Inflammation plays an important role in the pathogenesis of endometriosis. Infiltration of peritoneal macrophages and local proinflammatory mediators in the peritoneal microenvironment affect ovarian function and pelvic anatomy leading to the symptoms and signs of endometriosis. The identification of a noninvasive marker for endometriosis will facilitate early diagnosis and treatment of this disease.
Endometriosis is considered a chronic inflammatory disease. It is defined as the presence of endometrial stroma and glands outside the uterine cavity. The prevalence of endometriosis in women of reproductive age is about 10%.Although the exact etiology of endometriosis is still controversial, Sampson’s retrograde menstruation theory is a widely accepted explanation for endometriosis. In order to ectopically implant on the peritoneal cavity, refluxed endometrial tissue processes through attachment, acute inflammation, macrophage infiltration, tissue remodeling, and neovascularization.
Local microenvironment inflammation in endometriosis
Endometriosis is a pelvic inflammatory process with altered immune surveillance in the local peritoneal microenvironment. The endometriosis-associated inflammatory responses are dependent on increased activated macrophages and their secreted cytokines in peritoneal fluid. A local inflammatory microenvironment will sustain the growth and maintenance of endometriosis through endometrial-peritoneal adhesion, invasion, angiogenesis, and proliferation. The inflammation process in endometriosis further causes pelvic pain and infertility, two major symptoms of endometriosis.
Cyclooxygenase-2 in endometriosis
Local proinflammatory mediators, such as interleukin (IL)-1β and tumor necrosis factor (TNF)-α, will activate nuclear factor κB (NF-κB) and hypoxiainducible factor (HIF)-1α signaling pathways, and then increase cyclooxygenase (COX)-2 expression in endometriosis.These cytokines act as autocrine and/or paracrine signals to regulate local immune and inflammatory responses. COX is a rate-limiting enzyme in the biosynthesis of proinflammatory prostaglandins (PGs).
In peritoneal macrophages, COX-2 expression was significantly increased in women with endometriosis, whereas expression of COX-1 was only increased in severe stages of endometriosis.
The proinflammatory cytokines in the peritoneal microenvironment will stimulate the implantation of ectopic endometriotic tissues. Elevation of COX-2 expression in normal and endometriotic stromal cells is regulated by proinflammatory cytokines such as IL-1βor PGE2 . In ectopic endometriotic stromal cells, IL-1β upregulates COX-2 promoter activity (transcriptional regulation), which is mediated via mitogen-activated protein kinase (MAPK)-dependent signaling pathways through binding to the cAMP-responding element site of a COX-2 promoter.
In addition, chicken ovalbumin upstream promoter-transcription factor II (COUP-TFII) regulates a subset of genesinvolved in endometrial stroma cell decidualization and plays a role in controlling the expression of inflammatory cytokines. Downregulation of COUP-TFII in endometriotic stromal cells mediated by proinflammatory cytokines including IL-1β, TNF-α, and transforming growth factor (TGF)-β1 via microRNA-302a results in COX-2 upregulation. These promote an inflammatory microenvironment for the development of endometriosis.
Endometriosis is also considered as an estrogen-dependent inflammatory disease.7 Inflammation and estrogen are lined in a positive feedback cycle which enhances the expression of aromatase, COX-2, and local estrogen production in ectopic endometriotic lesions. Estrogen receptor β (ERβ) in endometriotic tissue is increased and mediates the induction of COX-2 expression by estradiol.A lack of ERβ promoter methylation is associated with increased ERβ activity in endometriotic stromal cells, and hypermethylation of a CpG island at the promoter region silences ERβ gene expression in eutopic endometrial stromal cells. Increased ERβ in endometriotic stromal cells suppressed the expressions of ERα and progesterone receptors, leading to inflammation, progesterone resistance, and deficient inactivation of estradiol in endometriotic tissue. Similar epigeneticchanges in endometriotic tissue are found in the orphan nuclear receptorsteroidogenic factor (SF) 1, which mediates PGE2-dependent induction of estradiol production. In the MCF-7 human mammary adenocarcinoma cell line, hypoxic treatment induces activation of HIF-1α and upregulates inflammatory response genes such as COX2 expression.
