Pathophysiology Of Endometriosis Research Articles

Peritoneal environment, cytokines and angiogenesis in the pathophysiology of endometriosis.

Endometriosis, defined by the presence of viable endometrial tissue outside the uterine cavity, is a common condition affecting 2-3% of women of reproductive age. Today, a composite theory of retrograde menstruation with implantation of endometrial fragments in conjunction with peritoneal factors to stimulate cell growth is the most widely accepted explanation. There is substantial evidence that immunological factors and angiogenesis play a decisive role in the pathogenesis of endometriosis. In women with endometriosis, there appears to be an alteration in the function of peritoneal macrophages, natural killer cells and lymphocytes. Furthermore, growth factors and inflammatory mediators in the peritoneal fluid, produced mainly by peritoneal macrophages, are altered in endometriosis, indicating a role for these immune cells and mediators in the pathogenesis of this disease.

Abnormal interleukin-1 receptor type II gene expression in the endometrium of women with endometriosis.

Interleukin 1 (IL-1) is a major proinflammatory cytokine that is believed to play a central role in the pathophysiology of endometriosis. The IL-1 receptor type II (IL-1RII) is known to bind to IL-1 and to inhibit its biological effects. In our previous studies, we showed that human endometrium expresses IL-1RII, and we observed reduced expression of the protein in women with endometriosis. The aim of this study was to investigate IL-1RII mRNA in the endometrial tissue of normal women (n = 26) and of patients with various degrees of endometriosis (n = 53). In situ hybridization showed that IL-1RII mRNA expression was significantly decreased in endometriosis, particularly during the early stages of the disease (stages I and II). This was quite obvious in both glandular and stromal cells, and it was corroborated by reverse transcription-polymerase chain reaction analysis of IL-1RII mRNA in the endometrial tissue of women with (n = 10) and without (n = 8) endometriosis. The reduced levels of IL-1RII mRNA in the endometrium of women suffering from endometriosis reveals a profound defect in IL-1RII gene expression and, consequently, a reduced capability of endometrial tissue to down-regulate IL-1 activity. Defective IL-1RII gene expression during the early stages of endometriosis (stages I and II) may contribute to the etiology of the disease.

Peritoneal environment, cytokines and angiogenesis in the pathophysiology of endometriosis

Endometriosis, defined by the presence of viable endometrial tissue outside the uterine cavity, is a common condition affecting 2-3% of women of reproductive age. Today, a composite theory of retrograde menstruation with implantation of endometrial fragments in conjunction with peritoneal factors to stimulate cell growth is the most widely accepted explanation. There is substantial evidence that immunological factors and angiogenesis play a decisive role in the pathogenesis of endometriosis. In women with endometriosis, there appears to be an alteration in the function of peritoneal macrophages, natural killer cells and lymphocytes. Furthermore, growth factors and inflammatory mediators in the peritoneal fluid, produced mainly by peritoneal macrophages, are altered in endometriosis, indicating a role for these immune cells and mediators in the pathogenesis of this disease.

Immunohistochemical analysis of the angiogenic status in vasculature of endometriosis

Objective: The knowledge of the involvement of the angiogenesis in endometriosis pathophysiology raises the possibility of a new medical management, using anti-angiogenic controls. However, surprisingly, little is know about the angiogenic status of the vasculature in endometriosis. The present study was undertaken to evaluate the angiogenic status of the vasculature in peritoneal and ovarian endometriosis, using an immunohistochemical technique. In addition, we evaluated the effect of preoperative gonadotropin-releasing hormone agonist (GnRHa) treatment on the angiogenic status of ovarian endometriosis.

Eutopic endometrium from women with endometriosis express a novel 41 kDa protease and elevated stromelysin activity: implications into the pathophysiology of endometriosis.

Objective: Endometriosis is an enigmatic disease whose etiology is poorly understood. Matrix metalloproteinases (MMPs) have been localized to endometriotic tissue and are postulated to contribute to the development and progression of the disease. The mechanisms which lead to this elevated MMP expression are unclear but may be inherent to the eutopic endometrium. The objective of the current study was to determine if eutopic endometrium from women with endometriosis exhibits greater MMP activity compared to the endometrium of women without the disease.

