Macrophages inhibit and enhance endometriosis depending on their origin.

Macrophages are intimately involved in the pathophysiology of endometriosis, a chronic inflammatory disorder characterized by the growth of endometrial-like tissue (lesions) outside the uterus. By combining genetic and pharmacological monocyte and macrophage depletion strategies we determined the ontogeny and function of macrophages in a mouse model of induced endometriosis. We demonstrate that lesion-resident macrophages are derived from eutopic endometrial tissue, infiltrating large peritoneal macrophages (LpM) and monocytes. Furthermore, we found endometriosis to trigger continuous recruitment of monocytes and expansion of CCR2+ LpM. Depletion of eutopic endometrial macrophages results in smaller endometriosis lesions, whereas constitutive inhibition of monocyte recruitment significantly reduces peritoneal macrophage populations and increases the number of lesions. Reprogramming the ontogeny of peritoneal macrophages such that embryo-derived LpM are replaced by monocyte-derived LpM decreases the number of lesions that develop. We propose a putative model whereby endometrial macrophages are "proendometriosis" while newly recruited monocyte-derived macrophages, possibly in LpM form, are "antiendometriosis." These observations highlight the importance of monocyte-derived macrophages in limiting disease progression.

The role of small extracellular vesicle-miRNAs in endometriosis.

Endometriosis is defined by the presence of extrauterine endometrial-like tissue, which can cause pain and infertility in 10% of reproductive-age women. To date, the pathogenesis is poorly understood resulting in significant diagnostic delays and poor therapeutic outcomes in many women. Small extracellular vesicles (sEVs) (<200 nm) are cell-derived vesicles containing molecules that can influence gene expression and behaviour in target cells. One such cargo are microRNAs (miRNAs), which are short, non-coding RNAs mostly 19-25 nucleotides in length that regulate post-transcriptional gene expression. This mini-review focuses on the role of sEV-miRNAs, which are conceivably better biomarkers for endometriosis than free miRNAs, which reflect the true pathophysiological state in the body, as sEV-encapsulated miRNAs are protected from degradation compared to free miRNA and provide direct cell-to-cell communication via sEV surface proteins. sEV-miRNAs have been implicated in the immunomodulation of macrophages, the proliferation, migration and invasion of endometrial cells, and angiogenesis, all hallmarks of endometriosis. The diagnostic potential of sEV-miRNA was investigated in one study that reported the sensitivity and specificity of two sEV-miRNAs (hsa-miR-22-3p and hsa-miR-320a-3p) in distinguishing endometriosis from non-endometriosis cases. Only three studies have explored the therapeutic potential of sEV-miRNAs in vivo in mice-two looked into the role of sEV-hsa-miR-214-3p in decreasing fibrosis, and one investigated sEV-hsa-miR-30c-5p in suppressing the invasive and migratory potential of endometriotic lesions. While early results are encouraging, studies need to further address the potential influence of factors such as the menstrual cycle as well as the location and extent of endometriotic lesions on miRNA expression in sEVs. Given these findings, and extrapolating from other conditions such as cancer, diabetes, and pre-eclampsia, sEV-miRNAs could present an attractive and urgently needed future diagnostic and therapeutic target for millions of women suffering from endometriosis. However, research in this area is hampered by lack of adherence to the International Society for Extracellular Vesicles 2018 guideline in separating and characterising sEVs, as well as the World Endometriosis Research Foundation Endometriosis Phenome and Biobanking Harmonisation Project protocols.

Efficacy of niclosamide on the intra-abdominal inflammatory environment in endometriosis.