Prostaglandins in endometriosis
Elevated PGE2 and PGF2α in peritoneal fluid due to the over-expression of COX-2 may contribute to the presence of symptoms of endometriosis such as dysmenorrhea and infertility.PGE2 is a polypotent eicosanoid that regulates many pathophysiological processes in the development of endometriosis, including cell proliferation, antiapoptosis, immune suppression, and angiogenesis. Due to its inherent nature,endometriotic stromal cells have more potential to migrate and invade than epithelial cells. Several proinflammatory cytokines induce COX-2 expression in peritoneal macrophages and endometriotic stromal cells, leading to increased PGE2 level in peritoneal fluid. Two positive feedback loops, COX-2-PGE2-estrogen in ectopic endometriotic stromal cells and COX-2-PGE2-pro-inflammatory cytokines such as IL-1β, TNF-α, and even PGE2 secreted by peritoneal macrophages will constantly keep an elevated PGE2 concentration in the peritoneal fluid of patients with endometriosis as a self-supported survival system to the growth of endometriosis.
Endometriosis is an estrogen-dependent disease. Two committed steps required for de novo synthesis of estrogen are steroidogenic acute regulatory protein (StAR) and aromatase. PGE2-induced StAR expression, which transports cholesterol across the mitochondrial membrane to the inner mitochondrial leaflet, is restricted in ectopic endometriotic stromal cells.Similar to the mechanism seen in StAR regulation, the induction of aromatase expression by PGE2 occurs only in ectopic endometriotic stromal cells, and is mediated via binding to EP2/EP4 receptor-coupled signalingpathways.Therefore, selective inhibition of EP2/EP4 receptors is considered as potential nonsteroidal therapy for women with endometriosis in human endometriotic cells.
Leptin is a multifunctional hormone with immunoregulatory, proinflammatory, and angiogenic effects and plays an important role in controlling reproductive functions and the development of endometriosis. Leptin is a prognostic factor of ovarian responsiveness after hyperstimulation. Elevated leptin concentrations may modulate embryo quality and be associated with poor outcome of in vitro fertilization/intracytoplasmic sperm injection cycle.
Leptin may modulate the functions of peritoneal macrophages and aberrant gene expression of stromal cells, which might contribute to the pathophysiological process of endometriosis. In addition, leptin significantly enhanced both eutopic and ectopic endometrial stromal cell proliferation. Hypoxic stress is associated with the growth and survival of endometriotic lesions.
Leptin is a cytokine which promotes CD4+ T helper I cell proliferation, macrophage phagocytosis, and the secretion of inflammatory cytokines. Elevated leptin levels in peritoneal fluid may contribute to the pathological process of endometriosis through the activation of peritoneal macrophages. It affects the presence of symptoms, such as menstrual pain, of patients with endometriosis.
NF-κB in endometriosis
NF-κB, a proinflammatory transcription factor, plays an important role in both physiological immunity and pathological inflammation. Peritoneal oxidative stress, secondary to iron overload in the peritoneal cavity during retrograde menstruation, may impair cellular function by increasing proinflammatory gene expression through the regulation of NF-κB activation.The activation of NF-κB involves macrophage migration inhibitory factor (MIF) gene expression in ectopic endometrial cells in response to IL-1β. Therefore, various inflammatory factors activate NF-κB, and NF-κB further stimulates the synthesis of proinflammatory cytokines to form an autoregulatory loop. Constitutive activation of NF-κB, involving p65- and p50-containing dimmers, in endometriotic lesions and peritoneal macrophages from patients with endometriosis promotes inflammation, invasion, angiogenesis, and cell proliferation, and inhibits apoptosis of endometriotic cells.The suppressed progesterone receptor (PR) and increased COX-2 levels is contributed to progesterone resistance and inflammation. PR-A and PR-B in the uterus play an anti-inflammatory role that antagonizes NF-κB activation and COX-2 expression.PR levels, particularly the PR-B isoform, are significantly decreased in endometriotic stromal cells, which is associated with a high ERβ-to-ERα ratio.This aberrant increased ERβ-to-ERα ratio in endometriotic stromal cells is also regulated by hypoxia, which may be associated with elevated NF-κB activation. Hypoxia-induced HIF-1α expression is blocked by NF-κB inhibition under hypoxic conditions at the translational level in cancer cells.The alteration of physiologic cyclic NF-κB-p65 subunit activation pattern in women with endometriosis is concurrent with progesterone resistance that could affect the implantation window and lead to infertility.