CD44s expression is reduced in endometriotic lesions compared to eutopic endometrium in women with endometriosis.

Immunohistochemical expression of the standard form of CD44 (CD44s) was examined in archival formalin-fixed endometriotic and matching eutopic endometrial tissue obtained from 25 patients in proliferative (N = 16) and secretory (N = 9) stages of the cycle. CD44s was expressed in most eutopic endometria and endometriotic tissue. Its expression was significantly higher in secretory than in proliferative phase endometrium. It was low but detectable in 13 of 16 proliferative phase biopsies. The majority of these endometria exhibited both glandular and stromal staining (63%). In the secretory phase, glandular cells exhibited a significantly greater intensity of staining compared to stromal cells. In endometriotic tissue, stromal cell CD44s expression did not differ between tissue types in either stage of the cycle. In contrast, glandular expression in endometriotic tissue during the secretory phase was reduced (p < 0.05) compared to eutopic endometrium. It was absent in 66% of cases and reduced in the remaining cases. Our results indicate a correlation between CD44s expression and secretory differentiation of endometrial glands in the cycle, suggesting hormonal regulation of its expression. This cyclic pattern of CD44s expression was lost in corresponding endometriotic tissue. Reduced expression of CD44s in endometriotic tissue may provide insight into the pathophysiology of endometriosis.

The possible anti-inflammatory role of circulating human leukocyte antigen levels in women with endometriosis after treatment with danazol and leuprorelin acetate depot.

Endometriosis is defined as an inflammatory condition of the female reproductive tract, a state often associated with infertility and miscarriage. Many exogenously administered factors (treatments) control the disease via as yet unknown pathways. Possible candidate molecules involved in these mechanisms could be the serum-soluble human leukocyte antigens (sHIA) that have been detected in a variety of human body fluids and that are associated with several diseases. We here examine how danazol and leuprorelin acetate depot treatments exert their anti-inflammatory action. It is plausible that subtle alterations mediated by these treatments and in relation to sHLA may explain the pathophysiology of endometriosis and provide insights towards new therapeutic protocols. Indirect enzyme-linked immunosorbent assay (ELISA), using specific monoclonal antibodies, determined serum-soluble class-I and class-II HLA levels. ELISA readings from treated women were compared with normal healthy subjects. Serum-soluble class-I and class-II HLA levels are statistically significantly lower (P < 0.001) in women with endometriosis than in the control groups. However, danazol but not leuprorelin acetate depot administration augments soluble HLA class I and class II (P < 0.01 and P < 0.001, respectively) to normal levels during the treatment period, an increase that may account for the anti-inflammatory effect and the remission observed. It is shown that one of the underlying causes of endometriosis may be the lack of both circulating class-I and class-II antigen levels. Danazol administration acts via an induced release of these antigens, whose presence correlates with the degree of the inflammatory alleviation obtained. We thus provide evidence that the inflammatory state of the disease appears to be associated with soluble HLA levels because, 3 months after ceasing therapy, the circulating antigens in the serum return to the same levels that correspond to the pathological condition.

Increased concentrations of soluble tumour necrosis factor receptor (sTNFR) I and II in peritoneal fluid from women with endometriosis.

Tumour necrosis factor alpha (TNFalpha), a proapoptotic cytokine, is known to be present in peritoneal fluid from women with endometriosis. An emerging view is that soluble TNF receptors (sTNFR) can modulate the effects of TNFalpha by acting as TNFalpha antagonists. To assess the relevance of sTNFRs in the pathophysiology of endometriosis, concentrations of sTNFR I, sTNFR II and TNFalpha in peritoneal fluid from women with endometriosis (n = 53) and without endometriosis (n = 40) were measured. Concentrations of both sTNFR I and sTNFR II in peritoneal fluid from women with endometriosis were significantly higher than in peritoneal fluid from women without endometriosis, both in the follicular and the luteal phases. TNFalpha concentrations did not differ in patients with and without endometriosis in both phases. When stratified by the stage of the disease, women with both stages I/II and stages III/IV exhibited significantly higher concentrations of sTNFR I and sTNFR II in peritoneal fluid, compared with women without endometriosis, whereas no appreciable difference in the concentrations was detected between stages I/II and stages III/IV. A significant correlation was found between the concentrations of sTNFR I and sTNFR II; while the correlations between TNFalpha and sTNFR I or sTNFR II, were either not significant or were very weak. Furthermore, mRNA for the membrane-associated TNF receptor type 1 and TNF receptor type 2, both of which convey the effects of TNFalpha, were shown to be expressed in endometriotic tissues as well as eutopic endometrium. Together, these findings suggest a possible involvement of sTNFRs in the pathophysiology of endometriosis.