Endometriosis, a common gynecological disease, causes chronic pelvic pain and infertility in women of reproductive age. Due to the limited efficacy of current therapies, a critical need exists to develop new treatments for endometriosis. Inflammatory dysfunction, instigated by abnormal macrophage (MΦ) function, contributes to disease development and progression. However, the fundamental role of the heterogeneous population of peritoneal MΦ and their potential druggable functions is uncertain. Here we report that GATA6-expressing large peritoneal MΦ (LPM) were increased in the peritoneal cavity following lesion induction. This was associated with increased cytokine and chemokine secretion in the peritoneal fluid (PF), as well as MΦ infiltration, vascularization and innervation in endometriosis-like lesions (ELL). Niclosamide, an FDA-approved anti-helminthic drug, was effective in reducing LPM number, but not small peritoneal MΦ (SPM), in the PF. Niclosamide also inhibits aberrant inflammation in the PF, ELL, pelvic organs (uterus and vagina) and dorsal root ganglion (DRG), as well as MΦ infiltration, vascularization and innervation in the ELL. PF from ELL mice stimulated DRG outgrowth in vitro, whereas the PF from niclosamide-treated ELL mice lacked the strong stimulatory nerve growth response. These results suggest LPM induce aberrant inflammation in endometriosis promoting lesion progression and establishment of the inflammatory environment that sensitizes peripheral nociceptors in the lesions and other pelvic organs, leading to increased hyperalgesia. Our findings provide the rationale for targeting LPM and their functions with niclosamide and its efficacy in endometriosis as a new non-hormonal therapy to reduce aberrant inflammation which may ultimately diminish associated pain.

Repurposing dichloroacetate for the treatment of women with endometriosis.

Endometriosis is a chronic pain condition affecting ∼176 million women worldwide. It is defined by the presence of endometrium-like tissue (lesions) outside the uterus, most commonly on the pelvic peritoneum. There is no cure for endometriosis. All endometriosis drug approvals to date have been contraceptive, limiting their use in women of child-bearing age. We have shown that human peritoneal mesothelial cells (HPMCs) recovered from the pelvic peritoneum of women with endometriosis exhibit significantly higher glycolysis, lower mitochondrial respiration, decreased enzymatic activity of pyruvate dehydrogenase (PDH), and increased production of lactate compared to HPMCs from women without disease. Transforming growth factor-ß1 (TGF-ß1) is elevated in the peritoneal fluid from women with endometriosis, and exposure of HPMCs to TGF-ß1 exacerbates this abnormal phenotype. Treatment of endometriosis HPMCs with the pyruvate dehydrogenase kinase (PDK) inhibitor/PDH activator dichloroacetate (DCA) normalizes HPMC metabolism, reduces lactate secretion, and abrogates endometrial stromal cell proliferation in a coculture model. Oral DCA reduced peritoneal fluid lactate concentrations and endometriosis lesion size in a mouse model. These findings provide the rationale for targeting metabolic processes as a noncontraceptive treatment for women with endometriosis either as a primary nonhormonal treatment or to prevent recurrence after surgery.

Endometriosis-Associated Pain - Do Preclinical Rodent Models Provide a Good Platform for Translation?

Pelvic pain is a common symptom of endometriosis. Our understanding of its etiology remains incomplete and medical management is limited by poor translation from preclinical models to clinical trials. In this review, we briefly consider the evidence, or lack thereof, that different subtypes of lesion, extra-uterine bleeding, and neuropathic pathways add to the complex and heterogeneous pain experience of women with the condition. We summarize the studies in rodent models of endometriosis that have used behavioral endpoints (evoked and non-evoked) to explore mechanisms of endometriosis-associated pain. Lesion innervation, activation of nerves by pronociceptive molecules released by immune cells, and a role for estrogen in modulating hyperalgesia are key endometriosis-associated pain mechanisms replicated in preclinical rodent models. The presence of ectopic (full thickness uterus or endometrial) tissue may be associated with changes in the spinal cord and brain, which appear to model changes reported in patients. While preclinical models using rats and mice have yielded insights that appear relevant to mechanisms responsible for the development of endometriosis-associated pain, they are limited in scope. Specifically, most studies are based on models that only resulted in the formation of superficial lesions and use induced (evoked) behavioral 'pain' tests. We suggest that translation for patient benefit will be improved by new approaches including models of ovarian and deep infiltrating disease and measurement of spontaneous pain behaviors. Future studies must also capitalize on new advances in the wider field of pain medicine to identify more effective treatments for endometriosis-associated pain.

Macrophage-derived insulin-like growth factor-1 is a key neurotrophic and nerve-sensitizing factor in pain associated with endometriosis.

