Peritoneal Fluid Of Endometriosis Patients Research Articles

Local cytokine production and reactive oxygen species in the peritoneal fluid of endometriosis patients: prospective controlled study.

Objective: The pathogenesis of endometriosis and its association with reproductive failure is not well understood. Not only is peritoneal fluid (PF) in close proximity to endometriotic lesions, but it is also the environment in which early reproductive events take place. Immunological factors may play a role in determining the occurrence of endometriosis as well as its heterogeneous symptoms. Reactive oxygen species (ROS) have been detected in many reproductive tissues. The objective of this study was to evaluate the role of PF interleukins and ROS in the pathogenesis of endometriosis. We also examined the correlation of PF interleukins with its ROS levels.

Nuclear Peroxisome Proliferator-Activated Receptors α and γ Have Opposing Effects on Monocyte Chemotaxis in Endometriosis

The peroxisome proliferator-activated receptors (PPARs) alpha and gamma are nuclear receptors that play important roles in inflammatory diseases like ulcerative colitis and arthritis. In this study, we examined the possible role of PPARs in macrophage attraction into the peritoneal cavity of patients with endometriosis. We identified PPAR-alpha and -gamma messenger RNA by RT-PCR and protein by immunoblotting of lysates of peritoneal macrophages and monocytic U937 cells. Using immunocytochemistry, we localized PPAR-alpha and -gamma within the nuclei of both cell types. Monocyte chemotactic activity of peritoneal fluid from patients with endometriosis was quantified in Boyden chambers. Migration of U937 cells was increased by WY 14643 and reduced by rosiglitazone. Peritoneal fluid from patients with endometriosis activated U937 cells transiently transfected with a PPAR-alpha/GAL4 luciferase reporter. By contrast, peritoneal fluid did not cause significant activation of PPAR-gamma/GAL4 constructs. The U937 cells transiently transfected with a PPAR response element luciferase reporter showed disease stage-dependent up-regulation when treated with peritoneal fluid from patients with endometriosis. Treatment with peritoneal fluid from healthy controls down-regulated PPAR response element transactivation. We conclude that peritoneal fluid of endometriosis patients contains activators of PPAR-alpha that stimulate macrophage chemotaxis. Inhibitors of PPAR-alpha or activators of PPAR-gamma could be developed for the treatment of inflammation associated with endometriosis.

Mouse embryo toxicity of IL-6 in peritoneal fluids from women with or without endometriosis.

To determine whether there is a factor (or factors) in the peritoneal fluid of endometriosis patients that impairs embryo growth and embryo implantation. Growth and development of two-cell mouse embryos which were cultured in media with peritoneal fluid from women with or without endometriosis and interleukin-1-beta (IL-1beta), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-alpha) levels in conditioned media were measured. The blastocyst rate in the non-endometriosis group was 46.4 +/- 31.1%, and that of the endometriosis group was 54.6 +/- 28.7%. Logistic regression analysis using the criteria of blastocyst development in 454 embryos, showed that the peritoneal fluid from endometriosis could promote (p=0.015) but IL-6 could arrest embryo growth to blastocyst (p=0.025). IL-1beta and TNF-alpha levels had no significant effect on blastocyst formation. Peritoneal fluid from women with endometriosis was not toxic to mouse embryo development. However, IL-6 in the peritoneal fluid deteriorated the growth and development of mouse embryos.

Tumor necrosis factor-alpha promotes proliferation of endometriotic stromal cells by inducing interleukin-8 gene and protein expression.

Endometriosis, a common disease among women of reproductive age, is characterized by the presence of endometrial-like tissue outside the uterus. We and others showed that several cytokine levels, including interleukin-8 (IL-8) and tumor necrosis factor-alpha (TNFalpha), are elevated in the peritoneal fluid of women with endometriosis compared with those in women without endometriosis. We also demonstrated that the addition of IL-8 to the culture medium stimulated the proliferation of cultured endometriotic stromal cells. TNFalpha is a multipotent cytokine that induces IL-8 production in various cell types. Therefore, we hypothesized that TNFalpha may also contribute to the pathogenesis of endometriosis by inducing the production of IL-8. To test this hypothesis, we analyzed the peritoneal fluid concentrations of IL-8 and TNFalpha using enzyme-linked immunosorbent assay (ELISA). We observed a significant correlation between the levels of TNFalpha and IL-8 in the peritoneal fluid of endometriosis patients. We also obtained the endometriotic stromal cells from chocolate cyst linings of the ovary. The expression of the receptors for TNFalpha (TNFR) was examined by RT-PCR. We observed the expression of both TNFR-I and TNFR-II genes in endometriotic stromal cells. The expression of IL-8 gene and protein was analyzed by Northern blot hybridization and enzyme-linked immunosorbent assay, respectively. TNFalpha induced the gene and protein expression of IL-8 in endometriotic stromal cells in a dose-dependent fashion. The addition of TNFalpha promoted the proliferation of the endometriotic stromal cells, and the stimulatory effects of TNFalpha were abolished by adding anti-IL-8 antibody. We demonstrated for the first time that TNFalpha stimulated proliferation of endometriotic stromal cells through induction of IL-8 gene and protein expression. We concluded that the TNFalpha may be one of the essential factors for the pathogenesis of endometriosis.