Epithelial Neutrophil-Activating Protein 78 (ENA-78) Is Elevated in the Peritoneal Fluid of Patients with Endometriosis

Objective: Immune cells are postulated to contribute to the pathophysiology of endometriosis. Chemokines of the α or CXC family have intrinsic angiogenic activity. Moreover, neutrophils themselves can secrete VEGF and may participate in endometrial vascularization (Mueller et al, in press). The current study tested the hypothesis that epithelial neutrophil-activating protein 78 (ENA-78), an α or CXC chemokine, might accumulate differentially in peritoneal fluid (PF) of women with and without endometriosis. Design: Case-control study. Setting: University hospital. Patients: Eighteen women with laparoscopic findings of endometriosis and eight women with no evidence of pelvic pathology. Main outcome measures: Peritoneal ENA-78 concentrations were measured blindly, in duplicate, by a specific quantitative sandwich enzyme immunoassay (sensitivity < 15 pg/mL). ENA-78 concentrations were compared among women with and without endometriosis (X2 and unpaired t-test). Results: Ten of the eighteen patients with endometriosis (56%) had elevated peritoneal ENA-78 values (Median: 65 pg/ml) whereas only one of eight subjects (13%) without pelvic pathology had detectable ENA-78 values (X2: P<0.05). Mean ± SD ENA-78 concentrations were higher in affected (44 ± 40 pg/ml) than control subjects (2.2 ± 3.8 pg/ml, unpaired t-test P<0.05). Conclusions: ENA-78 belongs to the ELR-CXC-chemokine class, known to have angiogenic activity. As angiogenesis occurs only when the balance of local factors promoting vascular growth exceeds those factors inhibiting angiogenesis, ENA-78 may contribute to the pathogenesis of endometriosis by promoting the neovascularization of ectopic endometrial implants. Immunohistochemical and in-situ hybridization studies are in progress to evaluate the cellular source(s) of ENA-78 production in endometriosis. (Supported by a grant of the SSMBS (MDM) and U54-HD37321 (DIL, RNT).

Do soluble cell adhesion molecules play a role in endometriosis?

Endometriosis is a chronic inflammatory disease associated with diverse immunologic disturbances. Cell adhesion molecules are essential for the development of immune and inflammatory reactions. This study was conducted to investigate whether or not serum and peritoneal levels of soluble cell adhesion molecules are altered in women with endometriosis. The study group comprised five women with moderate-to-severe endometriosis. Eight healthy women with a normal diagnostic laparoscopy served as controls. Serum and peritoneal fluid samples from both groups were analyzed for the soluble isoform of intercellular cell adhesion molecule-1 (sICAM-1). vascular cell adhesion molecule-1 (sVCAM-1), endothelial selectin (sES), and platelet selectin (sPS). Serum levels of sICAM-1 were significantly increased in women with endometriosis (median levels: 410.4 ng/mL; range: 233.9 ng/mL 598.4 ng/mL vs. 235.7 ng/mL; range: 187.4 ng/mL -323.7 ng/mL; P = 0.02). Although the levels of sVCAM-1, sES, and sPS in both samples were higher in the study group, the differences did not reach significance. Our results suggest a role of ICAM-1 in the pathophysiology of endometriosis. However. the role of other investigated cell adhesion molecules should be confirmed by further studies.

Estradiol amplifies interleukin-1-induced monocyte chemotactic protein-1 expression by ectopic endometrial cells of women with endometriosis.