Endometriosis is a common incurable inflammatory disorder that is associated with debilitating pelvic pain in women. Macrophages are central to the pathophysiology of endometriosis: they dictate the growth and vascularization of endometriosis lesions and more recently have been shown to promote lesion innervation. The aim of this study was to determine the mechanistic role of macrophages in producing pain associated with endometriosis. Herein, we show that macrophage depletion in a mouse model of endometriosis can reverse abnormal changes in pain behavior. We identified that disease-modified macrophages exhibit increased expression of IGF-1 in an in vitro model of endometriosis-associated macrophages and confirmed expression by lesion-resident macrophages in mice and women. Concentrations of IGF-1 were elevated in peritoneal fluid from women with endometriosis and positively correlate with their pain scores. Mechanistically, we demonstrate that macrophage-derived IGF-1 promotes sprouting neurogenesis and nerve sensitization in vitro. Finally, we show that the Igf-1 receptor inhibitor linsitinib reverses the pain behavior observed in mice with endometriosis. Our data support a role for macrophage-derived IGF-1 as a key neurotrophic and sensitizing factor in endometriosis, and we propose that therapies that modify macrophage phenotype may be attractive therapeutic options for the treatment of women with endometriosis-associated pain.-Forster, R., Sarginson, A., Velichkova, A., Hogg, C., Dorning, A., Horne, A. W., Saunders, P. T. K., Greaves, E. Macrophage-derived insulin-like growth factor-1 is a key neurotrophic and nerve-sensitizing factor in pain associated with endometriosis.

Relevant human tissue resources and laboratory models for use in endometriosis research.

Endometriosis is characterized by the growth of endometrium-like tissue outside the uterus, most commonly on the pelvic peritoneum and ovaries. Although it may be asymptomatic in some women, in others it can cause debilitating pain, infertility or other symptoms including fatigue. Current research is directed both at understanding the complex etiology and pathophysiology of the disorder and at the development of new nonsurgical approaches to therapy that lack the unwanted side effects of current medical management. Tools for endometriosis research fall into two broad categories; patient-derived tissues, and fluids (and cells isolated from these sources) or models based on the use of cells or animals. In this review, we discuss the literature that has reported data from the use of these tools in endometriosis research and we highlight the strengths and weaknesses of each. Although many different models are reported in the literature, hypothesis-driven research will only be facilitated with careful experimental design and selection of the most appropriate human tissue from patients with and without endometriosis and combinations of physiologically relevant in vitro and in vivo laboratory models.

Estradiol is a critical mediator of macrophage-nerve cross talk in peritoneal endometriosis.

Endometriosis occurs in approximately 10% of women and is associated with persistent pelvic pain. It is defined by the presence of endometrial tissue (lesions) outside the uterus, most commonly on the peritoneum. Peripheral neuroinflammation, a process characterized by the infiltration of nerve fibers and macrophages into lesions, plays a pivotal role in endometriosis-associated pain. Our objective was to determine the role of estradiol (E2) in regulating the interaction between macrophages and nerves in peritoneal endometriosis. By using human tissues and a mouse model of endometriosis, we demonstrate that macrophages in lesions recovered from women and mice are immunopositive for estrogen receptor ß, with up to 20% being estrogen receptor α positive. In mice, treatment with E2 increased the number of macrophages in lesions as well as concentrations of mRNAs encoded by Csf1, Nt3, and the tyrosine kinase neurotrophin receptor, TrkB. By using in vitro models, we determined that the treatment of rat dorsal root ganglia neurons with E2 increased mRNA concentrations of the chemokine C-C motif ligand 2 that stimulated migration of colony-stimulating factor 1-differentiated macrophages. Conversely, incubation of colony-stimulating factor 1 macrophages with E2 increased concentrations of brain-derived neurotrophic factor and neurotrophin 3, which stimulated neurite outgrowth from ganglia explants. In summary, we demonstrate a key role for E2 in stimulating macrophage-nerve interactions, providing novel evidence that endometriosis is an estrogen-dependent neuroinflammatory disorder.

A novel mouse model of endometriosis mimics human phenotype and reveals insights into the inflammatory contribution of shed endometrium.