Secretion of interleukin-6 by human endometriotic cells and regulation by proinflammatory cytokines and sex steroids.

Endometriosis is generally associated with an immunoinflammatory process that takes place in the peritoneal cavity of patients. Interleukin (IL)-6, a multifunctional cytokine involved in numerous immunological and proliferative processes, has been found at high concentrations in the peritoneal fluid of endometriosis patients. The purpose of this study was to investigate the ability of endometriotic cells to produce IL-y and to assess the regulation of its secretion by proinflammatory cytokines and sex steroids. Cultures of human endometriotic cells were exposed to different concentrations of cytokines and sex steroid hormones for varying periods of time. IL-6 secretion was measured using an enzyme-linked immunosorbent assay. Endometriotic cells spontaneously released IL-6 in culture. IL-1 beta and tumour necrosis factor (TNF)-alpha (0.1-100.0 ng/ ml) potentiated IL-y secretion in a time- and dose-dependent manner. Interferon-gamma (0.4-400 ng/ml) induced a dose-related increase in IL-6 secretion and showed a synergistic effect on that secretion in combination with TNF-alpha (10 ng/ ml). Either spontaneous or cytokine-induced IL-6 secretion was inhibited by progesterone (10(-8)-10(-5) M) and danazol (10(-6) M), whereas oestradiol (10(-8)-10(-5) M) had a limited inhibitory effect. The antiprogestin RU486 (10(-8)-10(-4) M) antagonized the inhibitory effects of progesterone and danazol, but showed agonist action when used alone. These findings indicate that endometriotic tissue may actively contribute to the biological changes observed in the peritoneal fluid of endometriosis patients. They also provide new insights into the mechanisms of action of progesterone and those of danazol and RU486 used in the treatment of endometriosis.

Elevated concentration and biologic activity of monocyte chemotactic protein-1 in the peritoneal fluid of patients with endometriosis

To estimate the concentration and the biologic activity of monocyte chemotactic protein-1 (MCP-1) in the peritoneal fluid (PF) of women with and without endometriosis. A case control study was conducted. Gynecology clinic and Laboratories of endocrinology of reproduction and immunology. Women presenting for infertility, pelvic pain, or tubal ligation in which endometriosis was diagnosed at laparoscopy (n = 36) and normal fertile controls presenting for tubal ligation (n = 21). Collection of PF via laparoscopy. Determination of PF concentrations of MCP-1 by an ELISA and evaluation of its monocyte chemotactic activity using a human hystiocytic cell line (U937). RESULTS. The concentration of MCP-1 (median, range of values) was increased in the PF of endometriosis patients (283, 0 to 1,930 pg/mL; conversion factor to SI unit, 0.155) compared with the control group (140, 0 to 435 pg/mL). The most significant elevation of MCP-1 levels was found in the stage II of the disease (371, 200 to 1,930 pg/mL). An increased chemotactic activity for monocytes (mean number of migrating cells/mm2 +/- SD) also was found in stages I (1,460 +/- 312) and II (1,541 +/- 336) of the disease when compared with fertile controls (393 +/- 56). Forty percent to 53% of this activity was inhibited in the presence of an antibody specific to MCP-1. These observations are consistent with previous data indicating increased leukocyte chemotaxis in the PF of patients with endometriosis and suggest that MCP-1 may play a relevant role in the peritoneal inflammatory reaction associated with the disease.

Effect of Peritoneal Fluid From Endometriosis Patients on Endometrial Stromal Cell Proliferation In Vitro

Late proliferative phase endometrial stromal cells grown in short-term culture were used as a model for the stromal component of endometriotic implants. Cells were grown in medium alone and in medium supplemented by 5, 10, and 20% concentrations of the cell-free fractions of peritoneal fluid obtained from patients with and without endometriosis. Nine fluid-sample pairs were matched based on the presence or absence of endometriosis at laparoscopy and the similarity of peritoneal fluid estradiol concentrations. Stromal cell proliferation as reflected by 3H-thymidine incorporation during sequential cell harvests over 72 hours was greater for cells exposed to endometriosis peritoneal fluid than for those exposed to non-endometriosis peritoneal fluid. This reached statistical significance with exposure to peritoneal fluid concentrations of 10 and 20% (P less than .05). A linear dose-response relationship between 3H-thymidine incorporation and peritoneal fluid concentration could be derived only for stromal cells exposed to fluid samples obtained from endometriosis patients (r = 0.51; P less than .001). In addition, proliferation over 72 hours was significantly greater for cells grown in 20% endometriosis peritoneal fluid than for those grown in nutrient medium alone (P less than .001). These data imply that factor(s) secreted into the peritoneal fluid of endometriosis patients may play a role in the proliferation or maintenance of disease implants.