Endometriosis, one of the most frequently occurring gynecological disorders, is estrogen dependent and is often associated with immunological changes. These include increased macrophage activation and infiltration into the endometriotic implants themselves as well as the peritoneal cavity where the implants often develop. Despite the critical role estrogens play in the development of endometriosis, the biochemical mechanisms of their action remain unclear. In the present study we report that estradiol (E2) enhances endometriotic cell responsiveness to the proinflammatory cytokine interleukin-1beta by up-regulating interleukin-1-induced monocyte chemotactic protein-1 (MCP-1) expression at the level of both protein secretion and messenger ribonucleic acid (mRNA) synthesis, whereas progesterone had no significant effects. According to mRNA half-life experiments, E2 action does not seem to be due to increased MCP-1 mRNA stability but, rather, to a higher level of transcription, as shown by run-on analysis. Interestingly, immunohistochemical analysis of MCP-1 expression in endometriotic tissue showed intense immunostaining in both epithelial glands and stroma regardless of the menstrual cycle phase, which is consistent with the cell culture data and indicates that MCP-1 expression is not subject to cyclic variation. The findings of the present study for the first time provide evidence that E2 up-regulates, although in an indirect way, the expression of a potent chemotactic and activating factor by ectopic endometrial cells, which may occur locally in the inflammatory site and contribute to peritoneal macrophage recruitment and activation, and reveal a new means of E2 action in the pathophysiology of endometriosis.

Immunohistochemical localization of insulin-like growth factor–binding protein-3 in eutopic and ectopic endometrial tissues

Objective: To evaluate the expression of insulin-like growth factor–binding protein-3 (IGFBP-3) in the eutopic endometrium and in endometriotic lesions. Design: Retrospective immunohistochemical study. Patient(s): Twenty-five normal women and 39 women with endometriosis. Intervention(s): Endometrial and endometriotic tissue biopsies obtained at laparoscopy. Main Outcome Measure(s): Expression of IGFBP-3 assessed by immunohistochemistry. Result(s): In the endometrium, positive immunostaining of IGFBP-3 was observed both in the stroma and the epithelial glands. The intensity of staining in the glands during the secretory phase was significantly higher in women with endometriosis compared with controls ( P=.018). An increased expression of IGFBP-3 over controls was found in stages I and II of the disease ( P=.018), whereas in stages III and IV, the difference between controls and women with endometriosis was not significant ( P=.300). In endometriotic tissues, a much-marked immunostaining of IGFBP-3 was noted in 90% of the glands and 67% of the stroma without apparent differences related to cycle phase. Conclusion(s): These data show intense staining of IGFBP-3 in endometriosis lesions and increased expression of the protein in the endometrium of patients with endometriosis compared to controls. This marked expression of IGFBP-3 could be related to its previous finding in the peritoneal fluid and to its potential involvement in the pathophysiology of endometriosis.

An update on the classification of endometriosis.

The challenge of creating a satisfactory classification of endometriosis remains to be answered. The ability of the current classification schemes to predict pregnancy outcome or to aid in the management of pelvic pain is recognized to be inadequate. Further revisions of the current classification scheme are anticipated as the understanding of how endometriosis contributes to infertility and pelvic pain evolves. In any revision of the classification system, use of empirically derived weights and breakpoints to define disease stages based on outcome data in larger clinical trials should be attempted. It is also possible that additional factors such as CA-125 level or lesion characteristics may be shown to play an important role in prognosis. If so, these will need to be accounted for in the classification scheme. Careful and consistent use of the recommendations of the American Society for Reproductive Medicine classification of endometriosis subcommittee should allow for collection of data for use in further revisions. It is quite possible that a classification scheme that is designed to predict outcome with respect to pregnancy may be totally inadequate in assessing patients who have endometriosis and pelvic pain. Factors found to be important in the assessment of pelvic pain may be different from those involved with the pathophysiology of endometriosis and infertility. The AFS form suggested for use in the management of endometriosis in the presence of pelvic pain allows for recording of variables such as depth of invasion, histology, as well as documenting adjunct investigations and preoperative physical findings. Such prospective data collection and review in large centers may provide a large clinical base from which to derive empirical point scores and breakpoints in a classification scheme.