Endometriosis is an estrogen-dependent inflammatory disorder characterized by the presence of endometrial tissue outside the uterine cavity. Patients experience chronic pelvic pain and infertility, with the most likely origin of the tissue deposits (lesions) being endometrial fragments shed at menses. Menstruation is an inflammatory process associated with a dramatic increase in inflammatory mediators and tissue-resident immune cells. In the present study, we developed and validated a mouse model of endometriosis using syngeneic menstrual endometrial tissue introduced into the peritoneum of immunocompetent mice. We demonstrate the establishment of endometriotic lesions that exhibit similarities to those recovered from patients undergoing laparoscopy. Specifically, in both cases, lesions had epithelial (cytokeratin(+)) and stromal (vimentin/CD10(+)) cell compartments with a well-developed vasculature (CD31(+) endothelial cells). Expression of estrogen receptor ß was increased in lesions compared with the peritoneum or eutopic endometrium. By performing experiments using mice with green fluorescent protein-labeled macrophages (MacGreen) in reciprocal transfers with wild-type mice, we obtained evidence that macrophages present in the peritoneum and in menses endometrium can contribute to the inflammatory microenvironment of the lesions. In summary, we developed a mouse model of endometriosis that exhibits similarities to human peritoneal lesions with respect to estrogen receptor expression, inflammation, and macrophage infiltration, providing an opportunity for further studies and the possible identification of novel therapies for this perplexing disorder.

Cancer-associated variant expression and interaction of CIZ1 with cyclin A1 in differentiating male germ cells.

CIZ1 is a nuclear-matrix-associated DNA replication factor unique to higher eukaryotes, for which alternatively spliced isoforms have been associated with a range of disorders. In vitro, the CIZ1 N-terminus interacts with cyclin E and cyclin A at distinct sites, enabling functional cooperation with cyclin-A-Cdk2 to promote replication initiation. C-terminal sequences anchor CIZ1 to fixed sites on the nuclear matrix, imposing spatial constraint on cyclin-dependent kinase activity. Here we demonstrate that CIZ1 is predominantly expressed as a predicted full-length product throughout mouse development, consistent with a ubiquitous role in cell and tissue renewal. CIZ1 is expressed in proliferating stem cells of the testis, but is notably downregulated following commitment to differentiation. Significantly, CIZ1 is re-expressed at high levels in non-proliferative spermatocytes before meiotic division. Sequence analysis identifies at least seven alternatively spliced variants, including a dominant cancer-associated form and a set of novel isoforms. Furthermore, we show that in these post-replicative cells, CIZ1 interacts with germ-cell-specific cyclin A1, which has been implicated in the repair of DNA double-strand breaks. Consistent with this role, antibody depletion of CIZ1 reduces the capacity for testis extract to repair digested plasmid DNA in vitro. Together, the data imply post-replicative roles for CIZ1 in germ cell differentiation that might include meiotic recombination - a process intrinsic to genome stability and diversification.

Spatial Transcriptomic Analysis Identifies Epithelium-Macrophage Crosstalk in Endometriotic Lesions.

The mechanisms underlying the pathophysiology of endometriosis, characterized by the presence of endometrium-like tissue outside the uterus, remain poorly understood. This study aimed to identify cell type-specific gene expression changes in superficial peritoneal endometriotic lesions and elucidate the crosstalk among the stroma, epithelium, and macrophages compared to patient-matched eutopic endometrium. Surprisingly, comparison between lesions and eutopic endometrium revealed transcriptional similarities, indicating minimal alterations in the sub-epithelial stroma and epithelium of lesions. Spatial transcriptomics highlighted increased signaling between the lesion epithelium and macrophages, emphasizing the role of the epithelium in driving lesion inflammation. We propose that the superficial endometriotic lesion epithelium orchestrates inflammatory signaling and promotes a pro-repair phenotype in macrophages, providing a new role for Complement 3 in lesion pathobiology. This study underscores the significance of considering spatial context and cellular interactions in uncovering mechanisms governing disease in endometriotic lesions.

ERß-dependent effects on uterine endothelial cells are cell specific and mediated via Sp1.

