peritoneal fluid (PF) of women with and without endometriosis and evaluated the in vitro effects of RBP4 on the viability, invasiveness, and proliferation of endometrial stromal cells (ESCs). RBP4 levels were significantly higher in the PF of the women in the endometriosis group than in the controls. RBP4 immunoreactivity was significantly higher in the ovarian endometriomas of women with advanced stage endometriosis than those of controls. In vitro treatment with human recombinant-RBP4 significantly increased the viability, bromodeoxyuridine expression, and invasiveness of ESCs. Transfection with RBP4 siRNA significantly reduced ESC viability and invasiveness. These findings suggest that RBP4 partakes in the pathogenesis of endometriosis by increasing the viability, proliferation and invasion of endometrial cells.

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Bovine herpesvirus 4 is tropic for bovine endometrial cells and modulates endocrine function

Reproduction ◽

10.1530/rep-07-0065 ◽

2007 ◽

Vol 134 (1) ◽

pp. 183-197 ◽

Cited By ~ 57

Author(s):

Gaetano Donofrio ◽

Shan Herath ◽

Chiara Sartori ◽

Sandro Cavirani ◽

Cesidio Filippo Flammini ◽

...

Keyword(s):

Epithelial Cells ◽

Stromal Cells ◽

Bovine Herpesvirus ◽

Endocrine Function ◽

Uterine Disease ◽

Endometrial Stromal Cells ◽

Persistently Infected ◽

Endometrial Cells ◽

Bovine Herpesvirus 4 ◽

Herpesvirus 4

Bovinepostpartumuterine disease, metritis, affects about 40% of animals and is widely considered to have a bacterial aetiology. Although the γ-herpesvirus bovine herpesvirus 4 (BoHV-4) has been isolated from several outbreaks of metritis or abortion, the role of viruses in endometrial pathology and the mechanisms of viral infection of uterine cells are often ignored. The objectives of the present study were to explore the interaction, tropism and outcomes of BoHV-4 challenge of endometrial stromal and epithelial cells. Endometrial stromal and epithelial cells were purified and infected with a recombinant BoHV-4 carrying an enhanced green fluorescent protein (EGFP) expression cassette to monitor the establishment of infection. BoHV-4 efficiently infected both stromal and epithelial cells, causing a strong non-apoptotic cytopathic effect, associated with robust viral replication. The crucial step for the BoHV-4 endometriotropism appeared to be after viral entry as there was enhanced transactivation of the BoHV-4 immediate early 2 gene promoter following transient transfection into the endometrial cells. Infection with BoHV-4 increased cyclooxygenase 2 protein expression and prostaglandin estradiol secretion in endometrial stromal cells, but not epithelial cells. Bovine macrophages are persistently infected with BoHV-4, and co-culture with endometrial stromal cells reactivated BoHV-4 replication in the persistently infected macrophages, suggesting a symbiotic relationship between the cells and virus. In conclusion, the present study provides evidence of cellular and molecular mechanisms, supporting the concept that BoHV-4 is a pathogen associated with uterine disease.

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The Role of the FOXO1/β2-AR/p-NF-κB p65 Pathway in the Development of Endometrial Stromal Cells in Pregnant Mice under Restraint Stress

International Journal of Molecular Sciences ◽

10.3390/ijms22031478 ◽

2021 ◽

Vol 22 (3) ◽

pp. 1478

Author(s):

Jiayin Lu ◽

Yaoxing Chen ◽

Zixu Wang ◽

Jing Cao ◽

Yulan Dong

Keyword(s):

Oxidative Stress ◽

Stromal Cells ◽

Restraint Stress ◽

Target Genes ◽

Mrna Levels ◽

Endometrial Stromal Cells ◽

Protein Levels ◽

Pregnant Mice

Restraint stress causes various maternal diseases during pregnancy. β2-Adrenergic receptor (β2-AR) and Forkhead transcription factor class O 1 (FOXO1) are critical factors not only in stress, but also in reproduction. However, the role of FOXO1 in restraint stress, causing changes in the β2-AR pathway in pregnant mice, has been unclear. The aim of this research was to investigate the β2-AR pathway of restraint stress and its impact on the oxidative stress of the maternal uterus. In the study, maternal mice were treated with restraint stress by being restrained in a transparent and ventilated device before sacrifice on Pregnancy Day 5 (P5), Pregnancy Day 10 (P10), Pregnancy Day 15 (P15), and Pregnancy Day 20 (P20) as well as on Non-Pregnancy Day 5 (NP5). Restraint stress augmented blood corticosterone (CORT), norepinephrine (NE), and blood glucose levels, while oestradiol (E2) levels decreased. Moreover, restraint stress increased the mRNA levels of the FOXO family, β2-AR, and even the protein levels of FOXO1 and β2-AR in the uterus and ovaries. Furthermore, restraint stress increased uterine oxidative stress level. In vitro, the protein levels of FOXO1 were also obviously increased when β2-AR was activated in endometrial stromal cells (ESCs). In addition, phosphorylated-nuclear factor kappa-B p65 (p-NF-κB p65) and its target genes decreased significantly when FOXO1 was inhibited. Overall, it can be said that the β2-AR/FOXO1/p-NF-κB p65 pathway was activated when pregnant mice were under restraint stress. This study provides a scientific basis for the origin of psychological stress in pregnant women.

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The scattered puzzle effect: implantation disorders in chronic endometritis

GYNECOLOGY ◽

10.26442/20795696.2020.6.200493 ◽

2020 ◽

Vol 22 (6) ◽

pp. 93-100

Author(s):

Victor E. Radzinsky ◽

Mekan R. Orazov ◽

Liliia R. Toktar ◽

Liudmila M. Mihaleva ◽

Pavel A. Semenov ◽

...

Keyword(s):

Uterine Cavity ◽

Special Role ◽

Inadequate Response ◽

Chronic Endometritis ◽

Vitro Fertilization ◽

Endometrial Cells ◽

Adverse Pregnancy ◽

Relationship Of

Chronic endometritis (CE) is defined as a state of inflammation localized in the endometrium, accompanied by edema, dissociated maturation of epithelial cells and fibroblasts, increased stromal density and the presence of plasma cell infiltrate in it. The connection between chronic inflammation in the endometrium and infertility deserves special attention. Inadequate response of immunocompetent endometrial cells, including impaired synthesis of proinflammatory cytokines, dysreceptiveness, disorders of proliferation and differentiation processes are the main links in the formation of infertility in patients with CE. Despite the fact that the presence of a normocenosis of the uterine cavity today is not in doubt this is a physiological norm, persistent bacterial infection of the endometrium is still called the main etiopathogenetic factor of CE and, therefore, the main point of application of therapeutic agents. Nevertheless, a number of works have emphasized the special role of not bacterial, but viral etiology of endometritis, especially in the context of infertility developing against this background. It seems that the role of viral endometrial infection in adverse pregnancy outcomes and in vitro fertilization programs is underestimated. Further research is needed to clarify the relationship of viral infection as a trigger of implantation failure in infertile women with CE.

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