Physiological and Cytogenetic Characterization of Immortalized Human Endometriotic Cells Containing Episomal Simian Virus 40 DNA

The study of misplaced endometrial cells, which abnormally implant and grow outside the uterine cavity, is of considerable interest for the understanding of the pathophysiology of endometriosis. However, endometriotic cells, particularly epithelial cells, required for primary cell culture are not easily available. We report here the characterization of an endometriotic cell line immortalized after infection of primary endometriotic cell cultures with simian virus 40. Transformed cells express T-antigen, and blot hybridization analysis showed that the viral genome is present as an episome. Cytogenetic analysis revealed a polyploid karyotype with numerical and structural rearrangements involving mainly the same chromosomes (6, 10, 11, 15, and 17). The cell line has been maintained in culture for over 80 passages and was still proliferating without any noticeable change in the biological properties investigated. Transformed endometriotic cells expressed both progesterone and estradiol receptors and were stimulated by these ovarian hormones to secrete monocyte chemotactic protein-1, a factor that may play an important role in the recruitment and activation of peritoneal macrophages. In addition, this response was enhanced in interleukin-1-treated cells. Taken together, these findings support the view that this cell line may be an interesting tool for the study of the pathophysiology of endometriosis.

The genetics of endometriosis

There is mounting evidence that endometriosis is inherited as a complex trait, like diabetes or asthma. This implies there are environmental factors, such as dioxin, that are interacting with multiple genetic susceptibility loci to produce the phenotype. The Oxford Endometriosis Gene (OXEGENE) study, an international collaborative project, seeks to identify the susceptibility loci using linkage analysis; the aim then is to use positional cloning techniques to identify genes that predispose women to the disease. Analysis of the biochemical function of the gene products will lead to a better understanding of the pathophysiology and aetiology of endometriosis. New therapies may be designed based upon knowledge of the gene function and disease associated genetic markers may be used to identify women at high risk of developing the disease.

Levels of Transferrin and Alpha 2-HS Glycoprotein in Women with and without Endometriosis

The study objective was to test the hypothesis that elevated levels of transferrin and alpha 2-HS glycoprotein occur in the peritoneal environment of patients with endometriosis that may lead to the observed autoimmunity to these proteins. We set up a double sandwich enzyme-linked inimunosorbent assay (ELISA) for measuring levels of transferrin and alpha 2-HS glycoprotein in the serum and peritoneal fluid samples from women with (n = 24–60) and without endometriosis (n = 35–49). Serum and peritoneal fluid levels of alpha 2-HS glycoprotein and peritoneal fluid levels of transferrin were significantly elevated in patients with endometriosis, in contrast to the controls. Serum levels of transferrin in patients, however, were significantly less in the patients than in controls. We conclude that transferrin and alpha 2-HS glycoprotein are present at high concentrations in the peritoneal fluids of patients with endometriosis. This may play a significant role in the autoimmune pathophysiology of endometriosis.

Endometriosis: a disease of oxidative stress?

Our central hypothesis proposes that oxidatively damaged red blood cells (RBCs), apoptotic endometrial cells or undigested endometrial tissue may signal the recruitment and activation of mononuclear phagocytes. Women with endometriosis are prone to respond to this stimulus with an inadequate macrophage scavenger receptor response although the secretory response is not impaired. Activated macrophages in the peritoneal cavity generate an oxidative stress, which consists of lipid peroxides, their degradation products, and products formed from their interaction with low-density lipoprotein (LDL) apoprotein and other proteins. The lipoproteins of the peritoneal fluid (interstitial fluid) have been shown to have lower vitamin E levels and to be more readily oxidized than plasma, so peritoneal fluid may actually contribute to the disease process actively rather than as a passive carrier of mediators of inflammation and growth. As a result of such a stress, a sterile, inflammatory reaction with secretion of growth factors, cytokines, and chemokines is generated, which is deleterious especially to successful reproduction. We propose that such a pro-oxidant environment (peritoneal fluid as well as activated macrophages) promotes growth of ectopic endometrium. The data presented in this review are just the beginning of exploring the role of oxidative stress in mediating the pathophysiology of endometriosis. Only by understanding the mechanisms involved in the pathogenesis of endometriosis can we develop the basis for new diagnostic and therapeutic approaches.