STUDY QUESTION: What are the in vitro effects of estrogen receptor ß (ERß) activation on the function of endothelial cells (ECs) from different vascular beds: human endometrial ECs (HEECs; endometrium), uterine myometrial microvascular ECs (UtMVECs; myometrium) and human umbilical vein ECs (HUVECs)? SUMMARY ANSWER: Studies conducted in vitro demonstrate that the ERß agonist 2,3-bis(4-hydroxy-phenyl)-propionitrile (DPN) has EC type-specific effects on patterns of gene expression and network formation. Identification of a key role for the transcription factor Sp1 in ERß-dependent signaling in uterine ECs offers new insights into cell-specific molecular mechanisms of estrogen action in the human uterus. WHAT IS KNOWN ALREADY: Estrogens, acting via ERs (ERα and ERß), have important, body-wide impacts on the vasculature. The human uterus is an estrogen target organ, the endometrial lining of which exhibits physiological, cyclical angiogenesis. In fixed tissue sections, human endometrial ECs are immunopositive for ERß. STUDY DESIGN, SIZE, DURATION: Cells were treated with a vehicle control or the ERß agonist, DPN, for 2 h or 24 h (n = 5) followed by gene expression analysis. Functional assays were analyzed after a 16 h incubation with ligand (n = 5). PARTICIPANT/MATERIALS, SETTING, METHODS: Analysis of DPN-treated ECs using Taqman gene array cards focused on genes involved in angiogenesis and inflammation identified cell type-specific ERß-dependent changes in gene expression, with validation using qPCR and immunohistochemistry. Molecular mechanisms involved in ERß signaling were investigated using bioinformatics, reporter assays, immunoprecipitation, siRNA and a specific inhibitor blocking Sp1-binding sites. The endometrium and myometrium from women with regular menses were used to validate the protein expression of candidate genes. MAIN RESULTS AND THE ROLE OF CHANCE: HEECs and UtMVECs were ERß+/ERα-. Treatment of ECs with DPN had opposite effects on network formation: a decrease in network formation in HEECs (P ≤ 0.001) but an increase in UtMVECs (P ≤ 0.05). Genomic analysis identified opposite changes in ERß target gene expression with only three common transcripts (HEY1, ICAM1, CASP1) in all three ECs; a unique profile was observed for each. An important role for Sp1 was identified, consistent with the regulation of ERß target genes via association with the transcription factor ('tethered' mechanism). LIMITATIONS, REASONS FOR CAUTION: The study was mainly carried out in vitro using ECs of which one type was immortalized. Although the analysis of the protein expression of candidate genes was carried out using intact tissue samples from patients, investigations into in vivo angiogenesis were not carried out. WIDER IMPLICATIONS OF THE FINDINGS: These results have implications for our understanding of the mechanisms responsible for ERß-dependent changes in EC gene expression in hormone-dependent disorders.

Cyclin E is recruited to the nuclear matrix during differentiation, but is not recruited in cancer cells.

Cyclin E supports pre-replication complex (pre-RC) assembly, while cyclin A-associated kinase activates DNA synthesis. We show that cyclin E, but not A, is mounted upon the nuclear matrix in sub-nuclear foci in differentiated vertebrate cells, but not in undifferentiated cells or cancer cells. In murine embryonic stem cells, Xenopus embryos and human urothelial cells, cyclin E is recruited to the nuclear matrix as cells differentiate and this can be manipulated in vitro. This suggests that pre-RC assembly becomes spatially restricted as template usage is defined. Furthermore, failure to become restricted may contribute to the plasticity of cancer cells.

Macrophages and small extracellular vesicle mediated-intracellular communication in the peritoneal microenvironment: Impact on endometriosis development.

Endometriosis is an inflammatory disease that is defined as the growth of endometrium-like tissue outside the uterus, commonly on the lining of the pelvic cavity, visceral organs and in the ovaries. It affects around 190 million women of reproductive age worldwide and is associated with chronic pelvic pain and infertility, which greatly impairs health-related life quality. The symptoms of the disease are variable, this combined with a lack of diagnostic biomarkers and necessity of surgical visualisation to confirm disease, the prognosis can take an average timespan of 6-8 years. Accurate non-invasive diagnostic tests and the identification of effective therapeutic targets are essential for disease management. To achieve this, one of the priorities is to define the underlying pathophysiological mechanisms that contribute to endometriosis. Recently, immune dysregulation in the peritoneal cavity has been linked to endometriosis progression. Macrophages account for over 50% of immune cells in the peritoneal fluid and are critical for lesion growth, angiogenesis, innervation and immune regulation. Apart from the secretion of soluble factors like cytokines and chemokines, macrophages can communicate with other cells and prime disease microenvironments, such as the tumour microenvironment, via the secretion of small extracellular vesicles (sEVs). The sEV-mediated intracellular communication pathways between macrophages and other cells within the peritoneal microenvironment in endometriosis remain unclear. Here, we give an overview of peritoneal macrophage (pMΦ) phenotypes in endometriosis and discuss the role of sEVs in the intracellular communication within disease microenvironments and the impact they may have on endometriosis progression.