Peritoneal proinflammatory cytokines and chemokines in endometriosis
The levels of cytokines and chemokines, such as IL-1β, TNF-α, IL-6, and IL-8 secreted by peritoneal macrophages and ectopic endometriotic lesions were abnormally increased in peritoneal fluid. These elevated peritoneal factors induce COX-2 expression and trigger PGE2 production, which forms a positive feedback loop to enhance the development of endometriosis. IL-1β is a major proinflammatory cytokine that is overproduced by endometriosis-derived peritoneal macrophages and is elevated in the peritoneal fluid of patients with endometriosis. Especially, IL-1β upregulates both COX-2 mRNA stability and promoter activity in ectopic endometriotic stromal cells. Similarly, IL-1β-induced COX-2 expression upregulates cell migrationand the invasion ability of endometrioma-derived ectopic endometrial mesenchymal stem cells.
TNF-α mediates inflammation by inducing inflammatory mediators IL-6, IL-8, granulocyte macrophage–colony-stimulating factor, and monocyte chemoattractant protein (MCP)-1 secretion in the endometrium.TNF-α can enhance endometrial epithelial cell proliferation in the ectopic environment of women with endometriosis. TNF-α-induced COX-2 overexpression in the eutopic endometrium of women with endometriosis is through NF-κB activation, which may play a critical role in the pathophysiology of endometriosis formation. Prolonged stimulation with TNF-α will induce partial methylation in the promoter region of PR-B with concomitant reduction of PR-B expression in immortalized epithelial-like endometriotic cells that is contributed to progesterone resistance
The activation of protease-activated receptor 2 (PAR2), as an important mediator of inflammation activated by enzymes from neutrophils and mast cells, stimulates proliferation and IL-6 and IL-8 secretion of endometriotic stromal cells through the involvement of MAPKs.IL-6 expression is highly associated with angiogenesis and migration. Inflammatory stimuli, IL-1β and TNF-α, first induce activin-A protein expression in endometrioma stromal cells. It further increases the expression of IL-6 and PAR-2 mRNA expression, and enhances the proliferation of endometrioma stromal cells. Additionally, TGF-β1 directly increases the expression of PAR-2 and increases IL-6 secretion in endometriotic stromal cells. IL-8 induces chemotaxis of neutrophils and is also a potent angiogenic agent. IL-8 may act as an autocrine growth factor in the endometrium promoting the vicious circle of endometrial cell attachment, cell growth, and further self-stimulation in the pathogenesis of endometriosis. In contrast to higher levels of leptin in early stages of endometriosis, elevated peritoneal fluid IL-8 levels in women with endometriosis is correlated with the severity of the disease. Sex steroidsstimulate the chemokine IL-8 expression in endometrial endothelial cellsfrom women with endometriosis. IL-8 increases endometrial stromal cell metalloproteinase activity and invasive capability that degrade extracellular matrix to help the endometrial cells invade the peritoneum and to develop an endometriotic lesion.
Macrophage-derived factors, such as migration inhibitory factor (MIF) and MCP-1, can promote angiogenesis in the development of endometriosis.MIF, with its potent proinflammatory and angiogenic properties, is markedly elevated in lesions from the early stage of endometriosis, particularly in endometrial implants with highly vascularized, active forms of the disease.The overproduction of IL-1β in endometriosis upregulates the expression of MIF in endometrial stromal cells and ectopic endometrial cells via NF-κB nuclear translocation that shows the immunomodulatory properties of MIF in endometriosis-associated inflammation, ectopic cell growth, angiogenesis, and tissue remodelling. MIF acts directly on ectopic endometrial cells to stimulate the synthesis of COX-2 and the production of PGE2 that triggers the coordinate activation of multiple enzymes in the PGE2 biosynthesis pathway.MIF, interacting with ectopic endometrial cells, further markedly upregulates the secretion of major angiogenic factors including vascular endothelial growth factor (VEGF), IL-8, and MCP-1 expression via CD44, CD74, and MAPK signaling pathways.
The peritoneal fluid level of MCP-1 is positively correlated with leptin level, especially in the early stages of endometriosis, which may play a role in the pathogenesis of endometriosis-associated infertility. The levels of VEGF are significantly increased in the endometrial stromal cells of women with endometriosis in the presence of MCP-1 chemokine and/or estradiol.In addition, the stimulation of MCP-1 expression by sex hormones in the endometrial endothelial cells in women with endometriosis may involve recruiting mononuclear cells and contributing to the inflammatory aspect of endometriosis.
Inflammatory factors play an important role in the pathophysiology of endometriosis, which will interact locally and systemically to affect symptomsand may be biomarkers for diagnosis or the target of treatment. Although treatment with COX inhibitors can improve the symptoms and signs of women with endometriosis, short half-life and unfavorable side effects affect the compliance of the treatment.