Identification of Various Exon-Deleted Progesterone Receptor mRNAs in Human Endometrium and Ovarian Endometriosis

We demonstrated the expression of various exon-deleted progesterone receptor (PR) variant mRNAs in human uterine endometrium and ovarian endometriosis using the reverse transcription-polymerase chain reaction-DNA sequencing analyses. In addition to PR wild-type mRNA, various exon-deleted PR variant mRNAs were identified in all samples analyzed. The sequence of these variants showed a perfect junction between exons surrouding the deletion area. PR wild-type, exon 6-deleted, exon 4-deleted, exon 5, 6-deleted and exon 4, 5, 6-deleted PR variant mRNAs were observed in all samples analyzed. Exon 4, 6-deleted PR mRNA was observed only in ovarian endometriosis. This is the first study to demonstrate the coexpression of various PR exon-deleted variant mRNAs with the wild-type in uterine endometrium and ovarian endometriosis. All resulting variant proteins might indicate functional diversity and modify the progestational action of wild-type PR, and thus be involved in the pathophysiology of ovarian endometriosis.

Decreased apoptosis and sensitivity to macrophage mediated cytolysis of endometrial cells in endometriosis.

Ectopic dissemination of endometrial cells and their subsequent implantation are the mechanisms involved in the development of endometriosis. While the process of dissemination appears to be a phenomenon common to all women, it is unknown what facilitates or prevents ectopic implantation of misplaced endometrial cells. Prior studies by our group and others suggest that cell-mediated immunity in patients with endometriosis is decreased. The present studies evaluated (i) peripheral blood monocyte (PBM) and peritoneal macrophage (PM) mediated cytolysis of autologous eutopic and ectopic endometrial cells and (ii) programmed cell death (apoptosis) in the eutopic and ectopic endometrium. PBM-mediated cytolysis was (mean+/-SD) 23.1+/-13% for the eutopic and 7.8+/-% for the ectopic endometrium (P < 0.004), while the corresponding percentages for PM-mediated cytolysis were 5.4+/-7 and 0.3+/-1 respectively (P < 0.04). This indicates that PBM are much more effective than PM in inducing cytolysis of both eutopic and ectopic endometrium and that ectopic endometrial cells are significantly more resistant to both PBM- and PM-mediated cytolysis. The apoptosis was significantly decreased in the eutopic endometrium of women with endometriosis as compared to fertile controls (0.375+/-0.17 versus 1.57+/-0.3, P < 0.0001). Furthermore, in matched samples apoptosis was significantly lower in the ectopic (0.149+/-0.075) than eutopic (0.375+/-0.17) endometrium (P < 0.001). We conclude from these studies that the decrease in the capacity of monocytes to mediate cytolysis of the misplaced endometrial cells in the peritoneal locations and an increased resistance of these cells to apoptosis are fundamental to the aetiology and/or pathophysiology of endometriosis.

Peritoneal fluid concentrations of interleukin-8 in women with endometriosis: relationship to stage of disease.

There is increasing evidence that immunological mechanisms play a role in the pathogenesis and pathophysiology of endometriosis. It was therefore of interest to study interleukin-8 (IL-8), a chemokine, in the peritoneal fluid and peripheral blood of women undergoing laparoscopic procedures. The presence and concentrations of IL-8 in relation to endometriosis, infertility and abdominal pain were evaluated. Samples of peritoneal fluid (n = 49) and peripheral blood (n = 50) were obtained from 50 consecutive patients undergoing laparoscopic surgery for various gynaecological indications (abdominal pain, infertility, sterilization). IL-8 was present in the peritoneal fluid of most women (87%). The concentration of IL-8 in the peritoneal fluid was higher in women with endometriosis compared to women without (P = 0.02). This difference was more pronounced in early (stage 1) endometriosis (P = 0.001). IL-8 concentrations in the peritoneal fluid were also higher in women with early endometriosis compared to women with later stages of the disease (P = 0.003). Peripheral blood concentrations did not correlate with peritoneal fluid concentrations of IL-8 and/or the presence of endometriosis. We conclude that IL-8 is an important factor that may contribute to the pathogenesis of endometriosis possibly by promoting neovascularization. This information can be a guide in the development of new therapeutic approaches for the treatment of endometriosis.