Ribosome Biogenesis Serves as a Therapeutic Target for Treating Endometriosis and the Associated Complications.

Ribosome biogenesis is a cellular process critical for protein homeostasis during cell growth and multiplication. Our previous study confirmed up-regulation of ribosome biogenesis during endometriosis progression and malignant transition, thus anti-ribosome biogenesis may be effective for treating endometriosis and the associated complications. A mouse model with human endometriosis features was established and treated with three different drugs that can block ribosome biogenesis, including inhibitors against mTOR/PI3K (GSK2126458) and RNA polymerase I (CX5461 and BMH21). The average lesion numbers and disease frequencies were significantly reduced in treated mice as compared to controls treated with vehicle. Flow cytometry analyses confirmed the reduction of small peritoneal macrophage and neutrophil populations with increased large versus small macrophage ratios, suggesting inflammation suppression by drug treatments. Lesions in treated mice also showed lower nerve fiber density which can support the finding of pain-relief by behavioral studies. Our study therefore suggested ribosome biogenesis as a potential therapeutic target for treating endometriosis.

Bioluminescent imaging in induced mouse models of endometriosis reveals differences in four model variations.

Our understanding of the aetiology and pathophysiology of endometriosis remains limited. Disease modelling in the field is problematic as many versions of induced mouse models of endometriosis exist. We integrated bioluminescent imaging of 'lesions' generated using luciferase-expressing donor mice. We compared longitudinal bioluminescence and histology of lesions, sensory behaviour of mice with induced endometriosis and the impact of the gonadotropin-releasing hormone antagonist Cetrorelix on lesion regression and sensory behaviour. Four models of endometriosis were tested. We found that the nature of the donor uterine material was a key determinant of how chronic the lesions were, as well as their cellular composition. The severity of pain-like behaviour also varied across models. Although Cetrorelix significantly reduced lesion bioluminescence in all models, it had varying impacts on pain-like behaviour. Collectively, our results demonstrate key differences in the progression of the 'disease' across different mouse models of endometriosis. We propose that validation and testing in multiple models, each of which may be representative of the different subtypes/heterogeneity observed in women, should become a standard approach to discovery science in the field of endometriosis.

Endometriosis-Associated Macrophages: Origin, Phenotype, and Function.

Endometriosis is a complex, heterogeneous, chronic inflammatory condition impacting ~176 million women worldwide. It is associated with chronic pelvic pain, infertility, and fatigue, and has a substantial impact on health-related quality of life. Endometriosis is defined by the growth of endometrial-like tissue outside the uterus, typically on the lining of the pelvic cavity and ovaries (known as "lesions"). Macrophages are complex cells at the center of this enigmatic condition; they are critical for the growth, development, vascularization, and innervation of lesions as well as generation of pain symptoms. In health, tissue-resident macrophages are seeded during early embryonic life are vital for development and homeostasis of tissues. In the adult, under inflammatory challenge, monocytes are recruited from the blood and differentiate into macrophages in tissues where they fulfill functions, such as fighting infection and repairing wounds. The interplay between tissue-resident and recruited macrophages is now at the forefront of macrophage research due to their differential roles in inflammatory disorders. In some cancers, tumor-associated macrophages (TAMs) are comprised of tissue-resident macrophages and recruited inflammatory monocytes that differentiate into macrophages within the tumor. These macrophages of different origins play differential roles in disease progression. Herein, we review the complexities of macrophage dynamics in health and disease and explore the paradigm that under disease-modified conditions, macrophages that normally maintain homeostasis become modified such that they promote disease. We also interrogate the evidence to support the existence of multiple phenotypic populations and origins of macrophages in endometriosis and how this could be exploited for therapy.

EP2 receptor antagonism reduces peripheral and central hyperalgesia in a preclinical mouse model of endometriosis.

Endometriosis is an incurable gynecological disorder characterized by debilitating pain and the establishment of innervated endometriosis lesions outside the uterus. In a preclinical mouse model of endometriosis we demonstrated overexpression of the PGE2-signaling pathway (including COX-2, EP2, EP4) in endometriosis lesions, dorsal root ganglia (DRG), spinal cord, thalamus and forebrain. TRPV1, a PGE2-regulated channel in nociceptive neurons was also increased in the DRG. These findings support the concept that an amplification process occurs along the pain neuroaxis in endometriosis. We then tested TRPV1, EP2, and EP4 receptor antagonists: The EP2 antagonist was the most efficient analgesic, reducing primary hyperalgesia by 80% and secondary hyperalgesia by 40%. In this study we demonstrate reversible peripheral and central hyperalgesia in mice with induced endometriosis.

Cortisol regulates the paracrine action of macrophages by inducing vasoactive gene expression in endometrial cells.

The human endometrium undergoes inflammation and tissue repair during menstruation. We hypothesized that the local availability of bioactive glucocorticoids plays an important role in immune cell-vascular cell interactions in endometrium during tissue repair at menstruation, acting either directly or indirectly via tissue resident macrophages. We sought to determine whether endometrial macrophages are direct targets for glucocorticoids; whether cortisol-treated macrophages have a paracrine effect on angiogenic gene expression by endometrial endothelial cells; and whether endometrial macrophages express angiogenic factors. Human endometrium (n = 41) was collected with ethical approval and subject consent. Donor peripheral blood monocyte-derived macrophages were treated with estradiol, progesterone, or cortisol. The effect of peripheral blood monocyte-derived macrophage secretory products on the expression of angiogenic RNAs by endothelial cells was examined. Immunofluorescence was used to examine localization in macrophages and other endometrial cell types across the menstrual cycle. Endometrial macrophages express the glucocorticoid receptor. In vitro culture with supernatants from cortisol-treated peripheral blood monocyte-derived macrophages resulted in altered endometrial endothelial cell expression of the angiogenic genes, CXCL2, CXCL8, CTGF, and VEGFC These data highlight the importance of local cortisol in regulating paracrine actions of macrophages in the endometrium. CXCL2 and CXCL8 were detected in endometrial macrophages in situ. The expression of these factors was highest in the endometrium during the menstrual phase, consistent with these factors having a role in endometrial repair. Our data have indicated that activation of macrophages with glucocorticoids might have paracrine effects by increasing angiogenic factor expression by endometrial endothelial cells. This might reflect possible roles for macrophages in endometrial repair of the vascular bed after menstruation.

Elevated peritoneal expression and estrogen regulation of nociceptive ion channels in endometriosis.

CONTEXT: Ovarian suppression is a common treatment for endometriosis-associated pelvic pain. Its exact mechanism of action is poorly understood, although it is assumed to reflect reduced production/action of estrogens. OBJECTIVE: The objective of the study was to measure the expression of mRNAs encoded by nociceptive genes in the peritoneum of women with chronic pelvic pain (CPP) with or without endometriosis and to investigate whether estrogens alter nociceptive gene expression in human sensory neurons. DESIGN: The study was performed using human tissue analysis and cell culture. SETTING: The study was conducted at a university research institute. PATIENTS: Peritoneal biopsies were obtained from women with CPP and endometriosis (n = 12), CPP and no endometriosis (n = 10), and no pain or endometriosis (n = 5). Endometriosis lesions were obtained from women with endometriosis (n = 18). MAIN OUTCOME MEASURES: mRNAs encoding ion channels (P2RX3, SCN9A, SCN11A, TRPA1, TRPV1) and the neurotransmitter TAC1 were measured in human tissue samples and in human embryonic stem cell-derived sensory neurons treated with estrogens. RESULTS: TRPV1, TRPA1, and SCN11A mRNAs were significantly higher in the peritoneum from women with endometriosis (P < .001, P < .01). TRPV1, SCN9A, and TAC1 were elevated in endometriosis lesions (P < .05). P2RX3 mRNA was increased in the peritoneum of women with CPP, with and without endometriosis (P < .05). Incubation of sensory neurons with 17ß-estradiol increased TRPV1 mRNA (P < .01). The estrogen receptor-ß-selective agonist 2,3-bis(4-hydroxy-phenyl)-propionitrile increased concentrations of TRPV1, P2RX3, SCN9A, and TAC1 mRNAs. CONCLUSIONS: Estrogen-dependent expression of TRPV1 in sensory neurons may explain why ovarian suppression can reduce endometriosis-associated pain. Strategies directly targeting ion channels may offer an alternative option for the management of CPP.